coating peptide

Technical Assistance TechAssist at
Fri Sep 13 13:34:59 EST 1996

moropeza at MAIL.MTY.ITESM.MX wrote:
> My name is Yuriana Oropeza, and I am a master degree student in
> Mexico. I am trying to produce monoclonal antibodies using a peptide
> as antigen. I was coupling the peptide to BSA and my questions are:
> I am not sure if the coupling was complete, do you know any protocol
> easy to perform to ensure this?
> I want to screen with ELISA, and I want to know the possibility to
> coat the peptide to an ELISA plate. Peptide molecular weight: 1430
> Do you know any protocol to do this?
> Thanks for your help.

The technique used to determine the coupling efficiency of peptide to BSA 
depends on how the peptide was conjugated. The common method of coupling 
through terminal cysteines makes coupling efficiancy determination simple. Just 
do an Ellman's assay (dtnb). 
Coating a peptide on an ELISA plate has a few inherent problems. To begin with, 
the interaction between plain polystyrene plates and small peptides can often 
cause degradation of the peptide. You may want to try a different aproach such 
as biotinylating the peptide and immobilizing it on a Neutravidin coated ELISA 
plate. This has the added benefit of inserting a speacer arm between the plate 
and the peptide, htereby increasing antibody recognition.

Tom Brotcke/Pierce Chem.

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