Bioreactors

Module 2 –– GROWTH OPERATIONS

A Practical Guide to Biopharmaceutical Manufacturing

CHAPTER 2 CULTURE TYPE
All types of micro-organisms are grown in suspension with the exception of cell culture, in which cells can also be anchorage dependent. Suspension –– the micro-organism can grow successfully whilst in a suspension Anchorage dependent –– the micro-organism requires a surface on whichto attach itself and grow Suspension based operation is favoured for large scale manufacture. Mammalian cells are naturally anchorage dependent. In comparison with suspension cultures, anchorage dependent cells are usually more genetically stable and have higher productivities. Modification of mammalian cell lines is possible to move from attachment to suspension based operation using one of threedifferent methods: Modification of the growth medium to prevent attachment, which may reduce productivity Selective use of cell lines to reduce attachment dependency Selective use of cells that will not attach to specially treated surfaces The second option is suitable for production cell lines, but it can be difficult to achieve. The third option is normally easier and more rapidly developed,allows operation at large scale, and enables attachment dependent growth at small scales, simplifying laboratory handling and development. 2.1 Suspension culture

Some micro-organisms grow freely suspended in culture fluid. Nutrients are supplied by the growth medium in which the micro-organisms are suspended. Oxygen is supplied by pumping air into the bioreactor. To support the high cell densitiesrequired for economic operation, the suspension must be agitated to ensure consistent and effective mixing so that all of the cells have access to nutrients and oxygen and that excess heat and exhaust gas are removed. The agitation method differentiates the two basic types of suspension bioreactor: Air lift –– agitation supplied by the movement of air through bioreactor Stirred tank –– mechanicalimpellers agitate the suspension For small volumes, it is possible to use alternatives such as shaker flasks, mixing bowls, and disposable wave bioreactors.

High cell densities in suspension cultures are achieved by consistent and efficient agitation to ensure that all the cells have access to the required nutrients and the removal of excess heat and exhaust gas

9

© Informa UK Ltd, 2006Module 2 –– GROWTH OPERATIONS

A Practical Guide to Biopharmaceutical Manufacturing

Irrespective of the agitation method, bioreactors are designed to avoid the introduction of any unwanted materials, or contaminants. This requires careful design of all connections, tubing, and the vessel itself to prevent areas that cannot be properly cleaned and sterilised. Bioreactors and associatedpipework surfaces are usually constructed of stainless steel and finished to a high standard to reduce attached growth and aid cleaning. Any entry or exit point from the vessel is a potential contamination route. The overall risk of contamination is often reduced by running the entire vessel slightly above atmospheric pressure, impeding the entry of external organisms. Cell cultures produce heat, andthe larger the working volume, the greater the heat load. The vessel is therefore cooled during the fermentation process to ensure that the process operating temperature remains at the optimum value. Cooling is often supplied by external cooling jackets or coils. Internal coils may also be used to allow more rapid cooling of vessels following sterilisation. This may be of greatest benefit if theprocess requires the sterilisation of growth media in situ. 2.1.1 Airlift bioreactors

Air-lift bioreactors achieve circulation of the growth medium and aeration by injecting air into the lower part of a bioreactor vessel (see Figure 2.1). The movement of the air mixes the growth medium and generates a re-circulating flow through the vessel as shown in Figure 2.1. The air reduces the density...