Recycling Cyanogen bromide activated sepharose affinity columns?

[Joe Mack]

In article <CKBILy.7pG at ucdavis.edu> szsclark at hamlet.ucdavis.edu (Sonya Clark) writes:
>Dear Netters - I am purifying antibodies from a polyclonal prep by 
>running them over a CNBr-activated Sepharose affinity column of my 
>antigen.  As CNBr-activated Sepharose is expensive &/or horrible to make, 

Well it isn't that expensive compared to the time saved by buying it (unless you're a graduate student whose time isn't valued much) and it isn't all that bad
to make (N-methyl-pyrrolidone/Na2CO3 method ) if you have a fume hood but anyhow

>I'd like to strip the column of the antigen currently bound, and re-use 
>the sepharose to make another affinity column. 

the real point is that the activated bond from the CNBr is used up by the 
coupling procedure. Even if you could get the protein off, you're back
to unactivated sepharose. 

	Joe Mack
	mack at ncifcrf.gov


 Does anyone have a protocol 
>to do this?  Would eluting the column with harsh eluents (eg. 
>Guanidine/urea/SDS) work? and would I then be able to re-use the sepharose 
>to couple another antigen? Any advice appreciated. Cheers.
>
>Sonya Clark.
>Botany Dept.,
>UCDavis
>szsclark at ucd.edu
>