Purification of protein from plasma - How to separate a 30kDa protein from serum albumin (Nov/20/2006 )
I am currently working on prufication of protein of interest (30kDa). When I run 1-D SDS PAGE and western blot, I find the majority of my protein appears complexed with the albumin bands. very little appears at the right size (30kDa). my question is, How can I completely solubilize my protein from albumin and separate it? In 1-D gels I used the regular 1X loading buffer. Are there any other protocols available to help separate the protein from albumin or any other complexes?
are you using reducing agent (2me, dtt) in your sample loading solution?
do you heat the solution prior to loading?
you will want to completely denature your sample. the interaction with bsa should not be covalent so these treatments should separate them.