Operating Manual 2019-09-04 Labfors 5 Bench-Top Bioreactor - Version for Microorganisms Dok-ID: 79341 / V.02.01 - Original Infors AG Headoffice, Schweiz Rittergasse 27 CH-4103 Bottmingen T +41 (0)61 425 77 00 F +41 (0)61 425 77 01 info@infors-ht.com service@infors-ht.com Infors GmbH Dachauer Str. 6 D-85254 Einsbach T +49 (0)8135 8333 F +49 (0)8135 8320 infors.de@infors-ht.com Infors UK Ltd The Courtyard Business Centre Dovers Farm, Lonesome Lane, Reigate Surrey, RH2 7QT, UK T +44 (0)1737 22 31 00 F +44 (0)1737 24 72 13 infors.uk@infors-ht.com Infors Sarl 2, rue du Buisson aux Fraises Bâtiment D13 F-91300 Massy T +33 (0)1 69 30 95 04 F +33 (0)1 69 30 95 05 infors.fr@infors-ht.com Infors Benelux BV Markweg 9-A, NL-6883 JL Velp (GLD) P.O. Box 125, NL-6880 AC Velp (GLD) T +31 (0)26 369 31 00 F +31 (0)26 369 31 09 infors.bnl@infors-ht.com Infors USA Inc. 9070 Junction Drive, Suite D Annapolis Junction, MD20701 T +1 301 362 3710 / T +1 855 520 7277 (toll-free USA) F +1 301 362 3570 infors.usa@infors-ht.com Infors Canada 8350 rue Bombardier Anjou, Quebec Canada H1J 1A6 T +1 514 352 5095 F +1 514 352 5610 infors.ca@infors-ht.com Infors Bio-Technology (Beijing) Co., Ltd. Room 505C, Building 106 Lize Zhongyuan Wangjing New Industrial Zone Chaoyang District, Beijing 100102 P.R. of China T +86 10 51652068 F +86 10 64390585 info@infors-ht.com.cn Infors South East Asia 16, 1st Floor, Taman City MY-51200 Kuala Lumpur Malaysia T +603 625 771 81 F +603 625 067 48 info@infors-ht.com.my Infors LATAM Ltda. Rua Dr. Alceu de Campos Conjunto 205 CEP: 04544-000 São Paulo – SP Brasil T +55 (11) 95304-0201 F +55 (11) 98585-5334 Infors.br@infors-ht.com Contact details of our local dealers worldwide can be found on our website. www.infors-ht.com Engineering and production in Switzerland Labfors 5 - Version for Microorganisms Table of Contents 04 September 2019 Page 1 of 216 1 General Information...............................................................9 1.1 About this Manual ..........................................................9 1.2 Explanation of Special Notices ......................................9 1.2.1 Warning Notices .............................................9 1.2.2 Other Notices................................................10 1.3 Equipment Identification (Standard Identification Plate)............................................................................10 1.4 Declaration of Conformity ............................................11 1.5 Customer Service and Services ..................................11 2 Safety and Responsibility ...................................................12 2.1 Intended Use, Incorrect Use and Misuse ....................12 2.2 Qualified Personnel .....................................................13 2.2.1 Provider ........................................................13 2.2.2 User..............................................................13 2.2.3 Operator .......................................................14 2.3 Unauthorised Persons .................................................15 2.4 Responsibility of the Provider ......................................15 2.5 General Hazards..........................................................15 2.5.1 Electrical Current..........................................16 2.5.2 Unauthorised Spare Parts and Accessories 16 2.6 Particular Hazards .......................................................16 2.6.1 Hot Surfaces.................................................17 2.6.2 Dangerous Gases ........................................17 2.6.3 Flammable or Explosive Substances ...........17 2.6.4 Corrosive or Toxic Substances ....................17 2.6.5 Bioactive Substances or Pathogenic Organisms ....................................................18 2.6.6 Overpressure or Vacuum .............................18 2.7 Warning Symbols on the Equipment ...........................18 2.8 Declaration of Decontamination...................................19 3 Setup and Function..............................................................20 3.1 Basic Unit.....................................................................20 3.1.1 Power Switch................................................21 3.1.2 Pumps ..........................................................22 3.1.3 Identification Plate ........................................23 3.1.4 Mains Connection and Equipment Fuses ....23 3.1.5 Water Connections.......................................23 3.1.6 Gas Connections..........................................24 3.1.7 Signal Connections ......................................25 3.1.8 Connections for Single or Double Walled Vessels.........................................................25 Labfors 5 – Version for Microorganisms Table of Contents Page 2 of 216 04 September 2019 3.1.8.1 Connections for Single Walled Vessels......................................... 26 3.1.8.2 Connections for Double Walled Vessels......................................... 27 3.1.9 Gassing Connection (Sparger).....................27 3.1.10 Sensor Connections (Sensor Cables) ..........28 3.1.11 Connections and Valve for Water Flow Regulation of the Exit Gas Cooler ................28 3.2 Operating Panel ...........................................................29 3.2.1 Monitor Keys.................................................30 3.2.2 Operating Panel Connections.......................31 3.3 Culture Vessels............................................................31 3.3.1 Double walled Culture Vessels.....................32 3.3.2 Single walled Culture Vessels ......................33 3.3.3 Top Plate ......................................................34 3.3.4 Ports in the Vessel Top Plate and their Configuration ................................................34 3.3.5 Top Plate DN 115 .........................................35 3.3.6 Top Plate DN 150 .........................................36 3.3.7 Top Plate DN 200 .........................................37 3.4 Temperature Control System.......................................38 3.4.1 Water Temperature Control..........................38 3.4.2 Temperature Control with Heater Pad and Cold Finger ............................................38 3.5 Stirrer ...........................................................................39 3.6 Gassing System...........................................................41 3.6.1 Gassing Strategy ..........................................41 3.6.2 Gasmix System.............................................41 3.6.3 Gas Entry......................................................41 3.7 Exit Gas........................................................................42 3.8 pH Control ....................................................................42 3.8.1 Measurement System...................................43 3.8.2 Calibration.....................................................43 3.8.3 Mounting of the Sensor ................................43 3.9 pO2 Control...................................................................44 3.9.1 Measuring System........................................44 3.9.2 Measurement and Calibration ......................45 3.9.3 Mounting of the Sensor ................................45 3.10 Antifoam Control ..........................................................46 4 Options..................................................................................47 4.1 Pump(s)........................................................................47 4.2 Servomotor (High Torque) ...........................................47 4.3 Level Measurement......................................................49 Labfors 5 - Version for Microorganisms Table of Contents 04 September 2019 Page 3 of 216 4.4 Exit Gas Analysis.........................................................50 4.4.1 Measuring Systems (Gas Sensors) .............50 4.4.2 Connecting the Gas Sensors .......................50 4.4.3 Calibration ....................................................52 4.4.4 Replacing the BlueVary Gas Sensor Cartridge.......................................................52 4.5 Multiplexer (Gas Switching Module) ............................52 4.6 Turbidity Measurement................................................53 4.6.1 Calibrating the Sensor..................................54 4.6.2 Mounting the Sensor ....................................55 4.6.3 Interferences Turbidity Measurement ..........56 4.7 Permissive Measurement ............................................56 4.8 Redox Measurement ...................................................57 4.8.1 Measuring Systems (Sensors) .....................57 4.8.2 Calibration ....................................................58 4.8.3 Mounting the Sensor ....................................58 4.9 pCO2 Measurement .....................................................58 4.10 Pressure Control..........................................................59 4.11 LabCIP, CIP/SIP Unit...................................................62 5 Accessories..........................................................................63 5.1 Base Tray.....................................................................65 5.2 Reagent Bottle and Pump Holder................................65 5.3 Sampling System Super Safe Sampler .......................67 5.4 Sparger ........................................................................69 5.5 Impellers ......................................................................71 5.6 Pocket for Temperature Sensor (Pt100)......................71 5.7 Baffles..........................................................................72 5.8 Lock Nut.......................................................................73 5.9 Blanking Plugs .............................................................74 5.10 Addition Port Adapters.................................................75 5.11 Inoculation Needles .....................................................76 5.12 Septum Collars ............................................................79 5.13 Dip Tubes.....................................................................80 5.14 Clamping Adapters ......................................................83 5.15 Electrode Holders ........................................................84 5.16 Threaded Adapter........................................................85 5.17 Adapter ........................................................................85 5.18 Heater Pad...................................................................86 5.19 Cold Finger ..................................................................87 5.20 Exit Gas Cooler............................................................89 5.21 Reagent Bottles ...........................................................91 5.22 Antifoam Sensor ..........................................................93 Labfors 5 – Version for Microorganisms Table of Contents Page 4 of 216 04 September 2019 5.23 Pump Heads ................................................................93 5.24 Sterile Filters ................................................................94 5.25 O-Rings and Gaskets...................................................95 5.26 Hoses and Accessories ...............................................96 5.27 Inoculation Accessories and Tools ..............................97 6 Transport and Storage.........................................................98 6.1 Transport......................................................................98 6.2 Storage.........................................................................99 7 Installation and Initial Operation ......................................100 7.1 General Location Requirements for Installation.........100 7.2 Minimum Distances....................................................101 7.3 Power Supply.............................................................101 7.4 Water Supply and Return...........................................101 7.5 Gas Supply.................................................................102 7.6 Exit Gas......................................................................103 7.7 Test Run.....................................................................103 7.7.1 Preparation Test Run .................................104 7.7.2 Filling the Water Circuit ..............................106 7.7.3 Stirring ........................................................107 7.7.4 Heating and Adjusting Temperature...........108 7.7.5 Gassing.......................................................108 7.7.6 End of Test .................................................109 8 Before Cultivation ..............................................................110 8.1 Preparing and Autoclaving the Culture Vessel ..........110 8.1.1 Mounting the Glass Vessel in the Holder ...110 8.1.2 Checking Gaskets (O-Rings)......................114 8.1.3 Mounting the Impellers ...............................115 8.1.4 Mounting Dip Tubes and Spargers.............115 8.1.5 Mounting the Baffles...................................117 8.1.6 Moistening/Filling the Culture Vessel .........118 8.1.7 Fitting the Vessel Top Plate........................119 8.1.8 Mounting a Threaded Adapter....................120 8.1.9 Mounting the Blanking Plugs......................121 8.1.10 Mounting Addition Port Adapters................122 8.1.11 Mounting the Pocket for Temperature Sensor (Pt100)............................................123 8.1.12 Preparing an Inoculation Needle ................123 8.1.13 Preparing the Dip Tube/Addition Port Adapter for Inoculation ...............................124 8.1.14 Equipping Port with Septum Collar and Septum for Inoculation................................124 Labfors 5 - Version for Microorganisms Table of Contents 04 September 2019 Page 5 of 216 8.1.15 Mounting and Preparing the Exit Gas Cooler .................................................125 8.1.16 Mounting the Cold Finger...........................127 8.1.17 Checking Lubrication of the Mechanical Seal .........................................127 8.1.18 Preparing the Sensors................................128 8.1.18.1 Calibrating the pH Sensor...........129 8.1.18.2 Mouting a Sensor into a 12 mm Port..............................................129 8.1.18.3 Mounting Sensors with Electrode Holder .........................130 8.1.18.4 Mounting the Antifoam Sensor....132 8.1.19 Preparing the Super Safe Sampler ............135 8.1.20 Mounting the Sparger Hose and the Inlet Air Filter ..............................................136 8.1.21 Preparing the Gassing Hose Line on the Basic Unit .............................................137 8.1.22 Calibrating the Pumps................................138 8.1.23 Preparing the Reagent Bottles, Pumps and Hoses ......................................138 8.1.24 Sterile Hose Connections...........................141 8.1.25 Removing the Pump Heads .......................141 8.1.26 Checklist Before Autoclaving......................142 8.1.27 Autoclaving.................................................143 8.2 Connecting the Culture Vessel and Preparing the Cultivation ..................................................................145 8.2.1 Connecting a Single Walled Vessel ...........145 8.2.2 Connecting a Double Walled Culture Vessel.........................................................146 8.2.3 Mounting the Pump Heads.........................147 8.2.4 Filling the Reagent Hoses ..........................148 8.2.5 Connecting the Gassing.............................149 8.2.6 Connecting the Exit Gas Cooler.................149 8.2.7 Coupling the Motor .....................................150 8.2.8 Filling the Culture Vessel............................150 8.2.9 Inserting the Temperature Sensor (Pt100) into the Pocket...............................151 8.2.10 Connecting the Antifoam Sensor................152 8.2.11 Connecting the pH Sensor .........................152 8.2.12 Connecting the pO2 Sensor........................154 8.2.13 Polarising the pO2 Sensor (METTLER)......155 8.2.14 Calibrating the pO2 Sensor.........................155 8.2.15 Checking the Hoses and Hose Connections................................................155 8.3 Preparing the Medium ...............................................156 Labfors 5 – Version for Microorganisms Table of Contents Page 6 of 216 04 September 2019 8.4 Sampling ....................................................................157 8.5 Inoculation..................................................................160 8.5.1 Inoculation with Inoculation Needle............161 8.5.2 Inoculation with a Syringe...........................162 8.5.3 Inoculation Using Dip Tube / Addition Port Adapter................................................162 8.6 Harvest.......................................................................163 8.7 Emptying the Culture Vessel......................................164 8.8 Emptying the Reagent Hoses ....................................164 8.9 Switching off the Equipment ......................................164 8.10 Autoclaving the Culture Vessel After Cultivation .......165 9 Cleaning and Maintenance................................................167 9.1 Cleaning Agent and Disinfectant................................167 9.2 Cleaning the Culture Vessel - Routine Cleaning .......167 9.3 Removing the Vessel Top Plate and Accessories .....169 9.3.1 Removing the Exit Gas Cooler ...................169 9.3.2 Removing the Sensors ...............................169 9.3.3 Removing Hoses, Filters and Pump Heads .........................................................170 9.3.4 Removing Blanking Plugs...........................171 9.3.5 Removing Threaded Adapters....................171 9.3.6 Removing the Inoculation Needle, Septum Collar and Septum ........................172 9.3.7 Removing Addition Port Adapters ..............172 9.3.8 Removing the Vessel Top Plate .................172 9.3.9 Removing the Pocket for Temperature Sensor (Pt100)............................................174 9.3.10 Removing the Sparger and the Dip Tube(s) .......................................................174 9.3.11 Removing the Baffles .................................174 9.3.12 Removing the Impellers..............................175 9.3.13 Removing the Stirrer Shaft .........................175 9.3.14 Removing the Glass Vessel from the Holder ...................................................176 9.4 Cleaning and Storing Individual Parts........................179 9.5 Cleaning the Sensors.................................................180 9.6 Cleaning the Hoses and Pump Heads.......................181 9.7 Cleaning the Super Safe Sampler .............................181 9.8 Cleaning the Exit Gas Cooler.....................................182 9.9 Cleaning the Basic Unit and Operating Panel ...........182 9.10 Cleaning the Heater Pad............................................182 9.11 Maintenance Plan ......................................................183 9.12 Lubricating the Mechanical Seal................................185 Labfors 5 - Version for Microorganisms Table of Contents 04 September 2019 Page 7 of 216 9.13 Decalcifying the Equipment .......................................186 9.14 Decalcifying Equipment via Vessel Jacket ................187 10 Interferences.......................................................................188 10.1 Interferences Basic Unit and Operating Panel ..........188 10.2 Interferences Drive System .......................................189 10.3 Interferences Temperature Control System ..............189 10.4 Interferences Gassing System...................................191 10.5 Interferences pH-System ...........................................192 10.6 Interferences pO2 System ..........................................193 10.7 Interferences Antifoam or Level Sensor and Antifoam Pump ...................................................194 10.8 Interferences Feed and Pump ...................................195 10.9 Replacing Equipment Fuses......................................196 10.10 Returning for Repair ..................................................196 11 Disassembly and Disposal................................................197 11.1 Disassembly...............................................................197 11.2 Disposal .....................................................................198 12 Technical Data....................................................................199 12.1 Dimensions 1 Unit......................................................199 12.2 Dimensions Master Unit and Satellite Units ..............201 12.3 Dimensions of Culture Vessels in Vessel Holder ......202 12.4 Weights (net)..............................................................203 12.5 Connection Values.....................................................204 12.5.1 Electrical.....................................................204 12.5.2 Water IN .....................................................204 12.5.3 Water OUT .................................................205 12.5.4 Gas(es) IN ..................................................205 12.5.5 Exit Gas......................................................205 12.6 Specifications.............................................................205 12.6.1 Operating Panel .........................................205 12.6.2 Culture Vessel ............................................206 12.6.3 Stirrer..........................................................207 12.6.4 Temperature...............................................208 12.6.5 Gassing ......................................................209 12.6.6 Antifoam .....................................................210 12.6.7 pH...............................................................210 12.6.8 pO2..............................................................211 12.6.9 Pumps ........................................................212 12.6.10 Sterile Filters...............................................213 12.6.11 Heater Pads for Single Walled Vessels .....214 12.7 Operating Conditions .................................................214 Labfors 5 – Version for Microorganisms Table of Contents Page 8 of 216 04 September 2019 12.8 Emissions...................................................................214 12.9 Utilities........................................................................215 12.10 Auxiliary Supplies.......................................................215 13 EC-Declaration of Conformity...........................................216 Labfors 5 - Version for Microorganisms General Information 04 September 2019 Page 9 of 216 1 General Information 1.1 About this Manual This manual enables the safe and efficient handling of the equip- ment. All the information and instructions in this operating manual comply with the current standards, legal regulations, the latest technological and scientific developments and the knowledge gained from the manufacturer’s many years of experience in this field. This operating manual is a component part of the equipment. It must be kept near to the equipment and must be accessible to the operators at all times. The users must read the operating manual thoroughly and fully understand its contents before beginning any work. Adhering to all the safety and operating instructions in this manual is essential to ensure that work is carried out safely. The scope of delivery may differ from the explanations, descriptions and figures in this operating manual due to special designs, additional options specified on ordering and the latest technical/mechanical modifications. This manual contains illustrations to aid general understanding. These may differ from the actual equipment as supplied. 1.2 Explanation of Special Notices 1.2.1 Warning Notices Warning notices in this manual are indicated by a coloured bar and begin with a signal word that signifies the degree of the hazard. DANGER The signal word “DANGER” indicates a dangerous situation that will lead to severe or even fatal injuries if not avoided. Labfors 5 - Version for Microorganisms General Information Page 10 of 216 04 September 2019 WARNING The signal word “WARNING” indicates a potentially dangerous situation that may result in severe or even fatal injuries if not avoided. CAUTION The signal word “CAUTION” indicates a potentially dangerous situation that may result in minor injuries if not avoided. 1.2.2 Other Notices ATTENTION The word “ATTENTION” on a blue bar indicates a situation that may result in significant damage to property if not avoided. INFORMATION Texts located below a grey bar bearing the notice “INFORMATION” provide useful tips and recommendations for ensuring efficient, fault-free operation of the equipment. 1.3 Equipment Identification (Standard Identification Plate) The identification plate is designed to allow clear identification of the equipment. It contains the following information:  Manufacturer name  Designation = Category of equipment  Type = Equipment type (name)  S/N = Serial number  Year = Year of manufacture  Mains = Nominal voltage and frequency  Current = Current consumption  Manufacturer address  CE marking Labfors 5 - Version for Microorganisms General Information 04 September 2019 Page 11 of 216 1.4 Declaration of Conformity The equipment is in compliance with the essential requirements of the following Directives:  Machinery Directive 2006/42/EC  EMC Directive 2014/30/EU The Declaration of Conformity according to EC Machinery Directive 2006/42/EC, annex II 1 A is attached to the operating manual, refer to chapter “EC-Declaration of Conformity”. 1.5 Customer Service and Services Our Customer Service is at your disposal for technical advice and specialist enquiries. For contact information, see page 2. Due to their familiarity with the potential applications of the equipment, the Customer Service team is able to provide information on whether the equipment can be used for a specific application or modified to handle the planned process. Experience of working with the equipment will be published semiregularly on the manufacturer’s website in the form of “application notes”. Furthermore, our colleagues are always interested in new information and experiences resulting from user’s applications for the equipment that may be valuable for the continued development of our products. Labfors 5 - Version for Microorganisms Safety and Responsibility Page 12 of 216 04 September 2019 2 Safety and Responsibility This section describes general considerations relating to user safety that must be taken into account when working with the equipment. In the remaining sections, warning notices are used only to highlight particular hazards directly arising from the actions being described in the section in question. It is essential to read the operating manual carefully – especially this section and the warning notices in the text – and to follow the instructions therein. This section also refers to areas that are the responsibility of the provider due to certain risks arising from particular applications for which the equipment is used deliberately and with full awareness of the associated risks. 2.1 Intended Use, Incorrect Use and Misuse The bench-top bioreactor Labfors 5 from INFORS HT is designed especially for running bio processes with microorganisms or animal cells for research and development in a biotechnology laboratory. The equipment is designed and constructed exclusively for the intended use described above. Intended use also includes following all the instructions in this operating manual, especially those relating to:  The installation site  User qualifications  Correct operation and maintenance  The use of undamaged tubing and glass vessels Any failure to observe the requirements specified in this manual shall be deemed incorrect use. Any use of the equipment outside the scope of the intended use as described above shall be deemed misuse. This also applies to applications for which the equipment is not designed, such as the use or production of explosive gases, which is not permitted because the equipment is not explosion-proof. Labfors 5 - Version for Microorganisms Safety and Responsibility 04 September 2019 Page 13 of 216 For use for special applications not covered by conventional, intended use, the equipment must be modified and certified accordingly by the manufacturer. Any use of the equipment outside of a biotechnology laboratory, i.e. in any environment in which the conditions required for the safety of the users cannot be fulfilled or cannot be fulfilled to their full extent, shall also be deemed misuse. 2.2 Qualified Personnel Due to the complexity of the equipment and the potential risks arising from its operation, the equipment may only be used by qualified, specialist personnel. 2.2.1 Provider The term “provider” applies to all persons who are responsible for making the equipment and the necessary infrastructure available. These persons may also be included in the group of people known as “users”, though this is not always the case. Irrespective of whether a provider is a member of the company’s board of management or a supervisor, they bear a special level of responsibility with regard to the processes and the qualification and safety of the users. 2.2.2 User General The term “user” applies to all persons who come into contact with the equipment in any way and perform work on or with it. This primarily applies to the following activities, which can be performed by the manufacturer’s own specialists or a variety of other persons (it is not always possible to distinguish clearly between the different types of person):  Assembly, installation and commissioning  Definition and preparation of the process  Operation  Troubleshooting and remedying of faults  Maintenance and cleaning (autoclaving, if necessary)  Service work and repairs  Disassembly, disposal and recycling Labfors 5 - Version for Microorganisms Safety and Responsibility Page 14 of 216 04 September 2019 Qualified personnel On account of their specific education, training and – in many cases – experience, the qualified personnel required for this work are able to recognise risks and respond accordingly to potential hazards. The qualified personnel (either internal or external) who cannot be categorised under the separate “operators” group are made up of the following groups of persons:  Electricians (electrical engineers)  Decontamination specialists  Repair specialists  Specialists in disassembly and (environmentally friendly) dis- posal  Recycling specialists 2.2.3 Operator The “operators” are a specific sub-group of users distinguished by the fact that they work with the equipment. They are the true target audience for this operating manual. Qualified technicians Only technicians who have been trained for working in a biotechnology laboratory can be considered for the role of operator. These include:  Process technicians in the fields of biotechnology and chemis- try  Biotechnologists (biotechnicians)  Chemists with a specialisation in biochemistry; chemists in the field of organic chemistry or biochemistry  Life scientists (biologists) with special education in cytology, bacteriology, molecular biology, genetics, etc.  Lab assistants (lab technicians) from various fields In order to be classed as a “sufficiently qualified technician” for the operation of the equipment, the persons in question must have received thorough training and have read and understood the operating manual. The operator must be informed in a training session provided by the provider of the tasks delegated to the operator and the potential risks of improper conduct. Tasks that go beyond the scope of operation under normal conditions may only be performed by the operator if this is specified in the manual and the provider has explicitly entrusted said tasks to the operator. Labfors 5 - Version for Microorganisms Safety and Responsibility 04 September 2019 Page 15 of 216 Technicians in training Persons in this group who are undergoing training or apprenticeships are only permitted to use the equipment under supervision and in accordance with the instructions of a trained and qualified technician. 2.3 Unauthorised Persons The term “unauthorised persons” applies to all persons who can access the work area but are not qualified to use the equipment in accordance with the aforementioned requirements. Unauthorised persons are not permitted to operate the equipment or use it in any other way. 2.4 Responsibility of the Provider The equipment is used for industrial and scientific purposes. As such, the provider of the equipment is individually liable with regard to the legal requirements relating to occupational health and safety in a biotechnology laboratory. In particular:  The provider is responsible for ensuring that the work and environmental regulations applicable in a biotechnology laboratory are observed.  The provider must ensure that the equipment remains in safe and proper working condition throughout its entire term of use.  The provider must ensure that all safety equipment is fully functional and is not disabled.  The provider must ensure that the equipment is only worked on by qualified users, and that said users receive sufficient training.  The provider must ensure that the protective equipment required for working with the equipment is provided and worn.  The provider must ensure that this operating manual remains in the immediate vicinity of the equipment throughout its entire term of use. 2.5 General Hazards This section covers general hazards and residual risks that are always present when using the equipment in accordance with normal, intended use. Labfors 5 - Version for Microorganisms Safety and Responsibility Page 16 of 216 04 September 2019 The following notices are general in nature. As such, with a few exceptions they are not repeated in the remaining sections. 2.5.1 Electrical Current The equipment runs on electrical power. There is an immediate risk of fatal injury if contact is made with live parts. The following points must be observed in order to avoid the risk of fatal injury:  In case of damage to insulation, disconnect the equipment from the mains immediately and arrange for it to be repaired.  Disconnect the equipment from the mains before commencing any work on the electrical system.  Always use qualified electricians for any work on the electrical system.  Keep moisture away from live parts. It may lead to a short cir- cuit. 2.5.2 Unauthorised Spare Parts and Accessories Incorrect or imitated spare parts and accessories as well as spare parts or accessories that have not been authorised by the manufacturer represent a significant safety risk. As such, we recommend procuring all spare parts and accessories from an authorised dealer or directly from the manufacturer. For the contact details of the manufacturer’s representatives, see page 2. 2.6 Particular Hazards This section covers particular hazards and residual risks that may arise when using the equipment for special applications in accordance with normal, intended use. Since the use of the equipment for such applications is deliberate, it is the responsibility of the operators and the provider to ensure that all personnel are protected from potential damage to health. The provider is responsible for ensuring that the appropriate protective equipment for such applications is provided, and that the necessary infrastructure is in place. Labfors 5 - Version for Microorganisms Safety and Responsibility 04 September 2019 Page 17 of 216 2.6.1 Hot Surfaces For processes that are carried out with temperatures over 55 °C, there is a danger of burns on hot surfaces. Since the equipment is intended for applications at high temperatures, it is the responsibility of the users to ensure that they have sufficient protection. The motor gets hot during operation. There is a risk of burns if it is touched. 2.6.2 Dangerous Gases The use or production of dangerous gases i.e. toxic or asphyxiant gases entails a significant health risk, especially in enclosed spaces. In order to prevent high emissions of dangerous gases, the following measures must be taken:  The gas connections on the equipment must be checked before any cultivation processes using dangerous gases are initi- ated.  The gaskets on the equipment must be checked at regular intervals and replaced if necessary.  Siphon off exit gas safely. 2.6.3 Flammable or Explosive Substances The use or production of flammable or explosive substances is not covered under “intended use” of the equipment, as the equipment is not explosion-proof. If the provider intends to use the equipment for such purposes, he must check its suitability for the planned application with the responsible local authorities. 2.6.4 Corrosive or Toxic Substances The use or production of corrosive or toxic substances entails a significant health risk. As such, special measures must be taken to protect the users for such applications. Since the equipment is used deliberately for such applications, it is the responsibility of the users to ensure that they have sufficient protection. Labfors 5 - Version for Microorganisms Safety and Responsibility Page 18 of 216 04 September 2019 2.6.5 Bioactive Substances or Pathogenic Organisms The use or production of bioactive substances, pathogenic organisms or genetically modified cultures entails a significant health risk. As such, special measures must be taken to protect the users for such applications. Since the equipment is used deliberately for such applications, it is the responsibility of the users to ensure that they have sufficient protection. 2.6.6 Overpressure or Vacuum Glass vessels may break or shatter when subjected to overpressure or vacuums. 2.7 Warning Symbols on the Equipment The following warning symbols (stickers) are attached to the equip- ment: Position Motor WARNING Illegible or missing warning symbols on the equipment will lead to the user being exposed to risks that the warning symbols in question were designed to make him or her aware of. It is the provider’s responsibility to ensure that all the stickers with warning symbols on the equipment are always intact. Labfors 5 - Version for Microorganisms Safety and Responsibility 04 September 2019 Page 19 of 216 2.8 Declaration of Decontamination When returning the equipment for repair, disassembly or disposal, it is required for the safety of all parties involved and because of legal provisions that a lawful declaration of decontamination is pre- sent. The following must be observed if this is the case:  The equipment, the component part or accessory must be entirely decontaminated before sending to the manufacturer  The provider is therefore required to completely and truthfully fill out a declaration of decontamination, and have it signed by the person responsible.  The declaration of decontamination must be affixed on the outer packaging in which the equipment is sent back.  These forms can be obtained from the licensed dealer or the manufacturer. See address on page 2. Important notice If the return shipment is not accompanied by a signed and complete declaration of decontamination or it is not affixed to the outer packaging, the shipment will be returned unopened to the sender at their expense (see also T&C). Labfors 5 - Version for Microorganisms Setup and Function Page 20 of 216 04 September 2019 3 Setup and Function 3.1 Basic Unit 1 Holder for operating panel, manometers and pressure reduction valves 2 Power switch 3 Sensor cables 4 Holder for gassing unit(s) 1) and optional pressure control 5 Pumps 6 Base plate 1) depending on the chosen gassing strategy this can be rotameter(s), mass flow controller(s), solenoid valve(s) 1 2 3 4 5 6 Labfors 5 - Version for Microorganisms Setup and Function 04 September 2019 Page 21 of 216 The whole measuring and control technology of the bioreactor is integrated in the basic unit. One operating panel with touchscreen software can be used to control up to six bioreactors (culture vessels) see also chapter “Operating Panel”. This means, one basic unit serves as the master unit and can control up to five more basic units, referred to as satellite units. 3.1.1 Power Switch The power switch, a green rocker switch, is located on t on top of the basic unit. It is labelled with POWER and lights up as soon as the equipment is switched on. In addition to normal switching on and off, the power switch also serves as an emergency switch. INFORMATION In the event of an emergency shutdown via the power switch during a running Batch (process), all settings are saved. After switching on via the power switch, the Batch continues with the same settings as before the emergency shutdown. This is also the case if the Batch is controlled via eve®, the platform software for bioprocesses. Labfors 5 - Version for Microorganisms Setup and Function Page 22 of 216 04 September 2019 3.1.2 Pumps Corrective reagent and feed solutions are added via four peristaltic pumps. The pumps are driven by stepper motors. The drive shafts of the pumps are situated on the upper front side of the console on the basic unit. Direction of rotation is clockwise by default. Rocker switches for manual operation of the pumps are provided above the drive shafts. They are labelled as follows (from left to right):  Acid  Base  AF (Antifoam)  Feed When the basic unit is switched on, pumps can be manually operated via the rocker switches:  Push and hold the rocker switch to the left: The pump drive shaft turns counter clockwise  Push and hold the rocker switch to the right: The pump drive shaft turns clockwise. The autoclavable pump heads are latched onto a mounting plate. The mounting plate is identically as the drive shafts labelled with the pump names. The pump heads together with the mounting plate can easily be plugged onto or pulled off the drive shafts. A transparent, not autoclavable (!) cover plate made of plexiglass (highlighted in red in the picture on the left) can be inserted into the holder of the mounting plate for protection during operation. Labfors 5 - Version for Microorganisms Setup and Function 04 September 2019 Page 23 of 216 3.1.3 Identification Plate The identification plate is located on the rear of the basic equip- ment. The data provided on the identification plate is specified in the main chapter “General Information”, chapter “Equipment Identifica- tion”. 3.1.4 Mains Connection and Equipment Fuses The mains connection is located on the rear side of the basic unit. It is labelled with Mains. The equipment is protected against excessive current consumption by two fuses. The equipment fuses are located directly below the mains connection. The country-specific power cable required for connection to the power supply is included in the scope of delivery. If the power cable is defective, replace it with a power cable of the same type. Before connecting the equipment, make sure that the voltage values of the equipment correspond to the local mains voltage. The mains connection must be easily accessible at all times so that the equipment can be disconnected from the power supply quickly in case of an emergency. 3.1.5 Water Connections The two hose nozzles for connection of water inlet and outlet of the basic unit are situated on the rear side at the bottom of the console. They are labelled with corresponding symbols:  Left: Water outlet  Right: Water inlet Labfors 5 - Version for Microorganisms Setup and Function Page 24 of 216 04 September 2019 3.1.6 Gas Connections The connection(s) for gas is/are located at the top on the back of the basic unit. Each connection is labelled accordingly, e.g. Air IN etc. A pressure reduction valve is installed after each connection. The factory settings of the pressure reduction valves must not be changed! Manometers are installed after each pressure reduction valve for pressure indication. They are labelled accordingly, e.g. Air etc. Labfors 5 - Version for Microorganisms Setup and Function 04 September 2019 Page 25 of 216 3.1.7 Signal Connections The following signal connectors are situated above the mains socket (from top to bottom): 1 25 pin Dsub Multi I/O: for connecting analogue and digital input and output signals (0/4..20 mA) 2 9 pin RS232: for connecting a balance or a switchbox with up to 7 balances 3 2 x iDDC bus: for connecting the touch screen operating unit and one or two satellites 3.1.8 Connections for Single or Double Walled Vessels Depending on the present temperature control system, different connectors for either the single or double walled vessels are located on the basic unit. 3 2 1 Labfors 5 - Version for Microorganisms Setup and Function Page 26 of 216 04 September 2019 3.1.8.1 Connections for Single Walled Vessels The socket for cable connection of the electrical heater pad is labelled and the water connectors for the cold finger are labelled with symbols. 1 Mains connection heater pad Heating mat 2 Water inlet cold finger 3 Water outlet cold finger 1 2 3 Labfors 5 - Version for Microorganisms Setup and Function 04 September 2019 Page 27 of 216 3.1.8.2 Connections for Double Walled Vessels The water connectors for the vessel jacket are labelled with corresponding symbols. 1 Water overflow vessel jacket 2 Water inlet vessel jacket 3 Water outlet vessel jacket 3.1.9 Gassing Connection (Sparger) The hose nozzle for connection of the gassing hose (sparger) is located below the holder for gassing unit(s) 1) on the right side of the basic unit. 1) depending on the chosen gassing strategy, this can be rotameter(s), mass flow controller(s), solenoid valve(s). 1 2 3 Labfors 5 - Version for Microorganisms Setup and Function Page 28 of 216 04 September 2019 3.1.10 Sensor Connections (Sensor Cables) The basic unit is equipped and configured by default for measurement of temperature, pH, pO2 and antifoam. This means, the temperature sensor (Pt100) and the cables for connecting these sensors are always present. The appropriate sensors for pH, pO2 and antifoam are separately available, they are not included in the standard package. From left to right: pH / Antifoam / Temperature (Pt100) / pO2. INFORMATION Depending on the chosen variant, the measuring system for pH and pO2 is equipped and configured either for analogue sensors from the manufacturer METTLER or digital sensors from the manufacturer HAMILTON. 3.1.11 Connections and Valve for Water Flow Regulation of the Exit Gas Cooler The water connections for the (optional!) exit gas cooler and the valve for the water flow regulation are located on the left front side of the basic unit. The connectors are closed with plugs and the valve is covered with a cap on delivery. Connections and valve are labelled with symbols. 1 Water flow regulation 2 Water outlet exit gas cooler 3 Water inlet exit gas cooler 2 3 1 Labfors 5 - Version for Microorganisms Setup and Function 04 September 2019 Page 29 of 216 The valve is factory adjusted, If needed, water flow can be manually adjusted here.  Open: turn it counter clockwise  Close: turn it clockwise A counter nut is provided to lock the valve in its desired position. 3.2 Operating Panel The operating panel has a 12" colour-touch screen with protection IP66. Up to six bioreactors (= culture vessels) can be individually or in parallel controlled by one operating panel. A detailed description of the touch screen software can be found in the separate manual. Labfors 5 - Version for Microorganisms Setup and Function Page 30 of 216 04 September 2019 3.2.1 Monitor Keys Four monitor keys are situated on the upper left side of the touch screen operating panel. 1 ON/OFF key 2 DECREASE brightness key: to set the display illumination darker 3 INCREASE brightness key: to set the display illumination brighter 4 DISPLAY key: to switch the display on/off Special details about the ON/OFF key The touch screen operating panel is switched on and off at the power switch on the basic unit. Therefore separate switching on at the ON/OFF key is not necessary. The symbol on the key is illuminated when the operating panel is switched on. INFORMATION The ON/OFF key is locked when a bioreactor (cultivation) is running. A dialogue box with the appropriate message appears when pressing the ON/OFF key by mistake during cultivation. When pressing the ON/OFF key when the bioreactor is stopped, a dialogue box for shutting down the system appears. 2 3 4 1 Labfors 5 - Version for Microorganisms Setup and Function 04 September 2019 Page 31 of 216 3.2.2 Operating Panel Connections Six connectors labelled with different symbols are situated on the rear side of the operating panel. 1 USB2.0 x 2: for backups and service purposes 1) 2 USB2.0 x 2: (Reserve) 3 Ethernet: for Ethernet cable 1) to connect with a network 4 COM2 (Reserve) 5 COM1: for iDDC bus cable 1) (display cable) 6 DC: for power supply cable 1) 1) Cable supplied with equipment 3.3 Culture Vessels Culture vessels consist of the glass vessel, the top plate with standard mounting parts and handholds and the vessel holder with handholds. The vessels are either made of double or single walled borosilicate glass, depending on the temperature control system used. For details see the chapter “Temperature Control System”. 1 2 3 4 5 6 Labfors 5 - Version for Microorganisms Setup and Function Page 32 of 216 04 September 2019 3.3.1 Double walled Culture Vessels 1 Handhold top plate 2 Handhold vessel holder 3 Water overflow vessel jacket 4 Water outlet vessel jacket 5 Glass vessel 6 Water inlet vessel jacket 7 Vessel holder 8 Top plate The picture above shows a double walled culture vessel with a total volume of 3.6 L and DN (diameter nominal = inner diameter) of 150 mm. Double walled vessels are readily equipped with silicone hoses with rapid couplings for connection to the basic unit ex-factory. 8 6 5 7 1 2 3 4 Labfors 5 - Version for Microorganisms Setup and Function 04 September 2019 Page 33 of 216 3.3.2 Single walled Culture Vessels 1 Handhold top plate 2 Handhold vessel holder 3 Glass vessel 4 Vessel holder 5 Top plate The picture above shows a single walled culture vessel with a total volume of 3.6 L and DN (diameter nominal = inner diameter) of 115 mm. Single walled culture vessels need an electrical heater pad and a cold finger for temperature control. For details see the chapter “Temperature Control System”. 5 3 4 1 2 Labfors 5 - Version for Microorganisms Setup and Function Page 34 of 216 04 September 2019 3.3.3 Top Plate Four knurled nuts are used to fixate the top plate to the glass vessel and the vessel holder. Drive hub and motor coupling are located in the middle of the vessel top plate. 3.3.4 Ports in the Vessel Top Plate and their Configuration The vessel top plate has different ports of different sizes to mount the different components such as sparger, blanking plugs, sensors etc. The number of ports in the top plate and its configuration depends on the diameter nominal (= inner diameter) of the culture vessel. Labfors 5 - Version for Microorganisms Setup and Function 04 September 2019 Page 35 of 216 3.3.5 Top Plate DN 115 1 Ground connector antifoam sensor 2 Ø 12 mm Pg13.5: Exit gas cooler 3 Ø 12 mm / Pg13.5: Sparger 4 Ø 19 mm: Addition port adapter, quadruple (for reagents & feed) 5 Ø 12 mm / Pg13.5: pO2 sensor 6 Ø 12 mm / Pg13.5: pH sensor 7 Ø 12 mm / Pg13.5: Inoculation needle 8 Ø 12 mm / Pg13.5: Dip tube for sampling 9 Ø 19 mm: Spare 10 Ø 10 mm: Antifoam sensor 11 Ø 10 mm: Temperature sensor (Pt100) 12 Fixation of baffle 12 11 10 9 8 7 6 5 4 3 2 1 Labfors 5 - Version for Microorganisms Setup and Function Page 36 of 216 04 September 2019 3.3.6 Top Plate DN 150 1 Ground connector antifoam sensor 2 Ø 19 mm: Spare 3 Ø 10 mm: Temperature sensor (Pt100) 4 Ø 10 mm: Antifoam sensor 5 Ø 19 mm: addition port adapter, quadruple (reagents & feed) 6 Ø 12 mm / Pg13.5: Sparger 7 Ø 12 mm / Pg13,5: Spare 8 Ø 12 mm / Pg13.5: Dip tube for sampling 9 Ø 19 mm: Inoculation needle 10 Ø 12 mm / Pg13.5: pH sensor 11 Ø 12 mm / Pg13.5: pO2 sensor 12 Ø 12 mm / Pg13.5: Exit gas cooler 13 Fixation of baffle (bact. cult.) 13 12 11 10 9 8 7 6 5 4 3 2 1 Labfors 5 - Version for Microorganisms Setup and Function 04 September 2019 Page 37 of 216 3.3.7 Top Plate DN 200 1 Ø 19 mm: Spare 2 Ø 10 mm: Temperature sensor (Pt100) 3 Ø 10 mm: Antifoam sensor 4 Ø 19 mm: Spare 5 Ø 12 mm / Pg13.5: Sparger 6 Ø 12 mm / Pg13.5: Dip tube for sampling 7 Ø 19 mm: Inoculation 8 Ø 19 mm: Spare or cold finger 9 Ø 19 mm: Spare 10 Ø 19 mm: addition port adapter, quadruple (corrective reagent & feed) 11 Ø 12 mm / Pg13.5: pH sensor 12 Ø 12 mm / Pg13.5: pO2 sensor 13 Ø 12 mm / Pg13.5: Exit gas cooler 14 Ground connector antifoam sensor 7 14 6 5 3 13 12 11 10 9 8 1 2 4 Labfors 5 - Version for Microorganisms Setup and Function Page 38 of 216 04 September 2019 3.4 Temperature Control System The following two temperature control systems are available:  Water temperature control for double walled vessels  Temperature control with electrical heater pad and cold finger for single walled vessels. Measurement of the temperature in the culture vessel takes place by a platinum resistance sensor (Pt100) for both systems. Water flow for either the circuit in the double walled vessel or in the cold finger is regulated via solenoid valve. 3.4.1 Water Temperature Control The water temperature control takes place in the vessel jacket, which is connected to the basic unit. The water is heated by the means of an electrical heating element in the basic unit. Cold water is fed into the circuit for cooling. The overflow in this open system prevents a dangerous overpressure in the circuit and allows pressure equalisation of the vessel jacket during sterilisation in the au- toclave 3.4.2 Temperature Control with Heater Pad and Cold Finger An electrical heater pad is put around the vessel and connected to the basic unit for heating. Labfors 5 - Version for Microorganisms Setup and Function 04 September 2019 Page 39 of 216 The water cooling is effected via a cold finger, which is connected to the basic unit and mounted into a port in the vessel top plate. Appropriate hoses with rapid couplings for connection to the basic unit are included. For further details see the main chapter “Accessories”, chapter “Cold Finger”. 3.5 Stirrer The stirrer shaft is rotated counter clockwise (top view) by a top drive. 1 Drive hub 2 Stirrer shaft 3 Mechanical seal The stirrer shaft is screwed on the drive hub in the vessel top plate and is sealed by a single mechanical seal. ATTENTION Manipulation on the mechanical seal may lead to its damage! 1 2 3 Labfors 5 - Version for Microorganisms Setup and Function Page 40 of 216 04 September 2019 The mechanical seal must be lubricated at any time. For this, two nozzles fitted with a piece of silicone hose are located on the drive hub. For details see chapter “Lubricating the Mechanical Seal” in the main chapter “Cleaning & Maintenance”. A brushless DC motor with mechanical coupling is used as a standard. To couple the motor, it is simply plugged onto the drive hub on the vessel top plate. Two Rushton impellers with six blades are provided as a standard for bacterial cultures. They are fixed by the means of grub screws to the shaft. Labfors 5 - Version for Microorganisms Setup and Function 04 September 2019 Page 41 of 216 3.6 Gassing System The basic unit is equipped with the appropriate gassing units (rotameter, solenoid valves, mass flow controllers and configured according to the chosen gassing strategy. The following gases can be used:  Air  Oxygen (O2)  Nitrogen (N2) 3.6.1 Gassing Strategy The following variants are available: Basic  Manual flow control via rotameter  Gasmix via solenoid valves Standard  Gas flow control via one electronic mass flow controller  Gasmix via solenoid valves High End Gas flow control and Gasmix via electronic mass flow controllers, 1 piece per gas. 3.6.2 Gasmix System The gas mixture takes place before entry into the culture vessel. The composition of the gas mixture is set and controlled in the touch screen software. A detailed description of the touch screen software can be found in the separate manual. 3.6.3 Gas Entry The gas or the gas mixture is led via a silicone hose from the gassing connection on the basic unit via sparger directly into the medium in the culture vessel. For details about the sparger see the main chapter „Accessories“, chapter „Sparger“. Labfors 5 - Version for Microorganisms Setup and Function Page 42 of 216 04 September 2019 3.7 Exit Gas Pressure in the culture vessel may be increased due to temperature increase or gas production also without actively gassing the culture. For this reason, installing an exit gas line to the vessel is mandatory for every cultivation process. Deviating the exit gas without exit gas cooler If no exit gas cooler is used, the exit gas can be deviated via addition port adapter or inoculation needle fitted with an exit gas filter. However, using this method may lead to blocking the exit gas filter due to humidity in the exit gas. For this reason, the use of an exit gas cooler is strongly recommended. The exit gas cooler is separately available, it is NOT included in the standard package. Deviating the exit gas via exit gas cooler The exit gas cooler dries the exit gas by condensation so that humidity cannot block the exit filter. This prevents from liquid loss in the culture medium at the same time. INFORMATION If strong build-up of foam is expected, a foam trap i.e. a bottle containing antifoam agent can be installed before the exit gas filter as an additional safety precaution. For details see main chapter “Accessories”, chapter “Exit Gas Cooler”. 3.8 pH Control pH in the medium is measured by the pH sensor and controlled by addition of reagents (acid, base). Addition of acid and base takes place via the two peristaltic pumps Acid and Base. Reagent bottles are filled with acid and base which are connected to an/the addition port adapter(s) in the vessel top plate and the two pumps by silicone hoses. Labfors 5 - Version for Microorganisms Setup and Function 04 September 2019 Page 43 of 216 3.8.1 Measurement System The measurement system for pH is either equipped and configured for analogue sensors from the manufacturer METTLER or digital sensors from the manufacturer HAMILTON. The pH sensors are separately available, they are NOT included in the standard packages. Variant METTLER  Traditional pH sensor (potential measurement against refer- ence)  Type: 405-DPAS-SC-K8S  Manufacturer: METTLER TOLEDO Variant HAMILTON  Traditional pH sensor (potential measurement against reference) with integrated electronics  Type: Easyferm Plus ARC  Manufacturer: HAMILTON HAMILTON pH sensors are preconfigured before dispatch by the equipment manufacturer INFORS HT. Replacement sensors of this type must be reconfigured before use! Details on technical data, use and maintenance of the pH sensors can be found in the separate documentation of the sensor manufacturer. Read and follow instructions stated in there. 3.8.2 Calibration As a general rule: Calibration of a pH sensor always must be carried out BEFORE sterilisation. This is executed on the touch screen operating unit. For details refer to the separate operating manual of the touch screen software. 3.8.3 Mounting of the Sensor pH sensors are mounted into a 12 mm / Pg13.5 port by the means of an electrode holder. For details see main chapter “Accessories”, chapter “Electrode Holder”. Labfors 5 - Version for Microorganisms Setup and Function Page 44 of 216 04 September 2019 3.9 pO2 Control The oxygen saturation of the (culture) medium is measured by the pO2 sensor, and can be adjusted as follows: Increasing the pO2 The content of the oxygen dissolved in the medium (pO2) can be increased using the following methods:  Increasing the stirrer speed  Increasing the gas volume flow rate (air and/or oxygen)  Increasing the oxygen content in the Gasmix. These approaches can also be combined. pO2 reduction In anaerobic processes, the vessel can be gassed using nitrogen. This displaces the oxygen dissolved in the medium. For details about pO2 control refer to the separate operating manual of the touch screen software. 3.9.1 Measuring System The measuring system for pO2 is either equipped and configured for analogue sensors from the manufacturer METTLER or digital sensors from the manufacturer HAMILTON. The pO2 sensors are separately available, they are NOT included in the standard packages. Variant METTLER  Traditional amperometric/polarographic pO2 sensor  Type: InPro 6820/25/080  Manufacturer: METTLER TOLEDO. Polarographic pO2 sensors must be polarised at initial operation or after disconnection from the voltage source. This means that the pO2 sensor must be polarised after sterilisation in the autoclave. For details refer to the separate documentation from the manufac- turer. Labfors 5 - Version for Microorganisms Setup and Function 04 September 2019 Page 45 of 216 ariant HAMILTON  pO2 sensor with integrated optical electronics  Type: Visiferm DO ARC  Manufacturer: HAMILTON HAMILTON pO2 sensors are preconfigured before dispatch by the equipment manufacturer INFORS HT. Replacement sensors of this type must be reconfigured before use! Details on technical data, use and maintenance of the pO2 sensors can be found in the separate documentation of the sensor manufacturer. Read and follow instructions stated in there. 3.9.2 Measurement and Calibration Generally speaking, the following applies: Unlike measurements such as pH, which are calibrated to absolute measurement values, the oxygen measurement is always calibrated to a relative reference point. For this purpose, the calibration is set to 100 % relative oxygen saturation, usually with air at max. stirring speed and maximum gas flow rate. The actual concentration of dissolved oxygen in mmol/L may therefore vary at 100 % saturation, depending on the process. INFORMATION Calibration is always carried out AFTER autoclaving. This is carried out on the touch screen operating unit. Depending on the specifications defined by the user, the pO2 sensor will be calibrated either before the vessel is filled with medium or afterwards, in the prepared medium. For details about calibration refer to the separate operating manual of the touch screen software. 3.9.3 Mounting of the Sensor pO2 sensors are mounted into a 12 mm / Pg13.5 port by the means of an electrode holder. For details see main chapter “Accessories”, chapter “Electrode Holder”. Labfors 5 - Version for Microorganisms Setup and Function Page 46 of 216 04 September 2019 3.10 Antifoam Control Foam hinders the exchange of gas between the medium and the gas phase in the head space. The exit gas filter can become clogged with foam, which causes a pressure build-up in the vessel. This can be prevented by adding antifoam agent. The antifoam agent is kept in a reagent bottle that is connected to the antifoam sensor and the antifoam pump via a hose. The sensor also acts as a dosing needle. When the sensor comes in contact with foam, the antifoam pump is activated and antifoam agent is fed into the vessel via the dosing needle. INFORMATION The antifoam sensor is separately available, it is NOT included in the standard packages. For details refer to main chapter “Accessories”, chapter “Antifoam Sensor”. Labfors 5 - Version for Microorganisms Options 04 September 2019 Page 47 of 216 4 Options The following options are available in addition to the equipment included in the scope of supply for the basic unit. 4.1 Pump(s) Additionally to the four peristaltic pumps, which are integrated on the basic unit by default, two analogue pumps are available. These are mounted to the side of the basic unit. Control is the same as for the standard analogue pump, it is adjustable within a range of 0 and 100 %. 4.2 Servomotor (High Torque) For high speed requirements, a powerful servomotor is available and can be used instead of the standard brushless DC motor. 1 Signal connector for (orange) signal cable 2 Power connector for (green) power cable The motor has a mechanic coupling and is coupled to the drive hub on the vessel top plate like the standard motor. 2 1 Labfors 5 - Version for Microorganisms Options Page 48 of 216 04 September 2019 The servomotor is operated by a separate AC servo amplifier. This is connected to the basic unit via two cables (not visible in the picture to the left). The green power cable and the orange signal cable are connected on the rear side of the amplifier box. Control of the servomotor takes place via touch screen operating panel like the standard mo- tor. Specification Stirrer with High-Torque Motor Description Value Drive Shaft with mechanical seal Motor Servomotor Nominal power: 330 W Nominal torque: 1.05 Nm Range of rotation speed with 2 impellers (Standard) 13 L / DN 200 vessel: 10 up to 1250 min-1 All other vessels: 10 up to 1500 min-1 Range of rotation speed with 3 impellers (Option) 13 L / DN 200 vessel: 10 up to 1000 min-1 All other vessels: 10 up to 1500 min-1 Accuracy Measurement: ± 5 min-1 at 100 - 1000 min-1 1 % setpoint at > 1000 min-1 Control: ≤ ± 5 min-1 at 100 – 1000 min-1 ≤ 1 % setpoint at > 1000 min-1 Direction of rotation Counter clockwise (top view vessel) Bearing Outside of vessel, in drive hub Impellers Rushton with 6 blades INFORMATION Rotation speed is valid for viscosity similar to water, without aeration. Vessels are equipped with two impellers by default. The max. adjustable rotation speed on the operating panel is set for 2 impellers. Labfors 5 - Version for Microorganisms Options 04 September 2019 Page 49 of 216 4.3 Level Measurement Detects liquid in the culture vessel, i.e. measures the level by means of the level sensor. When the level sensor comes in contact with liquid, a signal is generated which is displayed as 100 % in the touch screen software. If required, this signal can be used for level control in order to keep the working volume constant in the culture vessel. For this, e.g. a simple cascade can be setup to control the feed pump or one of the optional pumps, to either feed liquid into the vessel or to extract liquid from the vessel. Customized configurations are available on request. Level Sensor The type of level sensor with clamping adapter without O-ring for mounting in a Ø 12 mm / Pg13.5 port in the vessel top plate is supplied as standard. An additional appropriate adapter is supplied for mounting in a 19 mm port in a vessel top plate with DN200 (mm). 1 Connection for banana connector of sensor cable 2 Clamping adapter with slotted screw (A) 3 Sensor shaft with transparent insulation The level sensor is equipped with two NON-autoclavable protective caps. Level sensor types with clamping adapters for Ø 10 mm or 19 mm ports are available, too. For details about clamping adapters and adapters, refer to chapter “Accessories”. 1 2 A 3 Labfors 5 - Version for Microorganisms Options Page 50 of 216 04 September 2019 4.4 Exit Gas Analysis In order to allow the user to draw conclusions regarding the status of the culture while the bioprocess is still underway, the CO2 and O2 measurements are often taken and analysed in the exit gas flow of the bioreactor. 4.4.1 Measuring Systems (Gas Sensors) The following measuring systems are available for exit gas analy- sis:  INFORS HT Gas Analyser: gas analysis equipment (combined CO2 and O2 sensor) from the equipment manufacturer.  Gas sensors from the manufacturer BlueSens: combined CO2 and O2 sensors of the type BlueInOne Ferm, BlueInOne Cell or BlueVary Measurement ranges gas sensors Type gas sensor Vol. % O2 Vol. % CO2 Gas Analyser 0.1 – 25 1) 0 – 10 BlueInOneFerm BlueVary 1.0 – 50 1) 0 – 10 or 0 – 25 BlueInOneCell BlueVary 0 – 100 2) 0 – 10 or 0 – 25 1) only suitable for use in aerobic bioprocesses 2) suitable for use in aerobic and anaerobic bioprocesses For details on the safety, technical data, usage and maintenance requirements for the gas sensors, see the separate documentation provided by the sensor manufacturer. Read this documentation before using the gas sensor and follow the instructions contained therein. 4.4.2 Connecting the Gas Sensors In order to view measurements on the operating panel, the measuring system for exit gas analysis must be connected to the biore- actor. Labfors 5 - Version for Microorganisms Options 04 September 2019 Page 51 of 216 INFORS HT Gas Analyser Connection of the gas analysis equipment from the equipment manufacturer is described in details in its separate documentation. Gas sensors from manufacturer BlueSens The gas sensor must be connected to the sensor cable and the bioreactor’s exit gas must be led into the sensor via hose line. Generally, the cable connection is established only once during installation and remains then. The connection to the exit gas line must be re-established before each cultivation process The ideal connection conditions are detailed in the separate documentation provided by the manufacturer. Connecting the sensor cable The fixed sensor cable is pre-installed in the factory (rear of equipment). The cable has an 8-pin round plug connector. In order to connect the sensor, the plug connector is plugged into the socket marked Port A on the gas sensor. Due to the length of the sensor cable, the gas sensor can be positioned in a large number of possible locations. Establishing the hose connection 3 m of pressure hose, D = 8 x 14.5, and a hose clamp are provided with the equipment in order to establish the hose connection between gas sensor and culture vessel (exit gas filter). The hose connection between the culture vessel (exit gas filter) and the gas sensor must be designed in line with the direction in which the gas flows through the gas sensor. Proceed as follows: Procedure 1. Cut as short a piece as possible off the supplied pressure hose. 2. Push one end of the hose onto the hose nozzle (observe direction of flow) on the gas sensor’s flow adapter and fasten in place with the clamp. 3. Push the open end of the hose onto the exit gas filter on the exit gas cooler. INFORMATION Do NOT use a clamp here, as the hose must be easy to disconnect at this point, e.g. for autoclaving the culture vessel. Labfors 5 - Version for Microorganisms Options Page 52 of 216 04 September 2019 4.4.3 Calibration INFORS HT Gas Analyser The display of measured values on the bioreactor should be calibrated after the annual maintenance of the gas analyser has been carried out For details about maintenance of the gas analyser refer to its separate documentation. Gas Sensors from manufacturer BlueSens 1-point calibration must be carried out once per month and after installation in order to guarantee exact measurement results. This is done directly on the gas sensor itself. The procedure is described in the separate documentation provided by BlueSens. 4.4.4 Replacing the BlueVary Gas Sensor Cartridge The max. operating time of a BlueVary gas sensor cartridge amounts to 9000 operating hours. Once this limit is reached, measurement is no longer possible. I.e. there is no measured value output anymore and the display turns red. The gas sensor cartridge must be replaced by the sensor manufacturer. For detailed information refer to the separate documentation from the sensor manufacturer. 4.5 Multiplexer (Gas Switching Module) To be able to measure exit gas values - usually O2 and CO2 - while bio processes are running in parallel, every single bioreactor usually has to be equipped with the corresponding measuring system (see chapter “Exit Gas Analysis”). Using one or more of the equipment’s manufacturer’s Multiplexers makes it possible to use only a single measuring system for this. The exit gases from the individual bioreactors are directed sequentially to the measuring system used by means of the gas switching module(s). The determined values are assigned to the corresponding bioreactor. When using Multiplexers, the measurement information for the individual culture vessels is not available continually since one measurement sensor sequentially measures the exit gas of all culture vessels. The measurement interval achievable for each culture vessel depends, among other things, on the gas volume stream and the hose type used as well as its length. Labfors 5 - Version for Microorganisms Options 04 September 2019 Page 53 of 216 4.6 Turbidity Measurement Turbidity measurement can be used to draw conclusions regarding the biomass concentration in the culture. To determine the turbidity in the culture two measuring systems are available: Variant OPTEK  Sensor (Single channel light absorption) with transmitter integrated in the basic equipment  Type: ASD12-N with two optical path lengths  Version for microorganisms: OPL05 for higher cell densi- ties.  Version for cell culture: OPL10 for lower cell densities.  Manufacturer: Optek  Measures the absorption within a range of 0 to 4 CU The ASD12-N sensors supply a non-linearised turbidity measurement for the culture. This can be linearised manually using the soft sensor in eve®, for example, in order to determine correlation with factors such as the biomass concentration or optical density. INFORMATION If the temperature of the sensor rises above 50 °C during operation in the medium, an automatic switch-off takes place. After the medium has cooled down, the measurement continues auto- matically. Variant aquila biolabs  Sensor (non-invasive scattered light measurement) with transmitter integrated in the basic equipment (CGQ BioR gateway)  Type: CGQ BioR with two LEDs / measurement modes:  Green: (521 nm) for low cell densities  Infrared: (940 nm) for high cell densites Labfors 5 - Version for Microorganisms Options Page 54 of 216 04 September 2019 CAUTION The light emitted by the LEDs on the sensor plate is highly sensitive and can damage the iris or retina. The CGQ BioR sensor plate contains an infrared LED that emits high energy radiation in the invisible range. Sensor plates with this LED carry the warning symbol shown on the left. – Wear safety goggles and avoid direct contact of LEDs with eyes or skin. – Always keep a safety distance of >1 m from active sensor plates. – Pause or stop running measurements before all work within the safety distance.  Manufacturer: aquila biolabs  Measures within a range of 0 to 1000 INFORMATION CGQ BioR sensors are optimised for microbial bioprocesses. The sensors may be used in temperatures from 15 to 50 °C. The CGQ BioR sensors non-invasively measure the scattered light of the culture. This is proportional to the biomass concentration in the bioreactor, but can also be processed, e.g. by a soft sensor in eve®, in order to obtain a correlation with the optical density. Details and specifications of the sensors and their measuring principles as well as safety, use and maintenance can be found in the separate documentation of the manufacturers. Read these before using the turbidity sensor and follow the instructions. 4.6.1 Calibrating the Sensor Variant Optek Optek sensors are pre-calibrated ex-factory. Inserts are available for reference measurement. Due to the different light absorption of different media, zero point calibration should be performed before each cultivation process. This can be done on the operating panel, either before or after autoclaving, depending on the application in question. For more details, see the main chapter “Operation” chapter “Calibrating the Turbidity Sensor”. Labfors 5 - Version for Microorganisms Options 04 September 2019 Page 55 of 216 Variant aquila biolabs CGQ BioR sensors are pre-calibrated ex-factory. A new calibration is not necessary. 4.6.2 Mounting the Sensor Variant Optek For culture vessels with nominal widths of 90 and 145, Optek ASD sensors can be mounted directly into 12 mm/Pg13.5 ports. For culture vessels with a nominal width of 115, an electrode holder is used. For more details on the electrode holder, refer to main chapter “Accessories” chapter “Electrode Holder”. Note the following points for mounting:  Ensure that the sensor is fitted with an O-ring; fit an O-ring if necessary.  Mount the sensor by hand – do not use any tools!  If the mounting depth of the sensor is adjustable (mounting with electrode holder), make sure the mounting depth is set correctly prior to autoclaving, as later adjustment represents a contamination risk.  Mount the sensor in such a way that it cannot come in contact with other components or the glass vessel.  Mount the sensor in such a way that it has good access to the flow and there is no risk of bubbles collecting in the measurement gap. Variant aquila biolabs CGQ BioR sensors are always attached to the culture vessel with the strap attached to the sensor. For this purpose, the sensor with the measuring window is pressed against the glass vessel and fixed with the strap. Depending on the culture vessel, different positions of the sensor or attaching methods may be necessary. For mounting details, see separate documentation of the sensor manu- facturer. Note the following points for mounting:  Ensure that the sensor is not attached to markings or stickers on the glass vessel, this may affect the measurement.  Mount the sensor so that it is not in front of or in the direct vicinity of reflective steel parts (< 20 mm). Labfors 5 - Version for Microorganisms Options Page 56 of 216 04 September 2019  Ensure that the sensor is positioned in such a way that liquid is in front of the measurement window during the entire bio- process  Foam, high gas hold-ups and the use of antifoam agents can (significantly) interfere with the light scattering of growing cells. 4.6.3 Interferences Turbidity Measurement Interference Displayed measured value is not plausible / unusual Possible Cause Remedy By Sensor cable is twisted or kinked or not properly connected. Check and ensure that the sensor cable is not kinked or twisted. Connect the sensor cable properly as necessary. Operator Optek Sensor is not calibrated Calibrate the zero point Operator Optek Window fouling on the sapphire win- dows. Carefully clean the sensor Operator aquila biolabs Sensor is mounted in the wrong place / measures in foam Place the sensor at the level of the liquid. Make sure that there are no obstacles in front of the measuring window. Operator Faulty sensor cable Replace sensor cable Optek Qualified electrician aquila biolab Sensor manufacturer Faulty sensor cable Replace the sensor Operator 4.7 Permissive Measurement The sensor with the corresponding transmitters must be purchased directly from the manufacturer ABER. INFORS HT offers a connection to the transmitter on the basic unit. Calibration is performed according to the manufacturer’s guidelines directly on the transmitter.  Manufacturer: ABER  System: Futura  Measured parameters:  Permittivity in the 0 to 400 pF cm-1 range and:  Conductivity in the 0 to 40 mS cm-1 range Labfors 5 - Version for Microorganisms Options 04 September 2019 Page 57 of 216 Sensors of the ABER Futura systems measure the permittivity (also: capacitance) and conductivity of the culture. This measured data can be used to determine a correlation with the live biomass concentration, for example, using the soft sensor in eve® or data evaluation. All information about the ABER Futura system is available in the separate documentation provided by the manufacturer. 4.8 Redox Measurement Measures the reduction/oxidation potential (redox) in the medium. 4.8.1 Measuring Systems (Sensors) Two systems are available for redox measurement: METTLER version (analogue)  Classic combined sensor (oxidation reduction potential measurement against a reference)  Type: 405-DPAS-SC-K8S  Manufacturer: METTLER TOLEDO  Measures the reduced potential in the medium in the range from -2,000 mV to +2,000 mV. To use the sensor, the basic unit must be equipped with a corresponding connection. HAMILTON version (digital)  Classic combined sensor (oxidation reduction potential measurement against a reference) with integrated electronics  Type: Easyferm Plus ORP ARC  Manufacturer: HAMILTON  Measures the reduced potential in the medium in the range from -1,500 mV to +1,500 mV. If the basic unit is configured for Hamilton sensors, the redox sensor can be connected instead of the pO2 sensor. If the sensor is configured in addition to the Hamilton pO2 sensor or if the basic unit is configured for Mettler sensors, an additional connecting cable is required. Labfors 5 - Version for Microorganisms Options Page 58 of 216 04 September 2019 4.8.2 Calibration The redox sensor is usually not calibrated/adjusted. However, the calibration of the sensor can be checked using the appropriate redox buffer solution.  METTLER system: 1-point calibration in the calibration menu of the redox parameter in the touch screen software from the administrator user level.  HAMILTON system: Calibration using a Hamilton Arc Handheld or a Hamilton Arc USB cable, both of which are available separately from the sensor manufacturer. In case of a deviation of ± 10 mV from the buffer solutions, clean the redox sensor according to the description provided by the respective sensor manufacturer or contact the sensor manufacturer. For details on the technical data, usage and maintenance requirements for the redox sensors, see the separate documentation provided by the respective sensor manufacturer, i.e. METTLER TOLEDO or HAMILTON. 4.8.3 Mounting the Sensor Redox sensors (both variants) are mounted into a 12 mm / Pg13.5 port in the vessel top plate by the means of an electrode holder. An additional adapter is necessary for mounting the sensor into a 19 mm port. For details about the electrode holder see chapter “Accessories”, “Electrode Holder”. 4.9 pCO2 Measurement Measures saturation of dissolved carbon dioxide (CO2) in the culture in hPa. Measurement is done by a digital CO2 sensor with integrated temperature sensor. Measured values are displayed on the associated transmitter and in the touch screen software.  Type of sensor (digital): InPro5000i ISM  Measuring principle: potentiometric  Type of transmitter: M400  Manufacturer (sensor & transmitter): METTLER TOLEDO The displayed value of parameter pCO2 in the touch screen software is set to a range of 0 hPA to 1000 hPA analogous to the displayed value of the transmitter. Labfors 5 - Version for Microorganisms Options 04 September 2019 Page 59 of 216 If a CO2 gassing line is present and configured with a mass flow controller (parameter CO2 Flow), then this can be used e.g. via cascades for pCO2 control. Calibration of the sensor is carried out directly on the transmitter according the sensor manufacturer’s specifications. For details about technical data, use and maintenance of the pCO2 sensor and transmitter, refer to the separate documentation from the manufacturer METTLER TOLEDO. Mounting of the sensor pCO2 sensors are mounted into a 12 mm / Pg13.5 port in the vessel top plate by the means of an electrode holder. An additional adapter is necessary for mounting the sensor into a 19 mm port. For details about the electrode holder see chapter “Accessories”, “Electrode Holder”. 4.10 Pressure Control The standard equipment is built for pressure-free operation. It is mandatory to have a pressure control installed, if a cultivation process under pressure is foreseen. WARNING Overpressure in the glass vessel may cause the vessel to burst or break. Any build-up of pressure must be avoided without the pressure control option! Labfors 5 - Version for Microorganisms Options Page 60 of 216 04 September 2019 Pressure control consists of a solenoid valve (proportional valve) and a piezo-resistive pressure sensor with fixed O-ring (the O-ring is not visible in the picture to the left). Depending on the equipment configuration, the solenoid valve is situated next to or below the gassing unit on the right side of the equipment. CAUTION Risk of slight burns when touching the hot solenoid valve! The solenoid valve heats up when under electric voltage, which is the case as soon as the equipment is switched on. The pressure hose for connection to the exit gas filter during operation i.e. cultivation is installed when the equipment is stalled. Pressure control is set and activated in the touch screen software in parameter Pressure. Control is possible up to 400 mbar. If pressure control is switched off (parameter Pressure OFF), the valve automatically opens to prevent build-up of pressure in the vessel. For details about the touch screen software refer to the separate operating manual. The pressure sensor is mounted into a 19 mm port in the vessel top plate. A threaded adapter is used for this (refer to chapter “Accessories” for details). A plastic cap protects the steel membrane from damage. A screwed on steel cap protects the female cable plug during auto- claving. 1 Threaded steel cap 2 Hollow screw 3 Protective cap (plastic) Before autoclaving The pressure sensor is mounted into a 19 mm port in the vessel top plate and autoclaved with the culture vessel. 1 2 3 Labfors 5 - Version for Microorganisms Options 04 September 2019 Page 61 of 216 Proceed as follows for the mounting: Procedure 1. Fit a threaded adapter into the 19 mm port in the vessel top plate. 2. Carefully remove the protective cap from the steel membrane of the sensor. ATTENTION The steel membrane is very sensitive and can be damaged when knocking it on solid objects or when rubbed. Carefully mount the pressure sensor by hand. Do not use any tool! 3. Ensure the sensor is equipped with an intact O-ring, fit one if necessary. 4. Carefully screw the pressure sensor into the threaded adapter. 5. Screw the steel cap onto the female sensor connector. After autoclaving The exit gas filter (on the exit gas cooler) must be connected to the pre-installed pressure hose of the solenoid valve. This ensures that the exit gas is led via solenoid valve into the atmosphere i.e. into the exit gas line, which has to be installed on-site by the provider. The sensor cable must be connected to the pressure sensor, too. Proceed as follows: Procedure 1. Fit the pressure hose from the solenoid valve to the exit gas filter. 2. Unscrew the steel cap from the cable plug on the sensor. Labfors 5 - Version for Microorganisms Options Page 62 of 216 04 September 2019 3. Plug the male plug of the sensor cable into the female plug on the sensor. To do so, the red mark on each plug must be aligned. Maintenance Basically, the pressure sensor is maintenance-free. The recalibration interval depends on the operating conditions. However, annual recalibration is recommended by the sensor manufacturer. 4.11 LabCIP, CIP/SIP Unit The equipment is used for cleaning (CIP) and sterilisation (SIP) of the bioreactor, provided there is a certain equipment configuration. All information regarding safety, setup and function, operation, maintenance and technical data of the LabCIP is available in the separate operating manual from the equipment manufacturer. Labfors 5 - Version for Microorganisms Accessories 04 September 2019 Page 63 of 216 5 Accessories The kind and number of accessories included by default in every standard package are listed the following table. It is divided in columns according the diameter nominal (= inside diameter in mm) of the vessels and their different sizes (TV = total volume in L). Labfors 5 - Version for Microorganisms Accessories Page 64 of 216 04 September 2019 Accessory DN115 DN150 DN200 2.0 L TV 3.6 l TV 3.6 l TV 7.5 L TV 13 l TV Base tray 1 1 1 1 1 Sparger, L-shaped 1 1 1 1 1 Impeller, Rushton 2 2 2 2 2 Pocket for temperature sensor in Ø 10 mm port. 1 1 1 1 1 Baffle 3 3 3 3 3 Blanking plug for Ø 10 mm port 1 1 1 1 1 Blanking plug for Ø 12 mm / Pg13.5 port 5 5 5 5 4 Blanking plug for Ø 19 mm port (with fixed O-ring) 1 1 2 2 5 Addition port adapter, quadruple, for Ø 19 mm port. 1 1 1 1 1 Inoculation needle and septum collar for Ø 12 mm / Pg13.5 port 1 1 --- --- --- Inoculation needle and septum collar for Ø 19 mm port. --- --- 1 1 1 Clamping adapter for Ø 12 mm / Pg13.5 port with inside-Ø 6 mm (for sparger) 1 1 1 1 1 Dip tube, straight, Ø 6 mm for Ø 12 mm / Pg13.5 port 1 1 1 1 1 Threaded adapter for Ø 19 mm port 1 1 2 2 5 Sampling system Super Safe Sampler for Ø 12 mm / Pg13.5 port 1 1 1 1 1 Starter Set 1 1 1 1 1 Cold finger1 for Ø 19 mm port. 1 1 1 1 1 Heater pad1 1 1 1 1 1 Reagent bottle and pump holder for 250 mL reagent bottles 1 1 1 1 --- Reagent bottle and pump holder for 500 mL reagent bottles --- --- --- --- 1 1) For single walled vessels with temperature control system with electrical heater pad and cold finger. Labfors 5 - Version for Microorganisms Accessories 04 September 2019 Page 65 of 216 5.1 Base Tray Culture vessels are delivered with a base tray made of stainless steel. Dimensions:  373 mm × 373 mm for vessels with DN 115 / 150  423 mm × 423 mm for vessels with DN NW 200  437 mm x 612 mm for vessels combined with a scale underneath the vessel. 5.2 Reagent Bottle and Pump Holder The culture vessel is autoclaved along with reagent bottles, hoses and pump heads. For easy handling, a holder for reagent bottles and pump heads is available. There are two types available:  for 250 mL reagent bottles, purchased with vessels with DN 115 and 150  for 500 mL reagent bottles, purchased with vessels with DN 200 Labfors 5 - Version for Microorganisms Accessories Page 66 of 216 04 September 2019 The reagent bottle and pump holder can be fitted to the vessel holder. The mounting plate with the pump heads can be fitted to the reagent bottle and pump holder. The holder for 500 mL reagent bottles can be modified using the centering rings, which can be obtained as an optional modification set. The holder can be used then with 250 mL reagent bottles, too. Labfors 5 - Version for Microorganisms Accessories 04 September 2019 Page 67 of 216 5.3 Sampling System Super Safe Sampler Basically different systems and also individual components are available for sampling. This operating manual describes the operation and handling of the aseptic sampling system Super Safe Sampler combined with a dip tube. The use of the Super Safe Sampler prevents the culture vessel from contamination when sampling. Content of the set The set consists of a completely pre-assembled group of valves with hoses and two syringes. It is connected via silicone hose with a dip tube. Valve assembly 1 Sterile filter 2 Check valve 3 Luer activated sample valve 4 T-piece 5 Hose The valve assembly consists of a T-piece, two check valves, a Luer-activated automatic sample valve, a sterile filter, a length of hose as an adapter for the syringe and another hose for connection to the sample dip tube in the culture vessel. 1 2 3 4 5 Labfors 5 - Version for Microorganisms Accessories Page 68 of 216 04 September 2019 Principle of function The sample valve on the side arm of the T-piece opens by putting the Luer connector of the syringe into the valve and closes by removing the syringe. No further handling is necessary. Unintentional re-introduction of the sample material once it has been withdrawn is prevented by a check valve. Thus, contamination of the bulk culture is impossible. Following sampling, a second syringe can be fitted and air pushed in via the sterile filter, in order to displace culture solution from the sample hose and the dip tube of the vessel. With a conventional sampling system, the next sample cannot be taken immediately, as rinsing of the sampling hose and the immersion tube is necessary. By previously removing most of the culture in the sampling line, this sampling system can save culture volume, which is particularly important with small vessels and/or frequent sampling. The dead volume of the culture remaining in the group of valves after flushing with sterile air amounts to a few µl and is negligibly small. If the withdrawal of a very small sample volume is required, with minimum possibility of falsification, a small quantity of culture solution (e.g. 1 ml) can be introduced and rejected before the actual sample is taken. Designated use The Super Safe Sampler is designed for aseptic sampling of completely liquid samples. Solid parts in the sample may lead to clogging of the valves. Therefore, employing the Super Safe Sampler for solid media is not recommended. The Super Safe Sampler is autoclavable (not the syringes!) and for this reason reusable. Practical tips for the use of the Super Safe Sampler Sterility of the culture vessel is ensured at all times without the possible measures mentioned below. Labfors 5 - Version for Microorganisms Accessories 04 September 2019 Page 69 of 216 The use of a sterile syringe and sterile caps is only necessary if the sample has to be processed under sterile conditions. For sampling, the same non-sterile syringe can be used repeatedly, without fear of contamination of the culture vessel. Aseptic Sampling For each sample, use a new, sterile syringe with Luer Lock fitting, in order to ensure the sterility of the sample. Sterile syringes are consumables and therefore not included in the set. INFORMATION The use of another syringe is also possible. But a syringe with Luer lock prevents unwanted movement of the syringe.  Before fitting the syringe, disinfect the sample valve. Fort this, spray a commercially available disinfectant onto the valve.  After spraying and after each sampling, close the the sample valve with a sterile Luer-Lock cap (Dead End Cap) to keep the valve and sample sterile. The caps are not included in the kit. Very convenient to use are socalled combi-caps that fit on male and female connectors alike. Caps that are vented and made of steam sterilisable material can also be fitted during autoclaving. 5.4 Sparger The gas is fed directly into the medium via a sparger by default. The sparger is mounted in a 12 mm / Pg13.5 port in the vessel top plate using a clamping adapter, and connected to the gassing system on the basic unit via a silicone hose with a sterile filter. Labfors 5 - Version for Microorganisms Accessories Page 70 of 216 04 September 2019 Sparger, L-shaped Inside-Ø 4.0 mm Outside-Ø hose connection 6.0 mm Ring sparger Inside Ø 4.0 mm Outside-Ø hose connection 6.0 mm Labfors 5 - Version for Microorganisms Accessories 04 September 2019 Page 71 of 216 5.5 Impellers For details about impellers, refer to chapter “Setup and Function”, “Stirrer”. 5.6 Pocket for Temperature Sensor (Pt100) The pocket is a pipe with a sealed bottom end, and is used to insert the temperature sensor. Pocket Ø 10 mm Fitted with fixed O-ring. Two slotted screws are used for fixing in the 10 mm. The picture on the left does not show the full length of the pocket. Labfors 5 - Version for Microorganisms Accessories Page 72 of 216 04 September 2019 5.7 Baffles Culture vessels are equipped with three baffles by default. Baffle, 3 pieces Must be fitted with O-ring before mounting. A hexalobular screw is used for mounting in the vessel top plate. Labfors 5 - Version for Microorganisms Accessories 04 September 2019 Page 73 of 216 5.8 Lock Nut If no baffles are used, lock nuts must be fitted into the vessel top plate instead. They are disposable in a set of 3 pieces. Must be fitted with an O-ring before mounting. A hexalobular screw is used for mounting in the vessel top plate. Labfors 5 - Version for Microorganisms Accessories Page 74 of 216 04 September 2019 5.9 Blanking Plugs Blanking plugs are used to seal open ports. There are different blanking plugs for the different types of port. Blanking plug, Ø 10 mm Fitted with fixed O-ring. Two slotted screws are used for fixing in 10 mm port Blanking plug, Ø 12 mm Must be fitted with an O-ring before being mounted in the 12 mm/Pg13.5 port. Mounted using a thread. Labfors 5 - Version for Microorganisms Accessories 04 September 2019 Page 75 of 216 Blanking plug Ø 19 mm Fitted with fixed O-ring. A threaded adapter is used for mounting in 19 mm port. 5.10 Addition Port Adapters Addition port adapters are used for liquid addition into the culture vessel. They are mounted in a port in the vessel top plate and end in the headspace of the vessel. They have one or several hose connections and various models are available. The following can be connected to addition port adapters:  Reagent bottles with feed solution, antifoam agent or corrective reagent (base/acid).  If appropriate, exit gas line Addition port adapter, quadruple, Ø 19 mm Inside-Ø 3.0mm Outside-Ø hose connection 4.0 mm Fitted with fixed O-ring. Two slotted screws are used for mounting in 19 mm port. Labfors 5 - Version for Microorganisms Accessories Page 76 of 216 04 September 2019 Addition port adapter Ø 12 mm Inside-Ø 3.0 mm Outside-Ø hose connection 5.0 mm Muts be fitted with O-ring. Thread is used for mounting in 12 mm / Pg13.5 port. Addition port adapter Ø 12 mm Inside-Ø 6.0 mm Outside-Ø hose connection 8.5 mm Must be fitted with O-ring before mounting. Thread is used for mounting in 12 mm / Pg13.5 port. Addition port adapter Ø 19 mm Inside-Ø 3.0 mm Outside-Ø hose connection 5.0 mm Fitted with fixed O-ring. Threaded adapter is used for mounting in 19 mm port. 5.11 Inoculation Needles Inoculation needles are used for feeding liquids into the culture vessel, which cannot be autoclaved with the culture vessel. These liquids may be e.g. the inoculum or heat-sensitive reagents. If using an inoculation needle, then a septum (inoculation membrane) must be fitted along with a septum collar in the appropriate port. The inoculation needle is connected with the reagent bottle and autoclaved. The liquid, e.g. the inoculum, which shall to be added into the culture vessel, is shortly before addition filled into the reagent bottle under sterile condition. The septum is then Labfors 5 - Version for Microorganisms Accessories 04 September 2019 Page 77 of 216 pierced with the inoculation needle, which is screwed into the septum collar. The septum may be wetted e.g. with an alcohol solution that is lit up before the piercing. Inoculation needles are purchased including septum collar. They have an oblique tip to facilitate the piercing. The hose connection and the very sharp needle tip are covered with non-autoclavable(!) protective caps. INFORMATION For addition of autoclavable liquids, addition port adapters or inoculation needles without septum and without septum collar are suitable, too. These must be mounted in the ports and connected to the reagent bottles before autoclaving. Inoculation needle Ø 12 mm Inside-Ø 2.5 mm Outside-Ø hose connection 4.5 mm Must be fitted with O-ring. Thread is used for mounting in septum collar for a12 mm / Pg13.5 port. Labfors 5 - Version for Microorganisms Accessories Page 78 of 216 04 September 2019 Inoculation needle, Ø 19 mm Inside-Ø 2.5 mm Outside-Ø hose connection 4.5 mm Fitted with fixed O-ring. Thread is used for mounting in septum collar in 19 mm port. Inoculation needle, 19 mm Inside-Ø 4.0 mm Outside-Ø hose connection 6.0 mm Fitted with fixed O-ring. Thread is used for mounting into septum collar in 19 mm port. Labfors 5 - Version for Microorganisms Accessories 04 September 2019 Page 79 of 216 Inoculation needle, double, Ø 19 mm Inside-Ø 2.0 mm Outside-Ø hose connection 4.0 mm Fitted with fixed O-ring. Thread is used for mouting in septum collar in 19 mm port. 5.12 Septum Collars Septum collars are either used in combination with the corresponding inoculation needle with septum (piercing membrane) or with a syringe with injection needle and a septum. Inoculation needles are always supplied including a septum collar, but septum collars are separately available, too. Septum collar, Ø 12 mm With internal thread. A septum must be inserted in the 12 mm / Pg13.5 port before mounting. A thread is used for mounting. Labfors 5 - Version for Microorganisms Accessories Page 80 of 216 04 September 2019 Septum collar, Ø 19 mm With inside thread. Septum must be inserted in port before mounting in 19 mm port. Threaded adapter is used for mounting. Multi-inlet septum collar, system Roussel, Ø 19 mm With inside thread, for inoculation with syringe. Septum must be fitted in 19 mm port before mounting. Threaded adapter is used for mounting. 5.13 Dip Tubes Dip tubes are open at both ends and are mounted in a vessel top plate port with a clamping adapter. Dip tubes are used for a variety of purposes:  For filling the culture vessel after autoclaving. Using a dip tube prevents foaming.  For adding inoculum.  For sampling. The aseptic Super Safe Sampler system can be used for sampling.  For harvesting  For siphoning off medium during continuous cultivation  For draining the culture vessel Depending on the purpose, silicone hoses are connected to the dip tube via other vessels, sampling systems or, if necessary, hose trees. Multiple dip tubes can be used at any one time, providing that enough vessel top plate ports are available. Different types of dip tubes are available. Labfors 5 - Version for Microorganisms Accessories 04 September 2019 Page 81 of 216 Dip tube, straight, Ø 6 mm Inside-Ø 3.0 mm Hose connection outside-Ø 4.2 mm / 5.0 mm The dip tube does not reach as far as the bottom of the vessel. The illustration on the left shows only the upper section of the dip tubes. Dip tube, straight Ø 12 mm Inside-Ø 10 mm Hose connection outside-Ø 12 mm The dip tube does not reach as far as the bottom of the vessel. The illustration on the left shows only the upper section of the dip tube. Labfors 5 - Version for Microorganisms Accessories Page 82 of 216 04 September 2019 Dip tube, curved with horizontal tip, Ø 6 mm Inside-Ø 4,0 mm Hose connection outside-Ø 6.0 mm The dip tube does reach to the vessel bottom. Dip tube, curved with frit, Ø 6 mm Inside-Ø 2.0 mm Hose connection outside-Ø 4.0 mm Pore size of frit 40 µm The dip tube does reach to the vessel bottom. Labfors 5 - Version for Microorganisms Accessories 04 September 2019 Page 83 of 216 5.14 Clamping Adapters Clamping adapters are used for mounting the sparger, the various dip tubes and the antifoam/level sensors. The clamping adapter fixes the component part in place and can be used to adjust its mounting depth. The clamping adapter must match the outside diameter of the part to be mounted and the size of the port. Clamping adapter Ø 6 mm / 10 mm Fitted with fixed O-ring (B). Two slotted screws are used for mounting in 10 mm port. After loosening the slotted screw (A) the component part with Ø 6 mm can be inserted into/pulled out from the clamping adapter. The component part is fixed in the clamping adapter by fastening the slotted screw. Clamping adapter Ø 6 mm / 12 mm Must be fitted with O-ring before mounting. Thread is used for mounting in 12 mm / Pg13.5 Port. After loosening the slotted screw (A) the component part with Ø 6 mm can be inserted into//pulled out from the clamping adapter. The component part is fixed in the clamping adapter by fastening the slotted screw. Clamping adapter, Ø 6 mm / 19 mm Fitted with fixed O-ring (B). Threaded adapter is used for mounting in 19 mm port (see chapter “Threaded Adapter”) After loosening the hollow screw (A) the component part with Ø 6 mm can be inserted into/pulled out from the clamping adapter. The component part is fixed in the clamping adapter by fastening the hollow screw. A A A B B Labfors 5 - Version for Microorganisms Accessories Page 84 of 216 04 September 2019 Clamping adapter Ø 8 mm / 12 mm Must be fitted with O-ring before mounting. Thread is used for mounting in 12 mm / Pg13.5 port. After loosening the hollow screw (A) the component part with Ø 8 mm can be inserted into/pulled out from the clamping adapter. The component part is fixed in the clamping adapter by fastening the hollow screw Clamping adapter, Ø 12 mm / 19 mm Fitted with fixed O-ring (B) Threaded adapter is used for mounting in 19 mm port (see chapter “Threaded Adapter”). After loosening the hollow screw (A) the component part with Ø 12 mm can be inserted into/pulled out from the clamping adapter. The component part is fixed in the clamping adapter by fastening the hollow screw. 5.15 Electrode Holders Electrode holders are used to adjust the mounting depth of sensors (pH, pO2, etc.) in 12 mm/Pg 13.5 ports. The electrode holder, respectively the sensor must be fitted with an O-ring for mounting. The electrode holder comprises a sheath with a grub screw, a guide bar with a fork, and a hollow screw. The wrench for the grub screw is also included in the scope of supply. 1 Sheath 2 Grub screw 3 Guide bar 4 Fork 5 Hollow screw A A B 3 2 5 1 4 Labfors 5 - Version for Microorganisms Accessories 04 September 2019 Page 85 of 216 5.16 Threaded Adapter The threaded adapter has an inside thread and is used for mounting blanking plugs or clamping adapters with Ø 19 mm in (threadless) 19 mm ports in the vessel top plate. Two slotted screws are used for mounting. 5.17 Adapter The adapter is used for component parts with Ø 12 mm into a (thread less) 19 mm port in the vessel top plate. Fitted with a fixed O-ring. Labfors 5 - Version for Microorganisms Accessories Page 86 of 216 04 September 2019 5.18 Heater Pad The electrical heater pad is needed for heating up single walled vessels. It is fitted around the vessel and held together with the tensioning springs. 1 Power cable 2 Hook 3 Tensioning spring 1 2 3 Labfors 5 - Version for Microorganisms Accessories 04 September 2019 Page 87 of 216 5.19 Cold Finger A cold finger is provided for single walled vessels for cooling pur- poses. 1 Connection water inlet basic unit 2 Connection water outlet basic unit The cold finger is already equipped with pressure hoses for water inlet (upper connector) and water outlet (lower connector). Appropriate hoses with rapid couplings and plug-in nozzles for connection to the basic unit are included. 1 2 Labfors 5 - Version for Microorganisms Accessories Page 88 of 216 04 September 2019 Cold finger, Ø 19 mm Fitted with fixed O-ring. Threaded adapter is used for mounting in 19 mm port. Cold finger, Ø 12 mm Adjustable mounting depth, Must be fitted with O-ring before mounting. Thread is used for mounting in 12 mm / Pg13.5 port. Labfors 5 - Version for Microorganisms Accessories 04 September 2019 Page 89 of 216 5.20 Exit Gas Cooler The exit gas cooler is fully equipped ex-factory. 1 Exit gas cooler 2 Adjustable hose clamp 3 Exit gas filter 4 Rapid coupling 5 Plug-in nozzle The exit gas cooler is supplied with cooling liquid from the basic unit. The flow of the cooling liquid can be adjusted by the manual control valve on the basic unit. The exit gas cooler is already equipped with pressure hoses for water inlet (lower connector) and water outlet (upper connector). Appropriate hoses with rapid couplings and plug-in nozzles for connection to the basic unit are included. Another piece of pressure hose is fitted onto the exit gas pipe and equipped with a disposable exit gas filter. Hose connections and the filter are secured with hose clamps. The hose clamp securing the exit gas filter is equipped with a screw to loosen or tight it man- ually. INFORMATION The exit gas filter must be replaced after each cultivation. The exit gas cooler works with temperature control switched on only! 2 3 4 51 Labfors 5 - Version for Microorganisms Accessories Page 90 of 216 04 September 2019 Exit gas cooler Ø 12 mm Must be equipped with O-ring Its thread is used for mounting in 12 mm / Pg13.5 Port Exit gas cooler Ø 19 mm With fixed O-ring. A threaded adapter is needed for mounting in 19 mm port. Exit gas cooler with needle, Ø 19 mm Separately autoclavable Only available for vessel top plates with DN 50 and DN200 Exit gas cooler, swivelling sideways, Ø 19 mm With fixed O-ring. A threaded adapter is needed for mounting into 19 mm port. Labfors 5 - Version for Microorganisms Accessories 04 September 2019 Page 91 of 216 5.21 Reagent Bottles For addition of corrective reagent and feed solution (substrate) into the culture vessel, different sizes and models of reagent bottles made of borosilicate glass are available. Size Ø Hose Number of hose connectors 250 mL 1) 2 x 6 mm 2 250 mL 2 x 6 mm 3 + 1 500 mL 1) 2 x 6 mm 2 500 mL 2 x 6 mm 3 +1 1000 mL 3 x 5 mm 2 2000 mL 3 x 5 mm 2 5000 mL 3 x 5 mm 2 10'000 mL 3 x 5 mm 2 1) Thes two bottle sizes fit into the reagent bottle and pump holder. The reagent bottles are already equipped on delivery. 1 Cable tie 2 Silicone hose 3 Filter Reagent bottles with two hose connectors are provided as stand- ard. Two hose connectors are situated on the lid. One is equipped with a short piece of silicone hose with filter for pressure equalisation. The second connector is equipped with a piece of silicone hose on the inside of the lid. A piece of silicone hose is included to establish the hose line from the reagent bottle to the addition port adapter in the vessel top plate and to the pump head. 1 2 3 Labfors 5 - Version for Microorganisms Accessories Page 92 of 216 04 September 2019 The individual parts of the reagent bottle are: 1 Threaded cap, PBT 2 Plate with two hose nozzles, PVDF 3 Flat gasket, silicone 4 Laboratory bottle, Borosilicate The picture to the left shows the reagent bottle type with 3 + 1 hose connectors. 1 2 3 4 Labfors 5 - Version for Microorganisms Accessories 04 September 2019 Page 93 of 216 5.22 Antifoam Sensor If ordered, the antifoam model fitting into Ø 10 mm ports is supplied by default. A clamping adaptor with fixed O-ring is used for mounting. 1 Sensor head with port for banana connector (A) 2 Clamping adaptor with slotted screw (B) 3 Needle with transparent insulation The antifoam sensor is equipped with two NON-autoclavable protective caps. Models for Ø 12 mm / Pg13.5 or 19 mm ports with appropriate clamping adapters are available, too. Port Inside-Ø Outside-Ø hose connection Ø 10 mm 2 mm 4 mm Ø 12 mm / Pg13.5 3 mm 4 mm Ø 19 mm 3 mm 4 mm 5.23 Pump Heads The autoclavable pump heads are fitted with PharMed pump hoses prior to delivery. Three different hose diameters are available for different delivery rates:  1.0 mm (standard)  0.5 mm  2.5 mm For more detailed information about pumps and hoses refer to main chapter “Technical Data”, chapter “Specification”, “Pumps 2 3 1 A B Labfors 5 - Version for Microorganisms Accessories Page 94 of 216 04 September 2019 5.24 Sterile Filters Sterile filters are used to protect against contamination in both the gassing line and the exit gas line. In addition to this, all reagent bottles used for pressure equalisation must be fitted with a short piece of hose with a filter. All the sterile filters in the scope of supply are autoclavable, disposable filters with PTFE diaphragms. INFORMATION Sterile filters must be clean and dry at all times, and should thus ideally be replaced after each use. Ø 37 mm, marked red Application Supply air Retention rate 0.2 µm Ø 37 mm, marked green Application Exit gas Retention rate 0.3 µm dry 1.0 µm wet Ø 25 mm, not marked Application Super Safe Sampler Retention rate 0.2 µm Diaphragm PTFE Labfors 5 - Version for Microorganisms Accessories 04 September 2019 Page 95 of 216 Ø 25 mm, not marked Application Reagent bottles (pressure equali- sation) Retention rate 0.45 µm Diaphragm PTFE Ø 50 mm, Type ACro50 TF, no label Usage Inlet air 1) / Exit gas 2) Retention rate 0.2 µm 1) For high gas flows rates 2) For absolute sterility 5.25 O-Rings and Gaskets Description Ø mm Utilisation O-ring, EPDM 1.5 x 7.5 Gasket, port size 10 mm O-ring, EPDM 2.62 x 10.77 Gasket, port size 12 mm / Pg13.5 O-ring, EPDM 2.5 x 15.0 Gasket, port size 19 mm O-ring, EPDM 1.78 x 6.07 Drive shaft O-ring, EPDM 2.5 x 9.0 Baffles O-ring, EPDM 3.53 x 123.42 Top plate gasket, culture vessel DN115 O-ring, EPDM 3.53 x 158.34 Top plate gasket, culture vessel DN 150 O-ring, EPDM60 5.0 x 208 Top plate gasket, culture vessel DN200 Flat gasket, Sili- cone 32 x 42 x 2 Gasket for reagent bottle lid (for all sizes with two hose connections) Labfors 5 - Version for Microorganisms Accessories Page 96 of 216 04 September 2019 5.26 Hoses and Accessories Hose type Ø mm Application Pressure hose, fibre- glass-woven 10 x 17 Water outlet Pressure hose, fibre- glass-woven 8 x 14,5  Water inlet  Exit gas filter attachment (on exit gas cooler) Pressure hose, fibre- glass-woven 6 x 11,9  Gas connection(s)  Water supply and return, exit gas cooler Silicone hose 5 x 8 Gassing (sparger) Silicone hose, transpar- ent 2 x 6 Reagent bottles: 250 ml und 500 ml (hose lines for reagents) Silicone hose, transpar- ent 3 x 5 Reagent bottles: 1000 ml 2000 ml 5000 ml 10‘000 ml (hose lines for reagents) Silicone hose, 60° Shore, 10 x 16 Water connections vessel jacket (double walled culture vessels) Labfors 5 - Version for Microorganisms Accessories 04 September 2019 Page 97 of 216 Attachment Ø mm Application Hose clamp, 1 screw, INOX 14 Attachment hose(s) gas in- let(s). Hose clamp, INOX 17 Attachment hose water inlet Hose clamp, INOX 19 Attachment hose water outlet Hoffmann pinchcock, nickel-plated brass 12 For clamping off hose lines, e.g. unused addition port adapters, sparger hose etc. Cable tie, polyamide 2,4 x 85 Hoses for reagent bottles and pumps, inlet air filter, sparger, dip tube of sampling system. Double hose nipple, 3/32" x 1/16", PVDF --- Pump heads with hoses to reagent bottles with internal-Ø 1 mm Double hose nipple, 1/8" x 1/8", PVDF --- Pump heads with hoses to reagent bottles with internal -Ø 2.5 mm Rapid coupling plug nipple, DN12.7 hose nozzle 1/2" A --- water inlet and outlet hoses of vessel jacket (double walled culture vessels) Rapid coupling plug nipple DN12.7 hose nozzle 1/2" --- water overflow hose of vessel jacket (double walled culture vessels) 5.27 Inoculation Accessories and Tools Description Application Hexagon socket spanner, WAF 17 For blanking plugs in 12 mm/Pg13.5 ports Allen key, WAF 2, DIN911, For grub screws on impellers Septum (inoculation diaphragm), Ø = 16 mm MVQ Silicone, trans- parent For 12 mm/Pg13.5 ports Septum (inoculation diaphragm), Ø = 19 mm MVQ Silicone, trans- parent For 19 mm ports Labfors 5 - Version for Microorganisms Transport and Storage Page 98 of 216 04 September 2019 6 Transport and Storage The following specifications are based on transport and storage of an unpacked equipment at the provider’s site. 6.1 Transport WARNING Improper transport, the use of incorrect auxiliary equipment and careless handling of the equipment may lead to injuries and severe property damage. The following points must be observed when transporting the equipment internally (relocation):  Always work in pairs and use suitable auxiliary equipment when transporting the equipment.  The entire equipment (basic unit and culture vessel) contains delicate glass parts.  Especially when using auxiliary tools, it is important to observe that the equipment’s centre of gravity is not in the middle. WARNING The entire equipment (basic unit and culture vessel) is too heavy to be carried by one person alone. Even the basic unit on its own exceeds the weight that should be carried by one person alone. Labfors 5 - Version for Microorganisms Transport and Storage 04 September 2019 Page 99 of 216 6.2 Storage  Before each time they are put into storage, decontaminate, thoroughly clean and dry the culture vessel and all accesso- ries1.  Store the equipment and its components clean, dry and protected against dust, dirt and liquids.  Store the equipment and its components in a cool place with low air humidity but protected against frost.  Storage temperature: 5°C – 55°C  Relative air humidity, non-condensing: 10% - 95%.  Protect the equipment from aggressive media, direct sunlight and mechanical vibrations. 1) Maintain and store sensors produced by other manufacturers in accordance with the separate documentation. Labfors 5 - Version for Microorganisms Installation and Initial Operation Page 100 of 216 04 September 2019 7 Installation and Initial Operation Installation and initial operation of the equipment may only be carried out by qualified personnel from the manufacturer or personnel authorised by the manufacturer. WARNING Installation and initial operation requires qualified and experienced personnel. Faulty installation may lead to dangerous situations or severe loss of property. Only let carry out installation and initial operation by the manufacturer’s qualified personnel or authorised personnel by the manufacturer only. Therefore, the following sections only list the energies that must be provided and the connection requirements that are to be respected on site by the provider. Exception The basic functions of the bioreactor are tested and demonstrated at the same time to the operator in form of a short test run on site during installation by the qualified personnel. In order to become familiar with the basic functions of the bioreactor before the first cultivation or after a longer period of non-use of the equipment, the operator may afterwards carry out this short test run any time, too. For details refer to chapter “Test Run”. 7.1 General Location Requirements for Installation The following requirements must be met for the installation of the equipment:  The figures and ranges specified in the chapters “Technical Data, Connection Values” and “Technical Data, Operating Conditions” must be observed.  The equipment must only be installed inside a laboratory or a laboratory-like environment. Labfors 5 - Version for Microorganisms Installation and Initial Operation 04 September 2019 Page 101 of 216  The installation site must be level, sufficiently stable and able to bear loads.  There must not be any sources of electrical interference near the equipment. 7.2 Minimum Distances To operate and maintain the unit it must be installed with a minimum spacing of 150 mm from walls, ceilings or other equipment. 7.3 Power Supply The in-house electric power supply of the equipment must meet the following conditions:  Single-phase, constant power supply  Type 230 V / 50/60 Hz  Type 115 V / 60 Hz The power supply of the equipment must be made safe by the use of an FI-switch (or RCD – Residual Current Device) of the kind RCCB, Type B on the customer’s side. 7.4 Water Supply and Return The in-house water supply to the equipment, as well as the drainage of the water, must meet the following requirements  “Very soft” or “soft” water quality (CaCO3 concentration 0 mmol L-1 to 1.5 mmol L-1) ATTENTION Not observing the water quality requirements may lead to damage or failure of the equipment. The heating system has protection against dry running which is based on measurement of conductivity. Therefore the heating does not work or does not work properly when using demineralised or distilled water as cooling liquid!  Constant water supply at a pressure of 2 ± 1 bar Labfors 5 - Version for Microorganisms Installation and Initial Operation Page 102 of 216 04 September 2019  Inlet temperature 10 °C up to 20 °C  Manometer to check the primary pressure available  The drain is heat-resistant and without back pressure Hoses  Only use pressure-resistant and intact hoses.  Only use appropriate hoses, use adaptors as necessary.  Secure hoses with the appropriate clamps. 7.5 Gas Supply The in-house gas supply to the equipment must meet the following requirements:  Constant gas supply at a pressure of 2 ± 0.5 bar  Gas(es) is/are dry, clean and free of oil and dust  Recommended compressed-air quality as per DIN ISO 8573- 1: Class 1,2,3,4 ATTENTION The use of impure gases can lead to blockage of the sterile filter and damage the mass flow controller. Only use dry, clean and oil-free gases. Hoses  Only use pressure-resistant and intact hoses.  Only use appropriate hoses, use adaptors as necessary.  Secure hoses with the appropriate clamps. WARNING The use of inappropriate or damaged hoses and/or inappropriate fixing may lead to leakage of gases. Depending on the gas in question, there may be a danger of gas explosion and/or danger of suffocation as well as a hazard for the health of the oper- ator. Always close the gas supply before a hose is removed and when the equipment is not in use. Labfors 5 - Version for Microorganisms Installation and Initial Operation 04 September 2019 Page 103 of 216 7.6 Exit Gas On site, it must be ensured that:  the exit gas is safely led away by means of a suitable, gastight hose.  the working environment and/or the laboratory/laboratory-like facility is equipped with a sufficient ventilation system, depending on the application. 7.7 Test Run In order to become familiar with the basic functions of the bioreactor before the first cultivation or after a longer period of non-use of the equipment, a short test run can be executed. The test run comprises:  Temperature control (cooling / heating)  Stirring  Gassing Normal compressed air is used for gassing. To avoid calcium deposits, demineralised water is recommended for filling the vessel. The following description of the test run does not detail handling of individual components, e.g. stirrer, sparger etc. Detailed descriptions of their handling are given in the corresponding chapters of the main chapter “Before Cultivation”. ATTENTION Risk of damaging the glass vessel due to inappropriate handling of removing & mounting the vessel top plate! Strictly follow the instructions stated in the appropriately named chapters. For details on operation, refer to the separate operating manual of the touch screen software. Labfors 5 - Version for Microorganisms Installation and Initial Operation Page 104 of 216 04 September 2019 7.7.1 Preparation Test Run Before starting the test run, check and ensure the following:  All required services are available and activated  All services have the correct connection pressure The following work is to be executed before the test run: Procedure 1. Remove the vessel top plate (without tools!) and put it aside carefully. ATTENTION If the vessel top plate presses against long built-in components they may be distorted due to the weight of the top plate. Always position the vessel top plate so that it does not lie on top of components. 2. Fill the culture vessel with water – preferably demineralised – to the working level. 3. Ensure that the stirrer and sparger are mounted; if necessary, mount them. 4. Fit the top plate and Fix the knurled nuts cross-wise by hand! For this, strictly follow instructions stated in chapter “Fitting the Vessel Top Plate”. 5. Check the lubrication of the mechanical seal. For details about lubrication of the mechanical seal, refer to chapter “Cleaning and Maintenance”, “Lubricating the Mechanical Seal”. INFORMATION Lubrication of a new equipment is not necessary. When carrying out a test run on equipment which has not been in use for a longer period of time, controlling the lubrication of the mechanical seal, and if applicable, its lubrication, is imperative. If an exit gas cooler is used: 6. Screw the exit gas cooler into the port on the vessel top plate port. The exit gas cooler is equipped with a new exit gas filter in the factory. 7. Connect the exit gas cooler to the basic unit; follow the symbols on the basic unit: Labfors 5 - Version for Microorganisms Installation and Initial Operation 04 September 2019 Page 105 of 216 water inlet on bottom of exit gas cooler / water outlet at top of exit gas cooler. 8. Close all remaining open ports with blanking plugs. If no exit gas cooler is present or it is not used:  Close the control valve for water flow of the exit gas cooler on the basic unit.  One port in the vessel top plate must be open! 9. Connect the culture vessel to the basic unit: Single walled vessel: a) Connect the water inlet and water outlet of the cold finger to the basic unit; follow the symbols on the basic unit:  Connect the lower pressure hose of the cold finger to the connector with the symbol of the vessel jacket outlet.  Connect the upper pressure hose of the cold finger to the connector with the symbol of the vessel jacket inlet. b) Ensure the heater pad is dry and has no visible damages (cracks, kinks etc.) DANGER Danger of fatal electric shock! The use of a damaged or wet/damp heater pad can be perilous! c) Fit the heater pad around the vessel and connect it to the basic unit. Double walled (jacketed) vessel: a) Connect the water inlet, water outlet and overflow of the vessel jacket to the basic unit; follow the symbols on the basic unit. INFORMATION In order that the hose connections can be disconnected easily again, slightly moisten the rapid couplings on the silicon hoses prior to connecting them. Labfors 5 - Version for Microorganisms Installation and Initial Operation Page 106 of 216 04 September 2019 10. Equip the sparger with a piece of silicone hose (D= 5 x 8 mm) for gassing and a dry, clean inlet air filter (accessories, filter with red label). 11. Fit another piece of silicone hose for gassing (compressed air) to the hose nozzle on the basic unit. 12. Connect both hoses via the inlet air filter (connect the hose end to the hose nozzle of the inlet air filter). 13. Insert the temperature sensor as far as it will go into the pocket in the top plate. CAUTION Risk of burns and loss of property due to elevated temperature! The temperature control circuit will overheat without an inserted temperature sensor and/or without liquid in the vessel. This can lead to burns and loss of property. 14. Couple the motor. INFORMATION The motor is controlled via the basic unit. Its cable is plugged in during installation and will not be unplugged anymore during routing operation of the equipment. 15. Switch on the equipment on the power switch and wait until the system is booted. 7.7.2 Filling the Water Circuit To fill the water circuit, proceed as follows: Procedure 1. Set a low setpoint for parameter temperature (e.g. 10 °C), in order to activate the water supply into the vessel jacket or cold finger. 2. Start the bioreactor. All parameters except for Temperature remain switched off; switch them off if necessary. Labfors 5 - Version for Microorganisms Installation and Initial Operation 04 September 2019 Page 107 of 216 a) Temperature control system with electrical heater pad and cold finger: You should hear water flowing into the cold finger now. b) Water temperature control system: You should hear and see water flowing into the vessel jacket. If using an exit gas cooler: The water supply of the exit gas cooler is activated now. INFORMATION The exit gas cooler only works with activated temperature control (Parameter Temperature ON). 3. Use your hands to check whether the exit gas cooler is beginning to cool down. Open the control valve for water flow on the basic unit, if necessary. When the circuit is full, water escapes from the overflow on the vessel jacket or cold finger and flows into the water outlet. 4. Wait for min. 1 minute. This ensures that no air is left in the temperature control cir- cuit. 7.7.3 Stirring Bioreactor is running with temperature switched on To test the stirrer, proceed as follows: Procedure 1. On the operating panel for the parameter Stirrer, set a low set- point. For further information about the different ranges of rotation speed refer to the specification in main chapter “Technical Data”. 2. Switch the parameter on. For the rest of the procedure, allow the bioreactor to run with the temperature switched on and the stirrer running. Labfors 5 - Version for Microorganisms Installation and Initial Operation Page 108 of 216 04 September 2019 7.7.4 Heating and Adjusting Temperature Bioreactor is running with temperature and stirrer switched on To test the heating and adjust the temperature, proceed as follows: Procedure 1. On the operating panel, set a high setpoint for parameter Temperature, e.g. 45 °C.. The water supply for cooling is stopped; the system heats up. a) Temperature control system with electrical heater pad and cold finger: The heater pad starts to heat up. b) Water temperature control system: The temperature of the liquid (water) in the vessel jacket starts increasing. 2. Wait until the temperature has adjusted to the setpoint. For the rest of the procedure, allow the bioreactor to run with the temperature switched on and the stirrer running. 7.7.5 Gassing Bioreactor is running with temperature and stirrer switched on To test the gassing, proceed as follows: Procedure 1. If applicable, slowly open the rotameter needle valve. 2. If applicable, set a low setpoint in the appropriate parameter Flow (depending on the configuration) and switch the parameter on. 3. If applicable, ensure that all other gas parameters (e.g. Gasmix, GM Flow etc.) are switched off. If the gassing is working, air bubbles now form in the water in the culture vessel. Labfors 5 - Version for Microorganisms Installation and Initial Operation 04 September 2019 Page 109 of 216 7.7.6 End of Test After all parameter setpoints have been reached, the test can end here. Proceed as follows: Procedure 1. Stop the bioreactor on the operating panel and shut down the system. 2. Switch off the equipment on the power switch. ATTENTION Switching the equipment off at the power switch without previously stopping the bioreactor and shutting down the system on the operating panel may lead to damage of the operating panel! 3. Shut off the supply lines. 4. Let the motor cool down. CAUTION Risk of minor burns if the motor is touched during operation or its cooling phase! When the motor has cooled down: 5. Uncouple the motor from the vessel and place it on a clean and dry work surface. 6. Empty the culture vessel. Labfors 5 - Version for Microorganisms Before Cultivation Page 110 of 216 04 September 2019 8 Before Cultivation The following chapters describe all the preparatory work before starting the cultivation process. This essentially comprises:  Preparing and autoclaving the culture vessel:  Checking the gaskets (O-rings) on component parts and culture vessel  Mounting component parts  Filling or moistening the culture vessel  Preparing sensors and other accessories  Autoclaving  Connecting the culture vessel and preparing for cultivation:  Connecting the cables and hoses between the culture vessel and the basic unit  Filling the vessel if necessary  Preparing sensors and other accessories 8.1 Preparing and Autoclaving the Culture Vessel All accessories required for later cultivation must be prepared and mounted accordingly and autoclaved together with the culture ves- sel. 8.1.1 Mounting the Glass Vessel in the Holder The following procedure describes how to insert and fix the glass vessel in the vessel holder. It is necessary to strictly follow the procedure in order to avoid any damage to the glass vessel or other components. Proceed as follows: Labfors 5 - Version for Microorganisms Before Cultivation 04 September 2019 Page 111 of 216 Procedure 1. Position the vessel holder on a stable and flat surface. 2. Only for double walled vessels with DN 200: Fit a piece of silicone hose (20 mm) to the vessel jacket overflow to protect the glass olive. 3. Only for double walled vessels with DN 200: Place the O-ring around the vessel above the upper edge of the vessel jacket. The O-ring is used to protect the vessel jacket from being damaged. Labfors 5 - Version for Microorganisms Before Cultivation Page 112 of 216 04 September 2019 4. Place the black shock-absorbing ring under the collar of the vessel. 5. Bring the two parts of the vessel flange under the vessel collar and hold them in position. The bevelled edge of the two parts of the flange must point upwards. 6. Lower the glass vessel onto the vessel holder and insert the holder’s rods into the holes of the vessel flange at the same time. Carefully set the vessel down. 7. Only for vessels (single and double walled) with DN 200: Fix the two parts of the vessel flange on both sides with the connecting plates (8 x M5x10 screws). Use only the delivered original screws. 8. Place the white spacers on the rods of the vessel holder. Labfors 5 - Version for Microorganisms Before Cultivation 04 September 2019 Page 113 of 216 9. Place the second shock-absorbing ring (green) on top of the vessel collar. 10. Make sure that the shock-absorbing ring is correctly positioned on the vessel collar and is not shifted. The shock-absorbing ring must not overlap the outer edge of the vessel. 11. Position the stainless steel ring on top of the white spacers. Insert the rods of the vessel holder into the holes of the steel ring. 12. Fix the stainless steel ring with the four lock nuts. Screw down the lock nuts crossways by hand until they touch the steel ring. Do not tighten them and do not use any tool! ATTENTION If the lock nuts on the stainless steel ring are tightened too much, the glass vessel may be damaged. Only tighten the nuts by hand! It is imperative that they are not tightened with a tool under any circumstances. Labfors 5 - Version for Microorganisms Before Cultivation Page 114 of 216 04 September 2019 13. Place the O-ring (top plate seal) on top of the vessel. The Oring must rest completely on the slightly protruding edge of the glass vessel within the steel ring. Make sure that the O-ring is correctly positioned, otherwise the vessel will not be properly sealed. 8.1.2 Checking Gaskets (O-Rings) O-rings are used to seal all openings on the vessel and top plate. The top plate, its ports and all accessories are thus equipped with O-rings. Before every use, the O-rings must be checked that they are present, undamaged and correctly seated. Damaged O-rings must be replaced. INFORMATION Wet the O-rings with 70% alcohol or a little water to facilitate removing and replacing O-rings or accessories with O-rings. Do not use silicone grease; this can affect sterilisation results. Carry out this check as follows: Procedure 1. Check the vessel seal (O-ring) for damage and on correct seat: it must rest firmly on the slightly prominent vessel rim within the metal ring. Place it correctly as necessary. Labfors 5 - Version for Microorganisms Before Cultivation 04 September 2019 Page 115 of 216 2. Ensure that each mounting part is equipped with an intact Oring: check that O-rings are correctly positioned and undamaged. Replace or reposition, if necessary. The figure to the left shows a blanking plug with fixed O-ring for mounting into a 19 mm port with mounted threaded adapter. If mounting parts have to be fitted into other mounting parts (clamping adapter), an O-ring must be placed between them, too. INFORMATION Septum collars are sealed with a septum. No O-ring is used! 8.1.3 Mounting the Impellers To mount the impellers to the stirrer shaft, proceed as follows: Procedure 1. Slide the impeller onto the stirrer shaft. 2. Set the desired height. 3. Tighten the grub screws on the impeller with the Allen key. INFORMATION To avoid unnecessary foam formation, do not fit the impeller at the same height as the surface of the medium. 8.1.4 Mounting Dip Tubes and Spargers Straight spargers and dip tubes can be mounted to the outside of the vessel top plate. Curved spargers and dip tubes can only be mounted to the inside of the vessel top plate. Mounting to the inside of the vessel top plate is described here. This means that the vessel top plate is still removed. Labfors 5 - Version for Microorganisms Before Cultivation Page 116 of 216 04 September 2019 During mounting, ensure that the sparger or the dip tube does not come into contact with other mounting parts (stirrer). The sparger is positioned below the stirrer shaft. Proceed as follows: Procedure 1. Equip the clamping adapter with an O-ring and screw it into the port in the vessel top plate by hand. 2. Loosen the slotted screw head at the clamping adapter. 3. Insert the sparger/dip tube into the clamping adapter from be- low. 4. Set the desired mounting depth. 5. Tighten the slotted screw. Labfors 5 - Version for Microorganisms Before Cultivation 04 September 2019 Page 117 of 216 8.1.5 Mounting the Baffles To mount the baffles, proceed as follows: Procedure 1. Fit an O-ring to the baffles. 2. Fix the baffles with their hexalobular screws. 3. Ensure that the baffles are aligned correctly, so that they fit into the glass vessel (round bottom!). Labfors 5 - Version for Microorganisms Before Cultivation Page 118 of 216 04 September 2019 8.1.6 Moistening/Filling the Culture Vessel If in the culture vessel is to be autoclaved with the medium, the vessel can be filled before the top plate is put in position and the additional component parts are mounted. Note the following about filling the culture vessel before autoclav- ing:  Before autoclaving, only top up with heat-resistant media.  During autoclaving, evaporation may result in a loss of volume and thus to increased salt concentration in the medium. If necessary, top up with sterile water. INFORMATION Development of steam is not possible when autoclaving an empty and dry culture vessel. Successful sterilisation is not guaranteed. Ensure that there is liquid in the culture vessel (approx. 10 mL of water per litre of total volume). Labfors 5 - Version for Microorganisms Before Cultivation 04 September 2019 Page 119 of 216 8.1.7 Fitting the Vessel Top Plate Proceed as follows to fit and fix the vessel top plate: Procedure 1. Place the top plate carefully and with the correct alignment into position. ATTENTION If the knurled nuts on the top plate are tightened too much, the glass vessel may be damaged. Only tighten the nuts by hand! It is imperative that they are not tightened with a tool under any circumstances. 2. Fix the knurled nuts by hand (no tools!) as follows: Labfors 5 - Version for Microorganisms Before Cultivation Page 120 of 216 04 September 2019 a) Slightly screw down all four knurled nuts with two fingers until the nuts are touching the top plate. Do not tighten them at this stage! b) Tighten two nuts opposite each other (1) and (2) by 45° each (1/8 of a turn). c) Afterwards, tighten the other two nuts (3) and (4) by 45° too (1/8 of a turn). 3. Repeat steps b) and c) two times. At the end, all knurled nuts are tightened by 135°. 8.1.8 Mounting a Threaded Adapter To mount a threaded adapter into a 19 mm port, proceed as fol- lows: Procedure 1. Position the threaded adapter correctly aligned on the 19 mm port. 2. Fix it with the two slotted screws. 2 1 4 3 Labfors 5 - Version for Microorganisms Before Cultivation 04 September 2019 Page 121 of 216 8.1.9 Mounting the Blanking Plugs For mounting the different blanking plugs, proceed as follows: Ø 10 mm Ports Procedure 1. Insert the blanking plug with fixed O-ring into the port. 2. Fix it with both slotted screws. Ø 12 mm ports Procedure 1. Insert an O-ring into the port. 2. Insert the blanking plug and screw it in by hand. 3. Tighten it with the hexagon socket spanner hand tight. Labfors 5 - Version for Microorganisms Before Cultivation Page 122 of 216 04 September 2019 Ø 19 mm ports Procedure 1. Screw the blanking plug with fixed O-ring into the threaded adapter. 2. Tighten it by hand. 8.1.10 Mounting Addition Port Adapters Proceed as follows for mounting: Addition port adapter, quadruple, for Ø 19 mm port Procedure 1. Insert the addition port adapter with fixed O-ring into the port. 2. Fix it with the two slotted screws. Labfors 5 - Version for Microorganisms Before Cultivation 04 September 2019 Page 123 of 216 INFORMATION The mounting procedure of further models of addition port adapters (refer to main chapter “Accessories” for details) is the same as for blanking plugs into their corresponding ports. That is why their mounting is not repeatedly described here. 8.1.11 Mounting the Pocket for Temperature Sensor (Pt100) Proceed as follows: Procedure 1. Insert the pocket with the fixed O-ring into the 10 mm port. 2. Fix it with both slotted screws. 8.1.12 Preparing an Inoculation Needle To prepare an inoculation needle for later inoculation, proceed as follows: Procedure 1. Remove protective caps from the inoculation needle. 2. Keep the septum collar ready for use. Labfors 5 - Version for Microorganisms Before Cultivation Page 124 of 216 04 September 2019 3. Connect the inoculation needle via silicone hose with an appropriate container for the inoculum. 4. Put the inoculation needle in a sterile cover or wrap it up in some aluminium foil. 8.1.13 Preparing the Dip Tube/Addition Port Adapter for Inoculation If later inoculation is to be carried out by means of a dip tube or addition port adapter, proceed as follows: Procedure 1. Fit the dip tube with the clamping adapter or addition port adapter in the port. 2. Place a piece of silicon hose onto the dip tube/addition port adapter. 3. Equip the hose for a sterile hose connection. (Depending on the application: rapid coupling, sterile connector or weldable hose with sterile filter). 4. Secure the hose transition points with cable ties. 8.1.14 Equipping Port with Septum Collar and Septum for Inoculation A port in the vessel top plate must be appropriately prepared for later inoculation. For this, it must be equipped with septum collar and septum. Proceed as follows: Labfors 5 - Version for Microorganisms Before Cultivation 04 September 2019 Page 125 of 216 Procedure 1. Ensure that the port is not equipped with an O-ring, otherwise remove it. 2. Insert the septum into the port. 3. Screw the septum collar into the port by hand. INFORMATION If inoculation shall take place via 19 mm port, then mount a threaded adapter, first. The picture to the left shows the procedure with a 12 mm / Pg13.5 port. 4. Ensure the blanking plug is equipped with an O-ring, fit one, if necessary. 5. Screw the blanking plug into the septum collar by hand. If necessary, tighten it with the hexagon socket spanner hand- tight. 8.1.15 Mounting and Preparing the Exit Gas Cooler To mount the exit gas cooler and prepare it for the autoclave, proceed as follows: Procedure 1. Mounting the exit gas cooler into the port in the vessel top plate: a) Mounting in Ø 12 mm / Pg13.5 port Fit an O-ring to the thread of the exit gas cooler or fit it into the 12 mm / Pg13.5 port and screw the exit gas cooler into the port by hand. OR b) Mounting in Ø 19 mm port Equip the 19 mm port with a threaded adapter first and then screw the exit gas cooler (with fixed O-ring) into the threaded adapter by hand. Labfors 5 - Version for Microorganisms Before Cultivation Page 126 of 216 04 September 2019 2. Align the exit gas cooler to ensure that handling of other mounting parts is impaired as little as possible. 3. Check to ensure that the exit gas filter is fitted securely. 4. Cap the exit gas filter loosely with a little aluminium foil. INFORMATION A humidifier bottle with antifoam reagent can be installed between exit gas cooler and the exit gas filter if significant foam formation is expected. Take the following into account for autoclaving:  Only use a new, clean and dry exit gas filter and fix it in such a way that it cannot slip.  ALWAYS keep the exit gas line - hose at the exit gas cooler with secured exit gas filter - open. CAUTION If pressure equalisation does not take place via a top plate opening or the mounted exit gas cooler, overpressure or vacuum in the culture vessel may occur during autoclaving. Connecting the hoses for the exit gas cooler to the basic unit In order to connect the exit gas cooler to the water supply on the basic unit after autoclaving the culture vessel, the pressure hoses must be prepared as follows: Procedure 1. Plug the pressure hoses for water inlet and outlet according the symbols on the basic unit into their connectors. If no exit gas cooler is used, close the manual control valve for water flow on the basic unit or close the hose connectors with the plugs provided with the equipment. Labfors 5 - Version for Microorganisms Before Cultivation 04 September 2019 Page 127 of 216 8.1.16 Mounting the Cold Finger If applicable, i.e. temperature control system with electrical heater pad and cold finger is present (system for single walled culture vessels), the cold finger must be mounted in its appropriate port in the vessel top plate. Proceed as follows: Mounting in Ø 12 mm / Pg13.5 port Procedure 1. Equip the 12 mm / Pg13.5 port with an O-ring. 2. Insert the cold finger into the port and tighten it by hand. 3. Adjust its mounting depth as desired. Mounting in Ø 19 mm port Procedure 1. Equip the 19 mm port with a threaded adapter. 2. Insert the cold finger with fixed O-ring into the threaded adapter and tighten it with by hand. 8.1.17 Checking Lubrication of the Mechanical Seal The mechanical seal must be lubricated at any time. Therefore it must be ensured, that the silicone hose on the bottom of the drive hub is always entirely filled with glycerine. Top it up, if necessary. For details refer to chapter „Cleaning and Maintenance“, „Lubricating the Mechanical Seal“. ATTENTION Risk of loss of property due to the mechanical seal running dry! A mechanical seal, which has not been adequately lubricated, is destroyed when running dry. Labfors 5 - Version for Microorganisms Before Cultivation Page 128 of 216 04 September 2019 8.1.18 Preparing the Sensors All sensors that come into contact with the medium are mounted before autoclaving and are sterilised together with the culture ves- sel. Note the following about all sensors:  Mount all sensors by hand – do not use any tools!  Mount the sensors in such a way that they cannot come in contact with other components or the glass vessel.  If the mounting depth of is adjustable (mounting with electrode holder/clamping adaptor), make sure the mounting depth is set correctly prior to autoclaving, as later adjustment represents a contamination risk. pH sensor Calibrate the pH sensor on the operating panel before mounting and autoclaving. pO2 sensor Mount the pO2 sensor in such a way that it has good access to the flow and there is no risk of bubbles collecting. (Calibration is carried out AFTER autoclaving!) pH sensor & pO2 sensor METTLER Cover the sensor heads of METTLER pH sensor and pO2 sensors with aluminium foil during autoclaving. pH sensor & pO2 sensor HAMILTON ATTENTION Risk of damage to the (HAMILTON) pH and pO2 sensor. Covering the sensor heads with aluminium foil during autoclaving may lead to water gathering under the film, thus damage the contacts on the sensor head. pH and pO2 sensor heads should NOT be covered with aluminium foil during autoclaving. For details on the safety, technical data, usage and maintenance requirements for the pH and pO2 sensors, see the separate documentation provided by the manufacturers. Labfors 5 - Version for Microorganisms Before Cultivation 04 September 2019 Page 129 of 216 8.1.18.1 Calibrating the pH Sensor For a reliable pH measurement, a 2-point calibration with an upper and lower reference buffer must be carried out before each cultivation. The pH sensor must be calibrated before autoclaving. This is carried out on the operating panel and described in detail in the separate operating manual of the touch screen software. Procedure 1. Connect the sensor cable. (For more details, see the chapter “Connecting the pH Sensor”). 2. Switch on the equipment using the main switch. The operating panel is switched on automatically and the system is started. 3. Calibrate the pH sensor in accordance with the detailed description in the operating manual of the touch screen software. 8.1.18.2 Mouting a Sensor into a 12 mm Port Depending on sensor length and vessel volume, sensors can be directly screwed into 12 mm / Pg13.5 ports. To do so, proceed as follows: Procedure 1. Slide the O-ring onto the sensor. 2. Insert the sensor into the port. 3. Screw the sensor on its thread into the port by hand. Labfors 5 - Version for Microorganisms Before Cultivation Page 130 of 216 04 September 2019 8.1.18.3 Mounting Sensors with Electrode Holder To enable adjusting the mounting depth of a sensor in a 12 mm/Pg13.5 port, an electrode holder must be used for mount- ing. Proceed as follows: Procedure 1. On the electrode holder, lightly loosen the grub screw in the support guide with the key. 2. Pull the support guide from the guide bar. 3. Insert the sensor into the support guide and tighten it. Labfors 5 - Version for Microorganisms Before Cultivation 04 September 2019 Page 131 of 216 4. Insert the sensor into the hollow screw with the thread pointing in the downward direction. 5. Fit the fork of the guide bar into the groove of the hollow screw. 6. Push the hollow screw and the guide bar together upwards and insert the guide bar into the hole of the support guide. 7. Slide the O-ring onto the sensor and insert the sensor into the port. 8. Adjust the sensor to the desired height. Labfors 5 - Version for Microorganisms Before Cultivation Page 132 of 216 04 September 2019 9. Screw the sensor on the hollow screw into the port and tighten it. 10. Tighten the grub screw in the support guide with the key. 8.1.18.4 Mounting the Antifoam Sensor Note the following points for mounting:  The antifoam sensor is equipped with transparent insulation that must be intact, as otherwise a continuous signal „Foam/liquid detected“ may be generated. ATTENTION If the sensor is fixed too tightly in the clamping adapter, or the mounting depth of the sensor is changed while the screw on the clamping adapter is tightened, the sensor insulation may be damaged.  The sensor head must not touch the clamping adaptor, otherwise a continuous short-circuit is generated, indicating “Foam/liquid detected”.  The clamping adapter on the sensor must be equipped with an intact O-ring. Labfors 5 - Version for Microorganisms Before Cultivation 04 September 2019 Page 133 of 216 Mounting in 10 mm port Proceed as follows: Procedure 1. Remove the protective cap from the sensor. 2. Ensure the clamping adapter is equipped with an O-ring, fit one, if necessary. 3. Insert the sensor into the port. 4. Fix the clamping adapter with the two slotted screws. 5. Loosen the slotted screw at the clamping adapter. 6. Set the desired mounting depth of the sensor carefully. 7. Tighten the slotted screw carefully. Labfors 5 - Version for Microorganisms Before Cultivation Page 134 of 216 04 September 2019 Mounting in 12 mm / Pg13.5 or 19 mm port Proceed as follows: Procedure 1. Remove the protective cap from the sensor. Depending on type of clamping adapter: 2. Ensure the clamping adapter or port is equipped with an Oring, fit/place one, if necessary. If 19 mm port: mount a threaded adapter. 3. Insert the sensor into the port and screw it in by hand. 4. Loosen the hollow screw on the clamping adapter. 5. Set the desired mounting depth of the sensor carefully. 6. Tighten the hollow screw on the clamping adapter carefully by hand. Labfors 5 - Version for Microorganisms Before Cultivation 04 September 2019 Page 135 of 216 8.1.19 Preparing the Super Safe Sampler INFORMATION The following figures are for general purposes of comprehen- sion. In order to prepare the Super Safe Sampler sampling system for autoclaving, proceed as follows: Procedure 1. Attach the hose of the valve group on the dip tube. 2. Secure the hose with a cable tie. 3. Tighten the sample valve carefully by hand in a clockwise di- rection. This ensures that the non-return valve/sample valve screw connection is tight. Labfors 5 - Version for Microorganisms Before Cultivation Page 136 of 216 04 September 2019 4. Turn the sterile filter carefully by hand in a clockwise direction. This ensures that the non-return valve/sterile filter screw connection is tight. 5. Cover the valve group loosely with aluminium foil. 6. Clamp off the hose on the dip tube. 8.1.20 Mounting the Sparger Hose and the Inlet Air Filter The sparger must be equipped with the hose and inlet air filter before autoclaving. To do so, proceed as follows: Procedure 1. Cut a short piece of silicone hose. 2. Place the inlet air filter, marked in red, onto the hose end in the direction of the air flow. The nozzle with the red INLET marking remains exposed. Labfors 5 - Version for Microorganisms Before Cultivation 04 September 2019 Page 137 of 216 3. Place the silicone hose onto the sparger. 4. Secure the ends of the hose with the cable tie. 5. Clamp off the silicon hose with a hose clamp. 6. Lightly cap the inlet air filter with aluminium foil. 8.1.21 Preparing the Gassing Hose Line on the Basic Unit An appropriate hose line must be prepared on the basic unit in order to connect the sparger to the gassing after autoclaving. Proceed as follows: Procedure 1. Cut a piece of thick walled silicone hose (Ø 5 x 8 mm, in the starter kit). Choose its length so that the hose connection between sparger and gas supply on the basic unit does not have any tension or kinks. 2. Fit the hose to the hose nozzle of the gassing unit(s) on the basic unit. 3. Secure the hose with cable ties. Labfors 5 - Version for Microorganisms Before Cultivation Page 138 of 216 04 September 2019 8.1.22 Calibrating the Pumps Not calibrated pumps show the duration in seconds and the number of rotations. If the delivered volume (in mL) shall be indicated instead, the pumps must be calibrated before autoclaving. A detailed description of how to calibrate the pumps can be found in the separate operating manual of the touch screen software. 8.1.23 Preparing the Reagent Bottles, Pumps and Hoses ATTENTION Damaged hoses and/or clogged filters may lead to undesired pressure conditions in the reagent bottles. – Ensure each reagent bottle is equipped with an open pressure equalisation line with a clean and dry filter. – Only use clean, intact hoses and ensure they are firmly at- tached. The following sections contain a detailed description of how reagent bottles are equipped properly and connected to the pumps and culture vessel. Connecting the reagent bottles to the pumps and culture ves- sel Proceed as follows: Procedure 1. Cut two long silicone hoses with appropriate diameter (refer to table with hoses in chapter „Accessories“, „Reagent Bottles“)per pump/reagent bottle. INFORMATION The length of the silicone hoses must be selected to ensure that the hose connections between the reagent bottles, pumps and culture vessel do not have any tensions or kinks. 2. Thoroughly rinse the silicone hoses with distilled water. Labfors 5 - Version for Microorganisms Before Cultivation 04 September 2019 Page 139 of 216 3. Connect the silicone hoses and pump hoses of the pump heads with hose connectors. INFORMATION Note that the direction of rotation of the pumps is clockwise in operational state. 4. Secure with cable ties. Connection between pumps and culture vessel Proceed as follows: Procedure 1. Fit silicone hoses for base, acid and feed to the addition port adapter(s) and secure them with cable ties. 2. Attach the silicone hose of the antifoam pump to the mounted antifoam sensor in the culture vessel and secure it with a cable tie. Connection between reagent bottles and pumps Proceed as follows: Procedure 1. Ensure that a hose is fitted inside the reagent bottles at the exposed hose connection (without sterile filter); fit one if not: a) the end of the hose does not touch the bottom of the bottle, otherwise the hose may get sucked against the bottom and no longer be able to pump liquid. b) the end of the hose is cut diagonally. In this case the hose end can touch the bottom of the bottle. 2. Label the reagent bottles in accordance with their content. Labfors 5 - Version for Microorganisms Before Cultivation Page 140 of 216 04 September 2019 3. Depending on the application: Fill the reagent bottles with reagents and reclose them with their lid. ATTENTION Usage of the highly corrosive hydrochloric acid HCl as reagent leads to damage to components made of stainless steel such as e.g. component parts or the top plate. Use only non-corrosive acids, e.g. phosphoric acid, instead. INFORMATION Fill reagent bottles with heat-resistant reagents only. Sterilise non-heat-resistant feed solution separately and only transfer it to the reagent bottle after sterilising. 4. Place the reagent bottles in reagent bottle and pump holders. 5. Attach the correct silicone hoses to available hose connections of each reagent bottle and secure them with cable ties. 6. Close silicone hoses with clamps as close as possible to the hose connections of the reagent bottles to ensure that no reagent can flow into the culture vessel. 7. Ensure that:  each reagent bottle is connected with the appropriate pump according to its contents. (Base to base pump, etc.).  filters are clean and dry; short hose line is open. 8. Cap the filter loosely with aluminium foil. Labfors 5 - Version for Microorganisms Before Cultivation 04 September 2019 Page 141 of 216 8.1.24 Sterile Hose Connections If additional vessels are needed and these can only be connected to the culture vessel after autoclaving, such as vessels for the inoculum or bottles for sampling etc., rapid couplings (male/female), sterile connectors or – if weldable hoses are used – a hose welding device can be used to form a sterile connection. The connection pieces must be fitted to the appropriate hoses before autoclaving. Rapid couplings are connected after autoclaving in a sterile workbench. Sterile connectors and hose welding devices allow sterile connecting without a sterile workbench. 8.1.25 Removing the Pump Heads To remove the pump heads from the basic unit, proceed as fol- lows: Procedure 1. Pull up the pump cover plate and remove it from the holder. INFORMATION The pump cover plate serves only as a protective device in operating mode. It is not heat proof and therefore it may not be au- toclaved. 2. Pull the mounting plate with the pump heads off the drive shafts by the two handles. Labfors 5 - Version for Microorganisms Before Cultivation Page 142 of 216 04 September 2019 3. Fit the mounting plate with the pump heads onto the reagent bottle and pump holder. 8.1.26 Checklist Before Autoclaving Check and ensure the following items before autoclaving: Culture vessel All necessary O-rings are fitted. All unused ports are closed with blanking plugs Port for inoculation is equipped with septum, septum collar and blanking plug There is liquid in the culture vessel (autoclavable medium or approx. 10 mL water per litre working volume). Mechanical seal is lubricated. Drive hub is lightly capped with aluminium foil. Double walled culture vessels: The water overflow hose of the vessel jacket is open, it is NOT kinked or clamped off. Reagent bottles, hoses and pumps Reagent bottles are exclusively filled with autoclavable reagents, correctly labelled and connected with the culture vessel and the pump heads via hoses. Reagent bottles are equipped with sterile filters for pressure equalisation Reagent bottles are placed in reagent bottle and pump holder and pump heads are placed on the holder with a mounting plate. Inoculation needles Inoculation needle is connected to appropriate container for inoculum with a silicone hose. Inoculation needle is packed in sterile cover or wrapped in aluminium foil. Labfors 5 - Version for Microorganisms Before Cultivation 04 September 2019 Page 143 of 216 Super Safe Sampler The valve group is connected to the dip tube in the culture vessel by means of a hose. The valve group is lightly capped with aluminium foil. Sparger & exit gas cooler The sparger is equipped with a hose and an inlet air filter. The exit gas cooler is equipped with a new securely fastened exit gas filter. Filters & hoses All filters are clean, dry and lightly capped with aluminium foil. There are no open hose ends. All hose transition points are secured with an autoclavable cable tie or hose clamp to prevent them from slipping. Hoses on the reagent bottles, for sampling and the gassing system (sparger) are clamped off with hose clamps. The exit gas hose is NOT clamped off. The hoses are undamaged; the hose lines show no kinks and are not able to kink. Sensors All sensors required are mounted and, if necessary, calibrated. The antifoam sensor is mounted, set for the correct mounting depth and connected to the correct reagent bottle. The temperature sensor of the autoclave is inserted into the pocket for the temperature sensor of the culture vessel. pH and pO2 sensors: METTLER (analogue): are covered with aluminium foil. HAMILTON: are NOT covered with aluminium foil. 8.1.27 Autoclaving Before cultivation starts, the culture vessel is autoclaved in accordance with the application in question. The culture vessel can be autoclaved with or without medium. Adhere to the following:  Never autoclave the culture vessel dry; see also the chapter “Moistening/Filling the Culture Vessel”. Labfors 5 - Version for Microorganisms Before Cultivation Page 144 of 216 04 September 2019 INFORMATION Development of steam is not possible when autoclaving a completely empty and dry culture vessel. Successful sterilisation is not guaranteed. Ensure that there is liquid in the culture vessel (approx. 10 mL of water per litre of total volume).  If necessary, pump off any remaining water after autoclaving by means of the dip tube.  Sterilise all liquid, heat-instable components separately and add them after autoclaving.  If the medium is autoclaved in the culture vessel, you may then need to add sterile water to make up the volume. When transporting the culture vessel to/from the autoclave, note the following:  Always transport the culture vessel in the vessel holder.  Always transport the culture vessel to/from the autoclave in pairs and use suitable auxiliary equipment when transporting the culture vessel. WARNING Depending on the design, accessories and fill level, the culture vessel may be too heavy to be carried by one person alone. Proceed as follows to autoclave the culture vessel: Procedure 1. Place the culture vessel into the autoclave. 2. Ensure that the culture vessel and the accessories do not touch the inner wall of the autoclave. 3. Ensure that the exit gas filter is open. 4. Insert the temperature sensor of the autoclave into the pocket for the temperature sensor. 5. Select the program for liquids. 6. Autoclave the culture vessel in accordance with the operating manual of the autoclave manufacturer. Labfors 5 - Version for Microorganisms Before Cultivation 04 September 2019 Page 145 of 216 8.2 Connecting the Culture Vessel and Preparing the Cultivation As soon as the culture vessel with the accessories has cooled sufficiently, the various cable and tube connections between the basic unit and the culture vessel can be established. Depending on the present temperature control system, the following steps in the procedure are to be carried out:  Double walled vessels: connect the vessel jacket to the water connectors on the basic unit.  Single walled vessels: connect the cold finger to the water connectors on the basic unit, fit the heater pad around the vessel and connect it to the basic unit. 8.2.1 Connecting a Single Walled Vessel To connect a single walled vessel to the basic unit, proceed as fol- lows: Connecting the cold finger Procedure 1. Push the pressure hoses for water inlet and outlet (on cold finger and on basic unit) together via the rapid couplings. 2. Ensure the cold finger is connected to the basic unit as fol- lows:  Lower hose with connector: outlet of the cold finger  Upper hose with connector: outlet of the cold finger Labfors 5 - Version for Microorganisms Before Cultivation Page 146 of 216 04 September 2019 Connecting the heater pad Before doing so, check the heater pad and the cable connection for damages (cracks, breaks/kinks etc.) and ensure the pad is dry. Dry or replace the heater pad, if necessary. DANGER Danger of fatal electric shock! The use of a damaged or wet/damp heater pad can be perilous! Procedure 1. Fit the heater pad around the culture vessel and hook the tensioning springs. 2. Plug the cable into the connector labelled Heating mat. 8.2.2 Connecting a Double Walled Culture Vessel To connect a double walled vessel to the basic unit, the hoses for water inlet, outlet and overflow of the vessel jacket must be plugged into the appropriate connectors on the basic unit according the symbols. INFORMATION In order that the hose connections can be disconnected easily after cultivation, slightly moisten the rapid couplings on the silicon hoses prior to connecting them. Labfors 5 - Version for Microorganisms Before Cultivation 04 September 2019 Page 147 of 216 8.2.3 Mounting the Pump Heads To mount the pump heads to the basic unit, proceed as follows: Procedure 1. Pull off the mounting plate with the pump heads from the reagent bottle and pump holder. 2. Plug the mounting plate with the pump heads onto the pump motor drive shafts. 3. Insert the cover plate into the support. Labfors 5 - Version for Microorganisms Before Cultivation Page 148 of 216 04 September 2019 8.2.4 Filling the Reagent Hoses In order to prepare the reagent hoses for operation, they must be filled with reagent using the rocker switches on the basic unit. WARNING When using heavily corrosive reagents (acids and bases), it is particularly important only to use suitable and undamaged hoses. They must also be securely fastened. Furthermore, the exit gas filter must not be blocked. This ensures that no pressure builds up and no reagent escapes due to burst hoses. Observe the following points:  Remove the clamps from the reagent hoses, before filling.  Ensure that no reagent escapes into the culture vessel, if pos- sible. Proceed as follows for filling: Procedure 1. Switch the equipment on at the main switch. 2. Open the clamps on the reagent hoses. 3. Fill the reagent hoses individually, successively and manually via the appropriate rocker switch. Observe the following points:  Press the rocker switch to the right side: the pump runs forward (clockwise), reagent is sucked in from the reagent bottle and is pumped in the direction of the vessel.  Press the rocker switch to the left side: the pump runs backwards (counter clockwise), reagent is liquid is sucked in from the culture vessel and is pumped in the direction of the reagent bottle. Release the rocker switch just before liquid visibly enters the addition port adapter(s), respectively the dosing needle of the antifoam sensor so that no corrective reagent can enter the culture vessel. It is acceptable and time-saving to fill the reagent hoses automatically and simultaneously when operating several bioreactors. For details refer to the separate operating manual of the touch screen software. Labfors 5 - Version for Microorganisms Before Cultivation 04 September 2019 Page 149 of 216 8.2.5 Connecting the Gassing To connect the sparger to the gassing, proceed as follows: Procedure 1. Remove the aluminium foil from the inlet air filter. 2. Insert the gassing hose of the basic unit to the inlet air filter of the sparger and secure it with a cable tie. 3. Remove the hose clamp. 8.2.6 Connecting the Exit Gas Cooler To connect the exit gas cooler to the basic unit, proceed as follows: Procedure 1. Remove the aluminium foil from the exit gas filter. 2. Push the pressure hoses of the exit gas cooler and the basic unit together via the rapid couplings according the symbols on the basic unit. Labfors 5 - Version for Microorganisms Before Cultivation Page 150 of 216 04 September 2019 3. If necessary, adjust the standard setting for water flow on the manual control valve on the basic unit. The exit gas cooler only works with activated temperature control (Parameter Temperature switched ON). If no exit gas cooler is used, close the manual control valve for water flow on the basic unit or close the hose connectors with the plugs provided with the equipment. 8.2.7 Coupling the Motor For routine operation, it is not necessary to plug in and unplug the motor cable. The motor connected during installation is only coupled before cultivation. Proceed as follows: Procedure 1. Place the motor onto the drive hub with the groove aligned with the pin on the drive hub. The motor is held in its position. 8.2.8 Filling the Culture Vessel Depending on the application, the vessel can be filled after autoclaving. To prevent foam formation during filling, add the medium via a dip tube. To do so, proceed as follows: Procedure 1. Sterilise the medium separately. Labfors 5 - Version for Microorganisms Before Cultivation 04 September 2019 Page 151 of 216 2. If necessary, pump off any water that remains in the culture vessel. 3. Establish a sterile hose connection between the culture vessel and the medium container. 4. Pump the desired quantity of medium into the culture vessel. 5. Clamp off the medium hose; if necessary, apply a welded seal. 6. Disconnect the medium container from the culture vessel; if necessary, retain it as a harvest or waste container. INFORMATION If the stirrer is turning on the surface of the medium, foam will be formed. For this reason, only switch on the stirrer if it is fully covered by medium. 8.2.9 Inserting the Temperature Sensor (Pt100) into the Pocket The temperature sensor is not in direct contact with the medium. Procedure 1. Simply insert the sensor into the pocket in the vessel top plate as far as it will go. CAUTION Risk of burns and loss of property due to overheating of the temperature control circuit! The temperature control circuit will overheat without an inserted temperature sensor and without liquid in the vessel. This can lead to burns and loss of property. Labfors 5 - Version for Microorganisms Before Cultivation Page 152 of 216 04 September 2019 8.2.10 Connecting the Antifoam Sensor To connect the antifoam sensor, the two banana connectors of the sensor cable must be inserted as follows: Procedure 1. Insert the black banana plug into the earth connection in the top plate. 2. Insert the red banana plug into the connector on the sensor head. 8.2.11 Connecting the pH Sensor To connect the pH sensor, proceed as follows: METTLER pH sensor Procedure 1. Remove the protective cap or the aluminium foil from the sen- sor. Labfors 5 - Version for Microorganisms Before Cultivation 04 September 2019 Page 153 of 216 2. Place the connector of the sensor cable onto the sensor head. 3. Turn the ring on the connector and screw it tight. Ensure the sensor cable is not twisted or buckled. ATTENTION The cable shield can be damaged by buckling or twisting. This may lead to faulty measurements. HAMILTON pH sensor Procedure 1. Remove the red protective cap from the sensor cable. 2. Place the connector of the sensor cable onto the sensor head and screw into place. Ensure the cable is not twisted or buckled. Labfors 5 - Version for Microorganisms Before Cultivation Page 154 of 216 04 September 2019 8.2.12 Connecting the pO2 Sensor To connect the pO2 sensor, proceed as follows: METTLER pO2 sensor Procedure 1. Align the orange mark on the connector of the sensor cable with the orange mark on the sensor. 2. Plug the cable connector onto the sensor. 3. Turn the bajonet cap of the sensor cable clockwise und push gently towards the sensor. Ensure the cable is not twisted or buckled. ATTENTION The cable shield can be damaged by buckling or twisting. This may lead to faulty measurements. Labfors 5 - Version for Microorganisms Before Cultivation 04 September 2019 Page 155 of 216 HAMILTON pO2 sensor To connect the HAMILTON pO2 sensor, proceed in the same manner as with the HAMILTON pH sensor; see the chapter “Connecting the pH Sensor”, section “HAMILTON pH sensor”. 8.2.13 Polarising the pO2 Sensor (METTLER) Polarographic pO2 sensors must be polarised at initial operation or after disconnection from the voltage source. Correct calibration is not possible otherwise. This means that the pO2 sensor must be polarised before calibration. For polarisation, the sensor cable must simply be connected to the pO2 sensor and the equipment must be switched on at the main switch. Duration of polarisation (= polarisation time) depends on how long the pO2 sensor has been disconnected from the voltage source (= depolarisation time) As a general rule: if depolarisation time > 30 minutes, the minimum polarisation time is 360 minutes. More details about polarisation can be found in the separate documentation form the manufacturer METTLER TOLEDO. 8.2.14 Calibrating the pO2 Sensor Generally speaking, the following applies: The pO2 sensor should be calibrated after autoclaving has been performed because the sterilisation process may change the steepness of the pO2 sensor. As a rule, a 1-point calibration to 100 % is usually sufficient for exact measurement, and should be carried out before each cultiva- tion. Depending on the specifications defined by the user, the pO2 sensor is calibrated either before the vessel is filled with medium or afterwards, in the prepared medium. A detailed description on the calibration can be found in the separate operating manual of the touch screen software. 8.2.15 Checking the Hoses and Hose Connections Check and ensure the following items before each cultivation:  Hoses show no kinks and are not able to kink.  Hoses are undamaged and show no weaknesses.  Gas hoses and connections do not show any leaks. Labfors 5 - Version for Microorganisms Before Cultivation Page 156 of 216 04 September 2019  Hose lines are as short as possible.  Hoses are secured with cable ties and/or hose clamps.  Only the pressure hoses supplied by the equipment manufacturer are connected as supply lines (water, gas) between the in-house connections and the equipment. The following sections describe the work necessary for the performance of and after the completion of a cultivation, before the culture vessel with accessories is thoroughly cleaned and then prepared for another cultivation. This essentially comprises:  Preparing the medium, starting the bioreactor  Sampling  Inoculation  Harvest  Stopping the bioreactor, if necessary emptying the vessel  Autoclaving the culture vessel and accessories The requirement for the first item is that the culture vessel and accessories are autoclaved, cooled and connected to the basic unit. All cable and hose connections between the equipment and the culture vessel, including the reagent bottles, are present, pump heads are mounted and the reagent hoses are filled. Depending on the user specifications, the pO2 sensor is already calibrated. 8.3 Preparing the Medium Before the first sampling, which usually takes place as a ‘zero sample’ before inoculation, and before the inoculation itself, the medium must be warmed to the desired temperature. If necessary, the pO2 concentration and the pH are set. The time required for this depends on the working volume. Set and activate the desired setpoint of the parameters in question on the operating panel, and start the bioreactor. Depending on the specifications defined by the user, the pO2 sensor is calibrated either before the vessel is filled with medium or afterwards, in the prepared medium. Labfors 5 - Version for Microorganisms Before Cultivation 04 September 2019 Page 157 of 216 CAUTION If pressure equalisation does not take place via a top plate opening or the mounted exit gas cooler, overpressure in the culture vessel may occur during cultivation as a result of warming, gassing or fermentation processes. – Exit gas line - hose at the exit gas cooler with secured exit gas filter - ALWAYS keep open. – Only use clean and dry exit gas filters. For details about calibration and operation in general refer to the separate operating manual of the touch screen software. 8.4 Sampling Samples are taken from the culture vessel to gain material for offline analysis. The method of sampling can vary due to the different analyses carried out by the operator. The sampling procedure using the standard sampling system, Super Safe Sampler, is described below. Before starting, observe the following: WARNING Culture solution could emerge from the vessel if the sample valve mechanically fails. This could lead to serious health risks in the event of applications with pathogenic organisms. – When working with pathogenic organisms, always additionally clamp off the sampling hose with a metal (!) clamp. – Only remove the clamp when sampling. – Reattach the clamp before removing the syringe from the sample valve. WARNING Loose screws at components could lead to the penetration of unsterile air or contamination of the environment. Before and after autoclaving: Check that all screws are tightly screwed in and, if necessary, tighten them manually. Labfors 5 - Version for Microorganisms Before Cultivation Page 158 of 216 04 September 2019 If the sample is to be further aseptically processed, use a sterile syringe and sterile closing caps. For details, see the main chapter “Accessories” chapter “Sampling System Super Safe Sampler”, section “Aseptic Sampling”. Proceed as follows: Procedure 1. Check that all screw connections of the valve group are tightly screwed in. If necessary, gently tighten the screw connections with two fingers. 2. Remove the clamp from the sampling hose. 3. If present: Remove the closing caps. 4. If desired: Disinfect the sample valve. 5. Screw open the Luer-Lock syringe on the sample valve. 6. Pull back the syringe plunger to remove the desired sampling volume. If the dip tube was rinsed with air, air is sucked in first. Remove it as follows: a) Unscrew the syringe from the valve. Labfors 5 - Version for Microorganisms Before Cultivation 04 September 2019 Page 159 of 216 b) Hold the syringe with the plunger downwards so that the medium remains in the syringe. c) Push the air out of the syringe. d. Screw the syringe onto the sample valve. e) Draw in again. 7. Attach the clamp to the sampling hose. Rinsing the dip tube with sterile air The dip tube and its sampling hose can be filled with sterile air after taking a sample. INFORMATION Only use a clean and dry syringe to avoid blocking the sterile filter. This syringe can be reused as often as desired, since air is provided via a sterile filter. To do so, proceed as follows: Procedure: 1. Insert the syringe onto the hose at the sterile filter and push air through. The remaining liquid in the hose and in the dip tube is pushed back into the vessel. 2. Remove the syringe from the sterile filter to fill it with air again. 3. Repeat steps 1 and 2 as many times as necessary until bubbles rise out of the dip tube. Labfors 5 - Version for Microorganisms Before Cultivation Page 160 of 216 04 September 2019 Removing residual fluid To remove residual fluid from the system, proceed as follows: Procedure: 1. Hold the syringe with sample downwards, pull back the plunger. This removes all but a few µL of the residual fluid. 2. Hold the sample valve with one hand; unscrew the syringe with the other. 3. If desired: Place the closing caps on the sample valve and on the syringe with the sample. 8.5 Inoculation Check and ensure the following items before inoculation:  Medium has been filled.  Heat-labile, separately sterilised substances are present.  The reagent bottles are connected with the pumps and the culture vessel, and are filled with a sufficient amount of reagents and feed solution for the duration of the entire cultivation process.  The hoses of the reagent bottles are filled. Labfors 5 - Version for Microorganisms Before Cultivation 04 September 2019 Page 161 of 216  The correct operating temperature has been reached.  The required stirring speed is set.  The sensors are calibrated and the control is correct (or not yet switched on).  All clamps have been removed (except for sampling system).  Utensils for the inoculation and vessels with inoculum are ready. Methods There are a number of ways to add medium or inoculum before and during cultivation:  With inoculation needle and septum collar with septum  In a small volume, with the syringe via the septum  Via an addition port adapter from the reagent bottle (a sterile hose connection is required for this method).  Via a dip tube from the reagent bottle (a sterile hose connection is required for this method). These methods are described in the following sections. The implements for inoculation with inoculation needle are standard accessories for the equipment. This inoculation method is particularly suitable for all vessel sizes of the equipment. If separate addition of several different solutions is necessary, a second inoculation needle can be used with mounted septum collar in a spare port. 8.5.1 Inoculation with Inoculation Needle Proceed as follows for inoculation: Procedure 1. Fill the inoculum under sterile conditions into the prepared container. 2. Unscrew the blanking plug from the septum collar. 3. If appropriate, place a few drops of Ethanol (70 %) on the septum before piercing the septum. If appropriate, briefly flame the septum collar. 4. Remove sterile cover/aluminium foil from the inoculation nee- dle. 5. Briefly flame the inoculation needle. Labfors 5 - Version for Microorganisms Before Cultivation Page 162 of 216 04 September 2019 6. Immediately pierce the inoculation needle through the septum. 7. Screw the inoculation needle into the septum collar. 8. Transfer the desired volume of inoculum into the culture ves- sel. 9. Clamp off the silicone hose. Or: Pull the inoculation needle out and close the septum collar with the blanking plug. But this method is not secure regarding preventing contamination. 8.5.2 Inoculation with a Syringe Proceed as follows for the inoculation: Procedure 1. Fill the syringe with the required amount of inoculum. 2. Unscrew the blanking plug from the septum collar. As a possible additional protection against contamination: Before piercing, drop a few drops of ethanol (70 %) on the septum. 3. Pierce the septum and inject the inoculum. 4. Remove the needle from the septum and close the septum collar with a blanking plug. 8.5.3 Inoculation Using Dip Tube / Addition Port Adapter Proceed as follows during inoculation: Procedure 1. Fill the inoculum under sterile conditions into the prepared container. 2. Create a sterile hose connection with the dip tube/addition port adapter. 3. Transfer the desired volume of inoculum into the culture vessel. Pump it, if necessary. 4. Clamp off the hose with the hose clamp, weld it if necessary. Labfors 5 - Version for Microorganisms Before Cultivation 04 September 2019 Page 163 of 216 8.6 Harvest The culture can be harvested at the end of the cultivation. To prevent possible sedimentation from the culture, the stirrer can be switched on during harvesting. If necessary, activate gassing for sensitive cultures. However, all other parameters should be switched off, provided there are no other specifications for the user. The following possibilities exist for the harvest: a) Transfer to another vessel To transfer the contents of the vessel to another container in a laminar flow cabinet. b) Pump-down via a sterile hose connection To do so, proceed as follows: Procedure 1. Make a sterile connection between the hose at the dip tube for harvest and the new vessel. 2. Connect the hose to one of the pumps on the equipment or to an external pump. 3. Pump the desired amount of culture into the new vessel. INFORMATION Only switch on the stirrer, if it is fully covered by medium, as foam otherwise forms. 4. Switch off all parameters on the operating panel, i.e. stop the bioreactor. ATTENTION Switching the equipment off at the power switch without stopping the bioreactor and shutting down the system on the operating unit beforehand may lead to damage of the operating unit! Labfors 5 - Version for Microorganisms Before Cultivation Page 164 of 216 04 September 2019 8.7 Emptying the Culture Vessel Depending on the user specifications, the culture vessel can be emptied either before or after autoclaving. A previously emptied and culture vessel filled only with water for autoclaving is easier to clean afterwards. For emptying the culture vessel, the same options as for harvesting are available. For more information, see the “Harvest” section. If the culture will not be used further, it must be inactivated according to the current in-house instructions (e.g. by autoclaving or by lowering the pH value), and subsequently disposed of in an environmentally sound manner according to the local regulations. 8.8 Emptying the Reagent Hoses Before autoclaving the culture vessel with accessories, all reagent hoses must be completely emptied using the corresponding pump. This can either be done manually and individually for each pump using the rocker switch on the basic unit or all pumps are emptied time-controlled at once via the operating unit. It is also recommended to thoroughly rinse the hoses with water after emptying and before autoclaving. For details about automatic emptying of all pump hoses refer to the separate operating manual of the touch screen software. 8.9 Switching off the Equipment When the harvest is finished or the culture vessel has been emptied and the reagent hoses are also empty, the equipment can be switched off. Proceed as follows: Procedure 1. Ensure that the bioreactor has been stopped. If necessary, stop it in the touch screen software on the operating panel. 2. Shutdown the system on the operating panel. 3. Switch off the equipment at the power switch. Labfors 5 - Version for Microorganisms Before Cultivation 04 September 2019 Page 165 of 216 ATTENTION Switching the equipment off at the power switch without previously stopping the bioreactor and shutting down the system on the operating panel may lead to damage of the operating panel! 4. Close the supply lines (water, gas). 5. Let the motor cool down. 6. Autoclave the vessel, component parts and accessories as per the user-specific specifications and then clean them. 8.10 Autoclaving the Culture Vessel After Cultivation After emptying the culture vessel and before cleaning, the culture vessel must be autoclaved with all accessories. When doing so, do not autoclave the culture vessel when completely dry and observe the same safety regulations as when autoclaving before cultivation. Before starting, ensure:  There is liquid in the culture vessel (autoclavable medium or approx. 10 mL water per litre working volume).  Reagents and feed solution have been pumped back out of the hoses.  The equipment is switched off.  The motor has cooled down. Proceed as follows to prepare the culture vessel and accessories for autoclaving after cultivation: Procedure 1. Clamp off the hoses of the reagent bottles. 2. Clamp off the hose of the sparger. 3. Remove all cable and hose connections between the basic unit and the culture vessel: a) Uncouple the motor and place it to the side. b) Unplug the sensor cables. c) Pull the temperature sensor out of the pocket. d) If an exit gas cooler is in use: disconnect the water inlet and water outlet hoses from the exit gas cooler. e) Depending on the temperature control system: Labfors 5 - Version for Microorganisms Before Cultivation Page 166 of 216 04 September 2019  disconnect the water inlet / outlet /overflow hoses of the vessel jacket from the basic unit. OR:  disconnect the water inlet and outlet hoses of the cold finger. Unplug the heater pad and remove it from the vessel f) Remove the gassing hose (emerging from basic unit) from the inlet air filter on the sparger. 4. Lightly cover all filters and the drive hub with aluminium foil. ATTENTION HAMILTON pH and pO2 sensors: Do not cover with aluminium foil! 5. Open the pump cover. 6. Remove the mounting plate with pump heads from the drive shafts on the basic unit and place on the pump holder. 7. Check and ensure that the exit gas filter is free and dry and the exit gas hose is OPEN. 8. Insert the temperature sensor of the autoclave into the pocket on the culture vessel and autoclave the culture vessel. Labfors 5 - Version for Microorganisms Cleaning and Maintenance 04 September 2019 Page 167 of 216 9 Cleaning and Maintenance The following sections describe in detail how the culture vessel and accessories and the basic unit are cleaned and, as required, stored. In addition, the section contains a maintenance plan and corresponding descriptions for the procedures to be performed by the operator. 9.1 Cleaning Agent and Disinfectant Intended use Allowed products/tools Culture vessel Water and a non-scratch, non-abrasive sponge or washing-up brush; lab washer with special washing agent (for industry and lab use) Cleaning agent for denaturation of proteins (e.g. exit gas cooler) 0.1 N NaOH Cleaning agent for smaller component parts (e.g. exit gas cooler, dip tube) Ultrasonic bath Cleaning agent for sur- faces Water Disinfectant for surfaces Ethanol, 70 % Decalcifier for the equip- ment Phosphoric acid or citric acid (max. 5 %) 9.2 Cleaning the Culture Vessel - Routine Cleaning The culture vessel and accessories can be cleaned as soon as they have cooled down after autoclaving. ATTENTION Household washing-up liquid and soap (in particular cream soaps) can collect in glass pores and impair later cultivations. Never clean culture vessels and accessories with household soap and use special cleaning agent (for industrial and lab use) in the lab washer. Labfors 5 - Version for Microorganisms Cleaning and Maintenance Page 168 of 216 04 September 2019 The following method describes a routine cleaning between two cultivations. It takes place with the culture vessel completely assembled and the accessories completely mounted. This does not include the sensors, with the exception of antifoam or level sensors from the equipment manufacturer. To avoid damaging the other sensors during the routine cleaning, they are first removed and then cleaned separately according to the third-party manufacturer guidelines and then stored, if necessary. Also see the “Removing Sensors” section and “Cleaning Sensors”. Proceed as follows to carry out a routine cleaning of the culture vessel: Procedure 1. Carefully unscrew the sensors (except antifoam/level sensors) by hand (no tools!) from the vessel top plate ports and place them to the side for separate cleaning according to the manufacturer guidelines. 2. Completely fill the culture vessel with 0.1 N NaOH. 3. Fit the top plate on the vessel and secure it. 4. Connect the culture vessel to the basic unit (vessel jacket or cold finger and heater pad). 5. Couple the motor. 6. Switch on the equipment at the power switch. 7. At the operating panel in the touch screen software, start the bioreactor and stir strongly for 2 hours with the stirrer function (parameter Stirrer). INFORMATION It is recommended to warm the 0.1 N caustic soda to 60 °C and to prolong the duration of stirring for dealing with persistent residue of foam or protein. 8. Stop the bioreactor in the touch screen software at the operating panel. 9. Shutdown the system at the operating panel. 10. Switch off the equipment at the power switch. 11. Let the motor cool down. When the motor has cooled down: 12. Uncouple the motor. 13. Remove the top plate and carefully place it so that it does not(!) lie on top of components. 14. Empty the culture vessel. 15. Thoroughly rinse the culture vessel with distilled water. Labfors 5 - Version for Microorganisms Cleaning and Maintenance 04 September 2019 Page 169 of 216 9.3 Removing the Vessel Top Plate and Accessories All accessories must be removed for thorough cleaning of the individual parts of the culture vessel. This is described in the following sections. The cleaning itself is described in the chapter “Cleaning and Storing Individual Parts”. The cleaning of the hoses with pump heads, the basic unit, operating panel and the exit gas cooler are described in separate sec- tions. Sensors from third-party manufacturers are cleaned according to their manufacturer’s specifications. 9.3.1 Removing the Exit Gas Cooler Proceed as follows: Procedure 1. Unscrew the exit gas cooler from the vessel top plate port by hand. Ensure that the O-ring does not get lost. 2. Gently release the hose clamp with the hand wheel, pull off the exit gas filter and dispose of it. 3. Remove the pressure hose piece to thoroughly clean the exit gas cooler. (For details see the chapter “Cleaning the Exit Gas Cooler”.) 9.3.2 Removing the Sensors Sensors are either mounted directly into the ports or by means of an electrode holder or clamping adapter. For removal of the sensors, proceed as follows: Sensors without holder/clamping device Procedure 1. Carefully unscrew the sensor by hand (no tools!) from the vessel top plate port. Sensor with electrode holder Procedure 1. Carefully loosen and unscrew the sensor on the hollow screw of the electrode holder from the port by hand. 2. Carefully unscrew the sensor from the guide bar and gently pull it out of the electrode holder. Labfors 5 - Version for Microorganisms Cleaning and Maintenance Page 170 of 216 04 September 2019 Sensor with clamping adapter (antifoam and level sensor) Procedure 1. Loosen both slotted screws next to the sensor in the port. INFORMATION This step is only necessary for the type of clamping adapter for 10 mm ports. All other clamping adapters can be directly unscrewed and pulled out from the port or threaded adapter in the port by hand. 2. Carefully pull the clamping adaptor together with the sensor out of the port / threaded adapter in the port. 3. Ensure the O-ring of the clamping adapter does not get lost. As required: 4. Depending on the type of clamping adapter, loosen the slotted screw or the hollow screw and carefully pull the sensor out of the clamping adapter by hand. 5. Ensure that the insulation on the sensor does not get dam- aged. 9.3.3 Removing Hoses, Filters and Pump Heads To later clean reagent hoses and pump heads, they must be removed from the reagent bottles and from components of the culture vessel. INFORMATION To avoid damage, never dismantle the pump heads. Always replace a damaged pump head along with the pump hose, and vice versa. Proceed as follows: Procedure 1. Remove cable ties (e.g. with a side cutter) so that the hoses are not damaged. 2. Pull hoses off the culture vessel and the reagent bottles. 3. Remove and dispose of filters for pressure equalisation and hoses from reagent bottles. 4. Ensure that the inlet air filter is clean, dry and not blocked. If this is not the case, dispose of it. Labfors 5 - Version for Microorganisms Cleaning and Maintenance 04 September 2019 Page 171 of 216 INFORMATION If the filter for pressure equalisation and the corresponding hoses have been used several times, ensure that the filters are always dry and clean. 5. Dispose of the exit gas filter (see also chapter “Removing the Exit Gas Cooler”). 9.3.4 Removing Blanking Plugs Proceed as follows: Blanking plugs in 10 mm ports Procedure 1. Loosen both slotted screws next to the blanking plug in the port. Ensure the screws do not get lost. 2. Pull the blanking plug out of the port by hand. Ensure that the O-ring on the blanking plug does not get lost. Blanking plugs in 12 mm/Pg13.5 ports Procedure 1. Loosen the blanking plug with the hexagon socket spanner and remove it by hand. Ensure that the O-ring does not get lost. Blanking plugs in 19 mm ports Procedure 1. Loosen the blanking plug with the hexagon socket spanner in the threaded adapter in the port and remove it by hand. Ensure that the O-ring does not get lost. 9.3.5 Removing Threaded Adapters Proceed as follows: Procedure 1. Loosen both slotted screws next to the threaded adapter in the 19 mm port. Ensure the screws do not get lost. 2. Remove the threaded adapter. Labfors 5 - Version for Microorganisms Cleaning and Maintenance Page 172 of 216 04 September 2019 9.3.6 Removing the Inoculation Needle, Septum Collar and Septum Proceed as follows: Procedure 1. Unscrew the inoculation needle from the septum collar by hand. Ensure that the O-ring does not get lost. 2. Unscrew the septum collar out of the port or threaded adapter (in 19 mm port) by hand. 3. Remove the septum from the port and dispose of it. 9.3.7 Removing Addition Port Adapters Proceed as follows: Addition port adapter, quadruple, in 19 mm port Procedure 1. Loosen the two slotted screws next to the addition port adapter. Ensure the screws do not get lost. 2. Remove the addition port adapter. Ensure the O-ring on the adapter does not get lost. INFORMATION The removal procedure of further models of addition port adapters (refer to main chapter “Accessories” for details) is the same as for blanking plugs in their corresponding ports. That is why their removal is not repeatedly described here. 9.3.8 Removing the Vessel Top Plate Proceed as follows to remove the vessel top plate: Procedure 1. As far as possible, remove mounted parts before lifting the top plate. 2. Loosen and remove the knurled nuts on the top plate by hand (no tool!) and place them to the side. Labfors 5 - Version for Microorganisms Cleaning and Maintenance 04 September 2019 Page 173 of 216 3. Carefully lift the top plate vertically upwards from the vessel until the stirrer shaft and other long built-in components can no longer come into contact with the glass vessel. ATTENTION If the vessel top plate presses against long components they could bend because of the weight of the top plate. Always position the vessel top plate so that it does not lie on top of components. 4. If necessary, now remove components that have not yet been removed. 5. Check the glass vessel for damage (cracks, fissures, scratches) and replace if necessary. Labfors 5 - Version for Microorganisms Cleaning and Maintenance Page 174 of 216 04 September 2019 9.3.9 Removing the Pocket for Temperature Sensor (Pt100) Proceed as follows: Procedure 1. Loosen both slotted screws next to the port. Ensure the screws do not get lost. 2. Push the pocket from the inside of the vessel top plate up, so that it can be pulled out of the port from the outside of the top plate. Ensure the O-ring on the pocket does not get lost. 9.3.10 Removing the Sparger and the Dip Tube(s) Straight spargers and dip tubes can be removed from the outside of the vessel top plate. Curved spargers and dip tubes can only be removed from the inside of the vessel top plate. Since this equipment uses ring spargers and straight dip tubes, removal from the inside of the vessel top plate is described here. This means that the vessel top plate is already removed. Proceed as follows: Procedure 1. Loosen the slotted screw on the clamping adapter. 2. Carefully pull the sparger/dip tube from the bottom out of the clamping adapter. 3. Unscrew the clamping adapter out of the port by hand. Ensure that O-ring does not get lost. 9.3.11 Removing the Baffles The four baffles can be removed, once the vessel top plate is re- moved. Proceed as follows: Procedure 1. Loosen and remove the hexalobular screw from the vessel top plate. 2. Pull the baffle off and put aside. Ensure that the O-ring does not get lost. 3. Proceed the same way with the remaining baffles. Labfors 5 - Version for Microorganisms Cleaning and Maintenance 04 September 2019 Page 175 of 216 9.3.12 Removing the Impellers Before removing the impellers, it is recommended to measure and record the positions to aid later mounting. Proceed as follows to remove: Procedure: 1. Loosen the grub screws on the impeller with an Allen key – do not remove! 2. Carefully remove the impeller from the stirrer shaft. 9.3.13 Removing the Stirrer Shaft If needed, the stirrer shaft can be removed for cleaning. Particular care shall be taken, to do so. ATTENTION The mechanical seal is sensitive to applied forces. Manipulation on it may lead to its damage! To remove the stirrer shaft from the vessel top plate, proceed as follows: Labfors 5 - Version for Microorganisms Cleaning and Maintenance Page 176 of 216 04 September 2019 Procedure 1. Insert an Allen key or a thin metal bar into the opening on the upper end of the stirrer shaft. This locks the stirrer in its position. 2. Position an adjustable spanner on the two recesses on the upper end of the stirrer shaft. 3. Unscrew the stirrer shaft with the spanner counter clockwise from the drive shaft. 4. Check the O-ring on the drive shaft for damages, replace as necessary. 9.3.14 Removing the Glass Vessel from the Holder The glass vessel and the individual parts of the vessel fixation in the vessel holder may also be cleaned individually in the event of heavy soiling or if internal regulations so require. For this purpose, the glass vessel must be removed from the holder. Proceed as follows: Procedure 1. Remove the silicone hoses from the glass olives for water inlet, outlet and overflow of the vessel jacket. Labfors 5 - Version for Microorganisms Cleaning and Maintenance 04 September 2019 Page 177 of 216 2. Remove the top plate seal (O-ring). 3. Loosen and remove the four lock nuts on the stainless steel ring by hand. 4. Remove the stainless steel ring. 5. Remove the green shock-absorbing ring from the vessel col- lar. Labfors 5 - Version for Microorganisms Cleaning and Maintenance Page 178 of 216 04 September 2019 6. Remove the white spacers from the rods of the vessel holder. 7. Only vessels (single and double walled) with DN 200: remove the connecting plates on both sides of the two-piece vessel flange before or after lifting the glass vessel out of the holder Loosen the screws (8 x M5x10) and remove them with the plates. 8. Lift the glass vessel out of the holder on the two-piece flange by pulling it off the rods of the holder. 9. Remove the two-piece vessel flange. 10. Remove the black shock-absorbing ring from the vessel collar. Labfors 5 - Version for Microorganisms Cleaning and Maintenance 04 September 2019 Page 179 of 216 11. Only double walled vessels with DN 200: remove the O-ring from the upper edge of the vessel jacket. 12. Only double walled vessels with DN 200: remove the piece of silicone hose, which has been fitted for protection of the glass olive of the vessel jacket overflow. 13. Clean the vessel and individual parts. 9.4 Cleaning and Storing Individual Parts The procedure described here applies to the following individual parts:  Vessel  Accessories such as blanking plugs, spargers, dip tubes, addition port adapters etc.  Reagent bottles  Vessel top plate, with regard to its particular characteristics Particulars when cleaning the top plate  Do not place the top plate on the stirrer shaft (if stirrer shaft is not removed).  Never remove the drive hub and the mechanical seal! This may only be carried out by qualified personnel.  NEVER clean the vessel top plate in the dishwasher! Labfors 5 - Version for Microorganisms Cleaning and Maintenance Page 180 of 216 04 September 2019 ATTENTION The mechanical seal is very delicate. No water/cleaning agent may enter it and the silicone hose on the bottom of the drive hub may not be pulled off during cleaning of the vessel top plate. INFORMATION Cleaning of the sensors, hoses and pump heads as well as the basic unit and the exit gas cooler are described in separate sec- tions. Proceed as follows for cleaning: Procedure 1. Clean parts with distilled water and a soft sponge or in the dishwasher (except for the vessel top plate!). Ensure that the deposits in the dip tubes are removed. Use 0.1 N caustic soda solution followed by distilled water as necessary. For this, see chapter “Cleaning the Culture Vessel”. 2. Dry all parts, including the inner parts of the dip tubes, spargers and feed needles. 3. Check all O-rings for cracks or damage. Replace them if nec- essary. 4. Store the vessel, vessel top plate and accessories in a clean, dry state in a location where they cannot be physically damaged (e.g. by falling), or prepare them for the next cultivation. 9.5 Cleaning the Sensors Apart from antifoam and level sensor, all sensors are cleaned and maintained according to the descriptions of the sensor manufac- turer. Procedure 1. Clean the sensors according to the sensor manufacturer guidelines. 2. Prepare the sensors for the next cultivation or, if necessary, service and/or store them according to the sensor manufacturer guidelines. Labfors 5 - Version for Microorganisms Cleaning and Maintenance 04 September 2019 Page 181 of 216 9.6 Cleaning the Hoses and Pump Heads Proceed as follows to clean the reagent hoses and pump heads: Procedure 1. Thoroughly rinse the hoses with the pump heads with water. 2. Carefully dry all hoses and, if necessary, blow out with cleancompressed air. INFORMATION To avoid damage, never dismantle the pump heads. Always replace a damaged pump head along with the pump hose, and vice versa. 9.7 Cleaning the Super Safe Sampler ATTENTION Risk of damage to the sampling system from unsuitable cleaning methods or cleaning agent (such as acids, bases or solvents, for example). – Only use water or a mild soap solution for cleaning. – The sterile filter must remain dry at all times. Proceed as follows to clean the sampling system: Procedure 1. Fill the culture vessel with water or a mild soap solution. Or: Remove the sampling hose from the dip tube and hold it in a vessel, e.g. a beaker, with water or a soap solution. 2. Place the syringe on the automatic valve and pull out the plunger to rinse the sampling system. When using a soap solution: 3. Then rinse the sampling system thoroughly with water. INFORMATION If the test record requires that the culture is killed off after cultivation by autoclaving the culture vessel, the valves of the sampling system may become stuck due to reside of the culture solution. In such a situation, it would be better to autoclave the sampling system separately in a beaker of water (hoses filled with water, filter removed). Labfors 5 - Version for Microorganisms Cleaning and Maintenance Page 182 of 216 04 September 2019 9.8 Cleaning the Exit Gas Cooler If the exit gas cooler is only lightly soiled, an ultrasonic bath for approx. 15 minutes is sufficient to clean it. However, if foam has entered the exit gas cooler during cultivation, it must be cleaned thoroughly. To do so, proceed as follows: Procedure 1. Put the exit gas cooler into 0.1 N NaOH for 4 hours. 2. Rinse the exit gas cooler thoroughly with water. 3. Put the exit gas cooler into an ultrasonic bath for 2 to 5 minutes. 4. Flush the exit gas cooler with ethanol (70%). 5. Thoroughly rinse the exit gas cooler with distilled water. 9.9 Cleaning the Basic Unit and Operating Panel Proceed as follows to clean the surface of the basic unit and the operating panel as required: Procedure 1. Switch off the equipment at the power switch. 2. Disconnect the equipment from the power supply. 3. Wipe all surfaces with a damp cloth. Clean with an appropriate disinfectant as necessary. 4. Clean the screen with a wipe suitable for computer or laptop screens. 9.10 Cleaning the Heater Pad DANGER Damp cleaning of a heater pad, which is connected to energised equipment, can be perilous. – Always disconnect the heater pad from the basic unit before cleaning. – Preferably clean the heater pad dry. – Always let the heater pad completely dry after cleaning with a damp cloth and before next use. Labfors 5 - Version for Microorganisms Cleaning and Maintenance 04 September 2019 Page 183 of 216 If required, clean the heater pad as follows: Procedure 1. Unplug the heater pad from the basic unit. 2. Wipe the heater pad with a dry cloth. If necessary, use a damp cloth (water only!). ATTENTION Cleaning the heater pad with soap or aggressive detergents may damage the silicone sheathing. 3. Let the heater pad completely dry after cleaning with a damp cloth. 4. Check the heater pad and cable for damages (cracks, kinks etc.), replace as necessary. 9.11 Maintenance Plan WARNING Non-compliance of this maintenance plan contains a high risk! It is the responsibility of the user, that this maintenance plan is complied with. Non-compliance will lead to exclusion of liability (see General Terms and Conditions). The required maintenance for reliable operation is described in the following chapters. Reduce the maintenance intervals in case increased abrasion is detected during regular checks. Contact the manufacturer for questions concerning maintenance. Labfors 5 - Version for Microorganisms Cleaning and Maintenance Page 184 of 216 04 September 2019 To be carried out by operator Interval Maintenance work Before each cultivation Check all hoses and hose connections. Replace hoses, if necessary. Supply hoses may be replaced by qualified personnel. Check cables for damage and kinks. Check that O-rings and gaskets are leak-proof, replace if necessary. Check the integrity of all glass parts (vessel, reagent bottles) and replace if necessary. Check all sterile filters and replace if necessary. Replace the exit gas filter. If necessary, calibrate the sensors. Single walled vessels: check heater pad and its cable connection on visible damages, replace the heater pad, if necessary. Check lubrication of mechanical seal and lubricate as necessary. After every cultivation Autoclave and clean the culture vessel and accessories. As required Clean the basic unit and operating panel. Single walled vessels: clean the heater pad. Equipment with water temperature control system: decalcify the equipment via the vessel jacket. To be carried out by qualified personnel Interval Maintenance work Every 6 months Check and calibrate measuring sections (temperature, pH, etc.) with a sim- ulator As required Replace supply hoses. To be carried out by operator, ONLY AFTER CONSULTATION OF MANUFACTURER Interval Maintenance work As required Decalcify the equipment (practicable for both temperature control system variants) Labfors 5 - Version for Microorganisms Cleaning and Maintenance 04 September 2019 Page 185 of 216 9.12 Lubricating the Mechanical Seal The two-part silicone hose on the drive hub must always be filled with liquid (Glycerine, for detailed information see main chapter “Technical Data”, chapter “Operating Materials”) to ensure the mechanical seal is lubricated. ATTENTION Risk of loss of property due to the mechanical seal running dry! A mechanical seal, which has not been adequately lubricated, is destroyed when running dry. Proceed as follows for lubrication: Procedure 1. Pull off the longer piece of hose from the coupling on the shorter piece. 2. Fill a syringe with glycerine and plug it onto the open hose end. 3. Fill Glycerine into the hose. 4. Plug the longer piece of hose onto the coupling of the shorter piece. If Glycerine has come off the tubing, wipe off as necessary. Labfors 5 - Version for Microorganisms Cleaning and Maintenance Page 186 of 216 04 September 2019 9.13 Decalcifying the Equipment Calcification could block mounting parts, lines or valves in the basic unit. It may be necessary to decalcify the equipment if certain faults occur in the temperature control system. Note the following points, before begin of the procedure: ATTENTION Inappropriate decalcifying of the equipment may lead to loss of property. Only decalcify the equipment AFTER CONSULTATION of the manufacturer or licensed dealer! INFORMATION A turbid or milky coloured appearance of the glass of the vessel jacket may be a sign of lime scale in the equipment with water temperature control system. A first possibly already sufficient procedure may be to decalcify the equipment via the vessel jacket. Refer to chapter “Decalcifying the Equipment via Water Jacket” for details in this case.  Use phosphoric or citric acid (max. 5 %) as decalcifying agent.  Be sure to respect the in chapter “Technical Data” specified inlet pressure.  To warm up the decalcifier and pump it into the basic unit, use a chiller or a water bath and an external pump.  During decalcification, the decalcifier flows in a circuit between the basic unit and the chiller/water bath. Proceed as follows: Procedure 1. If applicable, mount the exit gas cooler into the port of the vessel top plate and connect to the basic unit. 2. If applicable, mount the cold finger. 3. Connect the culture vessel to the basic unit. 4. Let the water drain off from the water outlet on the basic unit. 5. Fill the chiller/water bath with decalcifier. 6. Connect the chiller or water bath to the water inlet and outlet on the basic unit. Labfors 5 - Version for Microorganisms Cleaning and Maintenance 04 September 2019 Page 187 of 216 7. To open the corresponding valves in the basic unit, set the temperature on the operating panel to 5 °C (cool). 8. Set the chiller/water bath to 20 °C to 40 °C. 9. Switch on the pump at the chiller/water bath. 10. Let the decalcifier flow through the equipment for an hour. 11. Connect the water inlet hose on the equipment to tap water. 12. Hang the water outlet hose of the equipment at the spout. 13. Rinse the equipment for an hour. 9.14 Decalcifying Equipment via Vessel Jacket A turbid or milky coloured appearance of the glass of the vessel jacket may be a sign of lime scale in the equipment with water temperature control system. A first possible procedure may be to decalcify the equipment via the vessel jacket. If there are still some interferences observed with the temperature control system after doing so, then a thorough decalcification of the equipment may be necessary. Refer to chapter “Decalcifying the Equipment” in this case. Prepare the following things and note the following points before begin of the procedure:  Use phosphoric or citric acid (max. 5 %) as decalcifying agent.  Prepare a funnel and piece of hose for connection to the water overflow of the vessel jacket.  Be sure to respect the in chapter “Technical Data” specified inlet pressure. Proceed as follows: Procedure 1. Completely empty the vessel jacket. 2. Fit the piece of hose to the water overflow connection of the vessel jacket. 3. Fit the funnel to the hose end. 4. Fill the decalcifying agent into the vessel jacket. 5. Connect the vessel to the basic unit. 6. Completely fill the vessel jacket with water. 7. Heat the vessel at a temperature of 50 °C for an hour. 8. Decrease the temperature and thoroughly rinse the jacket with cooling water. Labfors 5 - Version for Microorganisms Interferences Page 188 of 216 04 September 2019 10 Interferences The following section describes possible reasons for interferences and how to resolve them. Reduce the service intervals in correspondence with the actual loads if interferences become increasingly common. Contact the manufacturer or licensed dealer for interferences that cannot be resolved by following the above instruc- tions. 10.1 Interferences Basic Unit and Operating Panel Interference Equipment does not work. Power switch is not illuminated; monitor of the operating panel remains dark. Possible cause Remedy By Equipment is not switched on. Switch on the equipment at the power switch. Operator Power supply of the equipment is in- terrupted.  Check if the plugs are connected  Check the mains connection. Operator Equipment fuse is blown. Replace the fuse. If the fault occurs more than once, contact the Infors representative. Operator Interference Power switch is illuminated; monitor of the operating panel remains dark. Possible cause Remedy By Monitor of operating panel is switched off. Press the ON/OFF key on the monitor. Operator Power supply cable of the operating panel is not connected. Connect the power supply cable to the DC connection of the operating panel. Operator Interference No communication between equipment and operating panel. Possible cause Remedy By iDDC-bus cable (display cable) is not connected. Connect the iDDC-bus cable: Plug the round connector in at the COM1 connection of the operating panel. Plug the flat connector in one of the two iDDC-bus connections on the rear side of the basic unit. Operator Labfors 5 - Version for Microorganisms Interferences 04 September 2019 Page 189 of 216 10.2 Interferences Drive System Interference Stirrer does not start. Possible cause Remedy By Parameter Stirrer is not activated. Activate the parameter. Operator Setpoint of parameter Stirrer = 0. Set setpoint > 0. Operator Parameter pO2 is activated and set to oxygen control via the stirrer (cas- cade). Switch cascade off and test operation via parameter Stirrer. Operator Interference Motor control is volatile, irregular or stops. Possible cause Remedy By PID settings of parameter Stirrer are not correct Reset to default settings Operator Interference Unusual sounds when the stirrer is activated. Possible cause Remedy By Stirrer is in contact with other vessel built-in components. Stop the bioreactor. Shutdown the system and turn off the equipment. Correctly mount the mounting parts of the culture vessel while observing internal safety regulations. Operator 10.3 Interferences Temperature Control System Interferences No temperature control. Possible cause Remedy By Parameter Temperature is not acti- vated. Activate the parameter. Operator Parameter Stirrer is not activated. Activate the parameter. Operator Labfors 5 - Version for Microorganisms Interferences Page 190 of 216 04 September 2019 Interference No heating or inadequate heating. Possible cause single-walled culture vessels Remedy By Heater pad is not connected. Connect the heater pad. Operator Faulty heater pad. Check function of heater pad. Check heater pad and cable connections for damages (cracks, kinks etc.). Replace heater pad as necessary Operator Possible cause double-walled culture vessels Remedy By Circulation not filled or air bubbles in circulation are obstructing the circulation flow Enter low setpoint for parameter temperature to open the cooling valve and to fill the circulation (check acoustically) then enter a higher setpoint to heat up. Fully open the water supply for a moment. Check whether there is sufficient primary pressure on the water supply, adjust as necessary. Operator Cooling valve is blocked Decalcify the equipment Operator Interference No cooling or inadequate cooling. Possible cause Remedy By No water supply or inadequate flow. Check the water supply and turn the supply tap if necessary. Operator Temperature sensor is not inserted. Insert the temperature sensor into the pocket in the vessel top plate. Operator Incorrect Negative factor in option PID of parameter Temperature Check Negative factor: Value must be positive. Adjust as necessary. Operator Interference Temperature fluctuations Possible cause Remedy By Incorrect PID settings parameter Temperature Check PID settings and adjust as necessary, especially P-term. Operator Labfors 5 - Version for Microorganisms Interferences 04 September 2019 Page 191 of 216 10.4 Interferences Gassing System Interferences No gassing / air bubbles in the culture vessel. Possible cause Remedy By The on-site gas supply has been in- terrupted. Stop the bioreactor. Check the on-site gas supply and switch it on, if nec- essary. Operator Depending on the configuration of the gassing system: The rotameter valve(s) is/are not open. And/or: The Flow parameter(s) is/are not ac- tivated. And/or: Setpoint in the Flow parameter(s) = 0. Or: Parameter GMFlow = 0 and/or GasMix is/are not activated. Slowly open the rotameter needle valve(s). And/or: Activate the Flow parameter(s). And/or: Set the setpoint(s) in the Flow parameter(s) > 0. Or: Set parameter GMFlow > 0 and activate parameter- GasMix. Operator Hose connection(s) between the basic unit and the culture vessel is/are kinked or clamped. Check whether the hose connection(s) is/are clamped; if necessary open the clamp(s). Check hose connection(s) for kinks, if necessary route them again or replace them under observation of the sterility requirements. Operator Inlet air filter blocked. Replace the inlet air filter under sterile conditions. Operator Interference The desired gas flow rate is not reached. Possible cause Remedy By Blocked holes on the sparger. Stop the bioreactor and clean the sparger. Operator Labfors 5 - Version for Microorganisms Interferences Page 192 of 216 04 September 2019 Interference Sudden increase in evaporation losses in the culture vessel. Possible cause Remedy By The exit gas cooler does not cool. The control valve for water flow is closed. Open the control valve. Operator The exit gas cooler does not cool, parameter Temperature is activated. Check the water supply to the exit gas cooler. Restore it, if necessary. The basic unit is calcified. Decalcify the equipment, if necessary. Operator 10.5 Interferences pH-System Interference No display or incorrect display of pH Possible cause Remedy By Sensor cable not connected or not properly connected. Connect properly if necessary. Operator Measuring system METTLER (ana- logue) Function Temperature compensation is not activated. Activate this function in option Setpoint of parameter pH1. Operator pH drift during long cultivation. Recalibrate pH with offline values. Refer to the separate operating manual of the touch screen software for details: (Analogue) measuring system METTLER: chapter “Recalibrating a METTLER pH Sensor”. (Digital) measuring system HAMILTON: chapter “HAMILTON pH Sensor Product Calibration”. Operator Faulty pH-sensor. Test calibration with pH 4 and pH 7 buffer. Regenerate or replace the sensor. Consult the documentation of the sensor manufac- turer! Operator 1) This does not apply to the measuring system HAMILTON (digital): this function MUST be deactivated there!! Labfors 5 - Version for Microorganisms Interferences 04 September 2019 Page 193 of 216 Interference No pH control. Possible cause Remedy By Parameter pH is not activated. Activate the parameter. Operator Incorrect dead band setting. Check the dead band (Dead Band in PID settings): Switch off or enter a small value. Operator No addition of reagents (acids and base). Check the reagent bottles: Refill if necessary. Check the hose connections between the reagent bottles and the culture vessel: Connect properly if necessary. Open/remove hose clamps if necessary. Operator Pump(s) (base/acid) do/does not operate properly. Check operation using the rocker switch. Operator Pump hose is damaged. Replace pump head. Operator Incorrect hose type connected. Replace if necessary. Operator Interference pH value drifts up and down over time or acid and base are added almost continuously in turn. Possible cause Remedy By Incorrect PID setting in pH parame- ter. Check the PID settings and adjust as necessary. Change the special proportional factor (Prop. Term) or Dead band setting. Operator Incorrect strength of reagents: Concentration is too weak or too strong. Check the strength of reagents. Adjust if necessary: 0.1 mol to 2.0 mol. Operator 10.6 Interferences pO2 System Interferences No display or incorrect display of pO2. Possible cause Remedy By Sensor cable not connected or not properly connected. Connect properly if necessary. Operator pO2 sensor is not calibrated Calibrate the pO2 sensor. Refer to the separate operating manual of the touch screen software for de- tails. Operator Measuring system METTLER (ana- logue) pO2 sensor is not polarised. Polarise the pO2 sensor Operator Faulty pO2 sensor. Check the calibration of the pO2 sensor. Consult the documentation of the sensor manufac- turer. Operator Labfors 5 - Version for Microorganisms Interferences Page 194 of 216 04 September 2019 Interference No pO2 control. Possible cause Remedy By The pO2 parameter and/or cascaded parameter is/are not activated. Activate parameters. Operator The cascade settings are incorrect. Check the cascade settings and change as neces- sary. Operator. No gas flow into culture vessel. Refer to interferences in the gassing system. Operator Interference pO2 value fluctuates strongly or the stirrer speed fluctuates. Possible cause Remedy By Incorrect PID settings in the pO2 pa- rameter. Check the PID settings (PID parameter option) and adjust as necessary. Special proportional factor (Prop. Term) and dead band. Dead band value must be 0 (zero). Operator 10.7 Interferences Antifoam or Level Sensor and Antifoam Pump Interference Foam/medium is not detected. Possible cause Remedy By Sensor is not properly connected. Check connections and connect properly as neces- sary. Operator Interference Foam/medium is always/frequently detected. Possible cause Remedy By Sheathing of sensor is damaged. Have the sheathing of the sensor replaced. For this, contact Infors representative Infors service technician Interference Antifoam pump does not work. Possible cause Remedy By Parameter Antifoam is not activated. Activate the parameter. Operator Dosing time of parameter Antifoam = 0 (zero). Set dosing time > 0. Operator Labfors 5 - Version for Microorganisms Interferences 04 September 2019 Page 195 of 216 Interference No antifoam agent or medium supply or inadequate flow. Possible cause Remedy By Reagent bottle is empty. Refill if necessary. Operator Wrong antifoam agent or incorrect concentration. Replace if necessary. Operator Hose line blocked or clamped. Check the hose connection between the reagent bottle and the culture vessel: If necessary, connect them correctly. Open/remove hose clamps if necessary. Operator Antifoam pump does not work. Check operation using the rocker switch. Operator The pump hose is damaged. Replace pump head. Operator Incorrect hose type connected. Replace if necessary. Operator 10.8 Interferences Feed and Pump Interference No feed solution or inadequate feed solution. Possible cause Remedy By Parameter Feed is not activated. Activate the parameter. Operator Setpoint of parameter Feed = 0. Set setpoint > 0. Operator Reagent bottle empty. Refill if necessary. Operator Hose line blocked or clamped. Check the hose connection between the reagent bottle and the culture vessel: If necessary, connect them correctly. Open/remove clamp if necessary. Operator Feed pump does not work. Check operation using the rocker switch. Operator The pump hose is damaged. Replace pump head. Operator Incorrect hose type connected. Replace if necessary. Operator Labfors 5 - Version for Microorganisms Interferences Page 196 of 216 04 September 2019 10.9 Replacing Equipment Fuses INFORMATION Equipment fuses may only be replaced by fuses of the same rating. For detailed information concerning the requirements for the fuses refer to main chapter “Technical Data”, chapter “Connection Values”, “Electrical” To replace a defective equipment fuse, proceed as follows: Procedure 1. Switch off the equipment and pull out the power plug. 2. Unlock the plug for the fuses by pressing together the two flaps and pull out the plug at the same time. 3. Remove the defective fuse. 4. Insert a new fuse with the correct number of Amperes. 5. Push the plug as far back in the opening as possible until it snaps in. 6. Re-establish the power supply to the equipment. 10.10 Returning for Repair The provider must return the equipment or the faulty component part(s) to the manufacturer if, after consulting the service department of the local dealer or the manufacturer, on-site diagnosis and/or repair is not possible. INFORMATION When returning the equipment, the component part or accessory for repair, it is required for the safety of all parties involved and because of legal provisions that a lawful declaration of decontamination is present. Refer to main chapter “Safety and Responsibility”, chapter “Declaration of Contamination” for details. Labfors 5 - Version for Microorganisms Disassembly and Disposal 04 September 2019 Page 197 of 216 11 Disassembly and Disposal The equipment must be disassembled and disposed of in an environmentally friendly manner if it is no longer in use. INFORMATION When returning the equipment for disassembly or disposal, it is required for the safety of all parties involved and because of legal provisions that a lawful declaration of decontamination is present. Refer to main chapter “Safety and Responsibility”, chapter “Declaration of Contamination” for details. 11.1 Disassembly Prior to disassembly:  Switch off the equipment and lock any isolation switch in the ‘off’ position.  Physically disconnect the main energy supply from the equipment and wait for components to fully discharge.  Remove and dispose of all additional consumable items, auxiliary components and/or spent processing material in an environmentally acceptable manner. Clean and disassemble component parts professionally with regard to any local regulations concerning employment and environmental protection. If possible, separate materials. Labfors 5 - Version for Microorganisms Disassembly and Disposal Page 198 of 216 04 September 2019 11.2 Disposal Recycle disassembled components if no agreement is made concerning reclaim or disposal.  Send metals for scrap.  Send plastic components for recycling.  Sort and dispose of the remaining components according their material composition. WARNING Electronic waste, electronic components, lubricants or other auxiliary materials/supplies are subject to hazardous waste regulations and may only be disposed of by registered specialist disposal firms. For disposal, the system units are to be disassembled and dismantled into individual material groups. These materials are to be disposed of according to the applicable national and local legislation. Local authorities or specialist disposal firms can provide information regarding environmentally acceptable disposal. If no special arrangements have been made for return, INFORS HT units with the required declaration of decontamination can be sent back to the manufacturer for disposal. Labfors 5 - Version for Microorganisms Technical Data 04 September 2019 Page 199 of 216 12 Technical Data 12.1 Dimensions 1 Unit Front view  To the left: equipment with operating unit, holder for up to 2 gassing units, 5 pumps, and standard base tray  Middle: equipment with operating unit, holder for up to 4 gassing units, 5 pumps, and base tray for vessels with DN1 200 mm  To the right: satellite unit, holder for up to 4 gassing units, 5 pumps and standard base tray 1) DN = diameter nominal (= inner diameter) Dimensions in mm Labfors 5 - Version for Microorganisms Technical Data Page 200 of 216 04 September 2019 Top view and side view  To the left: equipment with holder for up to 2 gassing units and standard base tray  Middle: equipment with holder for up to 4 gassing units and base tray for vessels DN200  To the right: equipment with operating unit and standard base tray Dimensions in mm Labfors 5 - Version for Microorganisms Technical Data 04 September 2019 Page 201 of 216 12.2 Dimensions Master Unit and Satellite Units Front view master unit with 5 satellite units  Above: 6 units with holder for up to 4 gassing units, 5 pumps and standard base tray  Below: 6 units with holder for up to 4 gassing units and base tray for vessels with DN200 Dimensions in mm Labfors 5 - Version for Microorganisms Technical Data Page 202 of 216 04 September 2019 12.3 Dimensions of Culture Vessels in Vessel Holder Top view The figures above show top view of the culture vessel with standard exit gas cooler and with swivelling exit gas cooler and, with / without reagent bottle und pump holder. Ø Total volume (L) / diameter nominal of vessels (mm) 2 / 115 3.6 /115 3.6 /150 7.5 / 150 13 / 200 Dimensions in mm D1 330 330 335 335 365 D2 300 300 320 320 365 D3 340 340 355 355 420 D4 365 365 380 380 435 D1 D2 D3 D4 Labfors 5 - Version for Microorganisms Technical Data 04 September 2019 Page 203 of 216 Side view The figures show the culture vessel with standard exit gas cooler (left) and with swivelling exit gas cooler (right), both with reagent bottle and pump holder. Ø and height Total volume (L) / diameter nominal of vessels (mm) 2 / 115 3.6 /115 3.6 /150 7.5 / 150 13 / 200 Dimensions in mm D1 250 250 250 250 290 H1 373 538 405 635 616 H2 453 618 487 717 700 H3 514 679 546 776 760 12.4 Weights (net) Description Value Unit Basic unit 25 kg Touch screen operating unit 5 kg Labfors 5 - Version for Microorganisms Technical Data Page 204 of 216 04 September 2019 Culture vessels Total volume L Diameter nominal mm Weight kg 1) 2.0 115 10 3.6 115 12 3.6 150 15 7.5 150 18 13.0 200 22 1) Empty weight of double walled culture vessels in delivery state (equipped with default mounting parts) 12.5 Connection Values 12.5.1 Electrical Description Type 230 V Type 115 V Unit Value Voltage 230 115 V Frequency range 50 / 60 60 Hz Max. current 4 8 A Fuses (5 x 20 mm, slow- blown) 4 8 A 12.5.2 Water IN Description Value Unit Connection pressure 2 ± 1 bar Inlet temperature 10 - 20 °C Connection: OD of hose nozzle 8.3 mm Max. flow cooling vessel 1.6 L/min Max. flow cooling exit gas cooler 1.6 L/min Water quality „Very soft“ / „soft“ (CaCO3 concentration 0 mmol L-1 to 1.5 mmol L-1 INFORMATION The heating system has protection against dry running, which is based on measurement of conductivity. The heating will not work when using demineralised or distilled water as cooling agent! Labfors 5 - Version for Microorganisms Technical Data 04 September 2019 Page 205 of 216 12.5.3 Water OUT Description Value Unit Connection pressure No back pressure Temperature up to 80 °C Connection: OD of hose nozzle 10 mm 12.5.4 Gas(es) IN Description Value Unit Connection pressure 2 ± 0.5 bar Connection: OD of hose nozzle 7 mm General gas quality Dry, clean and free of oil and dust Recommended compressed air quality Class 1,2,3,4 As per DIN ISO 8573-1 12.5.5 Exit Gas Description Value Unit Connection pressure No back pressure Connection: OD of hose nozzle 8 mm 12.6 Specifications 12.6.1 Operating Panel Description Value HMI Colour touch screen 12" Protection IP 66 Labfors 5 - Version for Microorganisms Technical Data Page 206 of 216 04 September 2019 12.6.2 Culture Vessel Description Value Form Cylindrical with round bottom Models Single walled: for temperature control with electrical heater pad and cold finger Double walled: for temperature control with wa- ter Material Glass vessel Borosilicate glass Top plate and mounting parts Stainless steel, AISI 316L, electro polished O-rings EPDM Vessel sizes TV 1) VV max. 2) VV min. 3) DN 4) mm Height mm 2.0 1.2 0.5 115 205 3.6 2.3 0.5 115 370 3.6 2.3 1.0 150 235 7.5 5.0 1.0 150 465 13.0 10.0 2.2 200 445 1). Total volume in L 2). Min. working volume in L 3). Max. working volume in L 4). Diameter nominal = inner diameter of the vessel Ports in top plate ∅ mm Thread Number per diameter nominal of vessel DN 115 DN 150 DN 200 10 None 2 2 2 12 Pg13.5 6 6 5 19 None 2 3 6 Labfors 5 - Version for Microorganisms Technical Data 04 September 2019 Page 207 of 216 12.6.3 Stirrer Description Value Drive Shaft with mechanical seal Motor Type: DC, brushless Nominal power: 140 W Nominale torque: 0.4 Nm Range of rotation speed with 2 impellers (Standard) 2 L and 3.6 L / NW 115 vessel: 80 up to 1500 min-1 3.6 L and 7.5 L / NW 150 vessel: 80 up to 1200 min-1 13 L / NW 200 vessel: 80 up to 700 min-1 Range of rotation speed with 3 impellers (Option) 2 L and 3.6 L / NW 115 vessel: 80 up to 1500 min-1 3.6 L and 7.5 L / NW 150 vessel: 80 up to 1000 min-1 13 L / NW 200 vessel: 80 up to 600 min-1 Accuracy Measurement: ± 5 min-1 at 100 - 1000 min-1 1 % setpoint at > 1000 min-1 Control: ≤ ± 5 min-1 at 100 - 1000 min-1 ≤ 1 % setpoint at > 1000 min-1 Direction of rotation Counter clockwise (top view vessel) Bearing Outside of vessel, in drive hub INFORMATION Rotation speed is valid for viscosity similar to water, without aeration. Vessels are equipped with two impellers by default. The max. adjustable rotation speed on the operating panel is set for 2 impellers. Labfors 5 - Version for Microorganisms Technical Data Page 208 of 216 04 September 2019 Impellers Type / Number Material Rushton impellers with 6 blades Standard: 2 pieces Option: 3 pieces 316L stainless steel, electro pol- ished Vessel A B C 2 L and 3.6 L / NW 115 46 mm 11 mm 11 mm 3.6 l and 7.5 L / NW 150 54 mm 11 mm 11 mm 13 L / NW 200 70 mm 13 mm 19 mm 12.6.4 Temperature Description Value Sensor Type: Pt100 1/3 DIN-B Heating Double walled vessels: Water circulation in vessel jacket. Pump and heating 500 W integrated in basic unit. Single walled vessels: With electrical heater pad Cooling Double walled vessels: With tap water via water circuit into vessel jacket Option: with circulation chiller Single walled vessels: With tap water via cold finger in vessel Range of measure- ment -5 °C up to +150 °C Labfors 5 - Version for Microorganisms Technical Data 04 September 2019 Page 209 of 216 Range of control System with water circulation: from 5 °C above inlet temperature up to 70°C System with electrical heater pad and cold finger: from 1 °C up to 95 °C Accuracy Measurement: ± 0.2 °C at +10 °C up to +70 °C Control: ≤ ± 0.2 °C 1) Temperature depends on energy output (stirrer, gassing temperature, chemical and biological reaction) 12.6.5 Gassing The entire gas entry takes place via the sparger. The specific gassing rate, calculated for the max. working volume is for each vessel size and each variant 2 min-1. Variant Basic Gas(es) Gas flow control Accuracy Rotameter Gas mix control Air 1 Rotameter ± 4 % ---- Air + O2 1 Rotameter 2 solenoid valves Air + N2 1 Rotameter 2 solenoid valves Air + O2 + N2 1 Rotameter 3 solenoid valves Variant Standard Gas(es) Gas flow control Accuracy MFC Gas mix control Air 1 MFC ± 1 % ---- Air + O2 1 MFC 2 solenoid valves Air + N2 1 MFC 2 solenoid valves Air + O2 + N2 1 MFC 3 solenoid valves Labfors 5 - Version for Microorganisms Technical Data Page 210 of 216 04 September 2019 Variant High End Gas Gas flow control Accuracy MFC Air 1 MFC ± 0.3 % (final value) ± 0.5 % (measured value) Gases Gas flow control and gas mix control Air + O2 2 MFC Air + N2 2 MFC Air + O2 + N2 3 MFC Measuring ranges of MFCs 1) and rotameters Vessel max. WV 2) (Litres) Measuring range Rotameter L min-1 (Variant Basic) Measuring range MFC L min-1 (Variant Standard) Measuring range MFC L min-1 (Variant High End) 1.2 0.25 up to 2.4 0.05 up to 2.5 0.025 up to 2.5 2.3 0.30 up to 4.7 0.10 up to 5.0 0.05 up to 5.0 5.0 0.50 up to 11.0 0.20 up to 10.0 0.10 up to 10.0 10.0 1.00 up to 22.0 0.40 up to 20.0 0.20 up to 20.0 1) Mass flow controllers 2) Working volume 12.6.6 Antifoam Description Value Sensor (optional!) Conductive with dosing needle Adjustable in height Control Peristaltic pump AF”(Antifoam) Range 0 / 100 % (ON/OFF) 12.6.7 pH Description Value Control 2 peristaltic pumps (acid, base) and/or CO2 valve Control range pH 2 up to 12 Measuring accuracy pH ± 0,1 Labfors 5 - Version for Microorganisms Technical Data 04 September 2019 Page 211 of 216 Variants Measurement system Sensor (optional) METTLER (analogue) Traditional pH sensor (potential measurement against reference) Manufacturer: METTLER TOLEDO Measuring range: pH 2 up to 12 HAMILTON (digital) Traditional pH sensor (potential measurement against reference) with integrated electronics, type Easyferm Plus ARC Manufacturer: HAMILTON Measuring range: pH 0 up to 14 INFORMATION HAMILTON pH sensors are preconfigured before dispatch by the equipment manufacturer INFORS HT. Replacement sensors of this type must be reconfigured before use! Details on technical data, use and maintenance of the pH sensors can be found in the separate documentation of the sensor manu- facturer. 12.6.8 pO2 Description Value Control Cascaded stirrer Cascaded flow Cascaded gasmix Cascaded O2 addition The functionality of the parameters depends on the hardware configuration of the equipment Control range 0 – 100 % Measuring accuracy 1 % FS Labfors 5 - Version for Microorganisms Technical Data Page 212 of 216 04 September 2019 Variants Measurement system Sensor (optional) METTLER (analogue) Traditional amperometric/polarographic pO2 sensor Manufacturer: METTLER TOLEDO Measuring range: 0 - 150 % HAMILTON (digital) pO2 sensor with integrated optical electronics of the type Typ Visiferm DO ARC Manufacturer: HAMILTON Measuring range: 0.05 % - 300 % air sat- uration INFORMATION HAMILTON pO2 sensors are pre-configured from the equipment manufacturer INFORS HT. Replaced sensors must be configured again before use! Details about technical data, use and maintenance of the pO2 sensors are in the separate documentation from the sensor manufac- turers. 12.6.9 Pumps Description Value Type Peristaltic Standard Digital (3 pieces) Acid Base AF = Antifoam Analogue (1 piece) Feed Rotation speed Digital 74 min-1 / fixed rotation speed Analogue 74 min-1 / max. rotation speed, adjustable within range of 0 % up to 100 % Accuracy ± 1 % FS Labfors 5 - Version for Microorganisms Technical Data 04 September 2019 Page 213 of 216 Pump hoses & flow rates Description Value Material PharMed BPT Standard iD: 1.0 mm Wall thickness: 1.1 mm Flow rate: 3.5 ml min-1 Option 1 iD: 0.5 mm Wall thickness: 1.15 mm Flow rate: 1.2 ml min-1 Option 2 iD: 2.5 mm Wall thickness: 1,0 mm Flow rate: 17.2 ml min-1 Flow rates All flow rates at 74 min-1 (100 % rotation speed) 12.6.10 Sterile Filters Description Value All filters Autoclavable Disposable Inlet air filter (red label) 1) Retention rate: 0.2 µm Diameter: 37 mm Inlet air filter (no label) 2) Retention rate: 0.2 µm Diameter: 25 mm Exit gas filter (green label) 3) Retention rate: 0.3 µm dry, 1.0 µm wet Diameter: 37 mm 1) for vessels with DN 115 and 150 2) for vessels with DN 200 3) for all vessel sizes Labfors 5 - Version for Microorganisms Technical Data Page 214 of 216 04 September 2019 12.6.11 Heater Pads for Single Walled Vessels Description Value Material Silicone IP protection class X4 Electrical voltage 230 V 115 V Dimensions mm Vessels Total volume (L) Diameter nominal (mm) Electrical power W Length Width 2.0 / 115 200 200 370 90 3.6 / 115 400 400 370 170 3.6 / 150 400 400 475 140 7.5 / 150 400 400 475 140 13 / 200 1000 500 655 250 12.7 Operating Conditions Description Value Temperature range 5 °C up to 40 °C Relative air humidity, non-con- densing 20 % up to 90 % Altitude metres above sea level max. 2000 m.a.s.I Degree of pollution (as per EN 61010-1) 2 Min. distance from walls, ceilings and other appliances 150 mm 12.8 Emissions Description Value Units Noise emission <70 dB (A) Labfors 5 - Version for Microorganisms Technical Data 04 September 2019 Page 215 of 216 12.9 Utilities Glycerine Lubricant for mechanical seal. Permitted product:  Medicinal Glycerine 85 %  Quality: PhEur Bottles of 150 mL are available. 12.10 Auxiliary Supplies pH Buffers pH buffers are used to calibrate the pH sensors. 250 mL bags are available for the following buffers:  pH 4.04  pH 7.01 EG-Konformitätserklärung EC-Declaration of Conformity Déclaration de conformité © Infors AG, Rittergasse 27, CH-4103 Bottmingen, T:+41 (0)61 425 77 00, F:+41 (0)61 425 77 01, E-Mail: info@infors-ht.com Doc. FO094- V. 11.0 Gemäss der EG-Maschinen-Richtlinie 2006/42EG 2006, Anhang II 1 A In accordance with the EEC machine directive 2006/42EG 2006, appendix II 1 A D´après la directive relative aux machines 2006/42EG 2006, annexe II 1 A Hersteller: Manufacturer: Constructeur: Infors AG Rittergasse 27 CH-4103 Bottmingen Bezeichnung: Designation: Désignation: Tischbioreaktor Bench-top Bioreactor Bioréacteur de paillasse Typ: Type: Type : Für Version: For Version: Pour Version: Labfors 5 Mikroorganismen, Zellkulturen, feststoffhaltige & enzymatische Bioprozesse und phototrophe Organismen Microorganisms, Cell Cultures, Solid Substrates & Enzymatic Bioprocesses, Phototrophic Organisms Microorganismes, Culture cellulaire, Substrats solides et bioprocédés enzymatiques & Organismes phototrophes Baujahr: Year of construction Année de construction 2019 Gültig ab Seriennummer: From serial number: A partir du numéro de série: S-000128630 Dieses Gerät entspricht den Grundlegenden Anforderungen der Richtlinien: This equipment is in compliance with the essential requirements of Directive: Cet appareil est conforme aux exigences essentielles de la Directive: Maschinenrichtlinie 2006/42/EG Directive on machinery 2006/42/EC Directive relative aux machines 2006/42/CE EMV-Richtlinie 2014/30/EU EMC directive 2014/30/EU Directive CEM 2014/30/UE Aussteller: Issuer: éditeur: Bevollmächtigter für die technische Dokumentation Person authorised to compile the technical file Person autorisée à constituer le dossier technique Infors AG Rittergasse 27 CH-4103 Bottmingen F. Berz Anschrift Address: Adresse: Konformitätsbeauftragter Representative for Conformity Responsable des certificats de conformités Bottmingen, 02. Jan. 2019 L. Gutzwiller Director Technical Support Ort, Datum: Place, date: Lieu, date: