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Concentrating protein lysates

Deitiker, Philip R via methods%40net.bio.net (by pdeitik from bcm.edu)
Tue Jun 30 11:10:55 EST 2015


Lyophilization of the lysates. 

You probably want to clarify the solution first by spinning and ultrafiltration. 
The caveats of lypohilization is that the protein needs to be freeze tolerant. Lyophilizers typically reduce the temperature well below freezing, inside the chamber is -50'C, the outside is below -20C typically. 

Many proteins can be resolubilized in pure water after lyophilization. The salt will concentrate however many fold and this will cause your gel lanes to sway. Therefore before lyophilization it is best to reduce the salt concentration if at all possible. One very fast way to reduce the amount of salt is a G-15 desalting column. This of course will also separate out small peptides, amino acids and the like. It can be done however at 4'C. 

Ultrafiltration on an amicon LWCO membrane is another way to get rid of the salt and concentrate the protein at the same time.  

Philip



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