2D gel question
Stephen P. Driska PhD
driska at astro.ocis.temple.edu
Fri May 12 14:10:20 EST 1995
Regarding the issue of whether 2D gels are possible and the
presence of SDS, I think it depends. For 15 years or so we have run
whole tissue samples (homogenates of arterial smooth muscle) that were
in 1% SDS, 25 mM dithiothreitol and glycerol. The sample volumes were
even quite large (200 microliters run into a 3 mm i.d. gel, 10 cm
long). What happens, apparently, is that the SDS solubilizes the
proteins in the tissue homogenate as well as anything, the protein-SDS
complexes run into the IEF gels, and the SDS forms mixed micelles with
the NP-40 or other detergent you might use. At the bottom of the gel
there is a pronounced swelling that is attributed to the SDS-micelles.
The reason we always did this is that homogenizeing tissue in SDS
seemed more effective at getting the protein into the first dimension
gel than did homogenizing in the original O'Farrell lysis buffer
(there was less trapping and streaking at the origin).
But of course I can't say how it would work for you with your
sample of interest......
Good luck with it,
More information about the Proteins