buffer for anion exchange chromatography of basic proteins - (Dec/03/2007 )
I want to do anion exchange chromatography for protein purification. I am interested in basic proteins (theoretical pI=8.5 and 9.3). I think I would need to use a buffer of pH of about 10.
After anion exchange chromatography purification, I want to precipitate proteins with acetone and use them for SDS PAGE and then the relevant fraction for reverse phase chromatography.
Do you have any experiences with a similar experiment/work?
What type of buffer did you use for anion exchange chromatography (of basic protein)?
It is recommended: ethanolamine, piperazine, propane 1,3 diamino. Would those three buffers not interfere with acetone precipitation?
Could ammonium bicarbonate be used at pH=10?
Thanks for advices.
carbonate has a pK at 10, so it could be used. but you might want to be a little above so that it could resist change towards your proteins' pI better.
on the other hand, why don't you use cation exchange instead? then you can use neutral pH buffers that you may feel more secure with during post column processing.