CELL-tainer - Single use bioreactor 20L Cell...
Transcript of CELL-tainer - Single use bioreactor 20L Cell...
OPERATION & INSTALLATION MANUAL: 2015-INS-CM2SCMA-V2.0final cell page 1/66
CELL-tainer® - Single use bioreactor 20L
Cell culture
OPERATION & INSTALLATION MANUAL
CM2SCEU/cell culture
February 2015
Document Number
OPS-INS- CM2SCMA-V2.0 final cell
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Chapter Table of contents page
Introduction ................................................................................................................................ 4 1. System survey ........................................................................................................................ 9 2. Front Panel ........................................................................................................................... 10
2.1 Gas mix system .......................................................................................................... 10 2.2 Mains switch............................................................................................................... 11 2.3 Safety system/Emergency stop .................................................................................. 11 2.4 Touch screen ............................................................................................................. 11
3. Side panel ......................................................................................................................... 12 3.1 Gas connections ........................................................................................................ 12 3.2 Gas outlet connection ................................................................................................. 12 3.3 Chilled water connections: .......................................................................................... 12 3.4 Communication .......................................................................................................... 13
4. Agitating Platform .............................................................................................................. 14 4.1 Manual Control ........................................................................................................... 14 4.2 Mechanical adjustment circular movement: ................................................................ 15 4.3 Taking the tray out...................................................................................................... 16 4.4 Bag positioning ........................................................................................................... 17 4.5 Connecting the hoses ................................................................................................. 20 4.6 Connecting the sensor cables .................................................................................... 21
5. Disposable Bioreactor-bag ................................................................................................ 22 5.1 Aeration ...................................................................................................................... 23 5.2 Foaming ..................................................................................................................... 24 5.3 Additions .................................................................................................................... 24
6 Monitoring and Control ...................................................................................................... 25 6.1 Temperature measurement and control ...................................................................... 25 6.2 Filter-heating .............................................................................................................. 26 6.3 pH Measurement and control ..................................................................................... 27 6.4 DO measurement and control .................................................................................... 28 6.5 Agitation control ......................................................................................................... 30 6.6 Feed control ............................................................................................................... 32
7. Installation ......................................................................................................................... 34 7.1 Requirements ............................................................................................................. 34 7.2 Base pump ................................................................................................................. 35 7.3 Gas regulation ............................................................................................................ 37 7.4 Dose monitoring ......................................................................................................... 39 7.5 Web interface ............................................................................................................. 40 7.5.1 LAN connection to PC or notebook: ........................................................................ 40 7.5.2 Connection to DACS (Data Acquisition Software) optional! ..................................... 43 7.5.3 Download files from PLC to PC ............................................................................... 44 7.5.3 Load data into Excel ............................................................................................... 46
8. Human Machine Interface (HMI) ....................................................................................... 47 8.1 Main Menu ................................................................................................................. 48 8.2 Process View, logging and alarm ............................................................................... 49 8.3 Process Parameters ................................................................................................... 51 8.4 Advanced Settings: PID menu, alarm levels, dead zone ............................................ 53 8.4.1 PID settings ............................................................................................................ 53 8.4.2 Alarm levels ............................................................................................................ 54
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8.4.3 Dead zone .............................................................................................................. 54 8.4.4 Constant output (bias): ............................................................................................ 55 8.5 System configuration .................................................................................................. 56 8.5.1 Set Date and Time .................................................................................................. 56 8.5.2 Calibration analog inputs (pH and DO) and temperature ......................................... 56 8.5.3 Set up Operator password ...................................................................................... 58 8.5.4 Diagnostic screen ................................................................................................... 59
9. PID control ........................................................................................................................ 60 9.1 control settings ........................................................................................................... 60 9.2 Output control ............................................................................................................. 62
10. Technical specifications .................................................................................................... 63 10.1 Specifications CM2SCEU/US CELL-tainer cell-culture application ............................. 63
11. Toubleshooting ................................................................................................................. 65
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Introduction
The CELL-tainer® single-use bioreactor (fig. 1) provides a flexible, single-use method to
cultivate mammalian cells or micro-organisms. Using a patented motion pattern and a
disposable, multi-compartment bioreactor bag with sensors, cells and micro-organisms can
be cultivated under high mass transfer rates and easy adjustable process conditions.
Temperature, pH, Dissolved Oxygen (DO) of the culture can be monitored and controlled.
The patented, sterile bioreactor bag is mounted on the platform of the CELL-tainer® and
consists of the following parts:
Sterile, plastic, 20L bioreactor-bag In-line sensors for pH and DO measurement,
conveniently mounted in the bottom of the bioreactor bag.
Sample line and drain C-Flex line 3/8” (one at both ends of the bag) equipped with
Luer connectors.
Air inlet and air outlet line, including filters, mounted on the top of the bag.
Three (3) medium C-Flex inlet lines 1/4” with Luer connectors, mounted on the top of
the bioreactor bag.
Expansion Channel
The Expansion Channel was
developed in order to reduce the
minimal volume to 150ml in an easy
and reliable way. Now, microbial
cultures of 150ml – 10L can be
grown in the same bag and cell
cultures can be grown in volumes of
150ml – 15L in the same bag.
Note: the temperature of the medium inside the bag is measured with a digital temperature
sensor positioned underneath the bag (a non-invasive contact sensor).
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The CELL-tainer® incorporates a closed PID control loop that controls the process variables
(pH, temperature and DO). The measured variables are compared to the (adjustable) set
point value and the controller will appropriately adjust the output to keep process variables at
the set point. The controller output signals that drive the actuators are on/off signals and
used to control pumps, (solenoid) valves, cooling valve and heating vents.
Mixing of the culture is facilitated by a patented two-dimensional motion of the platform, using
a servo motor (electrical). The horizontal movement is minimum 6 cm and maximum 20 cm:
the radius can be changed by 7 steps of 10mm. The range for the vertical movement goes
from 5° up to 17°. Steps of 1.7° can be manually adjusted. The speed (rpm = rocks per
minute) can be adjusted between 5 – 30 rpm.
Gentle movement of the platform will be required with certain cell cultures whereas higher
rpm settings can be useful with high oxygen demanding cultures (e.g. micro-organisms). The
unique two-dimensional movement of the platform yields a high mass transfer rate compared
to conventional disposable bioreactors.
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The HMI (Human-Machine Interface) is a colour touch screen controller. It is used to
effectively communicate with the CELL-tainer®. The controller of the CELL-tainer® also
utilizes a http server and an external Ethernet connection to remotely monitor and control the
CELL-tainer® using a PC or notebook.
Screen shot of the web browser when connected to the CELL-tainer®
http://192.168.168.20:9090/celltain.htm
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General overview of the complete CELL-tainer® system
Bioreactor-Tray emergency stop HMI
Mixing platform peristaltic pumps pressure gauge Rotameter
Easy access for positioning the Bioreactor-bag
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Schematic overview of the CELL-tainer® system – cell
culture
The ‘Operations Manual’ describes the operating procedure of the complete system (the
CELL-tainer®, the disposable bioreactor bag and the HMI). The first 6 chapters will be of
importance when operating the CELL-tainer® for the first time, the following chapters contain
more detailed descriptions of the controls of the CELL-tainer® and for installation and
configuration.
Optional:
Extra for cooling: cooling-plates (heat exchanger) underneath the bioreactor bag, which can
be connected to the chilled water supply or a chiller-system.
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1. System survey
Cell culture set-up:
cooling plate underneath the tray (optional)
cooling water in
cooling water out
Gas outlet
AIR O2 CO2 gas entries 3x
Peristaltic pump: 1x for alkaline
Standard items:
1x peristaltic pump: for alkaline
3x entries for gasses: AIR = 1 nL/min and O2 = 1 nL/min, CO2 = 1 nL/min
Optional: feed-pump
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2. Front Panel
2.1 Gas mix system
The gas mix system is equipped with pressure regulators/gauges, flow meters (manual
adjusted rotameters), non-return valves and solenoid valves, all with (Festo) quick-
connectors. The maximum input pressure is 3 barg, the minimum input pressure is 1 barg.
The reducers of the gas mix system are positioned inside the equipment, and can only be
reached when the panel is opened. The first reducer (at the left side of the panel, seen from
above) is meant for AIR , the second for O2 and the third for CO2
The maximum (adjusted) pressure from the reducers can be read on the pressure gauges
located on the front panel, above the flow meters.
The gas-flows (volume) can be adjusted by turning the needle valve on top of the rotameter.
Overview of the front panel of the CELL-tainer® system
alarm light HMI
ON light E-stop active (red)
Emergency stop
Peristaltic Pressure gauges
Rotameters
pump
Mains switch
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2.2 Mains switch
Never switch off the MAINS, when the motor is running!
The mains switches the complete system on/off. The mains switch
is provided with an automatic fuse.
Switching the system off will not discard the logging data or
process parameters of the current process.
2.3 Safety system/Emergency stop
The CELL-tainer® is equipped with an emergency stop (red
button), mounted on the front panel. When pressed, the
currently active process will be interrupted (all controller outputs
and the movement of the agitating platform stop). To disengage
the emergency stop simply pull out the button.
Furthermore, when opening of the door of the CELL-tainer® this
is detected as an event (DOOR OPEN), the movement of the agitating platform stops: safety.
During a process, the controller stores all these events and alarms in a CSV file, which can
be obtained through the Ethernet connection (as described in chapter 7.6).
2.4 Touch screen
A colour touch screen is used as Human Machine Interface (HMI).
Don’t use any sharp objects to press buttons on the touch screen.
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3. Side panel
Overview of the side panel of the CELL-tainer® system
3.1 Gas connections
The quick connectors (SMC), suitable for tubing with outer diameter of 6 mm, are located on
the right side of the system. The gas inlets are labelled. To detach the connected tubing,
press the white ring and pull the tube.
Label the gas tubes or use coloured tubing for easier reference and to avoid mix-ups.
Inlet pressure: 1- 3 barg!
3.2 Gas outlet connection
The quick connector, suitable for tubing with outer diameter of 8 mm, is located on the most
right side of the gas connections.
3.3 Chilled water connections:
Water in / water outlet: for cooling or chilled water
Connections: 6 mm quick-connectors with non-return
valve: Inlet pressure: 1- 2 barg!
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3.4 Communication
The controller is equipped with a http server. This interface makes it also possible to
remotely monitor and control the CELL-tainer® by connecting the Ethernet connection to a
Local Area Network or directly to a PC.
All process data that is requested from the
CELL-tainer® can be loaded into Excel.
To establish the communication between the
CELL-tainer® and a PC see chapter 7.5.
The second Ethernet connection can be used to communicate with a computer-platform or
other devices like Trace Analytic glucose/lactate analyser.
Web interface: http://192.168.168.20:9090/celltain.htm
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4. Agitating Platform
Be careful: after switching on the CELLtainer® the motor can automatically
start running!
4.1 Manual Control
For safety reasons it is not possible to turn the agitation on, when the door is opened. It is
possible to manually jog the platform by continuously pressing the ‘Jog’ button: only when
the door is closed.
Wait 10 seconds until the motor stops, before putting your hands in the machine!
In the ‘Manual mode’ view, the air valve and feed pump can be switched on/off. During the
open door state it is possible to activate the process-, logging- or alarm view. In this case the
upper right button in these views becomes an active blue button: Manual Control.
Activate key: Sample -> right side down, Drain -> left side down or Middle position -> tray
horizontal at the left side, ready to take the tray out.
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Picture of the agitating platform in the middle position
4.2 Mechanical adjustment circular movement:
Never switch off the MAINS, when the motor is running!
Adjust the angle of the tray with a hexagon key (4mm): release the two bolts, move the tray
to the right to make the circle smaller. There is a spring-bullet between the two bolts to
position the next step: every step is ~1.7°. Tighten the two bolts correctly in the new chosen
position.
Standard adjustment: 8.5° (sixth + eight pin-position from the right).
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4.3 Taking the tray out
After positioning the tray (with closed door)
into the Middle position: pull out the fixation
pin on the right side of the platform and
move the tray out
The fixation pin must be locked in the
hole at the back of the platform to prevent
the tray from moving (sliding backwards)
when the tray is taken out.
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4.4 Bag positioning
The removable tray can be used to transport the disposable from and to the CELL-tainer®.
The bag will be placed in a removable tray:
Never inflate the bag (fully)
before securing the tray to
the platform.
See also manual cell-culture bag: OPS- CB00010220-V1.x final cell
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Next: mount the 4 bars above the bag, for extra fixation of the bioreactor-bag:
Push the bars (4x) into the right
side of the tray.
Tighten the bolts (4x) on the
left side.
To secure the tray to the platform: 4 bolts are used, one at every corner of the tray:
Right side corner (back): short!
Front right side
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When the tray is back in position, check if the
locking pin is secured, to prevent the tray from sliding,
before closing the door and start the agitation!
Now the bag can be inflated.
Locked: = safe!
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4.5 Connecting the hoses
To prevent the hoses (C-flex) from wearing out, it is important to fix the hoses properly in the
slider-rail:
Remove the support bar and click
the hose in the rail.
Close the support bar.
Move the slider to the middle position and fix the
hoses as described above, with a little tension
between the filter-heater and the slider-rail.
connect the tubing to the
correct inlet, outlet or the
pump
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4.6 Connecting the sensor cables
Left side is for the bioreactor Temperature cable, the middle position is for the DO sensor
cable and the right side is for the pH cable.
Be careful: elements can be hot!
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5. Disposable Bioreactor-bag
Drawing of the single-use 20L bioreactor bag
Aeration filters (connection: outlet filter left; inlet filter: right)
Drain tube Inlet tubes
DO sensor and pH sensor (bottom)
Sample tube
Tube clamp
When installing the disposable it is of importance that the two outside bottom plugs fit
into the holes of the tray. Label at the back side.
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5.1 Aeration
The filter located in the centre of the disposable is to be used as exhaust filter. This position
is chosen to maximize the distance to the liquid surface: the risk of wetting the filter is
minimized by installing a liquid trap in the bioreactor-bag.
The exhaust-filter will be mounted in a
filter heater:
Both gas inlet and gas outlet filters are identical in specs. The filters mounted on the bag are
suited for a gas flow of 1-20nL/min. The (maximum) working pressure of the bags as
delivered by CelltainerBiotech is 0.2 barg, when using the filters as supplied by
CelltainerBiotech (Millipore vent SLFG05000 filters) the maximum flow rate is limited to ~2
nL/min.
To make it easy for the operator, both in and outlet hoses can be fixed permanently in the
‘swingarm’. These hoses must be replaced as least once a year or when damage or wear out
is visible. Saves time connecting the bioreactor bag in the CELL-tainer.
Dry gas is going in, saturated (with H2O) gas is going out. Evaporation occurs.
Evaporation/day (24 hrs) cellculture
gasflow evaporation
l/min ml/day 0,0 0,0 0,1 6,35 0,2 12,7 0,5 31,7 ml/day
1,0 63,4
gasflow in l/min
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5.2 Foaming
In order to prevent foaming, the movement profile can be changed by adjusting (decreasing)
the angle and motor speed.
5.3 Additions
3 x addition hoses are available: C-flex 1/8” x 1/4”, luer lock 1/8”
For example:
1x Inoculum
1x Alkaline addition
1x Feed
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6 Monitoring and Control
The following parameters are measured and controlled:
Temperature bioreactor-bag (T1)
Temperature Incubator (T2) -> optional control! Instead of control on T1
Temperature Filter heating (T3)
pH
DO (Dissolved Oxygen)
RPM
Temperature, pH and DO each are controlled by a PID control loop. Each individual regulator
can be switched ‘on’ and ‘off’. This allows for independent control of each of the regulators.
6.1 Temperature measurement and control
The system is equipped with three temperature sensors. One of these sensors is located
inside the CELL-tainer® cabinet, and prevents the system from overheating. The second
sensor is located on the moving platform and is in contact with the bioreactor bag. The third
sensor is for the Filter-heating.
The sensor output from the bioreactor-bag is the input of the controller that controls the
actual temperature of the media inside the disposable.
The other input of the controller is the set point (set in Parameter Menu from the HMI). The
difference between the two input values determines the error value. The PID control loop
uses this value to calculate the appropriate output to reach set point conditions. In this
process, the proportional, integral and differential constants are set from the HMI and play
part in the response characteristics of the output controller.
The output will control the heating device controlled by the duty cycle of a slow PWM (pulse
width modulated) signal to control the heating elements. The output also can be negative,
then natural cooling will cool down the system.
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For extra cooling, optional the system can be equipped with cooling-plates (can be
connected to a cooler or chilled water system) underneath the bioreactor-bag, thus providing
a higher cooling capacity.
Process Parameters menu: Temperature control
6.2 Filter-heating
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6.3 pH Measurement and control
To measure and control the pH level inside the bioreactor bag, one electro-chemical sensor
(fixed mounted in the bottom of the bag) is used. The pH sensor output from the bioreactor-
bag is the input of the controller that controls the actual pH of the media inside the
disposable.
The other input of the controller is the set point (set in Parameter Menu from the HMI). The
input of the controller uses the error value (set point minus measured value) to control the pH
value. The output will activate the CO2 valve or the base pump. If the output of the PID is
negative, the CO2 valve is activated. If the output is positive the base pump is activated. The
analog value from the PID output is proportional to a duty cycle of a PWM signal controlling
the base or acid pump.
Process Parameters menu; pH control
Dead-zone:
To prevent the pH-control from over-and undershooting the set-point most of the time a dead
zone is used instead of the Integral time: this means define a zone around the set-point
where the control is off. This gives the alkaline time to mix and stabilize the pH.
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6.4 DO measurement and control
To measure the Dissolved Oxygen (DO) level inside the bioreactor bag, a Polarographic
sensor (fixed mounted in the bottom of the bag) is used . The DO sensor output from the
bioreactor-bag is the input of the controller that controls the actual DO% of the media inside
the disposable.
The other input of the controller is the set point (set in Parameter Menu from the HMI).The
input of the PID controller uses the error value to control the DO value. The output will
activate the O2 valve.
The dead-zone for the positive PID zone is set to 200%. Output of PID is not active. If N2
valve is available and the output of the PID is positive the solenoid valve for the N2 addition is
activated. If the output is negative the valve for the addition of O2 is activated.
Process Parameters menu; DO control
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Standard DO-control:
- Controller Output range: +100% to -100%
- From 0% to 100% with O2
- (0 to -100% with N2: only microbial)
Advanced settings Cascade rpm DO-control
disabled
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6.5 Agitation control
The movement of the bioreactor bag is performed using the parameters:
speed (rpm) : adjustable 5 -30 rpm or in DO-ctrl cascade
horizontal displacement : adjustable (mechanically) 60 – 200mm (radius: 7x 10mm)
and is equal to
vertical displacement : adjustable 5- 17° (mechanically by 7 steps of 1.7°)
Settings are introduced from the HMI.
The diameter of the circle determines the movement profile of the tray. The rotation is
clockwise. This movement is described in our patent application. The mass transfer data as
published are also based on this movement.
Process Parameters menu; Motor control
Default settings movement profile for start cell culture:
Speed : 5 rpm
Circle : ~8.5°
air flow : 0.01 vvm)
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NB.:
When using only the middle compartment of the Bioreactor-bag e.g. 350-500ml:
Do not use maximum values for mixing, because of medium flow into the in- and outlet filter
tubing’s at the top of the bag -> causing filter blockage!
Prevent medium flow into the outlet filter tubing’s at the top of the bag -> causing filter
blockage!
Volume at 0.2 VVM aeration: 10L -> max. 30 rpm
15L -> max. 30 rpm
20L -> max. 15 rpm
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6.6 Feed control
Manual control menu
Parameter menu -> Advanced Settings
Pump:
Auto feed: means a fixed speed for the pump -> time pump is on/off = constant. The flow
must be calculated/calibrated: speed/hose diameter (ml/time unit).
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Constant feed (0.1 seconds accuracy):
Value = 0 = off
Value = 0.1 = 1 sec on every 1000 seconds
Value = 0.2 = 1 sec on every 500 seconds
Value =100 = on all the time
Value =20.4 = 20.4 seconds of 100 seconds on
Value =80 = 80 seconds on, 20 seconds off
Feed DO control:
The maximum flow (feed) must be calculated/calibrated: pump-speed/hose diameter (ml/time
unit).
When this option is activated, the feed is maximum when the actual value is higher than the
set point. As soon as the Actual value comes below the set point, the flow will be decreased
and will even stop when the DO reaches 0%.
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This part of the manual describes the installation of the CELL-tainer® system.
7. Installation
The CELL-tainer® will be delivered on a Frame/trolley (height = 650mm), with 4 (2 lockable)
wheels. After setting the system in place the frame can be set on the adjustable feet and be
levelled.
The total mass of the equipment is about 170 kg, without disposable.
7.1 Requirements
Electrical:
Mains: 230/115 Vac, 50/60 Hz. Maximum used power ≤ 1000W. Check whether the power
supply is in accordance with the indicated voltage of the machine. Check fuse before
operation: 8A for 230Vac; 15A for 115Vac.
Check if the cable cord fits properly.
Gas:
Maximum 3 Barg supply of Air/N2/O2 gas (depending on what is desired for the specific
culture). Check the connection of the correct gases to the machine. Minimum pressure of the
gas supply should be 1 Barg.
Warning: the gas pressure connected to the machine may not exceed the indicated
pressure. A too low pressure will lead to non proper functioning of the reducer valves.
Gas mix
Depending on the application the system takes a maximum of three different gas supplies.
The gasses are used to control the DO and pH value in the culture. The supplies have to be
connected to the three connectors, mounted on the side panel of the CELL-tainer®, using the
quick connectors (suitable for tubing with outer diameter of 6 mm.).
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7.2 Base pump
The CELL-tainer® is equipped with one Watson Marlow peristaltic pump. To connect the
pump, Norprene or silicone tubing can be used with an wall-thickness of 1.6 mm.
To control the pH, alkaline can be added to the medium with the pump. The pump head is
located behind the door on the front panel. Before connecting the pump to the bioreactor bag
make sure that every part is sterilized and that the connections are made under sterile
conditions. For more information see: manual cell-culture bag OPS- CB00010220-V1.x final cell
Connect the base pump using e.g. Masterflex Norprene tubing (with an inside diameter of 1.6
mm) that ends in a pump flask containing base solution (sterile), it is advised to use an
alkaline not exceeding 0,5 N. To do so, lift the top of the pump to open it and lead the tube
through the pump (from right to left) and close the top. Use the two adjustable clamps on the
side of the pump to tighten the connection. Attach the tube to one of the addition lines on the
bioreactor bag.
Installing the tubing in the pump
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Using tubing with other dimensions is possible but (maybe) not compatible with the
operation procedure as described in this manual.
Rotation: counter clockwise
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7.3 Gas regulation
Adjust the internal gas pressure within the CELL-tainer® by pulling and turning the pressure
regulators located inside the CELL-tainer®, on the right side behind the side-panel.
AIR pressure regulator:
Adjustment knob: pressure up turn clockwise
Lock: release -> turn counter clockwise
Pressure regulators for CO2 and O2:
Adjustment knob and lock.
Pull out adjustment knob to release.
Pressure up turn clockwise
Push back to lock
Factory set at 0.15 barg at 3 barg gas supply pressure.
From left to right, the regulators correspond to Air, CO2, and O2.
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The individual valves can be switched on/off through the ´Diagnostic Menu´ : (Advanced
View Menu System Configuration Diagnostic Menu). When the desired pressure is
achieved, slowly turn the regulator to both sides with a light pushing motion until it slips back
into position .
On the flow meters located at the right-side of the front panel, the flow can be monitored and
controlled by turning the knob at the top of the flow meter.
Take care the pressure does not exceed 0.2 Barg when a valve is opened, the
internal gas pressure within the bioreactor bag will build up with the risk of blowing it up, for
example when the exhaust filter is blocked.
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7.4 Dose monitoring
Dose monitoring can be accessed by pressing Advanced View Show Total sec/ml/percent
in the Process View screen. This will display information for a specific valve regarding the
time it has opened and the volume of gas/liquid that has passed.
Calibrated volumes (measured with pump and the pump hose to be used) can be entered in
screen: volume factors ml/s
Volume will be displayed in 3 digits e.g.: 99.9ml or 999ml
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7.5 Web interface
To remotely control and monitor the CELL-tainer® with a PC an unique IP address has to be
configured for each CELL-tainer®. Go to ‘System Configuration’ in the Main Menu and press
‘IP Address’ to enter the specific IP address you want to use for the CELL-tainer®
(192.168.168.xx).
7.5.1 LAN connection to PC or notebook:
- Open Network connections
- Choose: Local area connection or LAN connection (depending on OS)
- Choose: Internet protocol (TCP/IP) or Internet protocol version 4 (TCP/IPv4)
- Choose: properties
- Choose: Use the following IP address: 192.168.168.1
- Subnetmask will be filled in automatically after entering the IP address
255.255.255.0
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LAN connection PC
Internet protocol version 4 (TCP/IPv4)
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Using the configured IP address 192.168.168.xx with port 9090 (in web browser:
http://192.168.168.xx:9090) the CELL-tainer® can be accessed with the computer. As
depicted, the interface that opens in the web browser allows for control over the CELL-
tainer®. In addition, updated firmware can be uploaded to the CELL-tainer® and installed. An
Ethernet crossover cable (not included) has to be used for the connection between the
CELL-tainer® and computer.
Screen shot of the web browser when connected to the CELL-tainer®
http://192.168.168.20:9090/celltain.htm
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7.5.2 Connection to DACS (Data Acquisition Software) optional!
LAN settings are the same as described above.
Webbrowser:
http://localhost/HMI/home/index.php?p=pview
The system is developed to monitor product up to 8 CELL-tainer systems as well as
producing records to support product release. For pharmaceutical applications the system is
classified as GMP critical.
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7.5.3 Download files from PLC to PC
config.csv
alarm.csv
log.csv
Right click on one of the files and save as <name>.csv file:
comma separated value.
config.csv example:
Cell-Tainer Configuration
System Date 9-2-2012
System Time 9:32:50
Cell-Tainer ID 00.05.51.00.EB.CC
Process Time (sec) 1328779970
Process Inputs (hex) 00000F1F
Process Outputs (hex) 106
Process Flags (hex) 00000C60
Process Alarms (hex) 0
Process paraName TTT
Process start Date 8-2-2012
Process start Time 15:39:12
Ana Temp Measure 348
Ana Temp Setpoint 350
Ana Temp %output 5 etc
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alarm.csv example:
contains events and alarms:
Date Time Type Event MsgNr Message
8-2-2012 15:46:25 0 Event 16 MOTOR OFF
8-2-2012 15:46:22 0 Event 9 DOOR CLOSED
8-2-2012 15:46:15 0 Event 8 DOOR OPENED
8-2-2012 15:40:30 0 Event 9 DOOR CLOSED
8-2-2012 15:40:15 0 Event 8 DOOR OPENED
8-2-2012 15:40:00 0 Event 9 DOOR CLOSED
8-2-2012 15:39:59 0 Event 8 DOOR OPENED
8-2-2012 15:39:43 0 Event 9 DOOR CLOSED
8-2-2012 15:39:39 0 Event 8 DOOR OPENED
8-2-2012 15:39:14 0 Event 11 TEMPERATURE CONTROL ON
8-2-2012 15:39:13 0 Event 26 TROOM ACTIVE
8-2-2012 15:39:13 0 Event 21 MOTOR SPEED RPM 8.0
8-2-2012 15:39:13 0 Event 20 SETPOINT DO 5.0
8-2-2012 15:39:13 0 Event 19 SETPOINT pH 7.0
8-2-2012 15:39:13 0 Event 18 SETPOINT TEMPERATURE 35.0
8-2-2012 15:39:13 0 Event 4 NEW PROCESS STARTED
log.csv example:
sec Temp pH DO rpm Glucose Lactate TRoom
60 218 76 50 80 0 0 234
120 218 76 50 80 0 0 236
180 220 76 50 80 0 0 238
240 220 76 50 80 0 0 244
300 220 76 50 80 0 0 248
360 220 75 50 80 0 0 254
420 220 75 50 80 0 0 260
480 222 75 50 0 0 0 264
540 222 75 50 0 0 0 264
600 222 76 50 0 0 0 262
Except for the time all parameter values have to be divided by 10 to get the real
value: PLC does not recognize the decimal dot.
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7.5.3 Load data into Excel
Import to Microsoft Excel
To import this back into Microsoft Excel 2007, click on Data, then click the From Text button
in the Get External Data menu. This will bring up the Text Import Wizard.
In the first step, make sure to have Delimited set as the first option. Select which row to start
with. In this example, It starts with the first row.
In the second step, make sure to check the box next to Comma. (See screenshot for an
example.) This will then show the data in columns at the bottom. If clicked the check next to
Space, it will divide the address column into two lines, in this example. Sometimes this can
be helpful when trying to break down data during the import, as some programs tend to lump
data together.
The third step simply chooses the format of the data, such as numbers, text, or dates. After
done that, just click Finish and the data will be imported into the spreadsheet. Depending on
how big the .CSV file is, the import process could be instantaneous or take several minutes.
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8. Human Machine Interface (HMI)
Touch-screen with built-in processor, connected
with control-unit (PLC).
With exception of all physical controllable parts of the CELL-tainer®, such as the pressure
regulators, flow meters, the hood and emergency switch, everything else is controlled
through a Human-Machine Interface (HMI). A colour touch screen enables the operator to
control every functionality of the CELL-tainer®. In this chapter, the menu’s architecture, as
depicted below (fig. 11), will be explained.
Fig. 11: HMI architecture. Take note that certain menu features are left out in this diagram
because they are self-explanatory.
Main
menu
Process
view
Advanced
view
Logging
Alarms
Process
parameters
Advanced
settings
Dead
zone
PID settings
T/pH / DO
Cascade
Feed ctrl
System
configuration
Diagnostic
screen
Alarm
levels
Calibration
pH / DO
and temp
Filter
heating
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8.1 Main Menu
From the main menu the other views can be accessed:
- Process view
- Logging
- Alarms
- Process parameters
- Manual Control
- System configuration
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8.2 Process View, logging and alarm
Whenever a screen view is active, except for the ‘Process View’ and ‘Diagnostic menu’, the
system will automatically switch to the ‘Process View’ when no user action (screen-touch) is
detected during 5 minutes.
The ´Process View´ gives a symbolic overview of the CELL-tainer®. Open valves or activated
pumps are indicated by a green colour, while closed valves or pumps switched off, are
indicated by a yellow colour. Devices which are disabled, (turned off by the controller) are
indicated by a grey colour. At the bottom of the screen the current date and time and the
process time is shown.
When an alarm occurs, the top right button starts to flash red. Pressing this button will bring
you to the ‘Alarms’ view and confirms the alarm which stops the flashing. The device in
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alarm, in this case the temperature, is coloured red. At the bottom right of the ‘Process View’
the alarm-icon is shown. This will remain visible as long the alarm is still active regardless of
the confirmation. In case the process is not running the stop-icon is shown at the bottom right
of the screen.
Pressing the ‘Advanced view’ button will bring up the ‘Advanced View’ screen. In this view
the process data is represented in a table. Besides the actual measurements the ´Advanced
View´ also displays the set points and the output of the PID-regulated controllers. From the
‘Advanced View’ screen you can select the ‘Main Menu’, ‘Logging’, or ‘Alarms’ view or switch
back to the default ‘Process View’.
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The ‘Logging’ view gives a graphical view of the temperature, pH and DO parameters, based
on a time scale. The time-axis can be zoomed in or out and moved to the left (the past) or
right. The date and time shown in the info field displays calendar date of the zero-coordinate
of the time axis. Just below this date the process time of the zero-coordinate is shown. The
hh:mm on the time axis is displayed as process time.
The ‘Alarms’ view lists the date and time of alarms and events. These data are stored in a
(downloadable) log file. Alarms are displayed in red. The restore of an alarm will be marked
green and events, such as the opening of the hood or adjusting a set point, are displayed in
blue.
8.3 Process Parameters
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From the ‘Main Menu’ view, the operator can choose the ‘Process Parameters’ menu to
control all process parameters (listed below), start/end a process or save/load preset
parameters. To save a certain set of parameters, press ´Save Para.´ and give the preset a
name. To load a preset, press ´Load Para.´ and choose the preset from the list. Up to 60
programs can be saved.
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8.4 Advanced Settings: PID menu, alarm levels, dead zone
From the ´Process Parameters` menu, one can proceed to the ‘Advanced Settings’ menu (as
depicted below).
8.4.1 PID settings
In the ‘Advanced Settings’ menu the operator can choose to set the PID constants. In
addition, the operator can proceed to the ‘Alarm levels’ view to set minimum and maximum
parameter values that are allowed in the process. When parameters exceed the set alarm
levels an alarm will be documented in the ‘Alarms’ menu. In the ‘Dead zone’ menu, one can
enter values that represent the maximum allowed deviation from the given set point. Within
this zone PID control is turned off and will be turned on when the measured process value
falls outside the dead zone. PID control and default settings will be explained in Chapter 9.
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8.4.2 Alarm levels
Advanced settings Advanced settings: alarm levels
8.4.3 Dead zone
Advanced settings Advanced settings: dead zone
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8.4.4 Constant output (bias):
The PID output will be minimal the Constant Output (CO), if the PID output has the same
sign as the Constant Output.
This is even true if the PID control is switched OFF or the process is Paused or Ended.
Temperature control: with CO set to 10%
If the PID output would be 0% or more (heating is needed) then the heater will be set to 10%.
If the PID output would be 25% (more than CO) then the heater is set to 25%.
If the PID output would be -2% (negative means cooling is needed) then CO has no influence. Cooling
will be set to 2% and heater is off.
If the Temperature PID control is switched off (or process is ended, paused) the heater is set to 10%
regardless of the current temperature.
pH control: with CO set to -5%
If the PID output would be negative or zero (or switched off) then ACID valve will be on for at least 5%.
If the PID output is positive then the BASE pump will be activated (no ACID is added).
DO control: with CO set to 15%
If the PID output would be positive or zero (or switched off) then the O2 output would be set to 15%.
If the PID output is negative then the O2 valve is closed and N2 will be added.
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8.5 System configuration
The `System Configuration´ menu can be accessed through the ´Main Menu´. The menu can
be used to calibrate the temperature sensors, to set up general system configurations and to
enter the ´Manual Output Testing´ menu
8.5.1 Set Date and Time
It is not possible to change Date and Time when a process is running, to prevent a ‘gap’ in
the data-acquisition. When a process is ended, the input boxes (keys) for Date and time turn
blue and are accessible to correct Date and Time. When a process is started the input boxes
turn grey: no changes allowed.
8.5.2 Calibration analog inputs (pH and DO) and temperature
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8.5.3 Set up Operator password
In the `System Configuration´ menu, one can choose to set up an OPERATOR password
and/or an ADMINISTRATOR password. In this respect, an OPERATOR is able to change the
process parameters in every detail. Only the ADMINISTRATOR is allowed to change system
configuration settings.
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8.5.4 Diagnostic screen
When choosing the ‘Diagnostic screen’ menu in the ‘ System Configuration’ menu, the
operator will be able to manually control all PID-regulated outputs of the system. By pressing
the buttons the respective control will be turned on or off. When leaving this menu, all
controls will be automatically set back to its original position, in order to maintain the set
points.
Additional information is given for service purposes, all switches can be detected on their
position.
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9. PID control
9.1 control settings
The pH, temperature and DO control loops of the CELL-tainer® are controlled by preset PID-
parameters (table 1). However, depending on the conditions of the culture, the PID
parameters can be adjusted manually. For example, when the pH is allowed to vary within a
range of 1,0 pH unit and you want to limit the addition of alkali or acidic substances to keep
the osmolarity of the medium constant, you can choose to keep the PID constants low to
slow down the response of the output controller. This will be more likely of use in the
beginning of a culture (small volume), when the pH control will add a relative large volume
alkali or acid to compensate for the pH.
When a surplus of added alkali or acid is extremely harmful to the viability of the culture, it is
to be advised to use very low PID settings (for example, P = 5, I = 0,08) in order to reduce
NaOH or Acid output to 0-5% (visible in ‘Advanced View’ menu). When set point is reached
make sure the output is set to 0% to avoid unnecessary addition of alkali or acid.
If the volume of the culture expands and the amount of cells increase,
acidification/alkalinisation will be more likely to occur. In this case, increasing PID settings
will be necessary to keep the pH around set point.
Temperature pH DO
P 20 50 5
I 0,25 0 0,3
D 0 0 0
Deadzone
negative - 0.1 -
Deadzone
positive - 0.1 -
Table 1. Preset PID values.
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The P-value (proportional constant) is used to calculate to which extent the controller needs
to be activated to minimize the error. The error is the value of set point minus actual
(measured) value. When a set point is entered and the response of the output is too slow,
the P-value can be increased.
The I-value (integral constant) is used to calculate how fast the controller needs to change its
output when the error state does not resolve over the course of time. When the set point will
not be reached by a small amount, the I-value can be increased to reach the set point.
Both the P-and I-value can be increased at any time to higher the controller’s output, but it
has to be kept in mind that increasing these values will have its side effects in reaching the
desired set point (as pointed out in the table below).
The D-value (derivative constant) is used to reduce overshoot, when the measured value is
close to reach set point. However, this constant is only effective in fast-paced processes,
unlike cell cultures which show slow process activity.
Table 2 sums up the eventual effects of increasing each of the PID parameters.
Effects of increasing parameters
Parameter Rise Time Overshoot Settling
Time
S.S.
Error
Kp Decrease Increase Small
Change Decrease
Ki Decrease Increase Increase Eliminate
Kd Small
Change Decrease Decrease None
Table 2. Effects of increasing the parameters of the PID control.
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9.2 Output control
The total PID regulated output will act in cycles of 10 seconds. Within this cycle time, each
output component is turned on or off. The maximum output for the gasses for each output
component is 1/2 cycle time (5s). The remainder of the 10 seconds will be used for opening of
the air valve.
Example:
If the PID output for DO is set to +20% and the PID output for pH
would be -40% than 20% of 5s = 1s for activation of the O2 valve.
This is followed by 40% of 5s = 2s for activation of the CO2 valve
and this is followed by 7s for the AIR valve opening
Total 10s Cycle Time
In case the PID output would result in a 0.33 sec/cycle time, these time increments are
added until a whole second is reached. In this case the valve is activated for 1 second every
3 cycle times.
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10. Technical specifications
10.1 Specifications CM2SCEU/US CELL-tainer cell-culture application
Parameter Specifications
Dimensions unit W x H x D = 1400 x 560 x 810 mm (approx.)
Dimensions platform W x H x D = 510 x 660 x 100 mm (approx.)
Removable tray (bag holder) to be positioned using the 4
pillars on the platform
Movement profile Circular movement: horizontal displacement combined with
vertical displacement.
According EP 1739165A1
Horizontal: 60mm - 200 mm (adjustable: 7x 10mm=steps
of 1.7°)
Angle: ~5 -17 degrees (adjustable: see horizontal
movement)
Rotational speed 5- 30 rpm (cycles)
Drift Negligible, at least < 2 rpm/wk
Temperature control Directly under the bag: 1-wire Parasite-power digital
thermometer, 25 – 40 oC (+/- 0,2oC)
In the cabinet: 1-wire Parasite-power digital thermometer,
20 – 40 oC (+/- 0,3oC)
Heating capacity 2 x heating ventilator (230 W each)
DO measurement Polarographic DO-sensor
5 – 250 % air saturation (+/- 5% air saturation)
Air flow control by flowmeter 1 ln/min
DO control (PID) Flowmeter / solenoid valve combination:
0 – 1 ln/min O2
pH measurement Electro-chemical pH-sensor
pH = 2 – 10 (+/- 0,1 unit)
pH control (PID) Alkaline: peristaltic pump Watson Marlow 50 rpm, 24VDC
Flow: ~16 ml/min (using tubing 1.6x1.6mm)
Acid: Flowmeter / solenoid valve combination:
CO2 Flow: 1 ln/min
Air supply 3 barg
Max. operating pressure 0,2 bar
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Parameter Specifications
Process monitoring
and control
Single Board Computer & Display unit (SBC)
CPU ARM9 300MHz
32MB Flash / 32MB RAM
3 x RS232 Serial interfaces
1 x Ethernet UTP interface
1x USB
1 x SD-card slot Optional CAN 2.0 interface (DeviceNet) Embedded Linux OS
Human Interface HMI: 5.7 inch color CSTN graphical LCD
320x 240 pixels
4-wire touchscreen I/O Module MCU
MCU Microchip PIC24FJ256GB106 (32MHz)
256KB Flash / 16KB RAM
1 x RS232 Serial interface
4 x 1-Wire interface for DS18S20
temperature sensor
12 x Optical isolated digital inputs
18 x Relais outputs
4 x Analog inputs
C-programmable controller
Data storage and
operation
Measuring rpm, T, DO, pH up to 1/s
(adjustable sampling time)
Data storage / batch
Alarm logs: door open, change set-points, not reaching
set-point
Dose monitoring: Show Total sec/ml or percent in the Process
View screen
Manual operation of all outputs
Manual position of tray (jog, sample, drain, middle)
Data export Ethernet UTP interface (csv.file) to be approached from
web browser (Internet Explorer)
Safety Emergency stop.
Door opening / safety switch (stop) on all functions
Protection
classification
CE/IP54
Mains 230 Vac / 50 Hz – 115 Vac / 60 Hz
Cable cord To be specified / ordered
Manuals Operation & Installation / Service
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11. Toubleshooting
Be careful: after switching on the CELLtainer® the motor can automatically start
running!
1. If the PLC does not accept commands from the touch screen: restart the
system by switching off the mains power and switch on again
2. A. If the movement stopped: press and release the Emergency stop button for
resetting the motor controller.
B. Check if there is a process running: otherwise actuators are not active.
C. Emergency stop is active: release.
D. Door is not closed: safety switches are active.
3. If a process is stopped (end process) the movement profile cannot be
activated: start a new data logging; start process.
4. Delete saved parameters: use the Web Interface:
http://192.168.168.20:9090/cgi-bin/filelist.cgi
5. Reset Dose-monitoring: start new process.
6. Touch screen calibration: see Maintenance manual.
7. For Servicing the CELL-tainer® refer to the Maintenance manual.
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Celltainer Biotech BV will not be responsible for labor charges and/or damage
incurred in installation, repair or replacement, nor for any indirect, incidental or
consequential damages, losses, injury or costs of any nature relating to this
item. Except as provided by law.
The warranty, 1 year after delivery, covers replacement of all defective parts
and finishes. Is only valid when original parts are used. Damage due to
installation error, product abuse, product misuse, or use of cleaners containing
abrasives or other organic solvents, whether performed by a contractor,
service company, or yourself, are excluded from this warranty.
o © Copyright 2015 Celltainer BioTech BV, Winterswijk Brinkheurne, THE
NETHERLANDS and contains propriety and confidential information: Celltainer
BioTech BV does not allow reproduction, republishing, uploading, posting,
transmission or distribution in any way the contents of this presentation, for public or
commercial purposes, without written permission by Celltainer BioTech BV.
o Information presented in this document is not complete without an accompanying
verbal explanation and installation support and should not be considered open to
interpretation.
o No rights can be obtained from data as presented; neither Celltainer BioTech BV
assures the validity of these data.