KCl staining for protein
ED at micro.uct.ac.za
Thu May 6 02:51:07 EST 1993
> From: mm6y+ at andrew.cmu.edu (Morris F. Manolson)
> Subject: Re: KCl staining for protein
> Date: 5 May 93 15:26:02 GMT
> >Anybody out there have experience staining protein gels with KCl and
then > >electroeluting the band you want?
> KCl is used to precipitate SDS in the gel matrix leaving regions of low
> SDS (ie: high protein concentration) visible as clear bands. Following
...but it isn't very sensitive.... - much more sensitive is 0.3M CuCl2,
which is a negative stain, is permanent (if you want it to be), and is
easily destained (if you want to blot/elute/whatever). Simply soak gel
straight after elec in 0.3M CuCl2 for as long as it takes to get good
staining (+/-15min), then store in ddH2O for as long as you like: the
proteins are insolubilised, so won't go anywhere. You can cut out and
store bands (best visualised by indirect illumination against a black
background) until you want to use them, or store gels until you want to
blot them. To destain you simply wash a fewx5min in 250mM EDTA/500mM Tris
pH 8.0: stain leaches out, and gel goes clear and prots are soluble once
And, it turns out, the gels stain better with Coomassie if copper stained
first (with no destaining). It is the simplest, one of the cheapest, and
most convenient (and quick) ways of staining SDS protein gels.
Use and enjoy.
C. Lee, A. Levin and D. Branton: Copper staining: a five-minute protein
stain for sodium dodecyl sulfate- polyacrylamide gels; Anal.
Biochem. 166, 303-312; 1987
Gels may be stored in water for up to several months at room temperature
with no problem or fading: proteins are immobilised as a Cu-SDS-
polypeptide complex in the gel, which remains clear; the colour and
background opacity are due to a Cu-SDS-Tris complex. Gels may
be destained completely by repeated washing in 0.1-0.25 M
Tris/0.25 M EDTA pH 8.0, and then electroblotted, or eluted from
the gel for other purposes. NB: NATIVE PA GELS CAN ALSO BE
STAINED : this is a less sensitive procedure, and bands are
visible as faint blue opaque bands on a clear blue background.
ELECTROBLOTS CANNOT BE STAINED WITH COPPER.
| Ed Rybicki, PhD | "Lord, won't you buy me |
| (ed at micro.uct.ac.za) | |
| Dept Microbiology | A Mer-ce-des Benz..." |
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