antibody purification

Frank O. Fackelmayer Frank.Fackelmayer at uni-konstanz.de
Wed Jan 26 02:53:58 EST 2000


Dave Parcej wrote:

> hold on! you said in your first post that the antibodies were unlikely to
> be derived from an impurity in the sample.

sure,  but there ARE usually impurities in the sample, and those impurities are
bacterial proteins. When you use such protein preps for purifying antibodies, you
are unlikely to remove all unwanted antibodies (even though they were not made in
response to immunization but had been there before due to natural immune
responses).


> Still, I spose you could blot
> and cut out the band to use as the affinity ligand. :)
>

yes, that is of course a good possibility. I personally, however, prefer to
couple antigen to a column for purifying antibodies by affinity-chromatography,
because you can use more antigen than you could ever blot to a membrane. I
routinely use 1mg antigen coupled to 500ul of CNBr activated Sepharose or
sulfolink material.


Frank









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