modes of culture for high cell densities

8/12/2019 Modes of Culture for High Cell Densities

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Modes of culture for high celldensities

Chapter 10 The Basics


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What is a Batch Culture? Cells are inoculated

culture left for several

daysuntil final

densityis reachedNothing is added or

removedduring culture

Substrates get used up

and products are

secretedfrom cells


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Heterogeneous system


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Detrimental environment for cell growth

- Depletionof an essential nutrient

- Accumulationof an inhibitor

- Complete coverageof available growthsurface


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What is Fed-batch culture? Cell growthnutrient supply or removal of by-

products

Cell yield will reach high densities

In open system/Fed-batch cultureinvolves

controlled nutrient feeding

Partial media changesat regular intervals


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What is a Continuous Culture?Open systemcontinuous feed of medium and

removal of spent medium

Cell growth - longer period in CC>BC


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Types of Continuous Culture

Chemostat cultureCellsand spent

mediumare continuously removed

- State of culture is dependent upon flow rate of

fresh medium


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Types of Continuous CulturePerfusion cultureCells are retained in

fermenter

- Medium is pumped continuously

- Cells are retained

- Becoming popular for large-scale production

- Attains high cell density- Cell separator


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Cell immobilizationImmobilizationof cells on or inside particles

Allows attachment of cells to solid surface

Anchorage dependent cells

Entrapment of cells in small beads


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What are Microcarriers?Microcarriers are

microscopic particles(diameter = 200 m)

Maintained insuspension in liquidmedium

Dextran, Collagen andPlastic


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Characteristics of MicrocarriersSmallto maximize the available surface area

for cell growth

Lightto allow easy suspension in culture

medium

Transparentto allow easy observation of

cell attachment and growth

Chargedto allow cell attachment onto

surface


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Porous microcarriersDextran microcarriers (example: Cytodex)

are microporous

- Pore size is not sufficient to allow cells to

colonize the interior of beadsGelatin microcarriers (example: Cultispher)

are macroporous

- Increased surface area for attachment- Interior environment to protect cells against

adverse shear forces


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Extraction of cells from MicrocarriersDetachment by either trypsin or collagenase

treatment

Detached cells separated by sieving through a

nylon mesh


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Immobilization of nonanchorage-dependentcellsProtection against mechanical stress

Ease of continuous operation

Isolation of products


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Immobilization of nonanchorage-dependentcellsCell entrapment

- Mixing and agitating suspension of cells in

warm agarose with hydrophobic liquid

paraffin oil

- Forms solid beads of agarose (100-200 m)

containing suspended cells

- Secreted cellular productsseparated from

entrapped cells


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Immobilization of nonanchorage-dependentcells - Encapsulation Cellsenclosed in

semipermeable membrane

Cells + sodium alginate

drip into calcium alginate

Polylysinecreates an outer

semipermeable membrane

Monoclonal antibodies

accumulate inside the bead

matrixcan be extracted

easily


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modes of culture for high cell densities

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