Synthetic Peptide Antibody Questions
George L. Gerton
ggerton at obgyn.upenn.edu
Wed Dec 21 09:44:35 EST 1994
In article <shark.10.000D0137 at sam.neosoft.com>, shark at sam.neosoft.com
(John D. Valentich) wrote:
> We have recently started preparing polyclonal antibodies in rabbits
against synthetic peptides
> from annexins and phospholipase A2 activating protein (PLAP). Peptides were
> conjugated to the carrier BSA. We are wondering about the problem of
> antibodies generated against BSA reacting with serum albumin or albumin-like
> epitopes in other cellular proteins. Questions:
> 1. Is it likely (or inevitable) that our antisera will contain high
titer antibodies against BSA?
> Could they be expected to cross react with other non-albumin cellular
> 2. If so, is the best (or only?) solution to this problem affinity
> purification of annexin or PLAP antibodies from the antisera?
> 3. What are other possible solutions?
I think it is highly likely that you would produce antibodies against
BSA. Many people use peptides conjugated against KLH (keyhole limpet
hemocyanin) as their immunogen instead of peptide-BSA conjugates.
However, for most of your purposes, you should be able "adsorb" out the
anti-BSA antibodies by including BSA in your antiserum diluent (blocking
buffer). For western blots, you should test this by running BSA (purified
protein or the BSA in your molecular weight standards). If sufficient BSA
is included in the blocking buffer, you should not see reactivity against
the BSA run on the gel.
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