Polyclonal Ab Purification

Paul Kenneth Farmer gt7680b at prism.gatech.edu
Sat Apr 15 18:34:11 EST 1995


Dear Netters:

     I have tried affinity purifying polyclonal antibodies using Affi-Gel
10, but unfortunately the elution using acidic, basic, or chaotropic
conditions failed to remove most of the bound antibodies or at least had
inactivated them. I am considering three approaches:  (1) Inverting the
column and eluting in the opposite direction to prevent antibodies from
dissociating and then reassociating with the bound ligand; (2) Using a
batch method instead of column purification; and (3) Using electroelution
of bound antibodies off of the column. My best success to date has been
the straight acid elution using a glycine-HCl buffer, pH 2.5. However, the
activity and specificity of the eluted antibodies were still unacceptable.

     Does anyone have any suggestions on how to improve this procedure? In
particular, has anyone tried electroelution coupled with inverting the
column so as to place the immobilized antibodies closer to the anode?

     Any suggestions would be greatly appreciated!!!!

     My E-Mail address is: gt7680b at prism.gatech.edu





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