5 kDa protein detection

Rich Dudley rdudley+ at pitt.edu
Fri Dec 18 15:18:45 EST 1998


Rasmus Wendelbo Nielsen wrote:

> I am currently trying to purify a protein of 5 kDa, but are having
> problems detecting the protein. As it only contains tree tyrosines and
> no tryptophans the A280 is very low. I have tried running 18% SDS-PA
> gels and a 15 to 25% SDS PA gradient-gels, without luck, the protein
> probably diffuses out of the gel during staining.
> So I am looking for other (easy) ways to detect low MW proteins. Any
> ideas would be appreciated!
> I have heard that adding coomassie to the running buffer instead of
> post-staining the gels could be a solution. Anyone got experience with
> that?
> Thanks, Rasmus

Try going to FMC's web site (I think it's www.bioproducts.com); they have
an acrylamide that when run with a tricine buffer can separate proteins of
that mass.  I've never tried it, and just heard of it yesterday.  I use a
protocol by Schagger and von Jagow to make discontinuous tricine gels.  I
don't have the ref handy, but a pubmed search will turn it up.

Good luck!

rich


--
--- --- --- -- -- -- --- --- ---
Richard J. Dudley (rdudley+ at pitt.edu)
Research Specialist V
Dept. of Cell Biology and Physiology
University of Pittsburgh
http://www.cbp.pitt.edu
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