In article <4s19qb$guv at deft.cc.purdue.edu>,
wilkinst at deft.cc.purdue.edu (Thomas Wilkinson) wrote:
-->Newsgroups: bionet.molecules.peptides
-->Subject: gel electrophoresis of peptides...
-->Summary:
-->Followup-To:
-->Distribution: world
-->Organization: Purdue University Computing Center
-->Keywords:
-->
-->
-->Hello,
-->
-->I was wondering if somebody had some experience with SDS gel
electrophoresis of
-->peptides. I have a sample that I would like to analyze to see if it
contains
-->a short (12-residue) peptide, molecular weight = 1462.
-->
-->The sample is not some mixture of many species...the sample to be analyzed
-->either has the 12-mer, or else the sample contains no peptide whatsoever.
Thus,
-->this gel should be really easy to interpret. I also have a similar sample
that
-->I would like to analyze for peptide content, except in this case I am
looking
-->for the presence or absence of a 33-residue peptide (MW = 3487).
-->
-->I would like to run these samples out on an SDS gel, and then
coomassie-stain.
-->My question is that I don't know what the optimal percentage of
polyacrylamide
-->would be for this task. What percent gel is best? Is gel electrophoresis
of
-->peptides commonly done? Your suggestions are much appreciated.
-->
You are at the lower limit for SDS-PAGE and would need to use a Tris-Tricine
buffer system (schagger & von Jagow, Anal. biochem. 1987, 166, 368-379)
and a fairly high percentage gel (15%-20%) Even then the 12mer may be too
small.
Alternatively, if you just wish for a yes or no answer for the presence of
peptide, you could dot your sample onto PVDF and stain with amido black.
best of luck.
.d
* David Martin, PhD - Post-Doctoral Research Fellow *
* Atherosclerosis and Thrombosis research group *
* Biotechnologisenteret i Oslo *
* Postboks 1125 Blindern, N-0316 Oslo, Norway *
* Tel: +47 22 95 84 54 Fax: +47 22 69 41 30 *
* http://www.uio.no/~damartin/david.martin at biotek.uio.no *
