TCA and HIC protein pufn

Dr E. Buxbaum EB15 at le.ac.uk
Tue May 13 10:21:22 EST 1997


I.McFarlane at icrf.icnet.uk (Ian McFarlane) wrote:

>My protein of interest happens to be soluble in 10% TCA. I am going to use
>TCA as the first step in its pufn. Does anyone know if I can go on to do
>hydrophobic interaction chromatography without dialysing away the TCA.
>Obviously anion or cation exchange would require dialysis. Any
>suggestions?

I have never tried HIC from TCA solutions. However, if this does not 
work, there are at least two options other than dialysis. The first would 
be gel filtration, which at the same time as removing TCA can be used as 
second purification step (by MW, obviously). The other option would be 
ultrafiltration in a stirred cell, like the ones produced by Amicon. The 
solute is pressed through a membrane by nitrogen pressure, proteins above 
the cut off size of the membrane are retained. Simply concentrate your 
sample to, say, 1/10 or 1/20 of its original volume, add your starting 
buffer for the next step and concentrate again. This process is a lot 
quicker than dialysis (normally 2-3 h total) and gives you a concentrated 
sample, which is easier to work with (although this is not too relevant 
with HIC). 




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