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Industrial Microbiology Dr. Butler 2011
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Lecture 9 Animal Cell BiotechnologyScaling up the production process
Scaling up animal cell/microbial processes
• optimal physiological conditions obtained via small scale processes maintained for large scale operation
• must consider:
1. fermentor/bioreactor system
2. control of agitation, pH, temperature, dissolved oxygen
Lecture 9 Animal Cell BiotechnologyScaling up the production process
T-flasks
Lecture 9 Animal Cell BiotechnologyScaling up the production process
Spinner bottle
Lecture 9 Animal Cell BiotechnologyScaling up the production process
Multiple process vs. unit process• can have 1000 100 mL flasks or a single 100 litre
fermentor (same volume)
1. Multiple process• easy to replicate samples and conditions, large surface
area/volume ratio, tedious to take care of all samples, labor intensive
2. Unit process• good for large scale production of biological products,
ease of handling, decreased possibility of contamination
Lecture 9 Animal Cell BiotechnologyScaling up the production process
Butler, M. 2004. Animal cell culture and technology 2nd ed. London and New York:Garland Science/BIOS Scientific Publishers. P203.
Lecture 9 Animal Cell BiotechnologyScaling up the production process
Suspension vs. Anchorage-dependence
• suspension cells are readily scaled up to production level using similar fermentation vessels as microbial cells (with modifications)
• anchorage-dependent cells require a surface for attachment in order to grow
→ grow as static cultures (i.e. hollow fibres)
→ grow on microcarriers, in a stirred tank reactor (fermentor)
Cell culture processes• Anchorage-dependent systems
Microcarrier (dextran, collagen or plastic) : 200 µm
Advantages: -Higher cell yields - increased productivity. - ↑ surface ratio (100 -200 cells). - suspension cultures • Suspension systems: > Industrial culture
Cytopore microcarriers
Cell Culture system• Vero cells
• Why? Regulated by WHO
• already licensed for vaccine
production such as polio, influenza,
rabies, others
• Serum-free medium
• Why? Safety and reproducibility
• Microcarriers
• Why? Scalability and ease of infection
•
Macroporous microcarrier
Lecture 9 Animal Cell BiotechnologyScaling up the production process
Usually scale up 10x at a time:
1 → 10 → 100 → 1000 → 10,000 L
Factors limiting scale-up• supply of oxygen
• shear damage from mixing
• build up of toxic metabolites
Lecture 9 Animal Cell BiotechnologyScaling up the production process
during scale up decreased product yields may be experienced due to the following:
1. inoculation – must consider quality and quantity of inoculum used to start process
2. choice of medium – cheaper materials often used for large scale production due to cost limitations - omission / reduction of serum, antibiotics
3. large scale sterilization – longer sterilization times may result in degradation of heat labile compounds, reducing quality of media
4. development of gradients - larger fermentors/bioreactors may be subject to development of nutrient, temperature, pH, and oxygen gradients
5. other factors -scaling up may also alter the generation of foam, shear forces
$ $$
Lecture 9 Animal Cell BiotechnologyScaling up the production process:
The Stirred Tank Reactor
• bioreactor is another term for fermentor
• stirred tank reactor simplest and most widely used system (pot and paddle)
→ small vessels (<20 litres) made of glass
→ large vessels (>20 litres) made of stainless steel
Bioreactor (Fermenter)
• Vessel that allows the growth of cells.
• Stirred tank bioreactor (STR). Growth: Bacteria, yeast, mammalian cells.• Homogeneous solution mixed by an impeller: -Laminar stirred: Just one dimension to lift the liquid.
www.corning.com
Cultures >1L: Uneven oxygen exchange and poor nutrient distribution
Stirred Tank Reactor
• Shape: -curve bottom mammalian cells -square bottom poor mixing
• Materials used: Vessel up to 10 L (glass) Head plate steel stainless
• Heat control: (heat pad or jacket water) - Constant To -mammalian cell culture (37o C)
Lecture 9 Animal Cell BiotechnologyScaling up the production process:
The Stirred Tank Reactor
Butler, M. 2004. Animal cell culture and technology 2nd ed. London and New York:Garland Science/BIOS Scientific Publishers. P156.
Fig. 9.2
Fig. 9.3
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