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Guide to Operations

1. 1 AS MIX OPTIO l LL YEL lI eo i 1 LvES 10 4 E 17 El E LK HT E LI N TACH SET POINT CelliGen BLU M1363 0050 User s Guide 119 Figure 49g Control Schematics 120V DIP SW SETTINGS FFLLLLLL A 17 E T A n 1363 8017 Y TMFC ANALOG 1 0 NA gt 7 A 7 M1363 8017 TMFC A AS 1363 8017 Y TMFC M A a LEL Naeeeoir TMFC 1 M1297 7002 Analog I O Board ALL JUMPERS SET TO THIS LOCATION El Number SW SW SW SW TMFCs Pos l Pos2 Pos 3 Pos 4 NA ON ON ON ON 11 ON OFF ON ON MI363 2012 7 ON OFF ON OFF M1363 2013 4 oN OFF OFF_ OFF New Brunswick Scientific User s Guide 120 Figure 49h Control Schematics 120V ETHERNET ISB alias lle OL E TB2 2 Ds K BLK cov Eb 1 y 3LK rio NOT USEI N J TERMINATION BOARI LOAD CELL SCALE A M1361 IFTMFC 3 lt J4 NOT USEL AZA
2. Use this button to return to a previous screen New Brunswick Scientific User s Guide 94 Figure 44 Sample Loop Alarms Screen pH A Alarms Screen New Brunswick Scientific Cell Culture Mode Choose loops to be shut down wh Selected Loop Setpoint PY this alarm is triggered 7 0 Agit Temp pH 7 00 8 04 Pump1 Pump2 Pump3 DO Volume Air 1 Absolute 02 2 N2 3 Low Limit High Limit eee OviFlo OvIAir Ovlo2 2 i OvIN2 OvlCO2 Deviation Low Limit 2 00 r Enable Audible ysy Trend o Pumps Mams Le Setup EE summary ii Calib 5 1 Pressing any of these loop buttons will cause the selected loop s to shut down when an alarm is triggered for the loop whose Alarms screen this is in this case pH Step 3a If you wish to set an Absolute alarm enter an Absolute Low Limit here Step 3b and enter an Absolute High Limit here Step 4a Press the Enable checkbox to enable the Visual alarm Step 4b and check the Audible checkbox to enable the Audible alarm WIN oje 5 Ifyou wish to set a Deviation alarm use the Deviation pane and follow the same procedure as outlined in Steps 3a 4b above 6 Use the Scroll Back button lt lt lt to return to the main Alarms screen and follow these steps for any other alarms you wish to set or press the Summary button to return to the SUMMARY screen 14 3 A
3. 3 The Totalizer feature tracks and displays the accumulated flow for the associated loop here it measures accumulated air flow 4 Use the Reset Totalizer button at any time to return to 0 0 and begin counting again CelliGen BLU M1363 0050 User s Guide 77 o NOTE To reduce the risk of clogging the microsparge gas flow should not be set below 2 CCM Figure 33b Sample Cascade Screen with 4 TMFCs a Cascade Screen New Brunswick Scientific Cell Culture Mode EU BLU All Exhaust Heater Cascade From po Me To DO Start Out EndSetpoint DO End Out le Start Setpoir Enabl AirFlo 1 Ed 20 0 30 0 60 0 O2Flo 2 v 0 0 10 0 20 0 100 0 No N2Flo 3 gt SH oo 20 0 50 0 100 0 No No None e None hd Ed summary i cae od trena Pumps A narms 2 Setup 1 Press each loop s Enable button to activate automatic cascade control Cascade values are automatically controlled but first they need to be activated by selecting each loop s Enable button in the CASCADE screen see Figure 33b Default values are provided as starting points they may need to be modified for the optimization of individual processes When you set the system to the 4 Gas Auto control mode see Figure 33c on the next page the controller automatically adjusts the flow ratio of Air O2 CO and N based on the demands of pH and DO The individua
4. Limits Set Low Set High o0 100 0 Decimal Places Alarm Settings 0 0 0 0 0 0 EDN o cai Ez case Trena Pumps A Alarms 2 setup To fine tune your cascade settings you can adjust the Output and Setpoints in the Cascade screen see Figure 31e for a sample Cascade screen and Section 10 for details on Cascades Figure 31e Sample Cascade Screen with 3 or 4 Gas Mix 1 a Cascade Screen New Brunswick Scienti Cell Culture Mode CelliGen BLU Cascade From DO z To Enable Start Setpoint DO tart Out End Setpoint DOfEnd Out GasFlo gt 20 1000 30 0 o0 EE summary i cae gr Trend 0 Pumps A Alarms Le setup 1 DO starting Output 2 DO ending Output CelliGen BLU M1363 0050 User s Guide 73 O NOTE When your system is equipped with 1 TMFC and 4 solenoids you may see some fluctuation in the GasFlo loop PV process variable as the solenoids pulse between gases This is completely normal 11 3 Gas Control with 3 TMFCs If your system is equipped with three TMFCs there are no solenoids and you have access to Manual Cascade or 3 Gas Auto control modes in the gas gauge screens see Figures 32a amp 32c Figure 32 Controller Setup Screen 3 TMFCs Pe Setup Screen New Brunswick Scientific Cell Culture Mode CelliGen BLU Controller Setup Recipe Manager System Settings Hardware Setup Security Se
5. New Brunswick an eppendorf company Guide to Operations CelliGen BLU Single Use Bioreactor MANUAL No M1363 0050 Revision C January 18 2011 New Brunswick Scientific PO Box 4005 44 Talmadge Rd Edison 08818 4005 USA 1 800 631 5417 1 732 287 1200 bioinfo nbsc com www nbsc com CelliGen Excella Innova and BioCommand are registered trademarks of and are owned by New Brunswick Scientific Co Inc USA CLAVE NeedleFree is a registered trademark of ICU Medical Inc in San Clemente California USA Luer Lock is a trademark of Becton Dickinson and Company in Franklin Lakes New Jersey USA Tri Clamp is a registered trademark of Ladish Co Corporation Alfa Laval Inc Richmond Virginia USA Windows is a registered trademark of Microsoft Corporation in the United States and other countries New Brunswick Scientific has attempted to identify the ownership of all trademarks from public records Any omissions or errors are unintentional CelliGen BLU e M1363 0050 User s Guide CONTACT US New Brunswick Scientific maintains regional sales and service offices throughout the world to best serve you To locate the office nearest you see the Contact Us section of our website at www nbsc com or contact us at our world headquarters New Brunswick Scientific 44 Talmadge Road Post Office Box 4005 Edison New Jersey 08818 4005 USA Tel 1 732 287 1200 Toll free in North America 1 800 63
6. 29a 29b 29c amp 29d on the next pages New Brunswick Scientific User s Guide 66 Figure 28 Controller Setup Screen 0 TMFC I gt Setup Screen New Brunswick Scientific Cell Culture Mode CelliGen BLU Controller Setup Recipe Manager System Settings Hardware Setup Security Settings Unit Type Unit Name CelliCen BLU CelliGen BLU No of TMFCs 0 TMFC Range NA Installed Options V Gas Mix V Vessel Volume T7 Gas Overlay Module Y Overlay TMFC E summary Calib a Case K Trend 0 Pumps A narn EA See Section 11 5 for details on the Gas Overlay The system will automatically generate three or four gas process loops on the SUMMARY screen These loops are labeled Air 1 O2 2 N2 3 and CO2 4 Their numbers 1 4 correspond to the gas connections on the cabinet see Figure 2 When you open the gauge screen for any of these gas loops you will find a series of buttons for flow control The gauge screen for each of the gases will have the option for 3 or 4 gas mixing but no cascade feature to the GasFlo loop Total gas flow is regulated manually by means of an external rotameter In addition if 3 Gas Air O2 and CO or 4 Gas Air O2 CO and N2 mode is selected the controller will automatically maintain gas composition percentages based on pH and DO The Manual gas mix control button allows you to manually adjust the gas mixture see Figures 29c amp 29d Whatever mixture you
7. Gas Mix Selection T Air 02 C02 4Gas Mix Flow Cycle Times Seconds On Time 120 Off Time 0 i 2 1 7 Manual Limits Set Low Set High 0 0 100 0 Decimal Places EI Y e Casc Ls rena Pumps A narms Ao setup 1 Press the Manual button to control gas mix manually You must now control the 4 Gas Air O2 CO2 amp N2 percentages manually either by the use of cascades or by entering gas percentage values in each gas loop gauge screen 2 Set Mix Flow Cycle Times typically used in microcarrier applications by indicating here the number of seconds this gas should cycle On and Off CelliGen BLU M1363 0050 User s Guide 69 The flexibility of gas control through these gauge screens makes it easy for you to change gas mixes during the various phases of your cell culture run As you change mixes if you change the number of gases entering the vessel your choices will be reflected in the process loops that appear in the SUMMARY screen Systems with 0 or 1 TMFC will still allow you to set manual control of the gas mix and gas flow It is important to remember than when the system is set to Manual it will not automatically change the mix based on pH or DO and that whatever mixture you set Air always makes up any difference to complete the 100 total gas 11 2 Gas Control with 1 TMFC If your system is configured with 1 TMFC gas mixing is controlled thr
8. lt lt or gt gt gt button both are now red and active to move the line one click at a time for more precision see Figure 38 Figure 38 Selecting a Read Line Location 1 Press anywhere along the desired vertical axis to locate the read line 3 Press the Read Line button again to return to the regular trend graph If you wish to use the zoom mode see Section 12 1 1 and the read line at the same time you must enter Zoom mode first New Brunswick Scientific User s Guide 86 13 ABOUT PUMPS After assigning the pumps see Section 9 5 you will need to select a setpoint and a control mode for each calibrate their flow rates and select their pulse periods This section will walk you through those operations There are three standard pumps on the front right of your control cabinet see Figure 23 repeated here for easy reference Remember to set up any optional pumps you may have added to your system see Section 13 5 to install an external Barnant VS pump and see Section 0 Appendix A about optional stacked pumps Figure 23 Standard Pump Array pumps open AN D E y E e ze Dd 1 Pump1 14 RPM Pump2 109 RPM Pump3 109 RPM 13 1 Pump Setpoint To enter a setpoint for any pump 1 Open the PUMPS screen Gauges for Pumps 1 3 are displayed in this screen see Figure 39 If you ha
9. 58 Figure 26 Pump Assignment Pane Assignment 22 MN 3 Yol Add Vol Harvy a 4 1 Step 3 Press the button that corresponds to your choice of assignment for Pump1 Step 4 Repeat Steps 2 amp 3 for the other pump s to be assigned Vol Add Volume Addition see Section 9 7 1 Vol Harv Volume Harvest see Section 9 7 2 WIN Eh 5 Press the Summary screen access button to save the pump assignment s and to return to the SUMMARY screen 9 6 Pump Calibration Pump flow rates are provided in Table 8 Section 13 3 However more accurate flow rates through the various lines may be established by pre calibrating the pumps using the PUMPS screen for details see Section 13 3 This screen controls all pump parameters for the three standard fixed speed pumps supplied with the control cabinet and for any additional pumps added through the available analog input and output connections Using the PUMPS screen you can view total pump flow rate in ml second set the pump s cycle time and assign each pump to one of four functions None Base Vol Add and Vol Harv these last two are explained in Section 9 7 NOTE To assure the most accurate flow rate calibrate the pump see Section 13 3 each time you change tubing 9 7 Using Optional Scales to Program Pumps 9 7 1 Setting a Feed Pump to Add Liquid A feed pump can be set to add liquid whenever the optional load cell scale informs
10. 6 3 Enable Security Feature Automatic Log Off Time DS 5 E 4 EN Administrators Supervisors Operators ped Summary ca Ez case Trend Pumps A nam ES When security is enabled the User button appears in this corner of all main screens This block will show the name of your unit 1 2 3 Use the dropdown menu here to define the time before the system automatically logs off leaving only the SUMMARY SYNOPTIC and TREND screens available 4 Remove User button see text below Add User button see text below 5 6 Only a user in the Administrator group has access to this feature checking Enable Security Feature turns security on deselecting it turns security off In this screen a user with Administrator status can move users from the Operators group to the Administrators group or vice versa by highlight the user name in the Administrators or Operators pane then pressing the gt gt or lt lt button to move that user from one pane to the other An Administrator can also add users to or remove users from the system using the Add User and Remove User buttons To remove a user press the user name to select it in the pane where it appears then press the Remove User button To add a user press the Add User button use the keypad screen that opens see Figure 47 on the following page to type in the user name and assign a Password if desired if you
11. C and the DO probe was polarized For this study 3 0 L of sterile CD CHO serum free medium was prepared by pre warming at 37 C for 24 hours in a CO incubator During this time the DO probe was connected to the controller for at least 6 hours to enable polarization as per the manufacturer s recommendation Once the medium was warmed and the inoculum grown to sufficient starting density the CelliGen BLU bioreactor vessel was removed from its sterile packaging and the heat blanket supplied with the unit was wrapped around the outside of the vessel Next the vessel containing the cell culture medium was connected to one of the bioreactor vessel s inlet lines using a tube welder A tube welder is offered as an optional accessory to the CelliGen BLU A pre sterilized medium filter with an attached quick connect or Luer connection can also be used if a tube welder is not available Since this was a batch process all of the medium was pumped into the bioreactor vessel All additional connections to the controller including sparge overlay RTD pH and agitation were also made pH and DO were calibrated through the touchscreen controller and all process setpoints were entered on the touchscreen using the Control Setpoint values shown below Once the parameters were at their setpoints the inoculum flasks were connected to the addition line in a sterile manner using a tube welder and contents were pumped into the bioreactor vessel Operational
12. CelliGen BLU M1363 0050 User s Guide 17 Figure 4b Headplate Arrangement shown with tubes 8 connectors 1 inch CPC guick connect for harvest 7 0 2 um overlay filter use is optional 2 0 2 um exhaust filter 8 inch CPC quick connect for Addition 2 3 Thermowell port for RTD 9 inch CPC quick connect for Addition 3 4 Port for optical pH probe 10 inch Luer Lok connect for Addition 1 5 Port for polargraphic DO probe 11 Clave NeedleFree connector for sampling 6 0 2 um sparge filter Table 2 on the following page summarizes the type length and size inner and outer diameter of all the vessel headplate tubing and their connections New Brunswick Scientific User s Guide Table 2 Headplate Tubing amp Connections Use Tubing Type Length Size Connection Filter Addition 1 C FLEX 20 bn o Vs inch Luer Lok TM mn 6 4 mm OD cennet aion onee ing Ganmo donea e Sampe oree dm Gemme e Floros C FLEX room eamm Da stick eS Exhaust pean ce T eamm Dx 0 2um filter apa nes ee Emo amater Overlay a AN isin D 0 2um filter 4 6 3 Heat Blanket amp Exhaust Tube Heat Blanket As soon as you unpack the vessel from its protective covering wrap it with the heat blanket There are two different heat blankets one for each vessel size If you have both vessel sizes be sure to use the correct blanket for each Wr
13. Figure 20b repeated on the following page Use this feature to access rename save load and delete recipe files for your cell culture runs Recipes consist of all user definable variables available on the CONTROL screens When a recipe is saved all the current settings on the controller including but not limited to setpoints control modes alarms P amp I values and cascades are saved to the controller s memory CelliGen BLU M1363 0050 User s Guide 99 You can save this data with a unique name using the Save As button see Step 2b below or overwrite an existing recipe using the Save button Step 2a Figure 20b Recipe Manager Screen Ke Setup Screen New Brunswick Scientific Cell Culture Mode Mily Exhaust Heater Controller Setup sc ENE System Settings Hardware Setup Security Settings Selected Recipe HEK293 rcp 2 Available Recipe 3 1 CHO rep Save As 4 Delete 5 Load Default Be summary cau Ta case p lt Trena Pumps A Alarms Bd Step 1 Press the recipe file of choice in the list shown in this box Step 2a Press the Save button to save the recipe as is or Step 2b Press the Save As button if you wish to rename the file use the pop up touchpad to designate a new name Step 3 Press the Load button to load the Selected Recipe file 5 Step 4 To delete a recipe from the system select it see Step 1 then press the Delete button WIM E
14. OFF OFF OFF User s Guide 128 Figure 50h Control Schematics 230V ONITO THE ET e MICROCONTROLLER ay a Jac nr CM 9586 ae ONITO TO DB 15 O ERLA TMF LK 3LK VEL WHT 3LK IFTMFC 4 lt 1297 8 NOT USED gt 1297 70 IFTMFC 3 lt wi297 8 OT USEI 7 2 J6 IFTMFC 2 lt 1297 80 OT USED IETMEC 1 M1297 8015 Y NOT ED CelliGen BLU M1363 0050 TERMINATION BOARD nee E DIGITAL SCALE LK YEL User s Guide 129 20 1 List of Drawings Figure Description Page la Front 8 Rear Views of Control Cabinet 10 1b Touchscreen to Control Cabinet Connections 11 2 Control Cabinet Service Connections 12 3 Vessel Assembly 15 4a Headplate Arrangement shown w o tubes amp connectors 16 4b Headplate Arrangement shown w tubes amp connectors 17 5 Internal Vessel Components 19 5a pH 8 DO Headplate Ports close up 20 6 Inputs amp Outputs for Ancillary Equipment 23 6a Bag Hanger 24 7 Touchscr
15. The remaining air passes through the 0 2 um exhaust filter WARNING NEVER block the exhaust to pressurize the vessel see Section 4 6 CelliGen BLU M1363 0050 User s Guide 3 10 3 11 3 12 3 13 3 14 Pressing the Exhaust Heater button at the top right of the control screens see Figure 8 for example will turn the exhaust heater on or off It should always be on when gas is flowing into the vessel Sampling System This system consists of a CLAVE NeedleFree connector attached to a sampling tube that extends to the lower portion of the vessel Both the CLAVE NeedleFree connector and the syringe use Luer Lok connections Samples can be taken by aseptically attaching a needle free syringe to the connector and drawing a sample see Section 16 4 for details Overlay System The overlay system allows gas to be inserted directly into the headspace of the vessel through a port located on the headplate Overlays can be selected as manual flow with a rotameter or as automatic flow via a thermal mass flow controller TMFC Like the sparge system the overlay gas mixture can be composed of up to four gases and its composition can be controlled automatically by solenoid valves See Section 11 5 for details Scale optional An optional load cell scale can be added to your system to display the weight volume of your culture vessel or feed vessel Cascades and feeds can be set up to pump in or pump out liquid auto
16. Use Vessel CelliGen BLU single use vessels are offered in 5 0 and 14 0 L total volume capacities The vessels are delivered pre assembled with pitched blade impeller porous microsparge and all the necessary tubing filters and connectors and come sterilized ready for use right out of the package All components in product contact are made of materials that meet USP Class VI standards and have been tested for leachables and extractables making these vessels appropriate for cGMP environments In this protocol we describe use of a CelliGen BLU with 5 0 L vessel Controller CelliGen BLU s compact non disposable control station is designed to provide advanced process management and monitoring capability ranging from three fixed speed pumps for additions and harvesting to a powerful controller with 15 industrial color touchscreen monitor Multiple options including gas flow control a weight scale validation packages and more enable customization to your needs The control station used in this protocol was configured with one 2 100 cubic centimeters per minute ccm Thermal Mass Flow Controller TMFC for direct sparging of gases and an integrated gas overlay with 0 1 3 0 Standard Liters Per Minute SLPM flow rate also regulated by a TMFC Both the gas flow and gas overlay are capable of 4 gas mixing for automatic pH and Dissolved Oxygen DO control Pumps temperature control agitation as well as all of the other process l
17. actuators that maintain the various parameters at their setpoints Using temperature as an example the controller compares the output of a temperature sensor to the user entered temperature setpoint and generates a signal to activate either a heater or a cooler to maintain vessel temperature at the temperature setpoint The controller provides the logic that generates appropriate drive signals to various actuators so that process parameters remain at their setpoints 22 2 What is a Control Loop A control loop is the basic element of automatic process control Three components comprise one control loop a sensor a controller and an actuator Based on information from a sensor the controller generates an actuator control signal that maintains a parameter at its setpoint Control will fail if any element in the control loop fails CelliGen BLU M1363 0050 User s Guide 135 22 3 22 4 What is Probe Calibration In bioprocess control calibration generally refers to establishing a correspondence between a probe s output and the actual value of whatever that probe senses For example pH probes are often calibrated with pH 7 0 and pH 4 0 buffers to establish a zero pH 7 0 and a span pH 4 0 Other buffers can be used but the principle is always the same For any probe calibration two values a zero and a span are required for the controller to correctly translate inputs from that probe DO and pH probes are ro
18. and DO On day 3 and for the remainder of the run 5 10 ccm of gas were directly sparged into the system using a porous sparger and automatic 4 gas mixing The overlay gas flow in the vessel headspace was kept at the previous settings A built in sampling device enabled sterile sampling Daily off line measurements of glucose and lactate concentration were read using a YSI 2700 and cell density and cell viability was measured using an Automated Cell Counting System NBS NucleoCounterTM CelliGen BLU M1363 0050 User s Guide 143 24 4 Results and Discussion All data was logged via BioCommand Batch Control PC compatible Supervisory Control and Data Acquisition SCADA software New Brunswick Scientific As shown in Figure A the CHO cells in this study grew steadily reaching a maximum viable cell density of 5 55 x 10 cells mL on day 5 Cell viability shown in Figure B ranged between 97 1 97 9 through Day 5 until the nutrient source glucose was depleted from the medium as shown in Figure 3 Figure C shows glucose consumption vs lactate production during the process As expected lactate production steadily increased as the available glucose in the medium was consumed As glucose in the medium become exhausted consumption of lactate as a secondary carbon source also declined This data demonstrates that the CelliGen BLU bioreactor is an easy to use efficient system for the culture of CHO cells No effort was made
19. be the most recently visited of the two That is if you leave the SUMMARY screen to view the TREND screen for example the far left button will be labeled SUMMARY 2 It is very important to be sure the exhaust heater is on when you are actively growing cells to reduce the risk of clogging the exhaust filter 6 2 4 Synoptic Screen From any main screen press the far left SYNOPTIC button to open the SYNOPTIC screen see Figure 9 on the following page If the far left button says SUMMARY press it to open the SUMMARY screen The button will now be labeled SYNOPTIC press it again This screen provides a visual representation of all the loops their settings and current process values the sample screen below may not be exactly like the screen you see it depends on your system s options This screen provides all the functionality of the SUMMARY screen with the exception of the ability to add loops New Brunswick Scientific User s Guide 32 Figure 9 Sample Synoptic Screen of Synoptic Screen New Brunswick Scientific Cell Culture Mode uma Mil Exhaust Heater 1 pte Heater 50 00 1 00 A f cai Ta Case trena Pumps A Atarms 2 setup 1 LOOPS Each gauge indicates setpoint SP and process variable PV Title color indicate the loop s status Red OFF Green ON and Blue MANUAL Touch the loop gauge in this screen to open th
20. below for easy reference Figure 17 Trend Screen pS Trend Screen New Brunswick Scientific Cell Culture Mode 06 40 08 40 10 40 12 40 14 40 EE summary Li cai 8 case o Pumps A Atarms A setup 1 Step 2 To select the first loop you wish to display press the red Setup button New Brunswick Scientific User s Guide 82 Figure 35 Trend Setup Screen pe Trend Screen New Brunswick Scientific Cell Culture Mode None 1 Loop Display High 1200 3 Temp Pump1 Display Low 50 Sample Rate Secs 4 Maximum Log Time Hours GasFlo Display Color WW 2 i Step 3 Select the first loop The program will automatically place it in the red box Step 4 If you wish to change the color of this loop press the new color choice here WIN Step 5 Press the Display High box to enter using the touchpad the high limit for the Y axis then use the Display Low edit box to set the low limit 4 Step 6 Press the ramp up gt gt gt or ramp down lt lt lt button to select the desired data sampling interval 5 15 30 or 60 seconds 7 Press OK to save your choice and return to the TREND screen or Cancel to return to the TREND screen without saving any changes 8 Repeat Steps 2 7 selecting a different color for each loop up to a maximum total of 8 loops 9 With reference to Figure 36 a sample Trend Graph in progress and Table 6 acqua
21. change his her password ADMIN is the default password 2 When Security is enabled only the Synoptic Summary and Trend navigation buttons remain active the other navigation buttons will be greyed out and inaccessible New Brunswick Scientific User s Guide 104 16 PERFORMING A CELL CULTURE RUN 16 1 Preparing for Start Up 1 Add sterile filtered media Aseptically connect the addition port tubing to the addition vessel which contains the media This can be done using a tube welder or by making the conection under a hood Open the addition tubing clamps Using gravity or a peristaltic pump move the media into the vessel Close the addition tubing clamps Set temperature control to the desired working temperature Check that agitation is in OFF mode Connect the motor Set agitation to the desired speed ae ee Bel caurion Agitation can only be increased 25 RPM at a time any greater increase will cause the magnetic drive to uncouple 6 Set mode to AUTO 7 Draw a sample from the vessel see Section 16 4 for the sampling procedure 8 With an external pH meter measure the pH of the liquid noting its pH and temperature 9 Adjust the display reading of the pH function to read the value displayed on the external pH meter Make a correction for the pH value if the temperature of the vessel is different from that of the sample During prolonged cell growth it is advisable to take a sample
22. dies di 0 0 0 0 00 EE Calib bd Casc bs Trend 0 Pumps A Alarms Le Setup 1 If you select Air 02 CO2 here the 4 Gas Gas Mix Control button will change to 3 Gas allowing you the option of switching to automatic 3 Gas control New Brunswick Scientific User s Guide 68 Figure 29c Air 1 Gauge with 0 TMFC amp Manual 3 Gas Control fall Gauge Screen New Brunswick Scientific Cell Culture Mode CelliGen BLU Air 1 Setpoint 100 0 Set Py 1000 100 0 Gas Mix Selection M Air 02 C02 T 4Gas Mix Flow Cycle Times Seconds On Time 120 Off Time O a 2 1 Manual Limits Set Low Set High o0 1000 Decimal Places Alarm Settings 000 0 0 000 cano Casc pe Trena 9 Pumps A Alarms I gt setup 1 Press the Manual button to control gas mix manually You must now control the Air O2 amp CO percentages manually either by the use of cascades or by entering gas percentage values in each gas loop gauge screen 2 Set Mix Flow Cycle Times typically used in microcarrier applications by indicating here the number of seconds this gas should cycle On and Off Figure 29d Air 1 Gauge with 0 TMFC amp Manual 4 Gas Control full Gauge Screen New Brunswick Scientific Cell Culture Mode CelliGen BLU Air 1 Setpoint 100 0
23. error from PV current process value error meaning any readings that are above or below the programmed setpoint As the error discrepancy increases or as the duration of such a discrepancy remaining static increases the Output also increases 3 End Setpoint is the loop value the user defines as the maximum allowable value when the DO End Out is reached Typically this value will also be the same as the system s maximum allowable setpoint for the loop 4 DO End Out represents the DO output where the user wants the cascade to stop This value can be set to any integer from 0 to 100 as long as it is greater than the Start Out The greater it is than the Start Out the smoother the increase in setpoints O NOTE E It is important to remember that cascades are based on the loop whether DO or pH Out value posted in the SUMMARY screen These numbers are the basis for all cascades involving that loop See the examples below for more explanation It can be a very beneficial exercise to watch how the values on the SUMMARY screen change to reflect differences between the present value PV the setpoint and the DO output percentage Out When the PV is greater than the setpoint the system will be generating a negative Out because the controller senses a need to decrease DO New Brunswick Scientific User s Guide 62 LoopName Setpoint Out Control Mode Units Casc When the PV is less than t
24. for a GasFlo cascade directly from a gas gauge screen see Figure 31b The cascade will regulate gas flow from minimum to maximum based on the system s pH and DO demand If the system has no demand for increasing DO or adjusting pH the GasFlo will go to the GasFlo Cascade Min setting As soon as the system calculates a demand GasFlo will begin to increase Figure 31a Air 1 Gauge Screen with 1 TMFC amp 3 Gas Control ful Gauge Screen New Brunswick Scientific Cell Culture Mode CelliGen BLU Air 1 100 0 Setpoint 100 Gas Mix Selection M Air 02 C02 T 4 Gas Mix Flow Cycle Times Seconds Set Py Dut a 100 0 100 0 1000 SAGE On Time 120 Off Time O 100 0 100 0 EE Limits GasFlo Cascade Min Set Low Set High 0 0 0 0 100 0 GasFlo Cascade Max 1 100 0 Decimal Places 0000 00 00 Alarm Settings wu an 0 0 0 0 summary Wg cai 3 Casc Es renal Pumps Alarms 2 Setup 1 GasFlo Cascade minimum 8 maximum settings In 3 gas mode the controller automatically adjusts the gas Air Oz amp CO2 percentages to control pH and DO Total gas flow from the TMFC can be directly linked to a cascade from this screen using the default GasFlo Cascade values see tem l in Figure 31a or setting your own This allows you to minimize the amount of gas going into the vessel when DO correction is needed it also limits the total ga
25. normal operating conditions and red when there is an alarm condition Press a LoopName box to open that control loop s alarm screen ABSLow This column indicates the Absolute low limit you program for control loops An alarm is triggered if the loop PV falls below this point ABSHigh This column indicates the Absolute high limit you program for control loops An alarm is triggered if the loop PV rises above this point ABSEnable ABSAudible This column indicates whether the Absolute alarm limits have been enabled Active or not InActive for visible ABSEnable and or audible ABSAudible alarms DEVLow This column indicates any tolerance you have set below the control loops setpoints DEVHigh DEVEnable DEVAudible This column indicates any tolerance you have set above the control loops setpoints This column indicates whether the Deviation alarm limits have been enabled Active or not InActive for visible DEVEnable and or audible DEVAudible alarms Acknowledge All button Press this button to acknowledge and stop all alarms Current Alarms button Press this button to open a screen that addresses any current alarm condition History Button Scroll Up w or Scroll Down 0 Press this button to open the historical record of alarms for the current run Use this button to scroll upwards or downwards in the table onscreen Scroll Back lt
26. offered to help minimize downtime from unexpected malfunctions in equipment operation Speak to your NBS sales representative for more information CelliGen BLU e M1363 0050 User s Guide vii BIOREACTOR INFORMATION SHEET On this page record the information for your bioreactor and retain this for future reference MODEL NUMBER VOLTAGE SERIAL NUMBER The above information can be found on the electrical specification plate Purchased with the following installed options New Brunswick Scientific User s Guide viii 1 2 3 TABLE OF CONTENTS WARNINGS amp CAUTIONS iii diua Nad FFY iria reir uakeeeteedtconidoskece 1 1 1 WARIO a 1 1 2 CAUTIO NS EN Y TN A 1 INSPECTION UNPACKING OF EQUIPMENT ccssssscssssssscccsssssccsssssecssssscces 3 2 1 INSPECTION OF BOXES Wa Gen 3 2 2 PACKING LIST VERIFICATION ccccccccccccscsssssssscceseccceceesssseceeesecseeesenssaeceseceesesenenseaeeesecs 3 2 3 BASIC CCOMPONEN TS coi A A ERS bs FYW 3 INTRODUCTION amp OVER VIEW swvccssccssccsscceneoseccoondsscevnnsdvcesaadvosvensedaccsodaccvonsvescsense 4 3 1 SN AN 4 3 2 MESS NL ND UN a ONR DA E EN AIR Eiu FAN 4 33 AGITATION SYSTEM sn We VE GW DW FF ak os AS eet 4 3 4 TEMPERATURE CONTROL ais oa 5 3 5 AER ATON AA Y Ho ddd FY HR a 5 3 6 PUMP Sian id DG AOA MR 5 3 7 PHEC DINKHA VAAN NND REE A NU ead GR o 6 3 8 DO CONTRO E tua lts 6 3 9 EXHAUSE SYSTEM oda 6 3 10 SAMPLING SYSTEM sortida A DYD E 7 3 11 COMERE
27. set from this screen entering values in the GasFlo Cascade Min and Max edit boxes to automatically increase and decrease the overall gas flow based on pH and DO demand Figure 31c Air 1 Gauge Screen with 1 TMFC 8 Manual 3 Gas Control ful Gauge Screen New Brunswick Scientific Cell Culture Mode CelliGen BLU i o Air 1 100 0 Gas Mix Selection 100 0 Set Py Ou Mix Flow Cycle Times Seconds 100 0 100 0 100 0 On Time 120 Off Time 0 100 0 100 0 are Limits GasFlo Cascade Min Set Low Set High 0 0 oo 1000 GasFlo Cascade Max 100 0 Decimal Places 0 0 0 0 0 0 Calib T Casc p lt trena Pumps A Aams 2 Setup New Brunswick Scientific User s Guide 72 In Manual 4 Gas control mode when you press the Manual control mode button and select 4 Gas in the Gas Mix Selection pane of any gas gauge screen you must also manually enter setpoints for each gas in its gauge screen or create a user defined cascade to control the gas mix Figure 31d Air 1 Gauge Screen with 1 TMFC amp Manual 4 Gas Control ful Gauge Screen New Brunswick Scientific Cell Culture Mode CelliGen BLU Air 1 100 0 Gas Mix Selection T Air 02 C02 M 4Gas Setpoint 2 Off 100 0 3 Set Py Out 4 Gas 100 0 100 0 100 0 100 0 100 0 Mix Flow Cycle Times Seconds On Time 120 Off Time 0
28. the pump that an addition is needed to maintain level CelliGen BLU M1363 0050 User s Guide 59 Open the PUMPS screen Select the feed pump you wish to pump liquid from your addition system see Section 8 3 into the vessel and press that pump s ASSIGNMENT button to open the PUMP ASSIGNMENT pane see Figure 26 repeated here for easy reference NO Figure 26 Pump Assignment Pane Assignment OO oo Step 3 Press the Vol Add button Step 4 Press the Summary screen access button to save the pump assignment and to return to the SUMMARY screen Ni 9 7 2 Setting a Pump to Harvest Vessel Contents A pump can be set to harvest liquid when the scale informs the pump that the harvest setpoint has been reached 1 Open the PUMPS screen Select the pump you wish to harvest contents from your vessel after aseptically setting up your harvest system and press that pump s Assignment button to open the PUMP ASSIGNMENT pane see Figure 26 above 3 Press the Vol Harv button then press Summary to save the pump assignment and to return to the SUMMARY screen New Brunswick Scientific User s Guide 60 10 CASCADE CONTROL Cascades are control schemes in which the Output of one process control loop influences the setpoint of one or more other loops In other words it uses feedback from one parameter to influence others In New Brunswick Scientific s CelliGen BLU bioreactors the output v
29. vessel 5 Carefully tear open the bag at the top Do not use sharp blade of any sort 6 Carefully slide the vessel out of the protective wrapper keeping the dome in place to hold all the tubing 7 Place the vessel next to your control station and install the heater blanket see Section 4 6 3 to protect the sensor from ambient light 8 Remove the plastic dome and remove all cable ties from the tubing attached to the vessel Figure 3 Vessel Assembly 2 3 4 5 LITER 14 LITER 1 Removable reusable motor 5 Internal components see Section 4 6 4 2 Exhaust filter 6 Addition tubes sample tubes sparge overlay 3 Headplate see Section 4 6 2 use is optional 4 Vessel 7 Heat blanketed exhaust tube see 4 6 3 New Brunswick Scientific User s Guide 16 4 6 2 Headplate Penetrations Figures 4a amp 4b show the port penetrations on the vessel headplate The ports are the same on both 5L and 14L vessels they are also labeled on the headplate itself Figure 4a Headplate Arrangement shown without tubes amp connectors 1 Sparge port 8 Harvest port 2 Overlay port use is optional 9 Exhaust heater support 3 Motor coupling 10 Exhaust port 4 Addition port 2 11 Temperature port for RTD amp thermowell 5 Addition port 3 6 Addition port 1 12 pH port 7 Sample port 13 DO port Note notches for proper alignment
30. 00 0 0 00 On Time 120 Off Time 0 100 0 100 0 100 0 EN a 2 Min Combined Flow Set Low Set High 0 0 2 0 100 0 Max Combined Flow 100 0 Decimal Places Totalizer CC 20 20 0 0 j cano 3 case lt Trena Pumps A Alarms Ke Setup N2 has been deselected and the Gas Control Mode has changed to 3 Gas Auto Ni CelliGen BLU M1363 0050 User s Guide 79 11 5 Gas Overlay Mixing When the Gas Overlay option is present and set to Automatic Flow Control with at least one TMFC in the system two additional process loops will be present in the SUMMARY screen OvlMix Gas Overlay Mix and OvIFlo Overlay Gas Flow If no TMFC is present only the OvlMix process loop will be available and the flow must be controlled manually by means of a rotameter We recommend that you always use an overlay set to 4 Gas mix to help maintain your setpoints It is especially important early in the run when there is little demand from the cultures Open the OvlMix loop gauge screen see Figure 34 to set the parameters for the Gas Overlay option Figure 34 OvlMix Gauge Screen Gas Overlay New Brunswick Scientific Cell Culture Mode Gas Overlay Mixing Mix Percentage Air 1100 0 2 02 0 0 l N2 0 0 CO2 oo Settings Max CO2 100 0 3 EXI Y ea case trena Pumps A Avarms Ao Setup 1 Step 1 Set the operating mode for the Gas Overlay by pressing the button of choice see Ta
31. 000 or higher operating system With this software you will be able to establish or change the setpoints for temperature pH DO agitation speed and pump flow rate You will also be able to read and log the process values of any parameters temp pH DO air flow pump flow rate levels and agitation that are monitored The data can also be stored plotted and afterwards transferred to other commonly available programs to be manipulated and analyzed in various ways New Brunswick Scientific User s Guide Table 4 identifies the pin designations for this 25 pin RS232 422 connector Table 4 Modbus Com Port Pin Designation Pin Number Signal Comments 1 4 6 8 11 14 20 22 23 NC not assigned 2 TXD RS232 Data Output from bioreactor 3 RXD RS232 Data Input to bioreactor 7 GND Ground reference for all signals 12 IRXD 24 IRXD RS422 paired data input to bioreactor 13 ITXD 25 ITXD RS422 paired data output from bioreactor 21 IOS Open selects RS232 Grounded selects R8422 Unless otherwise requested the baud rate is factory selected at 19200 Modbus and the connector is configured as an RS232 port i e no jumper between pin 7 and pin 21 The factory set address for the machine is 8 CelliGen BLU M1363 0050 User s Guide 45 7 PREPARATION amp CALIBRATIONS 7 1 pH Calibration The pH probe provided with this system is a reusable probe which works in conjunction
32. 1 5417 Fax 1 723 287 4222 Email bioinfo nbsc com Website www nbsc com New Brunswick Scientific User s Guide WARNING High voltage Always make sure this equipment is properly grounded WARNING This product is not designed to contain gases within the range of their lower explosion limit LEL and their upper explosion limit UEL If your process requires or produces gases be sure to verify their LEL and UEL concentration range available online E CAUTION This equipment must be operated as described in this manual If operational guidelines are not followed equipment damage and personal injury can occur Please read the entire User s Guide before attempting to use this unit Do not use this equipment in a hazardous atmosphere or with hazardous materials for which the equipment was not designed New Brunswick Scientific Co Inc NBS is not responsible for any damage to this equipment that may result from the use of an accessory not manufactured by NBS CelliGen BLU e M1363 0050 User s Guide Copyright Notice New Brunswick Scientific Box 4005 44 Talmadge Road Edison New Jersey 08818 4005 O Copyright 2010 New Brunswick Scientific All Rights Reserved Reproduction adaptation or translation without prior written permission from New Brunswick Scientific is prohibited Disclaimer Notice New Brunswick Scientific reserves the right to change information in this document wi
33. 1363 0050 User s Guide 131 20 2 List of Tables Table Description Page 1 Service Connections 13 2 Headplate Tubing amp Connections 18 3 SUMMARY Screen Features 30 4 Modbus Com Port Pin Designation 44 5 Gas Overlay Operating Modes 80 6 Trend Graph Buttons 83 7 Pump Control Modes 87 8 Flow Rate per Tubing Size 88 9 Alarms Screen Features 93 New Brunswick Scientific User s Guide 132 21 APPENDIX A STACKABLE PUMPS A novel optional feature of the CelliGen BLU pumps is that you can add another identical pump onto the front of one of the pumps in the array The pump that serves as the base however must be specially ordered to have no lower front cover and an exposed screw face on which to mount the second pump The stacked pump will be twinned to in other words controlled in exactly the same way at the same time as its base pump To stack a pump with reference to Figures 51 amp 52 1 Remove the standard pump that you wish to stack by pressing the pump release lever turning the pump to the left until it clicks then pulling it away from its baseplate mating surface see baseplate in Figure 51 2 Carefully align the center hole on the back of the special base pump with the nose of the baseplate screw face also aligning the curved slots like parentheses on either side of the hole with their male counterparts on the baseplate screw face you will need to ti
34. 3 2 3 3 3 INTRODUCTION amp OVERVIEW System CelliGen BLU is a versatile benchtop bioreactor that provides a fully equipped system in one compact package with its Reactor Process Control RPC software in cell culture mode and a 15 inch color touchscreen monitor as the user interface It can be employed for batch fed batch or continuous culture with control of 4 gas mixing pH dissolved oxygen DO agitation temperature pump feed and additional analog digital inputs and outputs Vessels The CelliGen BLU cell culture vessel is a single use stirred tank bioreactor designed for the growth of mammalian cells Each 5 0 or 14 0 liter preassembled vessel consists of a vessel body headplate pitched blade impeller with shaft and a retaining ring which supports all the internal tubing Ports are provided in the headplate for the following purposes addition 3 ports a thermowell for a resistance temperature detector RTD a sparger a harvest tube a sampler an exhaust tube and dissolved oxygen DO and pH sensors There is also a port for optional overlay use All ports and tubing come pre installed in the vessel The magnetic drive coupling is also located on the headplate see Figures 4a amp 4b for more details Each vessel comes pre sterilized with all tubing and filters already attached The tubing used for addition sampling and harvesting lines can be safely welded in a tube welder Agitation System A non disposa
35. 4 TMFCs Po Setup Screen New Brunswick Scientific Cell Culture Mode dj Exhaust Heater Ss Recipe Manager Systom Settings Hardware Setup Security Settings Unit Type Unit Name CelliGen BLU CelliGen BLU No of TMFCs 4 TMFC Range 0 100 CCM Installed Options F Gas Mix Vessel Volume Gas Overlay Module F Overlay IMF C Save Changes ES Summary Callb Casc i Trend Pumps LA wn EN 1 See Section 11 5 for details on the Gas Overlay The gas process loops you will find in the SUMMARY screen are labeled AirFlo 1 O2Flo 2 N2Flo 3 and CO2Flo 4 Their numbers 1 4 correspond to the gas connections on the cabinet see Figure 2 0 NOTE Any loop name ending in Flo represents a certain gas e g AirFlo O2Flo etc with a dedicated TMFC If the gas loop name does not end with Flo it represents Figure 33a the presence of a gas solenoid valve rather than a TMFC AirFlo 1 Gauge with 4 TMFCs in Manual Cascade Mode Gauge Screen New Brunswick Scientific Cell Culture Mode fg Exhaust Heater AirFlo 1 0 0 CCM Siainoio CM Off 0 0 ie ai o 1 1000 100 0 100 0 p P mis Sot Low Set High 2 20 1000 3 A 4 CEM caw E cae Trend Pumps 3 nama J setup 1 Manual Cascade mode has been selected the button is green NOTE Gas flow limits are 2 100 CCM SLPM not percentages
36. 50 User s Guide 53 Bel caurion Proper pH control is critically dependent on tubing size which should be as small as possible Consult Table 8 for guidance 8 3 1 Addition Tubing Size pH can be controlled by automatic additions of liquid base and CO2 Additions are triggered by the RPC controller which is constantly comparing current pH value with the pH setpoint and making adjustments as necessary The concentration of base and the inner diameter of the base addition tubing where it passes through the peristaltic pumps are critical parameters in the proper operation of a P amp I pH control system If the tubing is too large excessive doses will be added The result is that the system will overcontrol alternating in close succession between adding base then CO providing little or no change in pH reading A user selected deadband value is an aid to control pH within the user assigned range nothing will be added when the pH value falls within the deadband tolerance above or below the setpoint 5 normal solutions make a good trade off between moderate addition volume and good control characteristics The correct tubing diameter varies a little with process but inside diameters as small as 0 2 mm sometimes eliminate overcontrol while supplying sufficient liquid during high demand culture phases o NOTE Whatever the tubing ID the tubing wall thickness must be 1 16 inch 1 6 mm NBS suggests that you begin with t
37. AY SYSTEMS ida 7 3 12 SCAEETOPTTONAUT id E Gua CY FNAN 7 3 13 BAG HA NGER Stilton 7 3 14 RECOMMENDED ACCESSORIES amp SUPPLIES oocccccccnconnnononononocnononnnnononononccnononnnonanonconos 7 3 15 SUPERVISORY SOFTWARE cccessssssccsccecessessnsecececceseeesesseseeeseceesesssssaeeeseceeseeensnteaeeeeess 8 INSTALLATION Y ela nda 9 4 1 PHYSICAL LOCATION ii iia 9 4 2 ENVIRONMENT ui YW Y WYT Aa 9 4 3 INSTALLING THE CONTROL CABINET ccsccccccsccscessessaeceseccceesessnsececeecesesenssseseeesecsenenes 9 4 4 INSTALLING THE TOUCHSCREEN ccssessessececesccccsssesssececescesenessseaecesecsesssessnseeseesees 10 4 5 WTTELTES LS RW EII FWA Main AA RNa Ne WN Y Fo FN 11 4 5 1 Electric l Requirements neeaaeo a teenies 13 4 5 2 Gas Connections AAA DEG A a aba ea 13 4 6 MESSELJASSEMBIY E E E T E E E 14 4 6 1 Unpacking the Vesta E A O AA a s 14 4 6 2 HendplateP net Hain Su as cC Ud dd ds YB Did SM 16 4 6 3 Heat Blanket amp Exhaust Tube Heat Blanket YYYYYYYY YY nenn 18 4 6 4 INTERE CORDONES y dd Y Go y aati 19 4 6 5 inspect the DH Probe a ee 19 4 6 6 mil the PHD FOPE naaa dc idad 19 4 6 7 Inspecting the DO POD spices id NR wd tas ecelatet dU 20 4 6 8 DO Probe PEPITO RC awe aaa 20 CelliGen BLU e M1363 0050 User s Guide 4 6 9 Install DO Probe sive ete a a 21 4 6 10 Install Temperature RTD Probe ninas das 21 4 6 11 Install Messi iia 21 4 6 12 Install Motor ASS ENT Vasa FU FO SEA ARA 21 4 7 MAIN POWER SWITCH ii
38. Add User Defined Loop Screen E User Deninea Loop setup New Brunswick Scientific Cell Culture Mode Add User Defined Loop Remove Name Units Input Device Output Device None Loop Typo Analog In 1 Analog Out 1 Aux Analog Device Analog In 2 Analog Out 2 C Chart Rec Analog In 3 Analog Out 3 art Recorder Analog In 4 Analog Out 4 1 VS Pump 0 5V Analog In 5 Analog Out 5 Analog In 6 Analog Out 6 VS Pump 4 20mA Analog In 7 Analog Out 7 2 RS232 Input Device RS232 Output Device a Sawer SP Low Limit 0 0 RS232 Input Output SP High Limit 100 0 Case Trend Pumps A Alarms A Setup Selections for variable speed 0 5V and 4 20mA pumps Each TMFC in the system will utilize one of the 0 5V Input Output devices on the board If for example this system was configured with 1 TMFC that was assigned as l O Device 4 you would find 1 04 to be missing from the Input Device amp Output Device lists Ni CAUTION Be sure to set the dip switches correctly when using either 4 20mA or 0 5V inputs outputs New Brunswick Scientific User s Guide 92 14 ABOUT ALARMS 14 1 ABS and DEV Alarms There are two types of alarm modes you can set Absolute ABS and Deviation DEV An Absolute alarm is triggered when the control loop s Process Variable falls below the absolute Low limit or rises above the absolute High limit that you set A Deviation alarm is triggered when t
39. BLU Please take a moment to acquaint yourself with the content of each message Page numbers are provided so you can review the message and its application within its overall context 1 1 WARNINGS WARNING Page High voltage Always make sure this equipment is properly grounded iv 13 This product is not designed to contain gases within the range of their LEL amp UEL NEVER block the exhaust to pressurize the vessel Do not use this equipment in a hazardous atmosphere or with hazardous materials for which the equipment was not designed 11 14 NEVER OVER PRESSURIZE A CULTURE VESSEL Always turn your CelliGen BLU off and disconnect the power cord before performing maintenance 54 109 111 NO ONE BUT A PROFESSIONAL SERVICE PERSON should touch electric or electronic parts or assemblies in the electrical cabinet 1 2 CAUTIONS CAUTION 110 Page This equipment must be operated as described in this manual If operational guidelines are not followed equipment damage and personal injury can occur Please read the entire User s Guide before attempting to use this unit Do not use this equipment in a hazardous atmosphere or with hazardous materials for which the equipment was not designed NBS is not responsible for any damage to this equipment that may result from the use of an accessory not manufactured by NBS Do not expose the pH sensor within the vessel to direct light or t
40. E 5 1 Dip switches 1 3 to toggle from 4 20mA 4 USB connections are provided for up to 0 5V down for analog input and updates or data export output 2 These switches 1 3 can be set to 4 30 5 The MODBUS connection is provided for mA or 0 5V switches 4 7 are for 0 5V the use of NBS BioCommand see only Section 4 8 3 Capped as standard not used at this 6 Currently not available for use for future time expansion New Brunswick Scientific User s Guide 24 4 10 Install the Bag Hanger If you plan to use the bag hanger provided with the system install it with reference to Figure 6a on the following page on the two hinge pins located on the righthand side of the controller Figure 6a Bag Hanger p LN 1 0 4 3 i Es di a EM 4 1 Bracket hinge 3 Hinge pin on controller 2 Thumbscrew 4 Adjustable foot 1 With the three hooks at the top of the bracket and the adjustable foot at the bottom orient the long side of the bracket against the side of the controller sliding its two hinges onto the pins that are mounted on the controller 2 The bracket swings freely on the hinges unless you choose to fix it in place at the desired angle by tightening its two thumbscrews against the controller 3 Be sure to tighten the adjustable foot to secure the bracket on the benchtop before you add any contents to the bag s
41. FERMENTATION OB 297 7000 RE ME ae TE M LK 0 o co Ll un te case ae dnn Tpe AS MIX DVERL i EL patos a ae AS OVERLAY OPTIO gt lt Leesa tS RASE 1 gt lt lt _______ A LK pl Noy OVERLAY OVERL M Z AS MIX TMF RE l y y LVES e j LK E A lt DE E LK lt E een Ito ED GA ed aoa Cie l ES Ma rs a ee a a Y nhs ae J 1 IX OPTIO l aL EL II tt MIX LVE 0 E REI E 3LK HT E Li N N J TECH SET POINT CelliGen BLU M1363 0050 User s Guide 127 Figure 50g Control Schematics 230V O 11 io i TB2 2 OM LK TBe RESEN E 491 BASIC FERMENTATID B pH SENSOR a DD MP1 JMP2 JMP3 MP4 J Oo 0 3 30 LAMAN faa DO 410 ALL O IA S DO gt gt HRE lona DIP Sw SETTINGS AA L mi363 8017 TMFC ANALOG 1 0 N 1363 8017 TMF C M1363 8017 TMFC EL waea snt TMFC New Brunswick Scientific M1297 7002 Analog I O Board 2 ALL JUMPERS SET TO THIS LOCATION E Part Number SW SW Sw SW Numbers TMFCs Pos 1 Pos 2 Pos 3 Pos 4 IM1363 2010 N A ON ON ON ON IM1363 2011 ON OFF ON ON M1363 2012 3 ON OFF ON OFF IMi363 2013 4 ON
42. Guide 123 Figure 50c Control Schematics 230V 0420 32 RE CEM TB1 1 230V 3 3LK E JORG END GAR FU HT 3 mozal NHT TE TACH A N Lk e E HT e TACH New Brunswick Scientific User s Guide 124 Figure 50d Control Schematics 230V FILTER RF 002 Ol O RH eo RFI FILTER El HT CelliGen BLU M1363 0050 User s Guide AAA AO E 125 Figure 50e Control Schematics 230V Q D E 0 K ED 24N Tpo d ll Ll OM LK 4 ar o Be E HEATE LK u EX HEATER 3 TS RE EXH4UST 7 T LK HEATE J E ED LK LK 11 YEL E Tpo a U z E ED EDR Lt 3LK 8 E ES A 3LK a 3 ln 15253 T E LK 3LI M1363 TI ne LK ha Ll H TEMPERATURE i SENSO t E RTI ll TT Ll a ES o New Brunswick Scientific BASIC FERMENTATION User s Guide 126 Figure 50f Control Schematics 230V OY BASIC
43. NOTE Figure 27 is an example of a DO cascade for a system configured with 0 or 1 TMFC Gases are represented in ranges between 0 1009c Figure 27 Sample DO Cascade a Cascade Screen New Brunswick Scientific Cell Culture Mode CelliGen BLU Cascade From b4 To Enable StartSetpoint DO Start Out End Setpoint DO End Out loz 2 B oo 50 0 Mily Exhaust Heater 100 0 100 0 summary caw os K Trena Pumps A Alarms J setup 1 O2 Start 8 End Setpoints are set to increase between 0 100 2 This column indicates DO Output at the beginning of the cascade Assuming that DO output ranges from 100 to 100 a cascade setup that is enabled as explained below will cause the air and oxygen setpoints to vary with DO output If the air alone cannot maintain the DO setpoint O2 will increase to add oxygen or a combination of air oxygen In the cascade shown above once the DO Output increases to 50 the PV is below the SP the oxygen loop begins to increase from its staring setpoint of 0 to an ending setpoint of 100 As the O increases the Air will decrease by the inverse percentage The total Gas will remain 100 Alternately N2 can be used in a cascade loop to drive the DO down Using this method when the DO output decreases below zero nitrogen is directed to lower the DO Cascades like this can also be set up for pH where t
44. New Brunswick Scientific User s Guide 98 Figure 20a Controller Setup Screen 2 Le Setup Screen New Brunswick Scientific Cell Culture Mode CelliGen BLU Controller Setup Recipe Manager system Settings Hardware Setup Security Settings Unit Type Unit Name ColliGen BLU CelliGen BLU No of TMFCs 1 TMFC Range 0 100 CCM Installed Options Gas Mix Vessel Volume T7 Gas Overlay Module Overlay TMFC EE Summary Us Calibrate F Cascade LS Trend Pumps A narm setun 1 Press inside the Unit Name edit box to open a touchpad The name you write in this box will appear on a colored button tub on the top menu line 2 Step 2 The new Unit Name button tab appears here 3 Step 3 Press the Save Changes button to save the new Unit Name 4 Installed Options The controller automatically detects all factory installed options present and reflects them here with checks in the appropriate checkboxes M Name the bioreactor using the Unit Name edit box touchpad You may wish to simply name the unit by its default designation in the Hardware Setup screen see Section 15 4 Unit 1 The Save Changes button saves your new Unit Name If you leave this screen and wish to save any change you made be sure to press the Save Changes button before you move to another screen 15 2 Recipe Manager Press the second tab in the SETUP screen to open the Recipe Manager screen see
45. OLARIZATION ccsssssccssecccscsesssseceeecccseessesssseeesecsesesessseeeseceesenessnseaeeesees 47 7 3 DO PROBE CALIBRATION SETTING ZERO oooococccccccnonnnnanoncncnconononnnnonononconononnnnonanoncnnos 47 7 4 DO PROBE CALIBRATION SETTING SPAN cccccccccscssssssececeseceesessnssecesececeesensnseseeesees 48 T5 SCALE VOLUME CALIBRATION ua dida 49 8 CONNECTING THE VESSEL ASSEMBLY cccsssssscssssssscssssssccsssssscscssssccsseees 51 8 1 CONNECT THE VESSEL ASSEMBLY cccccsccsscsccecsesesseceeesececsesessaecesecccsesessnneseeesecseeeees 51 8 2 LOAD PUMP TUBING 2205 55 35 5035 os 51 8 3 INSTALL LIQUID ADDITION SYSTEMS coccccccncnonononononononononononononononononononononononanononananona 52 8 3 1 Addition Tubing Siz A e HYNNY 53 8 4 CONNECT GASES aain enea Rh Mk dd WYF YND ean Ae 53 Y GETTING STARTED andalocio bir vad cede FY UO Yny 55 New Brunswick Scientific User s Guide 9 1 CONTROEMODES ss facies dese AW NW WY IAI AG a 55 9 2 SETTING P SELVATUES Ai a Ltd YND IN O 55 9 3 LOOPS ETPOIN TS a A Nod GL di das dao tao E 55 9 3 1 EMECH HB ADOS a FA GY A AO O FRON 55 9 3 2 Medi IES EOS ASA GA A O O A dls 57 9 4 CASCADE SYSTEM sida 57 9 5 PUMP ASSIGNMENT tii A GY GAD Nid 57 9 6 PUMP CALIBRATION A FN WY NON 58 9 7 USING OPTIONAL SCALES TO PROGRAM PUMBS coconncoccccconononnnnnnnncconocnnnononanicicicnnanons 58 9 7 1 Setting a Feed Pump to Add Liquid iii 58 9 7 2 Setting a Pump to Harvest Vessel Contents oooooconcccccncc
46. P button the screen that opens is actually the first tab the CONTROLLER SETUP see Figure 20a on the following page screen CelliGen BLU M1363 0050 User s Guide 41 Figure 20a Controller Setup Screen te Setup Screen Controller Se Unit Type CethGen BLU 3 No of TMFCs 1 Installed Options F Gas Mix Overlay TMF MM summary canbrate New Brunswick Scientific Unit Name Cell Culture Mode cemcen ouu ily Exhaust Heater WJ Rocipe Manager System Settings Hardwaro Setup Security Settings CelliGen BLU _ _ 2 TMFC Range 0 100 CCM P Vessel Volume FP Gas Overlay Module C Cascade gt Trend O Save Changes Pumps A Alarm Controller Setup TAB Ni type in the desired name The Unit Name can be user selected Press this box then use the pop up touchpad to 3 The TMFC Thermal Mass Flow Controller Range and the number of TMFCs 0 means manual gas flow usually by rotameter are factory set Figure 20b Recipe Manager Screen 1 Pe Setup Screen New Brunswick Scientific Cell Culture Mode Selected Recipe Controller Setup REZIJA EE System Settings Hardware Setup Security Settings al HEK293 rcp Available Recipe TE D Load Default HE summary caw Ey Case b lt Trend 0 Pumps A narm BC Use the Recipe Manager screen to save and l
47. Parameters Cultivation of animal cells in an environment optimal for manufacture of desired end products require monitoring and control of a substantial number of physical and chemical parameters Physical parameters include temperature fluid flow gas flow and liquid flow rates and agitation rates Chemical parameters include the dissolved oxygen DO concentration and pH New Brunswick Scientific User s Guide 142 Control Setpoints Temperatur lt ee 37 C A eE EN 7 0 DO te hire tenet irinen 40 Apia corsarios 80 rpm pH Control pH control was set to Auto mode which automatically adds base solution or CO gas to the system based on culture demands Deauband sur 0 10 PID VANES escri dis Factory set default values Base Solution Sodium bicarbonate 7 5 solution Transfer tubing Narrow bore silicone tubing with Luer connection 118 inch ID amp 4 inch OD Vessel Il ai 1 8 inch inlet tubing in the vessel headplate Dissolved Oxygen Control DO control was set to Auto mode which automatically regulates gas mixing based on culture demand PID values factory set default values Gas Control The gas control was set to 4 Gas mode which automatically maintains DO and pH The gas flow rate was based on the vessel size Up until day 3 gases were introduced into the vessel headspace only through the overlay port at a rate of 0 30 L min using 4 Gas mixing to maintain pH
48. R MCCC mesr ans pl BU IFTMFC 1 Mt 15 NOT USED NV CelliGen BLU M1363 0050 User s Guide 121 Figure 50a Control Schematics 230V Al OWER ENT 50 HZ L HS lt A al ES x E a D 3 DY E i I D xl EL 4 y TEL LI une L FLT eo 2 STAGE 4 DOWER FILTE 10 F New Brunswick Scientific Li una L y L 4 a 2 lt o I a Ui MH ES BOOL 3 SR E ov E g a TI 7 Xx MOV r i I gt o TEL TBI ba ae E _ OR gt de y BL LL BL AA LL P OWE goe ver gy SUPPL TB1 2 LENE BLI BLI t a a ha _ D D D no 3 B2 4 T32 5 TB2 6 TB2 7 TB2 8 TB2 DM o T DC FAN 1 himi a AN a w REI i BLK B 0125 025 0125 User s Guide 122 Figure 50b Control Schematics 230V MONITOR A C CABLE REWORK AS NEEDED 01072 WHT TEI HSCR BLK _ uc DISPLA TA a a TB a a o o TED TOUCH HEATER HEATER BLANKET 010 ELI SSROL RED BL TBI 07 a HT 10 BL El TB CelliGen BLU M1363 0050 User s
49. Range 6 8 pH Control Non invasive optical pH probe Digital display in 0 1 increments 0 200 Control 3 or 4 gas P amp l control Sensor Non invasive polargraphic probe continued New Brunswick Scientific User s Guide 26 CelliGen BLU System Assignable peristaltic pump Fixed speed 14 RPM or variable duty cycle Available control modes Off On Prime Assignable peristaltic pumps Fixed speed 109 RPM or variable duty cycle Available control modes Off On Prime Utilities Gas_______ 6PSIG maximum Electrical 100 120VAC 50 60 Hertz Single phase 15 Amps Requirements 200 240VAC 50 60 Hertz Single phase 15 Amps Control Station 88 Ibs 40 kg with touchscreen 15 Ibs 6 8 kg Vessel empty without motor 25 in wide X 24 in deep X 34 in high Touchscreen 63 cm W X 61 cm D X 86 cm H 18 in wide X 24 in deep X 28 in high Touchscreen 46 cm W X 61 cm D X 71 cm H Port supplied for remote connection of interface computer Port supplied for connection of BioCommand supervisory host computer One 5A glass tube fast acting fuse Fuses Regulatory Compliance See Section 5 1 Ambient Operating Conditions 10 30 C up to 80 relative humidity non condensing See Table 8 Section 13 3 for pump flow rates according to tubing size Vessel weight does not include probes exhaust condenser or other options 5 1 Validation Documentation All pertinent validation documents are avail
50. Scientific User s Guide 110 18 2 1 Motor Assembly Replacement Z WARNING NO ONE BUT A PROFESSIONAL SERVICE PERSON should touch electric or electronic parts or assemblies in the control cabinet If the motor assembly should require replacement call for an authorized NBS service technician 18 3 Fuse Replacement There is one replaceable 5 Amp fast acting glass tube fuse CelliGen BLU M1363 0050 User s Guide 111 19 SERVICE If any problems occur with your CelliGen BLU system do not attempt to perform any service on it Unauthorized servicing may void the warranty Please contact your local NBS Service Department or your local NBS distributor In any correspondence with NBS please refer to the Model Number CelliGen BLU and the Manufacturing Part Number and Serial Number of the unit 19 1 Troubleshooting WARNING Always turn your CelliGen BLU off and disconnect the power cord before performing maintenance As with any equipment difficulties sometimes arise If you experience a problem with the operation of your CelliGen BLU consult the following list of symptoms You may be able to resolve the situation easily and quickly yourself If the problem is not listed below or if the suggested solutions do not work please call your NBS representative to request a service technician Other than the solutions proposed below do not attempt to fix the equipment yourself Problem Pos
51. Step 5 Press the appropriate Input Device designation 5 Step 6 Press the appropriate Output Device designation 6 Step 7 Input the desired Control Settings Setpoint low amp high limits 7 Step 8 After making all of your selections press the OK button to save them Figure 12 LoopName Touchpad LoopName CO2 off gas Ed EN EN E E A A ngng 00 E3 EZ EM ES O ES ES EA ES EI EI ES ES EI EN ES ES EI EM ES E2 ES EI ES l s ll IE E 4 3 1 Press Caps Lock to shift to CAPITAL letters Press it again to shift back to lower case 2 Press Cancel to return to the Gauge screen without saving work done with the touchpad 3 Press OK to return to the Gauge screen saving the work done with the touchpad 4 Press BackSp to backspace cancelling one character at a time 5 Press Clear to clear the LoopName edit box in this touchpad allowing you to begin again CelliGen BLU M1363 0050 User s Guide 35 6 2 7 Deleting Loops Only user added loops can be deleted If you wish to delete a loop 1 In the SUMMARY screen press the LoopName box for the loop you wish to delete 2 In the loop s GAUGE screen and if this is not a pump control loop press the UserSettings button see Figure 13 Figure 13 Deleting a Control Loop full Gauge Screen New Brunswick Scientific Cell Culture Mode i User Setti CO2 off gas 0 0 ai 1 Proportional Normalizing o I
52. a ha dw ddyd 106 16 4 SAMPLING THE VESSEL ccssssesceccccccccsssssscscccccescesssscscecccseeeuessssecesccsseuueaensncesess 106 16 5 SHUTDOWN estoi tds 107 17 CLEANING once dida 108 18 MAINTENANCE csssccscccssccesscteccsccses ccccccccscacectsccuscceasoccescevcscecccecacesdescucscsasedceessveuceasen 109 18 1 PROBE MAINTENANCE AND STORAGE cccscsceccccccccesssssesccccccceusessscecccccseeusauensscesess 109 18 2 PERIODIC INSPECTION ccccccssssesseccccccccseessscsccccceseusssescsccccceceuueussesceccssseuuuaensscecess 109 18 2 1 Motor Assembly Replacements dd wt ada iat 110 18 3 FUSE REPLACEMENT 00 DF Moats ete bites A dI NO NF 110 19 SERVICE sscicdcccscccscoscceccsssdcccssoaccdedsscscsancosscecescsdccosseaccdedoss ctorcedccdsescebsce sosesedsceseoscdesses 111 19 1 TROUBLESHOOTING c cccccccccssssesceccccccccuusssscecceccecessussecsccecesceuueussscescesseuauaenencesess 111 19 2 PRODUCT RETURN a deeded e DU 112 20 DRAWING Suerrin ser ae eei oeei aosa a aoatra aS Corka Sa 113 20 1 JEISSTOFDRAWINGS E EE CHW AE A DA AAE 129 20 2 EIST OF TABLES uc We Y AF GN ti 131 21 APPENDIX A STACKABLE PUMBS o ocococococonosononononinoninininonininonicinicininososisosonoss 132 22 APPENDIX B SOME GENERAL CONCEPTS Q 00 cccsssssssssssssssssssssesssesesess 134 22 1 WHAT IS A CONTROLLER cccccececcccccccsssssssececcccccsessssesceccsscessuusassseccesceseuuenensncesess 134 22 2 WHATIS A CONTROL LOOP ou cece cc ccsssse
53. able online at www nbsc com BLU With the vessel s lot number in hand see bottom of vessel double click on the PDF icon for CelliGen BLU Validation Guide in the lefthand column 5 2 Certifications The CelliGen BLU has been tested to ETL standards to comply with these safety standards UL 61010A 1 UL 61010A 2 010 CAN CSA C22 2 No 1010 1 CAN CSA C22 2 No 1010 2 010 As attested in the CE Declaration of Conformity reproduced on the following page they also conform to the appropriate CE standards CelliGen BLU M1363 0050 User s Guide 27 New Brunswick aneppendorf company C C DECLARATION OF CONFORMITY New Brunswick Scientific hereby declares that the product s listed below conform to the European Union d rective and standards identified in this declaration Product s CeliiGen BLU EU Directive s Low Voltage 2006 95 EC Electromagnetic Compatibility 2004 108 EC Standard s EN61010 1 EN61000 4 3 EN61010 2 010 EN61000 4 4 EN55011 EN61000 4 5 EN61000 3 2 EN61000 4 6 EN61000 3 3 EN61000 4 11 EN61000 4 2 The conformity assessment procedures were performed at the following notified body Intertek Testing Services 41 Plymouth Street Fairfield NJ 07004 Advanced Compliance Laboratory 6 Randolph Way Hillsborough NJ 08844 New Brunswick Scientific 44 Talmadge RD Edison NJ 08818 The technical documentation relevant to the above equipment will be held at New Brunswick Sc
54. ace it under the vessel 4 6 12 Install Motor Assembly 1 Position the motor assembly on top of the coupling using the locating notch to orient it properly 2 Connect the motor cable to the receptacle on the face of the control cabinet New Brunswick Scientific User s Guide 22 4 7 Main Power Switch The main power switch is located on the lower righthand side of the control cabinet see Figure la Front View E CAUTION Before turning on the main power switch make sure that the power cord is properly connected to the control cabinet and plugged into a suitable power outlet 4 8 Optional BioCommand Software If you are using NBS supervisory software be sure to consult your BioCommand user s manual for installation and start up instructions in addition to the general instructions provided below A 25 pin RS232 422 Modbus com port is provided on the rear panel of the control cabinet see Figure 6 to connect the CelliGen BLU to a supervisory host computer Communications to BioCommand software are via an optional RS 232 interface cable 1 Connect the 25 pin end of the RS 232 cable to the Modbus port and ensure that the connection is secure 2 Hand tighten the thumbscrews 3 Refer to the BioCommand user s guide for instructions on connecting the RS 232 interface cable to the supervisory host computer 4 9 Inputs Outputs for Ancillary Devices Each thermal mass flow controller TMFC uses one 0 5V analog inp
55. add a Password you will be prompted with Confirm PW to type it again to confirm it then press the OK button CelliGen BLU M1363 0050 User s Guide 103 Figure 47 Security Keypad PK Setup Screen New Brunswick Scientific Cell Culture Mode User Name User 2 ttttt E 0000000000 ES EZ EJ ES EI EI EU EU ES ES O EZ EZ EE EJ EZ EI EI EI EZ EB E2 ES EZ EZ E2 EZ EJ EN EN EA CO IC CA E EN summary f com GB case 2 wena row y WED 1 Password Confirm PW 1 When the User Name and Password are entered as desired press the OK button As shown in the Security Settings Screen on the previous page and in the Summary Screen below when security is enabled the User button appears in the top left corner of all major screens Figure 48 User Button y Setup Screen New Brunswick Scientific Cell Culture Mode 1 User Mig Exhaust Heater Controller Setup Recipe Manager System Settings Hardware Setup F Enable Security Feature Automatic Log Off Time 5 Minutos v z Administrators Supervisors Operators 2 m Summary Y cam FE case Trena Pumps A a ew 1 When a user presses the User button that user can use the popup buttons to Log Off or to Log On by pressing the appropriate button If the user has Administrator status the Change Password button will also be available Pressing it will open the security keypad to
56. alibration Screen New Brunswick Scientific Cell Culture Mode Allg Exhaust Heater Loops Do Calibrating Loop Volume Current Value Raw Value CA FRE El case Tren Pumps A nas 2 Setup Loops available for calibration Input boxes User s Guide 38 For details on probe calibration see Sections 7 1 pH probe and 7 3 DO probe For details on volume calibration see Section 7 5 6 2 10 Cascade Screen A cascade is a control function that uses the output of one loop to influence the action and output of one or more other loop s This screen see Figure 16 allows the user to set up cascades to view current cascade settings and to change those settings Figure 16 Cascade Screen 1 fab Cascade Screen lly Exhaust Heater Cascade From Stant Setnoirt DU Stan Ox End Setpowt DO End Ox 3 o 2 14 zZ o 3 o K Z o 3 BEREA z o 2 o El m summary TO cfe trena Q pumps A narma J setup 1 Loop being cascaded from 2 Loop s being cascaded to 3 User definable variables For details on setting cascades see Section 10 1 6 2 11 Trend Screen This screen see Figure 17 on the following page allows the user to set the parameters for plotting trend graphs and to view the graphs that track the activity of up to 8 selected loops during an
57. alue is mathematically determined by evaluating the error between measured present values and desired setpoints and integrating these values into a PID based control algorithm The CelliGen BLU s RPC controller allows cascading from any loop to as many as five other loops DO and pH are the most commonly cascaded from loops oxygen and nitrogen commonly receive the cascade from DO and CO and Base pump usually receive the cascade from pH altering their respective setpoints to correct errors in DO When more than one loop is configured as the recipient of a cascaded loop they may respond in parallel at the same time or in series one after the other depending on how the cascade has been set up Cascades set up to run in series generally give more predictable control responses Sometimes a small region of overlap where two loop setpoints vary simultaneously is used to smooth the transition from one loop to another To enable cascades gas loops must be in Manual Cascade mode which can be selected from the GAUGE screens and all other loops involved in the cascades must be set to Auto or in the case of pumps set to ON When systems are configured with multiple TMFCs and Manual Cascade mode is selected the controller automatically populates default cascade values for the gases being used but it is important to make sure all loops involved in the cascades are set to Manual Cascade mode Controllers configured with 0 or 1 TMFC are not auto
58. ample Alarms History Screen A Alarms History New Brunswick Scientific Cell Culture Mode Mil Exhaust Heater Alarm History LoopName Error Time Acknowledge Time Description 30 Jun 2009 06 24 00 30 Jun 2009 06 26 08 Power Restored 4 2 pH 30 Jun 2009 07 05 53 30 Jun 2009 07 05 56 Absolute High Alarm EE summary Calib be Casc bc Trend Oy Pumps Ke Setup 2 In the control loop s Alarms Screen that opens press the Alarm History button 3 Press the Scroll Down o or Scroll Up w button to read through the data 4 Press the Scroll Back lt lt lt button to return to the ALARMS screen To access the Alarms History screen to purge the history 1 Press the desired control loop s LoopName box in the main ALARMS screen 2 In the control loop s Alarms Screen that opens press the Alarm History button 3 Press the Purge button to erase all records O NOTE You cannot delete one record at a time you can only purge all records simultaneously 4 Press the Scroll Back lt lt lt button to return to the ALARMS screen CelliGen BLU M1363 0050 User s Guide 97 15 USING THE SETUP SCREEN The SETUP screen see Figure 20a repeated below for easy reference has one feature that you will use with frequency the Recipe Manager see Section 15 2 You can also use this screen to change Controller Setup see Section 15 1 to adjust System Settings select o
59. anannncnanonn nc conan Ynn 59 10 CASCADE CONTRO Disiasiccesiscesssecsusccccsstccccouscsccscccussceocsusecccssoccececasccsceveaccscscosenccseencs 60 10 1 CREATING A CASCADEL A A GANADA 61 10 2 CONTROLLING DO BY CASCADE occcocnnnnnonccnnnonononnnnanonononnnnonnnnnnonononnonononannanecinncnnnonnnnns 63 11 GAS MIXING FOR PH amp DO CONTROL occcccccccccccconononononcncnconononocononococonocononos 65 11 1 GAS CONTROL WITH O TMEC ona a A EA T TE A 65 11 2 GAS CONTROL WITH 1 TMEC eniai ee n a a aiaa ariaa i aiaa 69 11 3 GAS CONTROL WITH 3 TMECS 0oooococncnnnnonnnnnnnnnnccnnnnnononnnanacncnncnononnnnanccnnccnononnnnnninocos 73 11 4 GAS CONTROL WITH 4 TMECS 0oooococncncnncnonnnnninnccnnnonononannnocncnnonononnnnanoconocnnnnnnananinicos 75 11 5 GAS OVERLAY MIXING woiece cc cc cc ccesescececccccccesssssceccecccsseussssceccescessusunssseccssseseusenensssesess 79 12 PLOTTING TRENDS vessscscscesccscccssecscescccscecuccccsssccusscscececcesaccesuccsuccocsssscesisscedencessccenae s 81 12 1 CREATING A TREND GRAPH cccccccccccccccssssssscecccccccsunsssscecceccessuuensscscceccessuueaensncesess Sl 12 1 1 Using the Export BUON a 84 12 1 2 Using the Zoom BUON siat 84 12 1 3 OS 85 13 ABOUT PUMPS visissccecceiistecsidececsdeiestetadessesscsscecscaldecsedcasesestedacdse tess cssscabdecscseaveecscabasees 86 1301 BUMP SELPOINT2O Gu GW EN A Sd DYN See ees doe 86 13 2 PUMP CONTROL MODE ooooccccccnonononenocncnnnnonononenennconnnononnnnnnnoronnononnnnaninororcrnononnn
60. ap the blanket around the vessel securing it using the Velcro straps Be sure to position the hole in the blanket away from the pH sensor tube to minimize its exposure to light CAUTION Never turn on the heat blanket or exhaust tube heat blanket without first plugging in the RTD and inserting it into the thermowell inside the vessel see Section 4 6 10 Attach the exhaust tube heater by wrapping it around the exhaust tube see Figure 3 for reference and bracing it with the support rod The exhaust heater should always be on when gas is flowing into the vessel CelliGen BLU M1363 0050 User s Guide 19 4 6 4 Internal Components As illustrated in Figure 3 and in Figure 5 below the vessel comes completely assembled and sealed for sterility Figure 5 Internal Vessel Components 1 Retaining ring for support 5 pH tube 2 Pitched blade impeller upflow 6 Harvest tube 3 Sample tube 7 RTD thermowell 4 DO tube with silicone cap 8 Porous 7 12u sparge 4 6 5 Inspect the pH Probe Inspect the pH probe for possible shipping damage When the probe is plugged in and the control cabinet is powered on a purple light should pulse from the tip of the probe 4 6 6 Install the pH Probe 1 Make sure the probe cable is not connected to the control cabinet To protect the sensor from excessive use attach the cab
61. ble removable agitation motor located on top of the bearing housing on the headplate is connected to the agitation shaft with a magnetic coupling The magnetic coupling on the headplate has a notch to ensure correct and easy orientation as the motor is installed The motor can provide a speed range from 25 to 200 RPM The process control software ensures agitation speed accuracy throughout the speed range Default P amp I proportional amp integral values are preset at the factory We strongly recommend that you maintain the factory set parameters CelliGen BLU M1363 0050 User s Guide 3 4 3 5 3 6 Temperature Control The culture temperature setpoint is controlled within the range of 5 C above ambient temperature to 40 C by the process control software The media temperature is sensed by a Resistance Temperature Detector RTD submerged in the thermowell Temperature will be maintained or adjusted by ambient cooling and heating through the use of a silicone heat blanket Default P amp I proportional amp integral values are preset at the factory We strongly recommend that you maintain the factory set parameters Aeration Aeration is controlled by inputting values through the touchscreen on the control cabinet Gas can be introduced by means of a porous sparger or overlay port the overlay system is optional Up to four gases including air nitrogen carbon dioxide and oxygen can be introduced into the media o
62. ble 5 2 Step 2 If you have selected Manual mode use these edit boxes to set the percentage for O2 N2 and CO2 as desired the system will automatically set Air so that all gases total 100 3 Step 3 If desired use the Max CO2 edit box to set a maximum CO2 percentage This is for 4 Gas mode only New Brunswick Scientific User s Guide 80 Table 5 Gas Overlay Operating Modes Operating Mode Description Off Press this button to turn the Gas Overlay option off Manual Press this button to program the mix percentage of each gas yourself See Figure 34 Step 2 4 Gas Press this button to leave automatic gas mixing to the controller This selection disables user input to the Mix Percentage pane see Figure 34 CelliGen BLU M1363 0050 User s Guide 81 12 PLOTTING TRENDS Opening the TREND screen allows you to plot and display a graph of ongoing culture data viewing from 30 minutes to 144 hours of input Up to 8 loops can be plotted on the graph each in its own distinctive user selected color The graph and data are only available while the bioreactor is running Data cannot be stored in the controller but can be archived remotely on an auxiliary PC via the RS232 422 Modbus interface see Section 4 8 for details or saved to a USB storage device 12 1 Creating a Trend Graph 1 From any screen press the TREND button to open the TREND screen see Figure 17 repeated
63. by gravity or by using a peristaltic pump if appropriate to the working volume O Note OTE If microcarriers are being used they should be placed in the addition vessel with cells and part of the media After dispensing this into the vessel the rest of the media should be placed in the addition vessel and dispensed into the vessel This rinses microcarriers from the addition vessel and tubing 95 e Close the addition tubing clamps e Aseptically disconnect the addition port tubing from the addition vessel 5 Turn the power switch ON 6 Set Agitation to the desired RPM and the mode to AUTO Bel caution Agitation can only be increased 25 RPM at a time any greater increase will cause the magnetic drive to uncouple 0 NOTE For microcarrier culture the agitation should initially be set at 30 50 RPM After the cells have attached to the microcarriers the agitation can be increased above 50 RPM depending upon the microcarrier and cell being used For suspension culture the speed should be 30 50 RPM 7 Set the DO and pH to the desired control value New Brunswick Scientific User s Guide 106 8 Set the DO and pH mode to AUTO 9 Connect the Gas Inlet tubing to the filtered Gas Delivery System being used 10 Turn on the gases to 5 PSIG pressure 11 Display gas screen set mode to 3 GAS or 4 GAS set cycle time NOTE If at the very beginning of operation the pH value of the media is higher than the s
64. ccording to the loop and process mode There are also operating modes for all of the pumps see Section 13 2 for details To change operating modes for any of the displayed loops in the SUMMARY screen 1 Press either the LoopName or the Control Mode box in the row for the appropriate loop to open that loop s GAUGE screen see Figure 10 on the following page CelliGen BLU M1363 0050 User s Guide Figure 10 Sample GAUGE Screen Cell Culture Mode New Brun kScientinc Aliily Exhaust Heater Gains Factory Detaua Proportional 2000 Integral Deadband 3 7 00 ue i ac 5 mn u a 7 00 gu uo Limits Set Low Set High 200 1200 Decimal Placos 2 Tz caib EY Case be Trend Pumps A Marmo Ja Setup 1 Step 2 Press the button that corresponds to the desired Operating Mode 2 Step 3 To save the new operating mode and return to the SUMMARY screen press the SUMMARY button 3 Deadband is a user definable pH value within which above or below the setpoint no response will be triggered NOTE 6 2 9 New Brunswick Scientific 1 pH Sections 6 2 9 6 2 14 introduce you to the other main screens accessible by pressing the blue navigation buttons at the bottom of the screens Calibration Screen This screen is used to calibrate the pH and the DO probes and vessel volume Figure 15 Calibration Screen C
65. ckage and swab the Luer connector with IPA 3 Attach the NeedleFree syringe to the CLAVE connector 4 Open the tubing clamp is one is attached to the sample tubing CelliGen BLU M1363 0050 User s Guide 107 5 Slowly draw the sample by pulling back the syringe plunger until the desired volume has been removed You may need to use one syringe to purge the line of old media then a fresh syringe to draw the sample 6 Aseptically disconnect the syringe 7 Swab the CLAVE connector again with IPA and reinstall the cap When you draw additional samples we recommend that you flush the sample line by pulling a small sample first with one syringe to remove any old media and cells from the line the standard sample tube holds approximately 5 ml of liquid then draw a second sample with a fresh syringe to be your actual sample 16 5 Shutdown 1 Turn off all gases 2 Set the control mode of all loops to OFF and disconnect the agitation motor 3 Turn the power OFF 4 Disconnect the power plugs 5 Disconnect the probe cables 6 Remove the DO and temperature probes and store them properly see their respective manuals 7 Aseptically harvest the contents of the vessel 8 Clamp off all tubing connections 9 Properly dispose of the entire vessel assembly including the pH probe respecting all appropriate precautions regarding biohazards New Brunswick Scientific User s Guide 108 17 CLEANING At least once a m
66. cknowledging an Alarm When an alarm condition develops the LoopName box on the SUMMARY screen for the control loop involved will turn from blue to red as will the CelliGen BLU name box This is the Visual alarm A footnote written in red will also appear in order to identify the nature of the alarm e g Unit 1 Deviation Low Error The Visible alarm will remain onscreen until the alarm condition is rectified If the Audible alarm is also enabled beeping will occur until the alarm is acknowledged There are three ways to acknowledge alarms 1 one alarm at a time 2 all alarms for one control loop at a time and 3 all alarms for all control loops at a time for the rare occasion such a condition should arise CelliGen BLU M1363 0050 User s Guide 95 To acknowledge one alarm at a time Press the ALARMS screen button to open the ALARMS screen Press the red LoopName box to open that control loop s ALARMS screen Press the Current Alarms button to open the Current Alarms Summary screen Press the Index box for the alarm you wish to acknowledge It will turn green Press the Acknowledge button The alarm will be deleted from the screen Repeat Steps 4 8 5 for any other alarms recorded for this loop Press the Scroll Back lt lt lt button to return to the ALARMS screen Repeat Steps 2 7 for any other control loop alarms oe ee To acknowledge all alarms simultaneously for one control loop Press the ALARMS sc
67. cks and displays the accumulated flow for the associated loop here it measures accumulated air flow 4 Use the Reset Totalizer button at any time to return to 0 0 and begin counting again 0 NOTE To reduce the risk of clogging the microsparge gas flow should not be set below 2 CCM Figure 32b Sample Cascade Screen with 3 TMFCs rn Cascade Screen New Brunswick Scientific Cell Culture Mode CelliGen BLU Cascade From DO he 1 To Enabl Start int GDO Start Out End Setpoint DO End Out AirFio 1 gt Mee so 20 0 30 0 60 0 o2Flo 2 eR os 10 0 20 0 100 0 None I NO None a Ka None a Ka E Summary ti can EC ES Trend Gy Pumps A Aams A setup 1 Press each loop s Enable button to activate automatic cascade control CelliGen BLU M1363 0050 User s Guide 75 Cascade values are automatically controlled but first they need to be activated by selecting each loop s Enable button in the CASCADE screen see Figure 32b Default values are provided as starting points they may need to be modified for the optimization of individual processes When you set the system to the 3 Gas Auto control mode the controller automatically adjusts the flow ratio of Air O2 and CO based on the demands of pH and DO The Minimum and Maximum Combined Flow fields allow limits to be set to the total gas flow so you can prevent too much gas being inject
68. controllers TMFC and up to one overlay with or without a TMFC If you do not know how your system is configured this information can be found on the SETUP screen under the Controller Set Up tab The number of TMFCs and their flow rates are factory set and cannot be adjusted Your CelliGen BLU has the ability to adjust pH and DO by increasing or decreasing the individual gas concentrations within the vessel There is a wide variety of system configurations possible with respect to gas control This section will help explain these choices and will guide you through their operation Depending on how your system is configured you may find that the high limit for CO is below the high limit of its associated flow controller This limit is imposed to prevent oxygen starvation while adjusting pH with CO Your system is capable of operating in either 3 gas or 4 gas control mode 3 gas uses air and O to regulate DO and CO to regulate pH 3 gas mode does not include N2 4 gas mode uses air and Oz as well as N for the control of DO and CO to control pH 11 1 Gas Control with 0 TMFC If your system is configured with 0 TMFC gas mixing is controlled through a bank of 4 solenoids and a rotameter is used to manually control the flow The software will still control how the solenoids open and close and you will still be able to select a control mode from the gauge screen but the overall flow will be regulated manually by the rotameter see Figures 28
69. d Sec 10 EH Summary i caibrae Ta cascade lt rend Wd A alarm A Setup User s Guide 40 6 2 13 Alarms Screen This screen see Figure 19 allows the user to turn alarms on and off to read the alarm history and to acknowledge any alarm while it is active For details on using the ALARMS screen see Section 14 Figure 19 Alarms Screen A Alarm Summary New Brunswick Scientific Cell Culture Mode ABS Low ABS High ABS Enable DEVLow DEV High DEV Enable LoopName Agit 200 InActive 5 5 InActive 4 Temp 80 0 InActive 5 0 5 0 InActive Pump1 A 100 0 InActive 5 0 5 0 InActive A Pump2 100 0 InActive 5 0 5 0 InActive Pump3 A 100 0 InActive 5 0 5 0 InActive DO A 100 0 InActive 5 0 5 0 InActive pH 2 00 12 00 InActive 5 00 5 00 InActive Air 1 0 0 100 0 InActive 5 0 5 0 InActive Volume 0 00 5 00 InActive 10 00 10 00 InActive y 02 2 0 0 100 0 InActive 5 0 5 0 InActive y EB Summary ii cano y Casc bs Trena Q pumps PS A Setup 6 2 14 Setup Screen This master SETUP screen see Figures 20a 20b 20c amp 20d on the following pages is actually comprised of four screens accessed by tabs which are used to set up the controller recipe management system settings and hardware for the CelliGen BLU system This section will introduce you to those screens and their features For details on using the SETUP screen including a fifth tab that may be present see Section 15 When you press the SETU
70. diameter of your tubing Press and hold the pump mode Prime button or change the pump mode to ON at 100 setpoint and ensure that the pump operates smoothly A novel optional feature of these CelliGen BLU pumps is that you can add another identical pump onto the front of one of the pumps in the array see Section 0 Appendix B for details New Brunswick Scientific User s Guide 52 See Section 9 5 for details on pump assignment and Section 13 for details on pump set up and operation 8 3 Install Liquid Addition Systems Figure 24 is a simple depiction of a typical addition system Depending on the liquids base nutrients media to be added your system may be slightly different Figure 24 Typical Liquid Addition System 1 E Peristaltic pump 4 Plastic ties 2 Tubing 5 Addition bottle Breathing port with sterile filter 0 2um Addition port Be sure to read important CAUTION on the following page 1 Aseptically install if applicable a sterile 0 2um filter in one of the two penetrations on the addition bottle cap 2 Aseptically connect the tubing securing it with a plastic tie to the harvest tube in the addition bottle Clamp it off at the top 3 If you have not already done so thread the tubing through the selected feed pump 4 Connect the tubing securing it with a plastic tie to the appropriate addition port on the headplate 5 Remove the clamp CelliGen BLU M1363 00
71. e possibility to add more loops which are related to external auxiliary eguipment added via standard analog and optional serial RS232 inputs outputs located on the rear panel To add a new loop 1 Scroll down in the SUMMARY screen beyond the last pre assigned loop and press on a blank LoopName box 2 The Add User Defined Loop screen will open see Figure 11 on the following page User s Guide 34 Figure 11 Add User Defined Loop Screen 1 4 EE User Defined Loop Setup New Brunswick Scientific Cell Culture Mode Ad User Defined Loop Name Units Input Device Output Device gj Loop Type z Analog In 1 Analog Out 1 5 2 2 6 Aux Analog Device Analog In 2 Analog Out 2 c s Analog In 3 Analog Out 3 ER Si Analog In 4 Analog Out 4 C VS Pump 0 5V Analog In 5 Analog Out 5 j Analog In 6 Analog Out 6 VS Pump 4 20mA Analog In 7 Analog Out 7 3 RS232 Input Device Control Settings RS232 Output Device SP Low Link ha 6 RS232 Input Output SP High Limit hoo R Cao be Teena Pumps A Aarms Pe Setup 7 1 Step 3 Press here and use the LoopName Touchpad see Figure 12 to name the loop 2 Step 4 Press the appropriate option button The corresponding Unit of measurement will automatically appear in this sample screen 3 NOTE Options are not available grey if the system does not detect their presence 4
72. e Section 15 3 1 15 3 1 Resetting Date Time To reset the onscreen date and or time displayed in the lower righthand corner of every screen 1 In the System Settings screen press the edit box for the numeric parameter you wish to change 2 Use the pop up touchpad to input the new number and press the OK button 3 To change the month press the down arrow V and press the month you wish to select from its associated drop down menu 4 Press the Set button to save the new information You can do this after each change or after all changes have been made 15 3 2 Updating Software To update the system software obtain a new version of the software in a USB drive and plug the drive into the USB port on the control cabinet 1 In the System Settings screen press the Refresh button to update the current software status and to search for a new USB drive 2 The name of the new drive folder appears in the Update File box CelliGen BLU M1363 0050 User s Guide 101 3 Press the Update button to install the file The file will reboot twice this may take a little time 4 The Software pane will reflect the changes Updating software will not affect any previous user settings 15 4 Hardware Setup The CelliGen BLU system you purchased is preset in the factory as Unit1 with all the accompanying hardware In the Unit1 hardware list shown in the sample Hardware Setup screen Figure 20d below the system
73. e cause of the alarm condition and rectify 95 the situation to ensure proper operation of your CelliGen BLU Agitation can only be increased 25 RPM at a time any greater increase will 104 105 cause the magnetic drive to uncouple Never let a probe rest on its tip 109 CelliGen BLU M1363 0050 User s Guide 2 INSPECTION amp UNPACKING OF EQUIPMENT 2 1 Inspection of Box es When you have received your order from New Brunswick Scientific carefully inspect all parts of the shipment for damage that may have occurred during shipping Report any damage immediately to the carrier and to your local NBS Sales Order Department 2 2 Packing List Verification Verify against your NBS packing list that you have received the correct materials Report any missing parts to your local NBS Sales Order Department 2 3 Basic Components You should have at least the following components which will be described in greater detail later in this manual e Vessel complete assembly with tubing amp connectors e Control Cabinet e Probes RTD pH amp DO e Touchscreen e Motor e Heater Blanket e Exhaust Heater with support rod O NOTE The assembled Control Cabinet Touchscreen assembly is called a Control Station For purposes of clarity in this manual however the control cabinet which houses the controller and the touchscreen will be referred to separately by their component names New Brunswick Scientific User s Guide 3 1
74. e full loop gauge screen see Section 6 2 5 2 PUMPS Each gauge indicates setpoint SP and process variable PV Pump icon amp gauge title color indicates that pump s status Red OFF and Green ON 6 2 5 Gauge Screens Every loop has its own gauge screen To access it in the SUMMARY screen touch the screen inside the appropriate blue box in the LoopName column Your touch will open that loop s GAUGE screen see Figure 10 on the following page CelliGen BLU M1363 0050 User s Guide 33 6 2 6 New Brunswick Scientific Figure 10 Sample GAUGE Screen 1 2 3 Cell Culture Mode Exhaust Heater Factory Defaut lt 4 Gains Proportional 0 01 Integral eo Limits Set Low Set High 25 200 Decimal Places 7 6 00 00 Loop Name Process Variable present value Units the action of this loop Agitation is measured in RPM oi o noe P amp l values Limits Here you adjust the high amp low settings for this specific loop When adjusted the scaling for the gauge on the left of the screen will also be adjusted to reflect the high amp low limits selected Decimal Places Press the appropriate button to display values with O 1 2 or 3 decimal places Control Mode Setpoint and Units of measurement Adding Loops The CelliGen BLU comes to you with standard factory assigned loops and th
75. e intended for novices Be sure to refer to a textbook and consider utilizing the services of a technician e Allow sufficient time for the task Tuning is an iterative process It consists of configuring a loop with trial P I and D values evaluating loop response then readjusting the constants The process is repeated until the loop responds fully and without oscillation e One usually begins with a trial P setting I and D to zero After P is established a similar iterative process establishes I e Most bioreactor probes respond too slowly or are too noisy to utilize the D term to advantage In most cases D should remain at zero Agitation is sometimes an exception e The magnitude of the control signal depends on the P I and D constants It also depends inversely on a Normalizing Constant 1 For example Chinks F G Process Control Systems Application Design and Tuning McGraw Hill 1988 New York Auckland Bogota London Toronto Sydney Tokyo Montreal CelliGen BLU M1363 0050 User s Guide 137 22 6 What Do the Constants Mean The control signal Sy for a loop that has been N seconds in AUTO mode is expressed mathematically as Sn P en k amp 1 60 e k D n en 1 K Where P I andD are respectively the proportional integral and derivative constants e is the loop setpoint minus the process value or error k is a normalizing constant for the loop The controller reevaluates Sy ev
76. e outside container of the eguipment being returned to our facility In addition attach a duplicate copy of the completed form to the item being returned Returned Material Authorization RMA Number Eguipment being returned Model Number Serial Number Reason for its return This eguipment check all that apply New Product Biohazards O Never used Not used Used but decontaminated with Hazardous Chemicals Radioactive Materials Not used Not used Used but decontaminated with Used but decontaminated with I certify that the eguipment described above has been thoroughly cleaned and decontaminated of all chemical biological and radioactive contaminants and also certify that the returned unit is safe for unprotected human contact By Signature Print name Title Date Company Address Phone Fax Email Form 2847a CelliGen BLU M1363 0050 User s Guide 147 4 4 Gas Mix 75 A Add User 102 Adding Loops 33 Addition Tubing Size of 53 Aeration 5 Agitation Speed Precautions 104 105 Agitation System 4 Air Wash 80 Air 1 66 AirFlo 75 Airflow Control Automatic 5 Manual 5 Alarms ABS 92 Acknowledging 94 DEV 92 Alarms History 95 Alarms Screen 40 92 Analog Inputs amp Outputs 22 Bag Hanger 7 24 Installing the 24 BioCommand 8 22 43 106 Bioreactor Information Sheet v
77. e ready to use it The surface on which you place the CelliGen BLU bioreactor should be smooth level and sturdy Ensure that the surface can bear the weight of the bioreactor plus vessel contents and any applicable ancilliary equipment Also ensure that there is enough space around the back and the front of the CelliGen BLU for proper operation and access Allow at least 4 inches of clearance behind the unit for heat dissipation See Section 5 Specifications for weights and dimensions Environment The CelliGen BLU bioreactor operates properly under the following conditions e Ambient temperature range 10 C to 30 C e Relative humidity up to 80 non condensing Installing the Control Cabinet Position the CelliGen BLU control cabinet on a firm level surface in an area where utilities are readily available Level the horizontal surface of the base with four leveling glides if necessary Connect the power cord to the rear of the control cabinet At a later time once the unit is completely assembled and all connections have been made you will plug the power cord into a suitable electrical outlet New Brunswick Scientific User s Guide 10 4 4 Installing the Touchscreen With reference to Figures la for location amp 1b for installation align the monitor with the mounting rack on the cabinet and use the four screws provided with the monitor to securely fasten it to the rack The mounting rack swivels for easy acces
78. e si ifs Security Settings New Hardware A Module Status Unit1 Base Power Active Main ph DO Active MainAnalogl O Active Opto pH Active lt lt lt Scan Hardware SCADA 7 Mode NBS Modbus h Unit ID 6 EB Summary caib Ta Casc bc Trena 0 Pumps A nam T 1 Use the Hardware Setup screen to view hardware installed in the system and to set Unit IDs for software 2 Use this pane to choose software connections CelliGen BLU M1363 0050 User s Guide 43 6 3 RS232 422 Computer Interface An RS232 422 com port has been provided there is a 25 pin D connector located on the lower rear panel of the control cabinet see Figure 21 It is labeled Modbus Figure 21 Control Cabinet Rear Panel apo ETHERNET Analog Inputs amp Outputs are easily accessible Capped as standard not in use at this time Ethernet port for future expansion not in use at this time Modbus port for use with NBS BioCommand supervisory software 2 USB ports oO BR O h An NBS BioCommand advanced supervisory software program is available which will enable the operator to interface with a computer that has a Windows 2
79. ed in Table 7 Table 7 Pump Control Modes Control Mode Description Off The pump will receive no input and will not operate On The pump will operate according to the parameters you have set Prime This button toggles the pump on or off manually as long as you press the button the pump will run continuously When you release the button the pump will stop running 0 NOTE If pumps are linked to a cascade this may affect the ability to manually change setpoints and control modes To enter select a Control Mode for any pump press the appropriate button in the Control Mode pane of the PUMPS gauge screen see Figure 39 New Brunswick Scientific User s Guide 88 13 3 Pump Flow Rate amp Calibration Methods The pump will always run at the same speed but their flow rate depends on the diameter of the tubing you use Table 8 provides the pump flow rates according to various tubing diameters Table 8 Flow Rate per Tubing Size Tubing Wall Thickness 1 16 inch 1 6mm Tubing ID inch 1 32 1 16 mm 0 8 1 6 14 RPM Flow ml minute 0 84 109 RPM Flow ml minute 27 25 92 65 327 00 To calibrate any pump with the tubing you have selected 1 Load approximately three feet of the tubing into the pump head Set up a reservoir with water at the input end of the tubing and an empty graduated cylinder capable of measuring small quantities at the output end of the tubing 3 Read this step compl
80. ed should be medical grade No gas pressure should rise above 6 PSIG see also page iii Never leave a gas inlet open if no tubing will be connected keep the inlet plugged All gases should be regulated using a two stage regulator The scale of the regulator gauge for gases going into the bioreactor should be such that one can regulate pressure between 0 10 PSIG maximum 4 6 Vessel Assembly CAUTION Do not open any port in the vessel headplate until ready for use Prolonged exposure to light will adversely affect the pH sensor Never place the vessel in direct sunlight or in a brightly lit location Always keep the vessel wrapped with the heat blanket to avoid premature decay of the pH sensor 4 6 1 Unpacking the Vessel Because the CelliGen BLU vessel is shipped sterile it is important to carefully follow the directions below 1 Inspect the outer box for damage 2 Check the label on the box to make sure that the product has not exceeded its expiration period 3 Open the lid of the box then remove the cardboard insert by pulling directly up on the cut out handles 4 Remove the vessel in its protective opaque outer wrapper Do not open the wrapper yet Inspect the wrapper for damage CelliGen BLU M1363 0050 User s Guide 15 9 NOTE The outer wrapper is not only a secondary sterile boundary it is also opague to protect the pH sensor from light Do not proceed to Step 5 until you are ready to use the
81. ed while also maintaining a total positive flow Figure 32c AirFlo 1 Gauge with 3 TMFCs in 3 Gas Auto Mode ful Gauge Screen New Brunswick Scientific Cell Culture Mode ceticen atu BLU All Exhaust Heater AirFlo 1 0 0 CCM Gas Mix Selection 0 0 C02 Set Py Dutz Maniaienacade Mix Elo Cycle Times Coc 00 00 00 On Time 120 OffTime D 100 0 100 0 100 0 Min Combined Flow Set Low Set High 0 0 2 0 100 0 Max Combined Flow 2 100 0 Decimal Places 0000 00 00 Totalizer CC 0 0 Reset Totalizer EE E el O me 3 Gas Auto mode has been selected the button is green NOTE that these fields are not for GasFlo Cascade they are for Minimum and Maximum Combined Flow settings as explained in the text above this figure K Trend A Alarms Pe setup Ni 11 4 Gas Control with 4 TMFCs If your system is equipped with four TMFCs you have access to Manual Cascade 4 Gas Auto and 3 Gas Auto gas mix control in the gas gauge screens see Figures 33a 33c amp 33d With four TMFCs when you set any gas gauge to Manual Cascade see Figure 33a on the next page Air O2 CO and N setpoints can either be manually entered in their individual gas gauge screens or they can be controlled by cascades see Figure 33b New Brunswick Scientific User s Guide 76 Figure 33 Controller Setup Screen
82. een 28 8 Sample SUMMARY Screen 29 9 Sample SYNOPTIC Screen 32 10 Sample GAUGE Screen 33 37 56 11 Add User Defined Loop Screen 34 87 12 LoopName Touchpad 34 13 Deleting a Control Loop 35 14 Deleting a Pump Control Loop 36 15 Calibration Screen 37 45 15a Probe Data Screen 46 16 Cascade Screen 38 62 16a Cascade Screen detail 60 17 Trend Screen 39 77 18 Pumps Screen 39 57 19 Alarms Screen 40 88 20a Controller Setup Screen 41 93 94 20b Recipe Manager Screen 41 95 20c System Settings Screen 42 96 20d Hardware Setup Screen 42 97 21 Control Cabinet Rear Panel 43 22 Calibrating DO 48 22a Setting Zero for Scale Volume Calibration 49 22b Setting Span for Scale Volume Calibration 50 23 Standard Pump Array pumps open 51 82 24 Typical Liquid Addition System 52 25 Setpoint Touchpad 56 26 Pump Assignment Pane 58 59 27 Sample DO Cascade 63 28 Controller Setup Screen 0 TMFC 65 29a Air 1 Gauge with O TMFC 8 3 Gas Control 66 29b Air 1 Gauge with O TMFC and 4 Gas Control 66 29c Air 1 Gauge with O TMFC 8 Manual 3 Gas Control 68 continued New Brunswick Scientific User s Guide 130 Figure Description Page 29d Air 1 Gauge with 0 TMFC and Manual 4 Gas Control 68 30 Controller Setup Screen 1 TMFC 69 31a Air 1 Gauge Screen with 1 TMFC amp 3 Gas Control 70 31b Air 1 Gauge Screen wit
83. entire process run The data can be exported through the USB port in Excel format to a PC For details on using the TREND screen see Section 12 CelliGen BLU M1363 0050 User s Guide 39 Figure 17 Trend Screen do Trend Screen SP ou New Brunswick Scientific sn El UN PY Cell Culture Mode Ally Exhaust Heater 08 40 sm e o 10 40 1240 14 40 i gt ole ce 1 The user will assign a tracking color to each loop 6 2 12 Pumps Screen This screen see Figure 18 allows the user to access the pump gauges screens where the three standard pumps plus any optional pumps are displayed providing both current readings and the opportunity to change pump settings For details on using the PUMPS screen see Section 9 5 Figure 18 Pumps Screen O Pump Screen New Brunswick Scientific Cell Culture Mode Mily Exhaust Heater Pump1 14 RPM Setpoint 00 9b PV 00 Control Mode off Pump2 109 RPM Setpoint 0 0 P 00 Control Mode Off Pump3 109 RPM Setpoint 00 Ez PV 00 Control Mode off None Flow Rate mL Second Calibrate Flow Rate mL Second Calibrate Flow Rate mL Second Calibrate New Brunswick Scientific Calibrated Calibrated Calibrated 0 0000 0 0000 0 0000 ictal maad woal Me L ee J Period Sec 10 Period Sec 10 Perio
84. erates the desired control signal The operator has a choice of control modes the most common of which are ON OFF AUTO and MANUAL In cascaded control one sensor influences an actuator that is normally associated with a different sensor The onscreen control mode choice will be the name of the loop chosen to have influence on the actuator Setting P 8 I Values P amp I values are numbers that determine how the bioreactor responds to changing growth conditions and new setpoints These are listed in each loop s GAUGE screen You may need to modify P amp I values to suit your particular process To do so press inside the Proportional amp Integral edit boxes each time entering the desired value using the popup keypad 0 NOTE 9 3 9 3 1 If you change P amp l values you can return to the original settings at any time by pressing the Factory Default button see Figure 10 Loop Setpoints The setpoint is the value you want each loop to attain When the loop control mode is AUTO the bioreactor will automatically make appropriate adjustments to maintain the value at the setpoint Entering Setpoints To enter a setpoint for any loop follow these steps 1 Touch either the LoopName box or the Setpoint box for the desired loop on the SUMMARY screen In this example we have selected AGIT 2 The loop GAUGE screen opens see Figure 10 repeated on the following page for easy reference New Brunswick Scientific User s Gu
85. ery second I is divided by 60 so any value entered by the user should be in reciprocal minutes The normalizing constant k can be set to any non zero value but is usually set to the full scale reading of the loop For example if the range of expected temperatures is 0 to 125 setting k to 125 results in a P term value of P when the error is at a maximum i e P ey k P 125 125 P Similarly with a full scale error the I term after 1 minute and the D term will be I and D respectively New Brunswick Scientific User s Guide 138 23 APPENDIX C CORROSION RESISTANCE Websites such as www outokumpu com provide up to date information about the 316 type stainless steel used in your CelliGen BLU vessels CelliGen BLU M1363 0050 User s Guide 139 24 1 24 2 24 APPENDIX D PROTOCOL FOR GROWING CHO CELLS Abstract The study presents a typical protocol for the setup and operation of New Brunswick Scientific s new CelliGen BLU single use stirred tank bioreactor a versatile new benchtop system for the culture of a wide range of mammalian cells This bioreactor has been designed to provide research and production facilities with a single use vessel which combines the benefits of both traditional stirred tank design and single use technology capable of seamless process scale up The system can be operated in batch fed batch or continuous modes A procedure for culturing Chinese Hamster Ovarian CHO cells
86. etely before you do it with the input end of the tubing in the water reservoir prime the tubing line by pressing the pump s Prime button but allow it to run only until liquid starts to flow into the tubing DO NOT allow the liquid to run into the graduated cylinder yet 4 Ifyou are not using a scale skip to Step 5 If you are using a scale place the graduated cylinder with the tubing on the scale and press Zero on the scale 5 In the Flow Rate pane of the PUMPS screen see Figure 39 for that pump press the Calibrate button to open the Calibration pane see Figure 40 Figure 40 Calibrating the Pump Flow Rate Calib Run Time Seconds 60 120 300 2 Start Cancel I Amount Pumped mL 1 Step 6 Press your choice of Run Time 60 120 or 300 seconds that button will turn green 2 Step 7 Press the Start button The button will turn green and the pump will start running 3 Step 8 When the Run Time has elapsed record the amount mL of liquid accumulated in the cylinder then enter that number or the number registered on the scale in the Amount Pumped edit box 4 Step 9 Press the Set button to save this data to the PUMPS screen CelliGen BLU M1363 0050 User s Guide 89 0 NOTE Calibration must be performed at operating setpoint 0 NOTE Each pump and each tubing size will need its own calibration The pump is now calibrated As the pump runs you
87. etpoint by 0 3 or more it is recommended to set the control mode to Manual and add sufficient percentage of CO gas into the vessel to bring down the pH value Then change mode to 4 GAS 12 Adjust the airflow to the desired flow rate It is best to start at the lowest airflow rate at which the unit can control pH and DO and increase the flow rate only when the unit can no longer control pH and DO at that flow rate We recommend that you maintain pH and DO using an overlay then using overlay and sparge 13 Check to see that the flow is stable Check to see that all the gases are connected properly 16 3 Start BioCommand if present If you are using BioCommand supervisory software 1 Start BioCommand on your computer reset the EFT Elapsed Fermentation Time to zero make appropriate program selections to begin logging data 2 Make sure all gas pressures are 5 PSIG 3 If your CelliGen BLU has rotameter air flow control adjust the airflow to the desired rate Check to see that flow is stable and that all gases are properly connected 16 4 Sampling the Vessel A sample port with C Flex tubing a CLAVE NeedleFree connector and Luer Lok NeedleFree syringes are supplied as the sampling method for every vessel Follow this procedure to draw a sample from your vessel 1 Aseptically remove the cap from the CLAVE connector and swab the exposed area with sterile IPA 2 Open a sterile NeedleFree syringe pa
88. from the culture and measure its pH with an external pH meter If the readings are different the pH sensor s zero has drifted Readjust the zero on the Vessel pH function to match the reading of the external pH meter 10 When the vessel reaches the desired working temperature calibrate the DO probe see Section 7 3 amp Section 7 4 O NOTE When the media temperature in the vessel is much lower than the desired working temperature be careful not to set the temperature to the desired value at once as that could overheat the media continued CelliGen BLU M1363 0050 User s Guide 105 Generally set the setpoint about 1 C below the desired working temperature first As the temperature rises reset the setpoint several times Finally the desired temperature can be set and the vessel temperature will reach its desired value without substantial overshoot 16 2 Inoculation 1 All addition and harvesting vessels should be sterile The addition vessel should also be siliconized for microcarrier culture 2 Set Agitation and Temperature to OFF Turn the power OFF 4 Inoculate the vessel in the following way e Aseptically connect the addition port tubing to the addition vessel e Aseptically place the sterile media and cells and the sterile microcarriers if you are using them see important NOTE below in the addition vessel e Open the addition tubing clamps e Dispense the contents of the inoculation vessel into the vessel
89. g will stabilize Press the Set Zero edit box see Figure 22 use the touchpad to enter 0 press the OK button then press the Set Zero button Press N2 3 OFF or if in Step 3 you manually turned the N2 loop on now manually shut off the nitrogen flow to the vessel Figure 22 Calibrating DO 5 Calibration Screen New Brunswick Scientific Cell Culture Mode Volume Current Value Raw Value 1 6 921 Offset Zero Std Slope eso fo 0 04 TN 0 0 1 EH summary TN 2 Casc ES Trena Pumps A Alarms Lo setup The N2 3 On and N2 3 Off buttons will only appear on systems with 1 or 2 TMFCs 7 4 DO Probe Calibration Setting Span In the AGIT GAUGE screen set the AGIT speed to 50 RPM Set the AGIT mode to AUTO Vigorously sparge air into the vessel via the filter on the headplate until the display is stable for approximately 10 minutes this may take up to 30 minutes total In the CALIBRATION screen select DO Enter 100 in the Set Span edit box see Figure 22 then press the Set Span button CelliGen BLU M1363 0050 User s Guide 49 7 5 Scale Volume Calibration If you do not have the optional scale skip this section The scale is calibrated using a two point calibration method First make sure the scale is properly connected to the control station see Figure 2 optional Scale Connector 1 Press the Calib button to open the CALIBRATION screen then press Vol
90. h The controller is capable of storing up to 10 recipes You can retrieve these recipes by opening the Recipe Manager screen where all saved recipes are listed in the Available Recipes pane Select the desired recipe by following Step 1 in Figure 20b then load it as shown in Step 3 15 3 System Settings Press the third tab in the SETUP screen to open the System Settings screen see Figure 20c on the following page Use this feature select the onscreen language you prefer to reset the date and or time to update the software and to calibrate the CelliGen BLU touchscreen New Brunswick Scientific User s Guide 100 Figure 20c System Settings Screen 1 Le Setup Screen New Brunswick Scientific Cell Culture Mode Controller Setup Recipe Manager EE cui geg Hardware Setup Security Settings TouchScreen English 2 Date Time 30 JUN 2008 06 43 Software Revision CGD 13 Update Folder Refresh Backup Maintenance CGDCTRL Rey 11 Phoenix AwardBlOS v6 00PG y Pumps A narm Bd 1 Other languages are not available at this time To recalibrate the system s touchscreen press the Calib Button then touch the onscreen target each time it appears You will be guided through the process 3 Listed here are the current User Interface and Control Program versions To update the software see Section 15 3 2 4 To change the Date and or Time se
91. h 1 TMFC amp 4 Gas Control 71 31c Air 1 Gauge Screen with 1 TMFC amp Manual 3 Gas Control 71 31d Air 1 Gauge Screen with 1 TMFC amp Manual 4 Gas Control 72 31e Sample Cascade Screen with 3 or 4 Gas Mix 72 32 Controller Setup Screen 3 TMFCs 73 32a AirFlo 1 Gauge with 3 TMFCs in Manual Cascade Mode 74 32b Sample Cascade Screen with 3 TMFCs 74 32c AirFlo 1 Gauge with 3 TMFCs in 3 Gas Auto Mode 75 33 Controller Setup Screen 4 TMFCs 76 33a AirFlo 1 Gauge with 4 TMFCs in Manual Cascade Mode 76 33b Sample Cascade Screen with 4 TMFCs 77 33c AirFlo 1 Gauge with 4 TMFCs in 4 Gas Auto Mode 78 33d AirFlo 1 Gauge with 4 TMFCs in 3 Gas Auto Mode 78 34 OvlMix Gauge Screen 79 35 Trend Setup Screen 82 36 Trend Graph 83 37 Selecting Zoom Coordinates 84 38 Selecting a Read Line Location 85 39 Setting Pump Setpoint 87 40 Calibrating the Pump Flow Rate 88 41 Pump Period Sec 89 42 Variable Speed Pump 90 43 Rear Panel of CelliGen BLU Cabinet 90 44 Sample Loop Alarms Screen pH 94 45 Sample Alarms History Screen 96 46 Security Settings Screen 102 47 Security Keypad 103 48 User Button 103 49a h Control Schematics 120V 113 120 50a h Control Schematics 230V 121 128 51 Mounting Base Pump for Stacking 132 52 Stacking the Pumps 133 53 Stacked Pumps 133 A Cell growth over the 7 day run CHO cells 143 B Cell viability over the 7 day run CHO cells 144 C Glucose consumption lactate production 10 day run CHO cells 144 CelliGen BLU M
92. has the Base Power module the Main pH DO module the Main Analog module and the Opto pH module This system is also set to Modbus communication mode see the SCADA pane in Figure 20d and has the Unit ID number of 6 This is the unit s multidrop identification number Figure 20d Hardware Setup Screen I gt Setup Screen New Brunswick Scientific Cell Culture Mode Mil Exhaust Heater Controller Setup Recipe Manager System Settings ENERO Security Settings New Hardware Module Status pase cd Active ain ph D Active MainAnaioa O Active Active Unit1 SCADA Mode 2 NBS Modbus y Unit ID 6 EH Summary i Calib ml case lt Trena Pumps A narm The Scan Hardware function is reserved for use only by authorized Service Technicians Ni SCADA address 15 5 Security Settings The security feature on the BLU provides two user access levels Operators have access to routine operations but they change security settings Administrators have access to all operations including defining new users operators and administrators and setting security parameters New Brunswick Scientific User s Guide 102 Press the fifth tab in the SETUP screen to open the Security Settings screen see Figure 46 Figure 46 Security Settings Screen 1 2 le Setup Screen New Brunswick Scientific Cell Culture Mode Controller Setup Recipe Manager System Settings Hardware Setup
93. he PV and SP differences cause the percentage or flow rate of CO or a pump to react in response New Brunswick Scientific User s Guide 64 0 NOTE Regardless of DO cascaded from output the setpoint of any cascade to loop will not go below its own Minimum Setpoint value Minimum Output corresponds to the minimum value that will produce the minimum setpoint lower outputs will not affect setpoint Regardless of DO output the setpoint of any cascade to loop will not rise above its own maximum setpoint CelliGen BLU M1363 0050 User s Guide 65 11 GAS MIXING FOR PH amp DO CONTROL Due to the sensitivity of the cells pH and DO in cell culture processes are typically managed through gas mixing The addition of CO into the media through the sparge line is usually used to maintain pH The introduction of this gas causes the pH of the culture to decrease due to a slow accumulation of bicarbonate in the solution On the other hand decreasing the CO sparge rate allows the pH to drift up either due to the natural alkaline drift of the media or because of the accumulation of alkaline waste products in the solution DO is maintained in solution by introducing Air Oz and sometimes N2 in various combinations or at different flowrates The controller calculates which and how much of either of these gases is reguired to maintain the user selected DO setpoint Your system can come equipped with 0 1 3 or 4 thermal mass flow
94. he TREND screen It will turn red to indicate that the zoom mode is active 2 Press in succession two diagonal locations that would frame left to right the section of interest see Figure 37 for reference Figure 37 Selecting Zoom Coordinates 1 Press first here in the upper left corner of desired quadrant Press second here lower right corner of desired quadrant NOTE the rectangle does not appear onscreen it is indicated here for reference purposes only 3 The Trend view will display the data between the two points selected and will adjust the time axis to match the elapsed time represented by this close up 4 Press the Zoom button again to return to the regular trend graph Minimum axis time in zoom mode is 120 seconds If you wish to use the zoom mode and the read line see Section 12 1 3 at the same time you must enter Zoom mode first CelliGen BLU M1363 0050 User s Guide 85 12 1 3 Using the Read Line The Read Line mode allows you to read PV values from the graph displayed at the top of the screen at a position you select To use the Read Line 1 Press the Read Line button at the bottom of the TREND screen It will turn red to indicate that the read line mode is active and black vertical line will appear at the current time position on the graph 2 To move the line to a time of your choosing press the graph at the desired point You can also press the Read Line lt
95. he control loop s Process Variable falls below or rises above the control band that you specify around the loop s setpoint e g a tolerance of 10 rpm above or 5 rpm below the Agitation setpoint 14 2 Setting Alarms To set alarms 1 Press the ALARMS button to open the ALARMS screen see Figure 19 repeated here for easy reference and Table 9 which explains the features of this screen Figure 19 Alarms Screen la Alarm Summary New Brunswick Scientific Cell Culture Mode Unit 1 Afp Exhaust Heater LoopName ABS Low AHS High ABS Enable DEVLow DEV High DEV Enable Agit 25 200 InActive 5 5 Inactive 4 emp 10 0 80 0 InActive 50 5 0 Inactive Pump oo 100 0 InActive 50 50 Inactive A oo 100 0 InActive 50 50 InActive oo 100 0 InActive 50 50 inActive oo 1000 InActive 50 50 inActive 1 H 2 00 12 00 InActive 5 00 5 00 InActive olume 0 00 5 00 InActive 10 00 10 00 InActive v ir 0 0 100 0 InActive 50 5 0 InActive y 100 0 InActive 50 50 InActive ffi summary cano Case ke Trena Pumps Ja Setup 1 Step 2 Press the first loop for which you want to enable an alarm That loop s individual Alarms Screen will open For this example see Figure 44 we use the pH loop The instructions continue after Table 9 CelliGen BLU M1363 0050 User s Guide 93 Table 9 Alarms Screen Features Feature Name Description LoopName Like the CelliGen BLU name box this box is blue under
96. he setpoint the system will be generating a positive Out because the controller senses a need to increase DO LoopName Setpoint Out Control Mode Units Casc er en oe mm wo oe When the PV equals the setpoint the Out should be approximately 0 as the controller senses no need to make adjustments to DO LoopName Setpoint Out Control Mode Units Casc To create a cascade 1 Press the CASCADE button to open the CASCADE screen see Figure 16 repeated below for easy reference Figure 16 Cascade Screen 1 2 Cascade Screen New Brunswick Scientific Cell Culture Mode Cascade From DO Enable Start Setpoint DO Start Out End Setpoint 2 DO End Out 0 0 0 0 100 0 100 0 lt 3 EE oe E summary 3 Cano ba Trend Pumps A Alarms Po Setup Step 2 Use this W dropdown menu to select the Cascade From loop 2 Step 3 Use the first Y dropdown menu to select the first Cascade To loop 3 Step 4 Set the Start Setpoint DO Start Output End Setpoint and DO End Output values one by one by pressing the edit box entering the desired value on the touchpad and pressing the OK button 4 Step 5 Press the YES button to enable this cascade CelliGen BLU M1363 0050 User s Guide 63 10 2 Controlling DO by Cascade Example Cascading DO to Air and Oxygen In this example see Figure 27 DO is cascaded to O2 2 o
97. he supplied tubing which is correct for most applications If the system oscillates reduce the tubing ID where it passes through the pump Use commonly available step up step down adapters and narrower bore tubing to make the tubing modifications if reguired Consult Table 8 the flow rate tubing size chart for further information 8 4 Connect Gases Ensure that all gas lines air oxygen etc are routed to the appropriate ports and secured at both ends with plastic ties New Brunswick Scientific User s Guide 54 LA WARNING NEVER OVER PRESSURIZE A CULTURE VESSEL Always use eye protection and exercise caution in the vicinity of culture vessels If the vessel exhaust becomes blocked pressure can build up possibly shattering the vessel and endangering personnel Before opening the airflow valve s visually confirm that the vessel exhaust is not blocked by kinked tubing clamps or a wet filter After opening the airflow valve s verify by feel that air is flowing freely from the exhaust If not immediately close the valve s or turn off the air gas supplies Never intentionally block the exhaust to raise vessel pressure Use the minimum air gas pressure that will provide adequate airflow for the application Never exceed the maximum pressure specified in this manual CelliGen BLU M1363 0050 User s Guide 55 9 1 9 2 9 GETTING STARTED Control Modes A control mode is the logic by which a controller gen
98. he vessel CelliGen BLU M1363 0050 User s Guide 51 8 CONNECTING THE VESSEL ASSEMBLY 8 1 Connect the Vessel Assembly Me Position the vessel next to the CelliGen BLU control cabinet If you have not already done so carefully position the motor on the bearing housing on top of the vessel assembly using the notches to orient it properly 3 Connect the pH cable to the pH connector on the control cabinet 4 Connect the DO cable to the DO connector on the control cabinet 8 2 Load Pump Tubing The three standard pumps are located on the front of the control cabinet see Figure 23 Figure 23 Standard Pump Array pumps open 1 2 Before you insert tubing into the pump channel verify that the pump is in the OFF control mode Follow these steps to properly load tubing into the pump Pull the upper pump cover upward to gain access to the interior of the pump Select the desired tubing size see Table 8 in Section 13 3 for reference and cut a length sufficient to reach from the inlet source through the pump and to the outlet recipient allowing a few extra inches Noting that the pumps run in a counter clockwise direction place the tubing snugly across the pump channel Close the upper pump cover by pulling it back down until it clicks into place Use the pump wheel on the right side facing the pump to set the channel to the
99. ide 56 Figure 10 Sample GAUGE Screen ful Gauge Screen New Brunswick Scientific Cell Culture Mode Gains Factory Default Proportional Set Low Set High 25 200 Decimal Places 00 00 25 0 0 25 EE fg cano Casc pe Trend Pumps A Ararms 2 Setup 1 Step 3 Press inside the Setpoint box to open the touchpad see Figure 25 to continue 2 P amp I Values Adjusting these values will determine how your system responds to changes in your culture See Section 22 5 for more information Figure 25 Setpoint Touchpad T Gauge Screen New Brunswick Scientific Cell Culture Mode Agit Setpoint RPM 250 A 2 EE IC En EN EE EE EN EH EE EN EN p Calib E Casc kK Tren 3 Pumps A Alarms Le setup 1 Step 4 Use the touchpad number keys to enter the desired Setpoint Use the white Clear button at any time before Step 5 to empty the Setpoint edit box 2 Note that Agitation cannot be set higher than 200 RPM the system will default to 200 if you try to input a higher number 3 Step 5 Press the OK button to save the setpoint and to return to the GAUGE screen or press the Cancel button to return without saving the setpoint CelliGen BLU M1363 0050 User s Guide 57 9 3 2 9 4 9 5 Modifying Setpoints This process is the same as entering setpo
100. ientific Company PO Box 4005 44 Talmadge Road Edison New Jersey 08818 4005 U S A Tel 732 287 1200 Fax 732 287 4222 v O e Vio a Director of QA RA New Brunswick Scientific User s Guide 28 6 OPERATING CONTROLS 6 1 Touchscreen Y our primary interface with the CelliGen BLU is the touchscreen on the control cabinet Figure 7 Touchscreen IN 1 Touchscreen display 4 Pumps 2 Control cabinet 5 ON OFF power switch 3 Service Connections see Section 4 5 Figure 2 6 2 Display Screens 6 2 1 Touchscreen Calibration The first time you power up you will be prompted to calibrate the screen to your touch Follow the onscreen instructions to touch the target each time it appears Usually you will be prompted to touch the four corners of the screen twice in succession See important NOTE on the following page CelliGen BLU M1363 0050 User s Guide 29 o NOTE For optimal results be sure to stand or sit in the position from which you are most likely to work Height and angle of reach will affect calibration 6 2 2 Start Up Screen The Start Up screen which tells you which operating software version is installed in your CelliGen BLU is first screen you see each time you turn on the power if you have already calibrated the touchscreen see Section 6 2 1 This screen re
101. ii C Cabinet Cleaning of 108 Calibration of Touchscreen 28 100 Calibration Screen 37 Cascade Creating a 61 Cascade New Brunswick Scientific 20 INDEX Creating a 62 Cascade Screen 38 62 CAUTION Symbol for v Cautions Recap of 1 Certifications 26 27 CHO Cells Protocol for Growing 139 Cleaning after a Run 107 CO2 4 66 CO2Flo 75 Configuration Files 98 Contact NBS How to 111 Control Cabinet Installing the 9 Control Cabinet Connections 10 Control Loop Definition of 134 Controller Definition of 134 Copyright Notice v Corrosion Resistance 138 CRUSH WARNING Symbol for v D Decontamination Certificate 145 Decontamination Certificate 146 Deleting Loops 35 Description of Vessel 4 Disclaimer Notice v DO Control of 6 DO Cascade 63 DO Cascading 57 DO Probe 6 Inspection of 20 Installation of 21 Drawing Index 129 User s Guide 148 E Electrical Connections 13 15 20 21 Electrical Requirements 13 15 20 21 Exhaust Condenser 6 Exhaust System 6 Extended Warranty Option vi F Feed Pumps To Add Liquid 58 Fuse Replacement 110 G Gas Connections 13 Gas Control 0 1 TMFC 65 4 TMFCs 75 Gas Overlay Operating Mode 79 Gas Overlay Mixing 79 Gauge Screen 36 Growing CHO Cells 139 H Headplate Ports 16 HOT SURFACE Symbol for v Index of Drawings 129 Index of Tables 131 Information Sheet vii Inoculation 105 Inputs amp Out
102. in a 5 0 Liter L vessel using CD CHO serum free medium in a batch culture is described Introduction Historically stirred tank fermentors and bioreactors have been the trusted design for culturing all types of submerged cultures including suspension and anchorage dependent mammalian cells insect yeast plant and microbial cultures The tried and tested tank design offers scalability and proven reproducibility which is pivotal for cost saving process development and productivity In the last decade there has been an increasing acceptance and use of single use technologies due to their convenient operation and low start up cost Single use systems eliminate the need for cleaning and sterilization reduce validation requirements provide rapid turn around between runs and significantly reduce the risk of cross contamination and microbial contamination because the culture vessel is only used once and then discarded Although single use stirred tank systems in the 75 2000 L scale have been on the market for some time as have small scale single use bags that are gently rocked rather than stirred until now there has been no single use stirred tank system for small scale work The new CelliGen BLU fills that void offering a proven stirred tank design as well as the benefits of single use technology in a benchtop system New Brunswick Scientific User s Guide 140 24 3 24 3 1 24 3 2 24 3 3 Materials and Methods Single
103. inet ANALOG INPUT ANALOG OUTPUT g ea eeegu mA ua popa og oreo Ml less ea mE lesg less 3 123 5 123 E 12 4 7 12 4567 1 These dip switches are provided to switch connectors 1 3 between mA down and V up 2 These dip switches 1 3 are for mA or V_ 3 These dip switches 4 7 are for V only 3 The preferred connection for pumps is 4 20mA If you are using a 0 5V connection skip to step 5 To use 4 20mA connect the end of the green cable wire to one of the three 1 2 or 3 negative outputs at the bottom 4 Connect the end of the white cable wire to the positive output at the top be sure to use the same number 1 2 or 3 as you used for the green wire 5 If and only if you are using a 0 5V connection instead of 4 20mA connect the end of the green wire return to one of the four 4 5 6 or 7 negative outputs at the bottom and connect the end of the black wire 0 5V input to the matching positive outputs at the top 6 Setup the pump control loop using a loop for external equipment see Section 6 2 6 amp Figure 11 repeated on the following page for easy reference After a pump is added it will appear on the PUMPS screen CelliGen BLU M1363 0050 User s Guide 91 Figure 11
104. int yourself with the Trend Graph buttons at the bottom of the graph CelliGen BLU M1363 0050 User s Guide 83 Figure 36 Trend Graph pS Trend Screen New Brunswick Scientific Cell Culture Mode 00 200 06 39 06 54 07 09 1 MAAA e E AA UCH Pv 1 6 8 05 1 37 SP 0 0 7 00 0 00 ou 0 0 0 0 0 0 07 24 Trend Gy Pumps A Alarms Ke setup ES Summary cano a Case 2 1 Trend Graph buttons see Table 6 for more information 2 Timespan Indicator Table 6 Trend Graph Buttons Button Feature Description Summary Press this button to cycle through three summary display modes all eight loops at once loops 1 4 red fuschia and loops 5 8 dark green light green Single Press this button to display the graph for one loop at a time in the order left to right they are displayed in the colored buttons at the top of the screen Export Press this button to export a text file containing all of the Trend data See Section 12 1 1 for detailed instructions lt lt lt Ramp Down Press this button to select a lower Timespan or to move the Read Line toward the left of the screen Timespan Indicator Using the Ramp Down or Ramp up button on either side of this edit box select the timespan to display onscreen Preset increments range from 30 Minutes to 144 Hours gt gt gt Ramp Up Press this bu
105. ints See Section 9 3 1 above Cascade System Cascading brings several systems together to work jointly for the achievement of your goal An example of how this works would be controlling pH by using a cascade from a pump as the pH alters from the setpoint the program would cause the pump to turn on and off to compensate See Section 10 for details about setting cascades Pump Assignment The user has the ability to assign each pump present in the system To assign a pump 1 From any screen press the PUMPS button at the bottom to open the PUMPS GAUGE screen see Figure 18 Figure 18 Pumps Screen New Brunswick Scientific Cell Culture Mode Pump Screen Celligen D Pump1 14 RPM 0 0 Pump2 109 RPM 0 0 Pump3 109 RPM 0 0 Setpoint py Control Mode Off 0 0 Setpoint PY Control Mode 0 0 Setpoint PV Control Mode 1 Assignment None None None Flow Rate mL Second Flow Rate mL Second Flow Rate mL Second Calibrate Calibrate Calibrate Calibrated Calibrated Calibrated 0 0000 0 0000 0 0000 Total Total Total Period Sec 10 Period Sec 10 Period Sec 10 El Summary u calibrate T Cascade 24 Trend FONA A Aam 2 setup 1 Step 2 Press the Pump1 Assignment button to show the Pump Assignment pane see Figure 26 to continue New Brunswick Scientific User s Guide
106. l amp integral controller which operates peristaltic pumps assigned to add base or to use of gas es for this purpose The user can also select a deadband value to control pH within the user assigned range no base or gas will be added when the pH value falls within the deadband tolerance above or below the setpoint CAUTION 3 8 3 9 Do not expose the pH sensor within the vessel to direct light or to any liquid with less than 100mM ionic strength as these may adversely affect the performance of the sensor Cell culture pH control is typically done by the addition of base by means of a peristaltic pump or by sparging in CO to lower the pH Default P amp I proportional amp integral values are preset at the factory We strongly recommend that you maintain the factory set parameters DO Control DO is controlled and measured in the range of 0 200 by means of a non invasive non disposable polarographic DO probe It is sensed by the DO electrode and control is maintained by the P amp I controller by adjusting the gas mixture that is injected into the vessel Default P amp I proportional amp integral values are preset at the factory We strongly recommend that you maintain the factory set parameters Exhaust System The exhaust gases pass into the exhaust tube where the gases are heated up by the exhaust heat blanket any moisture present there is raised above the dewpoint and can easily pass through the filter
107. l Loops 32 Return Authorization 145 146 Returned Material Authorization Number 145 RMA 145 146 Rotameter 5 RS232 422 Computer Interface 43 S Sampling System 7 Saving a Process Configuration 135 136 Scales 7 Security Keypad 102 Serial Inputs amp Outputs 23 Service 111 Service Connections 10 12 Service Utility Electrical 13 15 20 21 Setting Alarms 92 Setup Screen 40 97 New Brunswick Scientific Shutdown after a Run 107 Siliconizing the Vessel 105 Specifications 25 Stacking Pumps 132 Start Up Screen 29 Supervisory Software 8 Synoptic Screen 31 T Table Index 131 Table of Contents xii Temperature Control 5 RTD 5 Setpoint 5 Temperature Control 104 Touchscreen Calibrating the 28 100 Installing the 10 Trend Graph 82 Creating a 81 Trend Graph Buttons 83 Trend Screen 38 Trend Setup Screen 82 Troubleshooting 111 U User Button 102 103 Utilities 11 V Validation Documentation 26 Vessel Description of 4 Installation of 21 Siliconizing the 105 Vessel Pressurization 14 W WARNING Symbol for v Warnings Recap of 1 Warranty vi Option for Extended vi User s Guide 150 Warranty Registration vi Z Zoom Coordinates 84 CelliGen BLU M1363 0050 User s Guide
108. l flow rates for each gas change but the total gas flow is maintained at a constant level set by adjusting the Total Combined Flow New Brunswick Scientific User s Guide 78 Figure 33c AirFlo 1 Gauge with 4 TMFCs in 4 Gas Auto Mode m Gauge Screen New Brunswick Scientific Cell Culture Mode CelliGen BLU AirFlo 1 0 0 CCM Gas Mix Selection 100 0 M N2 F CO2 Set Py Dut Kanae Aade Mix Flow Cycle Times Secon too f oo 100 0 On Time 120 OffTimefo 100 0 100 0 100 0 4 Gas Auto imi 1 Set Low Set High 20 100 0 Total Combined Flow 100 0 2 Alarm Settings Totalizer CC 20 20 0 0 i summary I ca T Casc p lt Trena Pumps A avarms Ao Setup 1 4 Gas Auto mode has been selected the button is green 2 Total Combined Flow box If N2 is deselected from the Gas Mix Selection pane the system responds by changing to 3 Gas Auto mix By changing to this mode users now have the ability to define a minimum and maximum combined flow cascade as if they had a 3 TMFC system see Figure 33c and Section 11 3 above Figure 33d AirFlo 1 Gauge with 4 TMFCs in 3 Gas Auto Mode ful Gauge Screen New Brunswick Scientific Cell Culture Mode CelliGen BLU AirFlo 1 0 0 CCM Gas Mix Selection 0 0 A ace 1 Set Py Ou ManuallCascade Mix Flow Cycle Times Seconds
109. le just prior to calibration see Section 7 1 New Brunswick Scientific User s Guide 20 2 Gently insert the probe into its headplate port see Figures 4a 4b amp 5 on the previous pages for reference and Figure 5a below 0 NOTE 4 6 7 4 6 8 The pH port is fluted and the fit is snug so gently turn the probe as you press it into the port to avoid breakage When the probe is fully inserted it becomes difficult to remove this ensures that the probe remains secure during the culture run Figure 5a pH amp DO Headplate Ports close up 1 DO port 2 pH port Inspecting the DO Probe Inspect the probe for possible shipping damage immediately report any damage you may observe to the New Brunswick Scientific Service Department Remove the protective cap from the electrode end The membrane is delicate and care must be exercised to prevent accidental damage Never rest the probe on its membrane DO Probe Preparation Default P amp I proportional amp integral gains are preset at the factory It is strongly recommended that you maintain the factory set parameters Nevertheless P amp I gains for the DO loop can be modified by an experienced operator using the touchpad on the front of the control cabinet Whether you choose to use as recommended the factory set values or to alter them it is highly unlikely that you will ever need to re set or change them unless you change your proce
110. llow six hours for polarization prior to calibrating the probe E ME cautions Never attempt to operate the system if the DO probe is not installed with its cable connected to the control cabinet 7 3 DO Probe Calibration Setting Zero There are two methods to obtain zero for calibrating the DO probe Review both methods and use the one you prefer Method 1 1 Remove the DO cable from the DO electrode 2 Go to the CALIBRATION screen see Figure 15 and select DO 3 Enter 0 in the Set Zero edit box see Figure 22 on the following page then press Set Zero 4 Reconnect the DO cable to the DO electrode O NOTE If you use Method 1 make sure the probe is not disconnected for more than two minutes Method 2 0 NOTE Nitrogen is needed for Method 2 There is an No gas inlet on the control cabinet for this purpose make sure that your nitrogen source is connected to this inlet New Brunswick Scientific User s Guide 48 Nn Connect the DO cable to the DO electrode and the control cabinet Go to the CALIBRATION screen see Figure 15 and select DO Press the N2 3 ON button see Figure 22 If your system has 3 or 4 TMFCs however this button will not be present In this case manually turn the N2 loop on from the SUMMARY screen and set it to the maximum setting depending on vessel size and flow controller Set Agitation to at least 50 RPM In approximately 10 30 minutes the current value readin
111. lt the pump a little to the left to catch all three mating surfaces see Figure 51 Figure 51 Mounting Base Pump for Stacking 1 Baseplate screw face 2 Exposed screw face of base pump 3 Pump release lever 3 Once all three mating surfaces are aligned push the pump back against the baseplate turning it until it is straight upright and snaps into place CelliGen BLU M1363 0050 User s Guide 133 4 Now add the standard pump in the same way to the screwface of the base pump see Figure 52 Figure 52 Stacking the Pumps 5 When you have stacked pumps load the tubing one pump at a time starting with the base pump If you open both at the same time the back pump channel will be blocked by the front pump s cover Figure 53 shows two pumps stacked Figure 53 Stacked Pumps New Brunswick Scientific User s Guide 134 22 APPENDIX B SOME GENERAL CONCEPTS NOTE In this section all discussions of P I D control are to explain the theory on which it is based This product uses only P proportional amp integral control not D derivative 22 1 What is a Controller The local process controller is a multi loop controller which means it can control several process parameters simultaneously It compares process values with setpoints and creates independent control signals for each controlled parameter The control signals are used to drive appropriate
112. mains in view for a few seconds then it is replaced by the SUMMARY screen 6 2 3 Summary Screen The SUMMARY screen see Figure 8 is command central it puts as many as 32 loops at your fingertips 2 Figure 8 Sample SUMMARY Screen 3 1 Summary Screen LoopName 11 Volume Air 1 O2 2 N2 3 CO2 4 OviFlo New Brunswick Scientific Cell Culture Mode Setpoint Out Control Mode Units Off 10 E 0 0 0 0 0 0 Calib E case bo trend 0 Pumps A Atarms A setup 30 Jun 2009 06 27 s _ 7 8 9 1 Screen Name and Icon 8 NOTE The dark blue button usually represents the screen being 2 Unit Tab displayed Here it shows a new screen SYNOPTIC that is 3 Operating Mode accessible from this screen see Section 6 2 4 for details 4 Exhaust Heater button 9 Any ALARMS will appear in this space 5 Scroll Up buttons 10 Screen Access buttons 6 Scroll Down buttons 11 Your CelliGen BLU comes with pre assigned loop names As you 7 Current Date amp Time add more use the Scroll Down buttons to see them New Brunswick Scientific User s Guide 30 Table 3 below identifies the other interactive features of the SUMMARY screen Table 3 SUMMARY Screen Features Parameter Column Description LoopName The system comes with standard factory assigned control loops e g Agitation Temperature
113. matically populated with cascade values when Manual Cascade mode is selected they must be entered manually Each cascade must be set to Enabled see Figure 16 in Section 10 1 The default cascade values can be overwritten and customer values can be entered using the procedures outlined in this section CelliGen BLU M1363 0050 User s Guide 61 10 1 Creating a Cascade Figure 16a below shows the headers from the CASCADE screen set to Cascade From DO with an explanation of the settings those headers represent Figure 16a Cascade Screen 1 2 3 4 A Cascade Screen New Bruhswick Scientific Cell Quiture Mode Cascade From DO MA To Enable Start Setpoint DO Start Out 02 2 y 0 0 0 0 100 0 100 0 Ramon a I End Setpoint DO End Out 1 Start Setpoint is the loop value the user defines for the system to be at when initial DO Start Out is reached Typically this value will be close to the normal operating setpoint 2 ODO Start Out represents the DO output value where the user wants the cascade to begin When this output 9e is reached the setpoint of the Cascade To loop will change to the value entered as Start Setpoint The current DO output can be found on the SUMMARY screen at the intersection of the Output column and the DO loop row This value is calculated by using the integrated Pl values It is essentially a mathematical calculation of setpoint
114. matically based on the scale s process loop information Bag Hanger A bag hanger is provided so that disposable addition harvest and or sample bags can easily be suspended for use in your process The hanger has three hooks which allow you to hang up to three bags at the right side of the controller see Section 0 for installation details Recommended Accessories amp Supplies Before you begin to assemble your CelliGen BLU it would be prudent to verify that you have all of the following accessories and supplies readily at hand Rubber gloves e Needle free syringes C FLEX tubing e Media Plastic tubing connectors e Antifoam agent A tie gun e Addition bottles New Brunswick Scientific User s Guide A user s kit is available from NBS with many of the commonly required items including a selection of tubing clamps filters connectors and addition vessels Speak to your NBS sales representative for more information 3 15 Supervisory Software In addition to the built in Reactor Process Control RPC software that you interface with through the touchscreen your CelliGen BLU system can be remotely controlled from a PC via NBS BioCommand optional Modbus supervisory software see Sections 4 8 and 16 3 Consult your NBS representative for details CelliGen BLU M1363 0050 User s Guide 4 1 4 2 4 3 4 INSTALLATION Physical Location E CAUTION Do not open your pre sterilized vessel package until you ar
115. n is not responding Calibrate touchscreen 19 2 Product Return If you should ever need to return your CelliGen BLU to NBS for servicing or any other reason you will need a product return authorization Please consult Section 25 Appendix E Product Returns for instructions CelliGen BLU M1363 0050 User s Guide 113 20 DRAWINGS Figure 49a Control Schematics 120V BLK 14 New Brunswick Scientific GRN YELU NR gys 2 oa z H ro i Jai a mm UN 11 TODT a TBI TBI I Ot E TBL TBL Bs Les x x e E DRC gg BLU El User s Guide 114 Figure 49b Control Schematics 120V g MONITOR A C CABLE REWORK AS NEEDED 010 WHT TBI IDUCHSCRE EN SR BLK z DISPLA ND B1 _ o a E E a MED TOUCH y HEATES HEATER BLANKET BE el Lo lt i ES AJ BLK SSSUL og HT z 110 El Ao Lo 010 HT TBi anne CelliGen BLU M1363 0050 U
116. naninicos 87 13 3 PUMP FLOW RATE amp CALIBRATION METHODS o oooooccccconononnnancncnnononononanacnacicnncnononnnnos 88 13 4 PUMP PERIOD bn ta A FI tt Ari sons 89 13 5 INSTALLING AN EXTERNAL VARIABLE SPEED PUMP cccoconnnnnnccccccnonononnnnnocccnnononononoss 89 14 ABOUT ALAR MS Y YG ddos SAS Sd rW ALYS SDN O AAWL NAS A FARA A FANNOD DOE od SA dU Shan 92 141 ABS AND DEV ALARMS o nennen a a a a a a a ai e 92 142 SETTING ALARM SS e ai RTE 92 14 3 ACKNOWLEDGING AN ALARM cccccssceccccccccssssssesccccceccussssssceccessesseseesscescesesseenenenss 94 14 4 ALARMS HISTORY 0 A tea chee AA 95 15 USING THE SETUP SCREEN sssssssssssssssssssosososossssssosssosososososssssssssssssssssssssssssso 97 15 15 CONTROLLER SETUP cui a A aaa Lat 97 15 2 RECIPE MANAGER cccccccccccccsssssscecccccceessssssesceccccessnsssesccscceeseussssescesccseeuauessescssess 98 15 3 SYSTEM SETTINGS WG Y TG GW wee AA Ea i ANG 99 15 3 1 est uu DU Fum ann A NO o FODA OO 100 CelliGen BLU e M1363 0050 User s Guide xi 15 3 2 U PAGING SON WATTS idad 100 15 4 HARDWARE SETUP nu ee E LS Maia destila 101 153 SECURITY SETTINGS tt NN 101 16 PERFORMING A CELL CULTURE RUN occccccccccnnnnnncnnonononononononononoconoconnnnnoss 104 16 1 PREPARING FOR START UP coconnnnnncncccononononnnnoncncnccnononnnnnnoroconnononnananorocccnononnnnnicicoss 104 16 2 INOCULATION te do ld lat da dla e ce pad als il 105 16 3 START BIOCOMMAND IF PRESENT A aislada DLWS d
117. ns not to exceed field wired to 15 Amp 110 disconnect in panel Process Air Oxygen Carbon Dioxide 4 5 1 Electrical Requirements 100 120 Volts 50 60 Hertz 15 Amp 200 240 Volts 50 60 Hertz 15 Amp O NOTE The electrical requirements vary depending on the part number that has been ordered Model Part Number and Electrical Power Requirements for each bioreactor appear on a metal label affixed to the rear of the unit just above the connection for the power cord CAUTION Before making electrical connections verify that the supply voltage matches the voltage and the power requirements marked on the electrical specification plate located on the rear panel of the cabinet and the control schematics supplied with the unit WARNING FN High voltage Always make sure this eguipment is properly grounded 4 5 2 Gas Connections Gas inlets for both the sparger and the overlay are located on the left side of the control cabinet see Figure 2 New Brunswick Scientific User s Guide There are push in connectors for air nitrogen oxygen and carbon dioxide These connectors accept flexible tubing which is supplied with the bioreactor Other soft flexible walled chemically inert tubing such as Marprene Pharmed etc may be used as well WARNING Do not use this equipment in a hazardous atmosphere or with hazardous materials for which the equipment was not designed All gases suppli
118. nscreen language when available change date amp time update software and calibrate the touchscreen see Section 15 3 and to check or change the Hardware Setup see Section 15 4 Figure 20a Setup Screen Ke Setup Screen New Brunswick Scientific Cell Culture Mode CelliGen BLU Mily Exhaust Heater Controller Setup Recipe Manager System Settings Hardware Setup Security Settings Unit Type Unit Name CelliGen BLU CelliGen BLU 1 No of TMFCs 1 TMFC Range 0 100 CCM 2 Installed Options 3 M Gas Mix V Vessel Volume 7 Gas Overlay Module T7 Overlay TMFC Save Changes 4 EH Summary Carat E Cascade pS trena Pumps A narm 1 The quantity of Thermal Mass Flow Controllers TMFCs installed 2 The flow range of your TMFC s 3 The factory installed options present on your system 4 Whenever you make a change be sure to press the Save Changes button In addition you may wish to access this screen if you need to contact NBS Customer Service about your CelliGen BLU to check the status of installed equipment and the firmware version which you also see briefly in the START UP screen by consulting the Hardware Setup pane 15 1 Controller Setup When you open the SETUP screen normally the Controller Setup screen will display first see Figure 20a above and on the following page If you find any other Setup screen in the display press the Controller Setup tab to open this screen
119. ntegral Set PV Out 100 0 100 0 100 0 Limits Set Low Set High 00 1000 Decimal Places 00 00 00 ii Calib Lc Casc kK Trena Pumps A Arms As setup 1 Press here to open the Add User Defined Loop screen 3 In the Add User Defined Loop screen press the Remove button in the upper righthand corner 4 Ifthe loop is a pump only optional pumps have a Settings button that provides access to their Remove button see Figure 14 on the following page New Brunswick Scientific User s Guide 36 Figure 14 Deleting a Pump Control Loop a Pump Screen EE Summary Pump1 14 RPM Setpoint 99 PV 00 Flow Rate oa Calibrated 0 0000 Total 0 0000 None Period Sec 10 Callb New Brunswick Scientific Flow Rate TN Calibrated 0 0000 Total 0 0000 None Period Sec 10 Casc kh Trend Cell Culture Mode Pump3 109 RPM Setpoint 0 0 EN PV oo LJ El Em Settings Flow Rate E Calibrated 0 0000 Total 0 0000 Control Mode 1 None A mmm se Press the Settings button to open the screen where you can press the Remove button to delete the pump loop If there is no Settings button in the pump gauge see Pump1 8 Pump2 for example that pump s control loop cannot be deleted 5 When you return to the SUMMARY screen the loop will be deleted 6 2 8 Selecting Loop Control Modes Control modes vary a
120. o YW YAWN 22 4 8 OPTIONAL BIOCOMMAND SOFTWARE ocooocccccccnonononnnnonononcnnononnnnonononconononanno no ncncnnononanos 22 4 9 INPUTS OUTPUTS FOR ANCILLARY DEVICES coooooccncnononnnonoonnancnconnanonocnannnnnononnannnconns 22 4 10 INSTALL THE BAG HANGER cccsessesscesececessessnecececcceseessseceeesecsesenssssaeeeseceeseeeneaaees 24 5 SPECIFICATIONS secissssceasccscteciesoccdoacosstasis RR HR t eseas isese os 25 5 1 VALIDATION DOCUMENTATION cccccsssessssececescescessenseaeeeseceesessnsaseeeseccesesensnseaeeesees 26 2 CERTIFICA TION Sila Y dd Ry Ng GST 26 6 OPERATING CONTROLS usina ini e 28 6 1 TOUCHSCREEN cui HYN A RES GG YY YW YN DO 28 6 2 DISPEAY SCREENS ina 28 6 2 1 Touchscreen CalibratiOiiici decida 28 6 2 2 STAR IN OA 29 6 2 3 SU A LEE dn 29 6 2 4 SVNODNC SOC A id 31 6 2 5 A E GR E a EE E E ER 32 6 2 6 Adlina PSA a 33 6 2 7 Deleting Loops A AS A AAA UON O AU 35 6 2 8 Selecting Loop Control Modes oi a cai 36 6 2 9 Calibration SCHON accesses else ta bah cc a Y YD ea iots 37 6 2 10 CAS CAES CER DIA a r ba i 38 6 2 11 TWU LEAD OA RIF HRN YCH eg 38 6 2 12 LYSH _ i GTA ATO AT RR YE 39 6 2 13 Alarms Screen eie tia ba 40 6 2 14 SOU IATA IEE EEEE YRU a EU GO FYR FO Y DEN 40 6 3 RS232 422 COMPUTER INTERPACE sicario ii NODD y0 43 7 PREPARATION amp CALIBRATIONS 9949999995999999959900959900888800888800900880000009090000000000 45 7 1 PH CALIBRATION 5 A GW NY WG 45 7 2 DO PROBE P
121. o any liquid with less than 100 mM ionic strength Do not open your presterilized vessel package until you are ready to use it Before making electrical connections verify that the supply voltage matches the voltage and the power requirements marked on the electrical specification plate located on the rear panel of the cabinet and the control schematics supplied with the unit 10 13 continued New Brunswick Scientific User s Guide CAUTION Page Do not open any port in the vessel headplate until ready for use 14 Never turn on the heat blanket or exhaust tube heat blanket without first 18 plugging in the RTD and inserting it in the thermowell inside the vessel Before turning on the main power switch make sure that the power cord is 22 properly connected Do not allow the combined weight of the bags and their contents to 24 exceed 15 kg 33 Ib Never attempt to operate the system if the DO probe is not installed 47 Proper pH control is critically dependent on tubing size 53 When selecting an external pump to operate with your system please consult with your local sales representative to be sure the model you choose is 89 compatible with your CelliGen BLU Be sure to set the dip switches correctly when using either 4 20mA or 97 0 5V inputs outputs Acknowledging alarms is NOT a replacement for correcting the condition that triggered the alarm Diagnose th
122. oad up to 10 recipes Recipes can be saved and loaded using these top three buttons WIN gt The Delete button removes the currently selected recipe the Load Default button restores factory settings A recipe consists of all setpoints controller settings control modes calibration data and cascades set on a system New Brunswick Scientific User s Guide 42 Figure 20c System Settings Screen 1 New Brunswick Scientific Cell Culture Mode Controller Setup Recipe Manager FE EC Hardware Setup Security Settings N i TouchScreen English Date Time a0 pN 2000 oe a3 Software Revision CGD 13 Update Folder CGDCTRL Rev 11 EZE Phoenix AwardBlOS v6 00PG Maintenance E Summary R cano FA Casc JE 0 Pumps A vo ET 1 English is the default language When other choices Francais Deutsch Espanol become available the user will select the language here Use this pane to calibrate the touchscreen see Section 6 2 1 for details WIN Use this pane to view the Software Firmware version installed and to update software via the USB port see Section 15 3 2 for details 4 Use this pane to change Date and Time see Section 15 3 1 for details Figure 20d Hardware Setup Screen 1 Le Setup Screen New Brunswick Scientific Cell Culture Mode Controller Setup Recipe Manager System Settings LEE Me
123. onth clean all the metal parts of your control cabinet Use a soft damp cloth moistened with water or mild detergent If a detergent is used remove all residue by rinsing them with clean water When you clean the heat blanket wipe it with a clean cloth moistened with isopropyl alcohol CelliGen BLU M1363 0050 User s Guide 109 18 MAINTENANCE Preventive maintenance keeps your equipment in proper working condition When performed routinely maintenance results in longer life for your equipment It also reduces time lost due to equipment failure LA WARNING Always turn your CelliGen BLU off and disconnect the power cord before performing maintenance 18 1 Probe Maintenance and Storage Use soft facial tissue to clean the DO and temperature probes When they are not in use in a vessel the DO and temperature probes should be stored At no time should either probe be allowed to rest on its tip E ME cautions Never let a probe rest on its tip 18 2 Periodic Inspection At three month intervals perform the following checks and inspections O NOTE Before you begin make sure that the power switch is in the OFF position and that the power supply has been disconnected 1 Check all controls and accessible items power switch connectors screws nuts and bolts to make sure they are properly tightened Tighten any loose item s 2 Check that all controls and connectors are free of dust New Brunswick
124. oops were controlled and monitored through the powerful Reactor Process Controller RPC firmware installed in the controller DO was monitored using a non invasive reusable polarographic DO probe and pH was monitored using a non invasive optical pH probe and fluorescence sensor Inoculum Preparation One 2 5 mL vial of CHO cells was thawed and used to inoculate a 125 mL shake flask which contained 25 mL of serum free CD CHO medium Invitrogen 10743 029 which was pre warmed to 37 0 C The flask was placed on an orbital biological shaker NBS Innova 2000 placed inside a CO incubator NBS Galaxy 170 R and set at 120 rpm CelliGen BLU M1363 0050 User s Guide 141 24 3 4 24 3 5 The CO incubator was programmed to provide a gas mix of 5 CO and 95 air at 37 0 C On day 4 when the viable cell density reached 1 5x 10 cells mL the cells were transferred into a 500 mL shake flask which contained 100 mL of freshly made pre warmed medium and allowed to incubate for 3 additional days at the same conditions as earlier The cells were then transferred to two 1 L shake flasks each containing 250 mL of the freshly made medium The inoculum was grown in the shake flasks until cell density reached 2 0 3 0 x 10 cells mL with greater than 90 cell viability sufficient for the bioreactor inoculation Bioreactor Set Up amp Inoculation One day before the cells reached inoculation density the growth medium was warmed to 37
125. ough a bank of 4 solenoids and the gas flow is controlled automatically by means of the TMFC GasFlo process loop The software controls how the solenoids open and close and you will still be able to select a control mode from the gauge screen but the overall flow will now be regulated by setting a value for the TMFC Figure 30 Controller Setup Screen 1 TMFC PK Setup Screen New Brunswick Scientific Cell Culture Mode EU BLU Ally Exhaust Heater Controller Setup Recipe Manager System Settings Hardware Setup Security Settings Unit Type 7 Unit Name CelliGen BLU CelliGen BLU No of TMFCs 1 TMFC Range 0 100 CCM Installed Options Gas Mix V Vessel Volume T7 Gas Overlay Module Overlay TMFC Save Changes EE summary Calibrate a Cascade pS Trena 0 Pumps A Nam 1 See Section 11 5 for details on the Gas Overlay The gauge screens for each of the gases will have the option for 3 or 4 gas mixing and in 3 Gas mode will also have the GasFlo cascade feature Gas mixing selections allow the system to automatically regulate gas composition to maintain pH and DO setpoints To activate the gas control it is important to set one of the gas gauge screens to 3 Gas 4 Gas or Manual see Figures 31a 31b 31c amp 31d New Brunswick Scientific User s Guide 70 The GasFlo cascade feature which is automatically enabled with 3 gas mixing allows you to define the high and low settings
126. ow and setpoints may never be reached Again this can be mistaken for defective components P I D constants are usually established by methodical trial and error New Brunswick Scientific User s Guide 136 22 5 What is P I D Tuning Tuning consists of establishing controller settings the proportional integral and derivative constants such that the controller provides proper control Ifthe P I D constants are incorrect the control signal may be too weak for the parameter to ever reach setpoint or at the other extreme the controller may respond excessively to small errors causing the actuator to oscillate between high and low values Usable P I D constants must be determined for each P I D loop The process is largely one of calculated trial and error All loops that are configured with the P I D control mode must be tuned When delivered as part of an NBS system P I D loops will have been tuned at the factory to work correctly with the NBS controlled instruments For other applications the user is responsible for P I D tuning Tuning can be a complex task for those unfamiliar with the process which is why a trained engineer or technician normally performs this task A number of textbooks that explain the theory and describe the process could be useful for the mathematically inclined novice The Ziegler Nichols method described in the footnoted reference is used at our production facilities The following suggestions ar
127. pH DO etc for the bioreactor There are also unassigned loops available to be named and set up by the user when adding external equipment for a maximum total of 32 loops PV Setpoint Process Variable here the display reflects the current value for each loop in comparison to its setpoint displayed in the next column The current setpoint default or user set for each loop Out Control Mode The current percent output for each loop This is an automatic control function to maintain current readings within the setpoint tolerance range Depending on the loop the control mode may be Off Auto Manual On or 4 Gas Unit of measure This is the unit of measure used for the PV and Setpoint Cascade If any cascades have been programmed they will be displayed here Summary This screen is command central it shows all your loops their current readings setpoints and what has been programmed for them Synoptic Calibration This screen is a graphical alternative to the SUMMARY screen It shows your loops their current readings and their setpoints It also displays the current state of the fixed speed pumps level probes and process valves This screen allows you to calibrate the DO probe the pH probe and vessel volume Cascade A cascade is a control function that uses the output of one loop to influence the action and output of one or more other loop s This screen allows you
128. puts 22 Inspection of Boxes 3 Installation Gas Connections 13 International Offices List of 111 L Level Probes Application of 58 Liquid Addition Systems 52 CelliGen BLU M1363 0050 Loading Pump Tubing 51 Location Environment 9 Physical 9 Loop Setpoints Entering the 55 Modifying the 57 Main Screen Features 29 Maintenance 109 Maintenance Inspections 109 Manual Conventions v Mass Flow Controller 5 Media Temperature 104 Modbus Com Port Pin Designation 44 Motor Assembly Installation of 21 Motor Replacement 110 N N2 3 66 N2Flo 75 NBS Offices 111 NOTE Symbol for v O 02 2 66 O2Flo 75 Operating Controls 28 Overlay Gas Mixing Option 7 OvlFlo 79 OvlMix 79 P P amp I Gains 20 P amp I Values Factory Settings 55 Setting 55 pH Control of 6 pH amp DO Control via Gas Mixing 65 pH Probe 6 Inspection of 19 User s Guide 149 Installation of 19 PID Explanation of Constants 135 Explanation of Tuning 136 Plotting Trends 81 Probe Calibration Definition of 135 Product Return Authorization 112 Product Returns 145 Protocol for Growing CHO Cells 139 Pump Array Standard 86 Pump Assignment 57 Pump Assignment Screen 59 Pump Calibration 58 88 Pump Control Modes 87 Pump Flow Rate 88 Pump Period sec 89 Pump Setpoints 86 Pumps Screen 39 R Read Line 85 Recipe Files 98 Regulatory Compliance 26 Remove User 102 Renaming Contro
129. r headspace The flow rate is controlled automatically by one three or four thermal mass flow controller s automatic gas mixing can be accomplished by a solenoid bank when only one thermal mass flow controller TMFC is present The TMFC is regulated automatically according to values set via the control cabinet touchscreen If you wish to use a rotameter there is an option for no TMFC The percentage of gas blended and injected into the vessel can be controlled manually by the user or automatically through the controller CO can be added to lower pH and nitrogen can be added to lower DO for further information on cascading see Section 10 Default P amp I proportional amp integral values are preset at the factory We strongly recommend that you maintain the factory set parameters Pumps Three Watson Marlow pumps are provided as standard on the right front of the control cabinet These pumps are very easy to load see Section 8 2 They are all fixed speed peristaltic pumps Pump 1 is set to 14 RPM and Pumps 2 amp 3 are set to 109 RPM For more information on selecting setpoints calibrating flow rates and selecting pulse periods see Section 13 for information about optional stacked pumps see Section 0 Appendix A New Brunswick Scientific User s Guide 3 7 pH Control pH is controlled in the range of 6 00 8 00 The pH is sensed by a non invasive optical pH sensor Control is maintained by a P amp I proportiona
130. reen button to open the ALARMS screen Press the red LoopName box to open that control loop s ALARMS screen Press the Current Alarms button to open the Current Alarms Summary screen Press the Acknowledge All button All alarms will be deleted from this screen Press the Scroll Back lt lt lt button to return to the ALARMS screen ae IA To acknowledge all alarms for all control loops at the same time 1 Press the ALARMS screen button to open the ALARMS screen 2 Press the Acknowledge All button All alarms will be deleted from this screen 3 Press the Scroll Back lt lt lt button to return to the ALARMS screen CAUTION Acknowledging alarms is NOT a replacement for correcting the condition that triggered the alarm Diagnose the cause of the alarm condition and rectify the situation to ensure proper operation of your CelliGen BLU 14 4 Alarms History Each time an alarm is triggered whether Visible and or Audible the controller records the event The controller also records each alarm acknowledgement You can access the Alarms History screen 1 to consult the data 2 to save the data to an optional auxiliary PC and or 3 to purge the records once the condition has been rectified To access the Alarms History screen see Figure 45 on the following page to consult data 1 Press the desired control loop s LoopName box in the main ALARMS screen New Brunswick Scientific User s Guide 96 Figure 45 S
131. s E CAUTION Before making electrical connections verify that the supply voltage matches the voltage and the power requirements marked on the electrical specification plate located on the rear panel of the cabinet and the control schematics supplied with the unit Figure 1a Front 8 Rear Views of Control Cabinet TA FWD pr neu l 1 M 2 a AE i u SEE i ji N 2 a Am 3 4 B d 5 tl D r 6 a 6 h a 7 a dl I El ue a 1 Touchscreen 5 Power cord connection 2 Heat vent 6 Service connections see Figure 2 3 Pumps Section 4 5 4 See Figure 6 Section 4 9 7 Power switch ON OFF With reference to Figure 1b on the following page connect the cabinet s power cord plug com port connector and VGA monitor connector to the bottom of the touchscreen monitor CelliGen BLU M1363 0050 User s Guide 11 Figure 1b Touchscreen to Control Cabinet Connections 1 Touchscreen rear view 3 Touchscreen bottom view 2 Attach the monitor to the control cabinet 4 Power cord plug mounting rack with the 4 screws 5 COM port connector provided using these 4 holes 6 VGA monitor connector 4 5 Utilities LX WARNING Do not use this eq
132. s flow to avoid foaming In this control mode flow will automatically adjust based on both pH and DO demands In 3 gas mode we suggest that you always use an overlay if possible to help maintain your setpoints In 4 Gas control mode see Figure 31b the controller automatically adjusts the gas Air O2 CO2 and N2 percentages to control pH and DO Total gas flow is digitally controlled and can either remain constant or be manually linked to a cascade in the individual gas gauge screens GasFlo cascade is not automatically available in 4 Gas mode CelliGen BLU M1363 0050 User s Guide 71 Figure 31b Air 1 Gauge Screen with 1 TMFC amp 4 Gas Control ful Gauge Screen New Brunswick Scientific Cell Culture Mode CelliGen BLU Air 1 100 0 Gas Mix Selection 100 0 F Mix Flow Cycle Times Seconds Set Pv Ou 4 6 100 0 100 0 M 1000 On Time 120 Off Time 0 100 0 100 0 FE Limits Set Low Set High 0 0 100 0 ee 1 Decimal Places 0 0 00 0 0 EXA 3 Casc kK Trena Pumps A a arms A Setup 1 GasFlo Cascade is not automatically available in 4 Gas mode In Manual 3 Gas control mode when you press the Manual control mode button and select Air O2 CO in the Gas Mix Selection pane of any gas gauge screen you must also manually enter setpoints for each gas in its gauge screen or create a user defined cascade to control the gas mix A GasFlo cascade can be
133. seeccccccccesssssscecceccessuusasceccescessuuenenecesess 134 22 3 WHAT IS PROBE CALIBRATION ccccccccccsssssesececcccccesssssscecceccessssssescscceccessuuensnsssesess 135 22 4 WHAT ARE P I D CONSTANTS NAAA 135 22 5 WHAT IS P D TUNNG nerin YF A YN AY 136 22 6 WHAT DO THE CONSTANTS MEAN ccsssseeccccccccssssssesceccescecssseessceccescessuneaensscesess 137 23 APPENDIX C CORROSION RESISTANCE cccccsssssssssssssssssssssssssssssssccces 138 24 APPENDIX D PROTOCOL FOR GROWING CHO CELLS 139 24 1 TABSTRA Cit GY EE EAS EE 139 DAD INTRODUCTION A Y yd dd a OEE EEN 139 24 3 MATERIALS AND METHODS cini naa ar N A A a ii A a 140 24 3 1 Y ER EA A AAN a aaa a FEN SAN FEED 140 24 3 2 GControll 22 on e ND YL T AA 140 24 3 3 Inoculum Preparation tas 140 24 3 4 Bioreactor Set Up amp TRI CUA a 141 24 3 5 Operational PATONES ES ON E aN daw GOR 141 New Brunswick Scientific User s Guide xii Zam RESUETS AND DISCUSS ON YW imesh E NE 143 25 APPENDIX E PRODUCT RETURNS somvccinoininonacia ocio dean OUT doin denia 145 25 1 RETURN PROCEDURE sunno lia 145 25 2 RETURN AUTHORIZATION AND DECONTAMINATION CERTIFICATE cccoooocccncoonncnncnnns 145 26 A Dn FN TT RH o HR GN A 147 CelliGen BLU e M1363 0050 User s Guide 1 WARNINGS amp CAUTIONS The following section is a recap of all WARNING and CAUTION messages contained in this manual This information is essential to the safe operation of your CelliGen
134. ser s Guide 115 Figure 49c Control Schematics 120V LK lo BLK F2 LK Bl MOTOR E ESA e E lt CONTROLER 4 p oH e du E le u i ee qe wn ESA OS TACH A HT A LI A E TACH New Brunswick Scientific User s Guide 116 Figure 49d Control Schematics 120V F002 LK M lt a Ol O7 ee E FI FILTE gt l SSY PCB CelliGen BLU M1363 0050 User s Guide 117 Figure 49e Control Schematics 120V m ose co RE E TE M or LI COM M y O TB2 E HEATE SLK NO 013 4 EX HEATE z E EXHAUST L HEATER PES El E ELK 3LK r YEL ZZER TR BARA E Pilea 3LK 3LK a z Y E A J ik 0 g 5 N TB2 3 v E LK J a uk TI ki LI ll TEMPERATURE l R ENSOR E TI ll r TT ll TT e New Brunswick Scientific User s Guide 118 Figure 49f Control Schematics 120V BASIC FERMENTATID CI a gt 4 gt RE
135. set Air always makes up any difference to complete the 100 total gas CelliGen BLU M1363 0050 User s Guide 67 Figure 29a Air 1 Gauge with 0 TMFC amp 3 Gas Control 1 full Gauge Screen New Brunswick Scientific Cell Culture Mode Celligen BLU Air 1 100 0 Gas Mix Selection Setpoint Off M Air O2 CO2 T 4Gas 3 100 0 m Mix Flow Cycle Times Seconds Set PY Dut 1000 1000 100 0 ER On Time 120 Off Time 0 4 100 0 100 0 100 0 2 EE Set Low Set High 0 0 100 0 Decimal Places i Calib El casc lt trena Pumps A narms A setup 1 Gas Mix Control buttons 2 Press the Manual button to control gas mix manually 3 If you select a 4 gas mix here the 3 Gas Gas Mix Control button will change to 4 Gas allowing you the option of switching to automatic 4 Gas control 4 Set Mix Flow Cycle Times typically used in microcarrier applications by indicating here the number of seconds each gas should cycle On and Off Figure 29b Air 1 Gauge with 0 TMFC and 4 Gas Control full Gauge Screen New Brunswick Scientific Cell Culture Mode Celligen BLU i O Air 1 100 0 Gas Mix Selection Setpoint Off F Air 02 C02 4Gas 1 Set WW o Bu mc Mix Flow Cycle Times Seconds 1000 1000 100 0 On Time 120 Off Time 0 100 0 100 0 100 0 Limits Set Low Set High oo 100 0 Decimal Places
136. sible Solution TEMPERATURE Readout is a negative value e Make sure the temperature probe is connected to the typically 225 C cabinet jack The unit will not heat up e Make sure the temperature probe is plugged into the vessel thermowell AGITATION Agitator does not turn or turns e The motor drive may not be seated securely on the only slowly coupling check the orientation of the notches e Make sure the motor is plugged into the cabinet receptacle TURN OFF MAIN POWER BEFORE CONNECTING THE PLUG continued New Brunswick Scientific User s Guide 112 Problem Possible Solution DO and pH PROBES DO probe readings are erratic Recalibrate the probe carefully following instructions in this manual Recharge the probe carefully following instructions in this manual Probe may need a new membrane and a refill of electrolyte Check for a secure connection Replace probe cable or DO probe pH probe readings are erratic Recalibrate the probe carefully following instructions in this manual Check for a secure connection GASFLOW There is insufficient gas flow Check that the air pressure is within the specified range Make sure the control mode for DO and for pH is set to AUTO not OFF Make sure that the GasFlow loop is ON Make sure that the Air loop is in O2 Enrichment mode Make sure that the DO cascades are Enabled GENERAL Touchscree
137. ss Even if the power fails during a run the P amp I values pre set if you do not change them or your settings when you do are stored in memory and should still be in effect when the power is restored For details regarding P amp I settings see Sections 22 4 amp 22 5 CelliGen BLU M1363 0050 User s Guide 21 It is always prudent to check these values at the beginning of a run 4 6 9 Install DO Probe 1 Wear protective gloves to protect yourself in case of accidental breakage 2 Gently insert the probe into its port see Figures 4a 4b 5 amp 5a on the previous pages for reference 0 NOTE The DO port is fluted and the fit is snug so gently turn the probe as you press it into the port to avoid breakage When the probe is fully inserted it becomes difficult to remove this ensures that the probe remains secure during the culture run 4 6 10 Install Temperature RTD Probe 1 Insert the RTD temperature probe with reference to Figures 4a 4b amp 5 for location into its port making sure the probe extends all the way to the bottom 2 Attach the RTD cable to the Temperature connector on the control cabinet see Figure 2 4 6 11 Install Vessel Position the vessel next to the control cabinet in the rounded cut out designed for vessel placement between pumps and connectors NOTE Remove puill off the plastic clips that immobilized the scale during shipping If you are using the optional scale pl
138. ter the Imin Imax pHO dpH and Cal T values in their respective edit boxes here by pressing the box using the popup keypad to type the number then pressing the keypad OK button 2 Step 5 When all values have been entered press the Apply button 3 See Step 7 below 4 See Step 8 below 5 See Step 9 below 6 Take a sample of media see Section 16 4 for the sampling procedure and measure the pH using the external meter 7 Press inside the edit box below the Set Zero button and using the popup keypad enter the value of measured pH from Step 6 Press the OK keypad button to save the value 8 Press the Set Zero button 9 Press the Close button O NOTE A Raw Value indication of 9999 means that the values you have entered are incorrect or that the pH probe is either not connected or not functioning properly CelliGen BLU M1363 0050 User s Guide 47 You can expect to see a very slight drift in your pH value over a period of days due to the properties of optical pH sensors We recommend that you recalibrate the probe every 3 5 days following the procedure specified above 7 2 DO Probe Polarization NOTE If the probe has been disconnected from a voltage source either the unit s O amplifier or a separate polarizing module for longer than 5 minutes it will need to be re polarized To re polarize the DO probe connect the probe to the operating O amplifier or polarizing module A
139. thout notice Updates to information in this document reflect our commitment to continuing product development and improvement Manual Conventions O NOTE CAUTION gt WARNING gt WARNING A WARNING CRUSH WARNING New Brunswick Scientific Notes contain essential information that deserves special attention Caution messages appear before procedures which if caution is not observed could result in damage to the equipment Warning messages alert you to specific procedures or practices which if not followed correctly could result in serious personal injury This particular Warning message represents a potential electrical hazard This particular Warning message whether found in the manual or on the unit means HOT SURFACE and therefore represents a potential danger to touch Crush Warning messages alert you to specific procedures or practices regarding heavy objects which if not followed correctly could result in serious personal injury User s Guide vi WARRANTY New Brunswick Scientific s equipment is protected by a comprehensive warranty The warranty covers faulty components and assembly and our obligation under this warranty is limited to repairing or replacing the instrument or part thereof which shall prove to be defective after our examination The NBS warranty does not cover loss of time or materials such as the loss of biological or biochemical by products caused by an
140. to optimize either the medium or the cell culture process control parameters This study was only intended to document a general guide to bioreactor setup and operation and present typical results you could expect to achieve with your mammalian cell line For protocols on other cell lines or for additional information on the CelliGen BLU see www nbsc com BLU Figure A Cell growth over the 7 day run 6 00 5 00 4 00 3 00 2 00 Viable cells x10e6cells ml 0 1 2 3 4 5 6 7 8 Culture time day New Brunswick Scientific User s Guide 144 Figure B Cell viability over the 7 day run Viable cells x10e6cells ml on o o 0 1 2 3 4 5 6 7 8 Culture time day Figure C Glucose consumption and lactate production over the 10 day run 7 00 T 6 00 e Glucose dD io 5 00 m Lactate 4 00 amp amp 3 00 o 2 00 o 3 o 1 00 0 00 0 1 2 3 4 5 6 7 8 Culture time day 1 Reference A single nutrient feed supports both chemically defined NSO and CHO fed batch processes Improved productivity and lactate metabolism by Ma N Ellet J Okediadi C Hermes P McCormick E amp Casnocha S Biotechnol Prog 2009 Jul 27 Authors of this article Guozheng Wang Sr Research Scientist R amp D Lab Wenying Zhang Research Scientist Assistant R amp D Lab Rich Mirro Prod
141. to set up cascades to view current settings and to make changes to those settings Trend This screen allows you to set the parameters for plotting trend graphs and to view the graphs that track the activity of the selected loops over a certain period of time Pumps This screen gives you access to the Pump Gauges screen where the three pump gauges are displayed providing both current readings and the opportunity to change pump settings Alarms In this screen you can turn alarms on and off read the alarm history and acknowledge any alarm Setup This screen allows you to load amp save recipes and to make changes to your system settings hardware setup amp controller setup 1 see the following page continued CelliGen BLU M1363 0050 User s Guide 31 Other Buttons Description Exhaust Heater Scroll Up Press this button to turn the exhaust heater on or off when it is connected to the control station The button is green when the heater is on Press this button to scroll upwards one loop at a time Scroll Down Press this button to scroll downwards one loop at a time 1 The far left navigation button at the bottom of all main screens is a toggle between the SUMMARY and the SYNOPTIC screens When viewing one the button will be labeled for the other Upon leaving either for one of the other screens the default selection shown on the button will
142. ttings Unit Type Unit Name CelliGen BLU CelliGen BLU No of TMFCs 3 TMFC Range 0 100 CCM Installed Options F Gas Mix V Vessel Volume 1 FV Gas Overlay Module Overlay TMFC Save Changes Es summary i Calib a Casc pS Trena 0 Pumps A alarms BC 1 See Section 11 5 for details on the Gas Overlay The gas process loops you will find in the SUMMARY screen are labeled AirFlo 1 O2Flo 2 or CO2Flo 3 Their numbers 1 3 correspond to the gas connections on the cabinet see Figure 2 With three TMFCs when you set any gas gauge to Manual Cascade see Figure 32a on the next page Air O and CO setpoints can either be manually entered in their individual gas gauge screens or they can be controlled by cascades see Figure 32b on the next page New Brunswick Scientific User s Guide 74 Figure 32a AirFlo 1 Gauge with 3 TMFCs in Manual Cascade Mode full Gauge Screen New Brunswick Scientific Cell Culture Mode CelliGen BLU AirFlo 1 0 0 CCM 0 0 Set lad Dut Manual Cascade 1 0 0 00 oo 100 0 100 0 100 0 3 Gas Auto Limits 2 Set Low Set High 2 0 100 0 Decimal Places Totalizer CC 4 20 20 0 0 Ut cai a case Trend Oy Pumps A Aims 2 setup 3 Manual Cascade mode has been selected the button is green 2 NOTE Gas flow limits are 2 100 CCM SLPM not percentages 3 The Totalizer feature tra
143. tton to select a higher Timespan or to move the Read Line toward the right of the screen Zoom Press this button to open an interactive mode where you can zoom in on a section of interest on one plot The button turns red when you touch it indicating you are in Zoom mode See Section 12 1 2 below for detailed instructions Read Line Press this button to open an interactive mode where you can move a vertical cross sectional line across the graph to aid in determining a particular reading See Section 12 1 3 below for detailed instructions New Brunswick Scientific User s Guide 84 12 1 1 Using the Export Button To export Trend data as a text file to a USB external memory device for use with a PC program e g Microsoft Excel 1 Install the USB external memory device into one of the USB connections on the back of the Control Cabinet 2 Open the TREND screen and push the Export button 3 In the screen that opens select the USB external memory device from the list of available drives 4 Touch the empty FileName box Using the touchpad that appears enter the desired file name then press the OK button Press the Save button to save the file to the USB external memory device 6 Remove the USB external memory device and use it to download the data to your PC r 12 1 2 Using the Zoom Button To zoom in on a particular section of one loop plot 1 Press the Zoom button at the bottom of t
144. uct Manager Vikram Gossain Applications amp Training Manager New Brunswick Scientific CelliGen BLU M1363 0050 User s Guide 145 25 1 25 2 25 APPENDIX E PRODUCT RETURNS Return Procedure As explained in Section 19 2 should you need to return your CelliGen BLU to NBS for any reason first contact Customer Service to obtain a Returned Material Authorization RMA number This number must appear on the outside of the shipping container otherwise NBS Receiving will refuse to accept the shipment In addition you must also certify that the instrument being returned has been thoroughly cleaned and decontaminated A form for this purpose is provided on the following page you can photocopy it and fill it out by hand It can also be downloaded from our website www nbsc com if you prefer to fill it out electronically A copy of the completed Return Authorization and Decontamination Certificate must be attached to the outside of the container with a second copy packed inside with the instrument Return Authorization and Decontamination Certificate A sample form for you to copy and fill out is provided on the following page New Brunswick Scientific User s Guide 146 New Brunswick aneppendorf company Eguipment Return Material Authorization RMA and Decontamination Certificate Contact New Brunswick Scientific for an RMA number prior to returning any eguipment then complete this form and attach it to th
145. uipment in a hazardous atmosphere or with hazardous materials for which the equipment was not designed The control cabinet assembly must be properly connected to gases and electrical power All gas connections are located on the lefthand side of the cabinet see Figure 2 on the following page The power connection is located on the back of the cabinet see Figure la New Brunswick Scientific User s Guide 12 Figure 2 Control Cabinet Service Connections 1 Sparge outlet connection 7 Inlets for overlay gases 2 GAS1 GAS4 inlets for gas connections 8 Scale connector optional 3 RTD temperature probe connection 9 Motor cable connection 4 Vessel heat blanket connection 10 DO probe cable connection 5 Exhaust heater connection 11 pH probe cable connection 6 Outlet to Overlay port on headplate Using standard plant practices and respecting all applicable codes connect services to the appropriate connections as recapped in Table 1 and explained in greater detail in Sections 4 5 1 4 5 2 CelliGen BLU M1363 0050 User s Guide 13 Table 1 Service Connections Service Utilit Requirement Connection 100 120 VAC 50 60 Hz Single Phase 100 120 VAC 1ph 15 Amp fluctuations not to exceed field wired to 15 Amp 110 disconnect in panel 200 240 VAC 50 60 Hz Single Phase 200 240 VAC 1ph 15 Amp fluctuatio
146. ume in the Loops pane 2 Place a graduated cylinder or Ehrlenmeyer flask on the scale and allow the reading to settle see Figure 22a Figure 22a Setting Zero for Scale Volume Calibration Calibration Screen New Brunswick Scientific Cell Culture Mode Celligen D DO Calibrating Loop a Volume Current Value 32 92 A A 1 EZ EZ E sunmary PRR E Cascade pe Trend Pumps A alarm J Setup 1 Allow the Current Value arbitrary reading time to settle 3 Press the Set Zero button 4 Fill the cylinder or flask to a known quantity of water e g 1 0 liter 5 Enter the volume in the Set Span edit box New Brunswick Scientific User s Guide 50 Figure 22b Setting Span for Scale Volume Calibration Calibration Screen New Brunswick Scientific Cell Culture Mode Celligen D DO Calibrating Loop Pa Volume Current Value 32 92 E EXA Set Span 5 Enter liquid volume here E summary PS E cascave e Trend Pumps A narm A Setup 1 Enter liquid volume here 6 Place the filled container on the scale then press the Set Span button 7 Remove the container from the scale and place the complete CelliGen BLU vessel assembly with motor heat blanket and exhaust heater but without media on the scale Press the Set Zero button The scale is now calibrated and will measure the weight of liquid added or removed from t
147. ut and one 0 5V analog output port Additional analog input and output ports are available on the control cabinet rear panel see Figure 5 for the connection of analog ancillary devices such as additional pumps gas analyzers and glucose analyzers After the inputs are connected to the control cabinet the collected information will be viewed and controlled via the touchscreen display Three of these additional analog input and output ports have dip switches to allow selection of either 4 20MA or 0 5V The other four are 0 5V dedicated CelliGen BLU M1363 0050 User s Guide 23 Two USB serial ports are available on the control cabinet rear panel see Figure 6 for the connection of serial ancillary devices such as scales for vessel and addition bottles You can connect a box with eight serial RS232 inputs and outputs to one USB port to allow you to connect and control up to eight scales or other ancillary equipment Figure 6 Inputs 8 Outputs for Ancillary Equipment ANALOG INPUT ANALOG OUTPUT I ced lt l nA mug ma y oa moo E 123 1 ane EES 12 4567 123 4567 a E 2 m o m a El D 4 He MODBUS ETHERNET O B
148. utinely calibrated before each use Most other probes need be calibrated only infrequently What are P I D Constants The mathematics of P I D control is familiar to most control and process engineers In P I D mode the controller creates a control signal that is based upon setpoint and input from a sensor The magnitude of the control signal is determined by a mathematical formula that can include proportional P integral I and derivative D terms The P I and D constants are three numbers that determine the relative sizes of the proportional integral and derivative terms respectively To use a temporal analogy the P or proportional part of the control signal reflects present deviations between setpoint and process value The I or integral component reflects past deviations and the D or derivative term anticipates future values of the error Generally with noisy or slow responding sensors such as dissolved oxygen and pH probes the D constant should be set to zero If the constants for a loop are too large that loop will oscillate displaying extreme swings in actuator output If for example agitation changes suddenly and frequently between minimum and maximum RPM one should suspect incorrect P I and D values for the agitation control loop This condition can easily be mistaken for a defective component when it actually results from incorrect settings If the constants are too small control response will be sl
149. ve one or more additional pumps press the gt gt gt button to continue past Pump 3 If you have a stacked pump it will be governed by settings for its base pump CelliGen BLU M1363 0050 User s Guide 87 Figure 39 Setting Pump Setpoint O Pump Screen New Brunswick Scientific Cell Culture Mode Celligen D Pump1 14 RPM Pump2 109 RPM Pump3 109 RPM 1 amp 2 Setpoint 9 Setpoint co 9 Setpoint oo Ez PV 00 PV o0 PV o0 gt 3 Control Mode Control Mode Control Mode re mci Flow Rate mL Second Flow Rate mL Second Flow Rate mL Second Calibrate Calibrate Calibrate Calibrated Calibrated Calibrated 0 0000 0 0000 0 0000 Total Total Total Period Sec 10 Period Sec 10 Period Sec 10 E Summary Calibrate T Cascade pS Trena WEL A arm As Setup Step 2 Press inside the Setpoint edit box for Pump1 Step 3 Use the touchpad that pops up to enter the desired setpoint then press the OK button in the touchpad to save the setpoint and to return to this screen or press the Cancel button to return to this screen without saving a setpoint Ni 3 If optional pumps are installed on your system these ramp up gt gt gt and ramp down lt lt lt buttons will be active allowing you to scroll to the next page or back 4 Repeat Steps 2 3 for each pump 13 2 Pump Control Mode There are three available control modes for each pump as explain
150. will see that the total will increase by this calibration standard 13 4 Pump Period At the bottom of each pump gauge is the Period Sec pane see Figure 41 Figure 41 Pump Period Sec Period Sec 10 Use this edit box and its associated touchpad to enter a pump cycle time in seconds For example if the pump setpoint is 30 setting a period of 5 seconds as illustrated will cause the pump to run 1 5 seconds stop for 3 5 seconds then cycle back on again 0 NOTE Running at a very low percentage renders the totalizer s results inaccurate We recommend the use of smaller tubing to avoid choosing a very low percentage for the pump setpoint 13 5 Installing an External Variable Soeed Pump CAUTION When selecting an external pump to operate with your system please consult with your local sales representative to be sure the model you choose is compatible with your CelliGen BLU 1 Connect the D connector of the pump s 3 wire cable at Interface on the rear panel of the pump see Figure 42 on the following page Your pump will be marked 115V or 230V depending on the electric supply you specified when you purchased the pump New Brunswick Scientific User s Guide 90 Figure 42 Variable Speed Pump 1 Use Interface to connect the cable provided 2 Locate the Analog Output Connections on the rear of the CelliGen BLU cabinet see Figure 43 Figure 43 Rear Panel of CelliGen BLU Cab
151. with a disposable optical pH sensor the light that blinks at the tip of the probe when it is plugged in to the control cabinet is not dangerous to your vision To ensure proper calibration of the sensor and to compensate for any discrepancy you will need an accurate external pH meter 1 Press the Calib Button at the bottom of the SUMMARY screen to open the CALIBRATION screen see Figure 15 repeated below for easy reference Figure 15 Calibration Screen Calibration Screen New Brunswick Scientific Cell Culture Mode Ally Exhaust Heater Calibrating Loop 1 pH 2 Current Value Raw Value 8 05 11008 3 Offset Zero Std 1003 8 E su SN ES case rena Pumps A Alarms Le Setup Step 2a Press pH here Step 2b and pH appears here Calibrating Loop box Step 3 Press the Probe Data button to open the PROBE DATA screen see Figure 15a 0O N the instructions continue on the following page New Brunswick Scientific User s Guide 46 Figure 15a Probe Data Screen 1 Calibration Screen New Brunswick Scientific Cell Culture Mode CelliGen BLU Probe Data can Calibrating Loop Imin 53 80 oH pHO 6 31 Current Value Raw Value dpH 0 53 74 740 CalT 104 0 Probe Data Set Zero Zero Std di e L ran 5 1 Step 4 As marked on the label on the bottom of your vessel and also on the vessel box en
152. y work interruption resulting from equipment failure it does not extend to equipment that has been subject to misuse neglect accident or improper installation or application nor does it cover any machine that has been repaired or altered by anyone other than an authorized NBS factory trained service representative without prior approval from your local NBS sales office Expendable items such as bearings and seals lamps probes sensors including incubator sensors glass filters etc are not covered The warranty begins on the date the equipment ships and extends through the period indicated in the chart below Instrument Parts Warranty Labor Warranty Innova 3 years 2 years Series 2 years 2 years Shaker Excella amp C 76 2 years 2 years Accessories 1 year 1 year CO Incubators Incubators 2 years 2 years Accessories 1 year 1 year 5 years Ficezais ULT Freezers vacuum insulation 2 years panels 12 years Accessories 1 year 1 year Fermentors Bioreactors amp all other 1 year 1 year NBS equipment 1 Chart recorders photosynthetic light banks etc 2 Stacking stand casters shelves etc 3 Chart recorders CO LN back up systems etc Warranty Registration To register your warranty with NBS complete the online form at www nbsc com under the How Can We Help tab or use the warranty card enclosed with your equipment Extended Warranty Option A variety of service plans are
153. you hang there 4 As needed feed the tubing from the bag s into the dedicated pump s and aseptically attach the tubing to the appropriate port connection addition harvest or sampler E CAUTION Do not allow the combined weight of the bags and their contents to exceed 15 kg 33 Ib CelliGen BLU M1363 0050 User s Guide 25 5 SPECIFICATIONS CelliGen BLU System Cell Culture Vessel Controller Control Station Controls up to 32 control loops stores 10 recipes 8 process variables for trend graphing Includes an industrial touchscreen monitor user interface 3 built in pumps amp connectors for all utilities amp communications signals Touchscreen 15 inch industrial monitor is standard with the Control Interface Displa Station Temperature Digital display in 0 1 C increments From 5 C above ambient temperature to 40 C Control P amp l for heating and cooling heating via the Heat Blanket and cooling via ambient temperature Platinum RTD probe Agitation Permanent magnet motor with high torque input Digital display in 1 RPM increments 25 200 RPM Control Pl controlled Optical photoplastic disc 500 lines rev with quadrature output Pitched blade 0 2um disposable filter Tube Heat blanketed Aeration 4 Gas System Up to 4 gases including air N2 CO 4 Oz delivered to sparger Sparger Porous Microsparger 7 12 0 2um disposable filter For calibration of DO probe Digital display in 0 01 pH increments

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