Problems with silverstaining

Bernard Murray, PhD spam at 127.0.0.1
Wed Jun 28 16:24:51 EST 2000


In article <3959FE10.EF0D4DC4 at casema.net>, Suzanne Hermeling
<deefsuus at casema.net> wrote:
 
> I have a question. I am working on protein purification and because I
> have very low protein concentration I use silverstaining to visualize
> the proteins on an SDS-PAGE gel.
> 
> But here comes the problem:
> One time we have a good pattern of proteins and the other time we have
> only a very dark background without any pattern.

If it is a contamination problem then make sure your glass plates for
electrophoresis are *really* clean (start with a nitric acid wash)
and also make sure your staining box is equally clean.
   If you are fixing your gel with acetic acid before staining
make sure you have *throughly* rinsed it with high quality water
before you start to stain.
   When making up the staining solution use very high quality water
(we actually use HPLC grade).

If it is a development problem then you have to stand there and
watch the pattern develop and stop it when it is optimal.  You
cannot simply time the process as it takes a variable period for
the bands to start to appear and the background will come up soon
after that.
   Good luck,
      Bernard

-- 
Bernard P. Murray, PhD
bpmurray at cgl . ucsf . edu
Department of Cellular & Molecular Pharmacology, UCSF






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