Release of intracellular components in downstream processing

Posted by admin on October 2, 2016 in Uncategorized |

Targets like recombinant proteins and many enzymes are intracellular in downstream processes.

Sometimes breaching of cytoplasm membrane and the cell wall can bring problems. This can occur

when the cell has a strong cell wall. The liberation of the DNA which can, in the long run increase

viscosity of suspension is some of the many problems that are associated with cell disruption. This

can affect further downstream processing. The DNAse addition can help to prevent this problem.

Heat is greatly released if the disruption that is involved is mechanical and this may need cooling

measures to be implemented. Products that are released by eukaryotes are subject to degradation by

the hydrolytic enzymes and this damage can be reduced by adding of enzyme inhibitors, rapid

processing and cooling of the cell extract.

Both mechanical and non mechanical means can be used to achieve cell disruption. The disruption

processes can be measured in terms of quantity by monitoring changes in absorbance, total protein

concentration, particle size or the activity of specific enzymes.

The following are methods for cell disruption in downstream processing

 Mechanical cell disruption

 Non mechanical cell disruption

There are several mechanical methods involved in cell disruption in downstream processing. There

are those methods based on solid shear and those that are based on liquid shear. The extrusion of

frozen cell preparations via a narrow orifice in high pressure is involved in solid shear. This method is

suitable in laboratory operations but not at factory level. Liquid shear methods are more effective.

Non mechanical cell disruptions are downstream processing methods that are alternative to

mechanical methods. This involves causing cells to be permeable. It can be achieved in several ways

including osmotic shock, autolysis, rupture with crystals or heat shock. More techniques have been

developed for small scale operations.

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