CELL-tainer - Single use bioreactor 20L Cell culture · 2019. 10. 28. · Introduction The...

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


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


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


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).


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.


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


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


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.


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


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


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.

http://www.bath.ac.uk/internal/bio-sci/Images/haz general warning.gif


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!


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



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.




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).



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.



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



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


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!



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


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!



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.



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



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


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.


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


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.


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


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


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)


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


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).


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%.


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.).



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


Using tubing with other dimensions is possible but (maybe) not compatible with the

operation procedure as described in this manual.

Rotation: counter clockwise


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.


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.



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


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


LAN connection PC

Internet protocol version 4 (TCP/IPv4)


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.xx:9090/


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.

http://localhost/HMI/home/index.php?p=pview


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


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 15:39:14 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.


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.


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


8.1 Main Menu

From the main menu the other views can be accessed:

- Process view

- Logging

- Alarms

- Process parameters

- Manual Control

- System configuration


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


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’.


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


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.


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.


8.4.2 Alarm levels

Advanced settings Advanced settings: alarm levels

8.4.3 Dead zone

Advanced settings Advanced settings: dead zone


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.


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



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.


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.


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.


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.


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.


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


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


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.

http://192.168.168.20:9090/cgi-bin/filelist.cgi



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.

Date post:	27-Aug-2020
Category:	Documents
Upload:	others
View:	4 times
Download:	0 times

CELL-tainer - Single use bioreactor 20L Cell culture · 2019. 10. 28. · Introduction The...

Documents