Affinity purification of antibody against MBP-fusion

John Leonard via methods%40net.bio.net (by jleonard.iii from gmail.com)
Wed Jun 4 16:31:39 EST 2008


I have an antibody against a protein I'm studying, and I'd like to purify it
from the serum to see if I can improve its specificity for my protein in
Western, EMSA, ChIP, etc.  I have a vector for an MBP fusion of my protein
(which I know the antibody binds very well), and I was hoping to use that to
affinity purify my antibody.  Ideally, I would like to express the bacterial
fusion, run it over an amylose column, then use the column-bound MBP fusion
to affinity purify my antibody.  I have seen similar protocols using His- or
GST-tagged proteins, but have not found a protocol for MBP fusions.

If anyone has a protocol for something like this, or has another idea for
how I can purify my antibody based on specific ligand interaction, I'd
greatly appreciate it!


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