Re: Antipeptide antibodies

Gottfried Proess (g.proess@eurogentec.com)
Thu, 29 Jul 1999 12:40:56 -0700

Latif Kazim wrote:
>
> Dear Colleagues:
>
> One of our researchers recently asked the following question about a method
> for making antipeptide antisera:
>
> >Do you have knowledge/experience with direct use of synthetic
> >peptides for producing antisera--particularly the 6xhis-tagged peptide
> >method?? Apparently the peptide is bound to Ni-NTA agarose and used
> >directly.
>
> Any responses would be appreciated.
>
> Thanks,
>
> Latif Kazim
>
> ----------------------------------------
> A. Latif Kazim, Ph.D.
> Biopolymer Facility
> Science-4
> Roswell Park Cancer Institute
> Elm & Carlton Sts.
> Buffalo, NY 14263
>
> E-mail: kazim@sc3101.med.buffalo.edu
>
> Phone: (716) 845-8923
> Fax: (716) 845-7621

Dear Latif,

having read the answer of <gsarath@unlnotes.unl.edu>, there is maybe a
misunderstanding somewhere. What I understood from your question is that
the peptide is synthesized with a 6His-tag, bound to Ni-NTA agarose and
injected like this for antibody production. Am I right or does your
question effectively concern the antiserum purification on a Ni-NTA
column on which the 6His-tag peptide has been fixed?

If I am correct, I cannot see any real advantage in doing this type of
antiserum production. The addition of 6 histidines makes the peptide
more expensive and maybe even more difficult to synthesize. The classic
method of coupling part of the untagged peptide to KLH works very fine
for antibody production and the remaining uncoupled peptide can be used
easily for subsequent antiserum purification on an affinity column using
again the same type of coupling reaction to an adapted matrix.

But if you want to try antibody production using the agarose beads, I
would not suspect any difficulties or problems in doing this. We did
several successful antibody productions with agarose-bound proteins, but
not yet with peptides.

I hope this helps a little.

-- 
Gottfried Proess
Tel.: + 32 4 366 01 50  Fax : + 32 4 365 16 04
E-Mail: g.proess@eurogentec.com