BIOEXCEL A

Animal Cell Culture?

Cell culture has become one of the major tools used in the life science today. Animal cell culture was first successfully undertaken by Ross Harrison in 1907, since then, steadily enlarged its research territory through development in science. First, there was the development of antibiotics that made it easier to avoid many of contamination problems that plagued earlier cell culture attempts. Second was the development of the techniques, such as the use of trypsin to remove cells from culture vessels, necessary to obtain continuously growing cell lines (such as HeLa cells). Third, using these cell lines, scientists were able to develop standardized, chemically defined culture media that made it far easier to grow cells. These three areas combined to allow many more scientists to use cell, tissue and organ culture in their research.

BIOEXCEL A Bioreactor!

Animal cells are more difficult to culture than microorganisms because they require many more nutrients and typically grow only when attached to specially coated surfaces. Animal cells are more difficult to culture than microorganisms because they require many more nutrients and typically grow only when attached to specially coated surfaces. Despite these difficulties, various types of animal cells, including both undifferentiated and differentiated ones, can be cultured successfully. Cell culture is used for model systems, toxicity testing, cancer research, virology, cell-based manufacturing, genetic counseling, gene therapy and drug screening and development. Its business area is getting widened into pharmaceutical and bio-therapeutical industries.
BIOEXCEL A system is a lab scale bioreactor designed for multipurpose animal cell culture. It is an outstanding animal cell culture bioreactor system based on interactive control system. And the bioreactor system can offer various types of vessels such as Single round bottom vessel, Spin filter and Depth filter vessel, Double welled vessel, Small volume vessel, ranged from 50ml to 19L volume. As BIOTRON can re-design these various vessels according to customers’ demand, customers can obtain the best animal cell culture lab environment. Moreover, we can offer additional accessory items and external device, including micro carrier and disposable bag, thus constructing inclusive animal cell culture system.

Characteristic of Animal Cell

Unlike bacteria with cell wall, animal cell has only weak plasma membrane. Thus it is very sensitive to shear forces by bioreactor agitation or continuous stimulation of bubbles by supplied air/gas. Size of animal cells are approximately 10-100µm, relatively bigger than bacteria and dT(double time) is very slow, 18-28 hours. (Bacteria has 0.5-5 hours dT). Hence, animal cell requires very strict cell growth environment.

 The most important property of cell defining cell culture method is whether it is adherent cell or suspension cell. Adherent cell needs to be attached on the surface in order to grow, while suspension cell can do suspension cell culture. Normally, a cell maintaining adherence is much more stable condition, and when a cell changes to be suspension, it obtains cancer cell property.  This is because most cells except blood fluid or lymph are adherent cells. Most cells are adherent cell. However, one of most frequently used cell in bioreactor, Chinese Hamster Ovary (CHO) Cell, is processed through medium modification and adaption and then can be suspension cell cultured. Hybridoma can naturally suspension cell cultured.

For Scale-up, suspension cells are usually used in order to maintain maximum cell density volume per unit. CHO cell, normally used for medicine production, is adherent cell at first, but can be adapted to be suspension cell. They are easy to be scaled up and maintaining high density cell concentration, but may not supply nutrient and oxygen to cell inside cluster formed during cultivation. Hence, for animal cell culture scale up, it should not damage cell and must supply oxygen well. Oxygen has very low solubility in the medium. So supplying oxygen effectively to the animal cell inside culture medium is pivotal.(oxygenation)

Characteristic of animal cell is its sensitivity to shear stress due to large size and no cell wall, unlike microorganism. Inside bioreactor, hydrodynamics shear stress by agitation or aeration does damage cell. Mechanisms can be cell crash by each other or stirrer and baffle, pressure difference between impeller front and back, and eddy. Among them, main causes are formation of eddy and air bubble burst. When air bubble bursts, cell around it experiences huge shear stress. Therefore, in order to prevent cell damage, low aeration rate and gentle agitation are crucial for large volume bioreactor. Hollow fiber reactor has numerous fibers with bundle shape. The space and stoma between fiber and fiber bind cells in large space for growth. Medium penetrates through hollow separated by membranes continuously, thus supplying to the cell. Also, the resultant produced during cell growth, lactic acid, or ammonia passes through membrane and comes out to the medium with proliferation. At this time, cell concentration reaches 108 cells/ml and this can be operated more than one month. It can be used not only adherent cell, but also suspension cell such as hybridoma cell.

In case of suspension cell, suspension cell culture is commonly used for efficient production of monoclonal antibody with hybridoma cell, and adherent cell uses immobilization cultivation using micro-carrier or equivalent carrier for mass culture. For micro-carrier culture, dozens of micro-carrier are currently researched and used commercially. The shape of bioreactor is a stirred tank, and maximum cell density is 2-3 X 106 cells/ml by using 3-5% of micro-carrier. Especially, macro-porous micro-carrier can increase maximum cell density to 107 cells/ml. In order to increase productivity, medium recycling type perfusion continuous culture is commonly used. In this case, the technological development which can trap cell in the bioreactor, continuously supply only medium, and collect them and high medium supply speed are required.

BIOTRON applies spin filter for recycling type perfusion continuous culture. In most case, lab can also achieve obtain above 107cells/ml and 300-500µg/ml of antibody recovery. However, for pilot scale bioreactor, stability of bio-process, contamination possibility, and cell retention technology are really imperative.

Description Of BIOEXCEL A [Animal Cell Culture Bioreactor] Vessels

SPIN FILTER VESSEL

In order to increase productivity, medium recycling type perfusion continuous culture is commonly used. In this case, the technological development which can trap cell in the bioreactor, continuously supply only medium, and collect them and high medium supply speed are required. BIOTRON applies spin filter for recycling type perfusion continuous culture.

DEPTH FILTER VESSEL

Hollow fiber reactor has numerous fibers with bundle shape. The space and stoma between fiber and fiber bind cells in large space for growth. Medium penetrates through hollow separated by membranes continuously, thus supplying to the cell.

SINGLE ROUND VESSEL

• Single round bottom vessel is used for animal cell fermentation as low speed agitation
• Single vessel is relatively cheaper than double vessel
• Low risk of glass damage from shock..
• Temperature: single vessel does not require outside temperature control equipment.

DOUBEL VESSEL

• Heat transfer is stable due to larger heat transfer surface area.
• Useful for quick cooling and decreasing temperature.
• Water Bath is required for temperature control.
• No congested area due to round shape bottom.

SMALL VOLUME VESSEL

• Small volume vessel below 1L are primarily for animal or stem cell cultivation.
• As surface area of head plate is very limited on vessel below 1L, pH, DO, Temp, Foam, ORP Sensors, and etc. cannot be equipped and used on the top of the vessel.
• In order to overcome such problem, sampling, condenser, feeding ports, and sensor ports are inserted in the side of the vessel as the form of several glass tubes with certain gradient.
• The number, as well as size and shape, of side glass tubes can be adjusted according to customer’s request.
• Customized design of the vessel can be offered according to the micro-carrier the user selected for high density cell cultivation.
• For stem cell cultivation, customized design is also available for assembling form of vessel with different heights of flat type or insertion of membrane inside the vessel.
• The vessel can be customized for both adherent cell and suspension cell culture. Also, spin filter or depth filter type is available.

BOWL VESSEL

• Bottom part is stainless.
• Low risk of damage from shock.
• Production capacity : Maximum total volume 19L.

Characteristics of heat :

• Heat transfer surface area is larger than single vessel
• Temperature cooling is faster than single vessel
• It is a suitable combined form between single vessel and double vessel, and requires temperature controlling equipment (chiller or water bath)
• Bottom stainless part is double jacket which can control vessel inside temperature by heating or cooling, so cold finger is not required. For huge volume, bowl vessel is most appropriate choice

Lab Scale Animal Cell Culture Bioreactor BIOEXCEL A

DO Control Mode

• Cascade Select
• RPM Control Setup, Gas Control Setup, Air Flow Control Setup,
• RPM, Gas, Air Flow,

DO Control Mode : RPM Setup

• Action Delay Time, Maximum RPM, Increase Step, Increase Time
• Stable RPM Return
• RPM After Gas Insert

DO Control Mode : Gas Setup

• O2 Gas On/Off Time setup, Use/Not Use
• N2 Gas On/Off Time setup, Use/Not Use
• CO2 Gas On/Off Time setup, Use/Not Use

DO stat fed batch

• Set Point, Active Level (High/Low), Delay Time, Feed Pump On/Off Time, DO Control Use/Not Use

pH Calibration

• 4 pH, 7 pH, 10 pH, Measuring pH, Temperature and mV
• Temperature Compensation, Auto Calibration, Auto Calibration Processing Status
• pH Calibration Status        : Use Buffer, Slope, Run Calibration Status, Last Calibration Date,
• pH Recalibration               : Process Value, Recalibration Set Value
• pH Calibration End            : Notice, 1 Point Calibration/pH is Not Calibration

DO Calibration

• Zero, Sat
• Temperature Compensation, Auto Calibration
• DO Calibration Status        : Last Calibration Date, Zero mV, Calibration Temp., Span mV, Max. DO

Temperature Calibration

• Present Value, Set Temperature Input

Pump Calibration

• Acid Pump Calibration       : Active Time Input, Input Volume(㎖), Pump On Start
• Base Pump Calibration      : Active Time Input, Input Volume(㎖), Pump On Start
• Foam Pump Calibration     : Active Time Input, Input Volume(㎖), Pump On Start
• Feed Pump Calibration      : Active Time Input, Input Volume(㎖), Pump On Start

Variable Speed Pump Calibration

• Tube Width, RPM Set, Flow Rate
• Time Control Use : On/Off Time, Test

Setup Menu

• pH Setup, DO Setup, Temperature Setup, RPM Setup, Foam Setup, Feed Setup, Alarm Setup
• pH Setup : pH Set Value
• Pump Set : Acid On/Off Time, Base On/Off Time
• DO Setup : DO Set Value Input
• Temperature Setup : Temperature Set Value Input
• RPM Setup : Agitation Set Value Input
• Foam Setup : Foam Set Value Input
• Pump Set : Pump On/Off Time
• Feed Setup : Pump Set On/Off Time

Alarm Setup

• pH Alarm : Low set value, High set value, [Notice : Screen, BUZ, Lamp]
• Acid Alarm Use : Set value(㎖)
• Base Alarm Use : Set value(㎖)
• Foam Alarm Use : Set value(㎖)
• Feed Alarm Use : Set value(㎖)
• DO Alarm : Low set value, High set value, [Notice : Screen, BUZ, Lamp]
• Temperature Alarm : Low set value, High set value, [Notice : Screen, BUZ, Lamp]
• RPM Alarm : Low set value, High set value, [Notice : Screen, BUZ, Lamp]

Data Log

• Data Log Setup, Report View, Real Time Graph Trend

Data Log Setup

• Data Log Destination : PC, Flash Memory, Recorder, Printer
• PC                                   : Data Log Time Interval, User Name or Description
• Data Log Item              : pH, DO, Temperature, RPM, Pressure, ACID, BASE, FEED, FOAM Detect, Set Value Display, Number, Date & Time
• Flash Memory              : Data Log Time Interval, User Name or Description
• Data Log Item              : pH, DO, Temperature, RPM, Pressure, ACID, BASE, FEED, FOAM Detect, Set Value Display, Number, Date & Time

Recorder

• Data Log Time Interval, User Name or Description
• Data Log Item                   : pH, DO, Temperature, RPM, Pressure, ACID, BASE, FEED, FOAM Detect, Set Value Display, Number, Date & Time

Printer

• Data Log Time Interval, User Name or Description
• Data Log Item  : pH, DO, Temperature, RPM, Pressure, ACID, BASE, FEED, FOAM Detect, Set Value Display, Number, Date & Time

Locking Warning   : Login ID/Pass Word

System

• Diagnostic, System Configuration, Advanced User Configuration

Diagnostic

Communication Test(Protocol Test), Temperature Test, Pump Test(Variable/Constant), H/W Port Test, Motor Test(Inverter/PWM), Flash Test, Inverter selection

System Test

Communication Test

• Status Display, Receipt Information Display, 1 Packet Submit, 1 Packet Receipt, ACK Code, NACK Code, Automatic Submit and Receipt, ID

Temperature Test

• PID Control Setting, Temperature control constant adjustment, Graphic Checking Display

H/W Port Test

• Pump                 : Acid, Base, Feed, Foam, Gas1, Gas2, On/Off Checking
• Temperature    : Heater, Cooler, On/Off Checking
• Motor                 : Set Value, RPM Read Checking

System Configuration

• pH Control Configuration, Temperature Control & Sensor Configuration, Anti Foam Configuration, DO Sensor & Gas Configuration, Motor Select Configuration, Pump(Kind) Configuration, Communication Configuration, Touch Calibration, Net Work Configuration, Control Board Configuration, Protocol Configuration, Remote Control Configuration, H/W Module, Use/Not Use

Advanced User Configuration

• pH Configuration, DO Configuration, Temperature Configuration, Other

System

• Diagnostic, System Configuration, Advanced User Configuration

Diagnostic

• Communication Test(Protocol Test), Temperature Test, Pump Test(Variable/Constant), H/W Port Test, Motor Test(Inverter/PWM), Flash Test, Inverter selection

Advanced User Configuration

• pH Configuration, DO Configuration, Temperature Configuration, Other