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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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