blot affinity IgG purification

Bipin K Dalmia bipin at
Tue Feb 22 16:25:57 EST 1994

i would like to get monospecific antibodies to a protein in my crude
extract. the polyclonals react with one major band and few minor ones
and i want to get the antibodies reacting with only one band. there is a
procedure described by smith and fisher (J. cell biol. 1984, vol 99,
20-28) that discusses transferring proteins from a prep gel to
nitrocellulose, cut a strip from the nitrocellulose and visualize the
bands with a total protein stain, cut out the section of the remaining
nitrocellulose with the protein band of interest, incubate it with the
polyclonals (crude serum), wash out the unbound antibodies, and then
elute monospecific antibodies at low pH.

if anyone has a detailed protocol or has done this and found any
possible sources of problems, please let me know.

bipin k. dalmia               the other night i was lying on my bed, looking
bipin at             up at the beautiful stars, and i said to myself, 
n2.bkd at     'where the F*CK is my ROOF !!'

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