[ Article crossposted from bionet.molbio.methds-reagnts ]
[ Author was Jens Krieger (krieger at mibm1.ruf.uni-freiburg.de) ]
[ Posted on 4 Jul 1996 21:55:29 GMT ]
Hello!
I'm staining SDS-PAGE with methylgreen, using the method of Cutting and
Roth (73), but I've problems with high background and no seriously fixed
proteins.
I fix the gels with sulfosalicylic acid (SSA, 10% w/v)) and CaCl2 to build
unsoluble calcium-phosphat complexes. So far, so good. With the last
step, rinsing with aequos ammonium-molybdat, there should be
phospho-molybdat complexes. I think it works so far, because if I stain
it with methylgreen (5% in 7% acetic acid) and then destain it with SSA,
i will see for a short time very sharp green bands, but there are still a
very high background. So I have to destain ti further on, and
suddenly....the bands disappear. The background is okay, but without any
bands.....
Does everyone has an idea, why the bands suddenly disappear? Another
destaining reagent, another fixing reagent? (i've tried TCA and normal
acetic acid; it doesnt work at any time).
Thanks in advance, Jens.
--
/~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~\
| Jens Krieger "Solange die Alpen noch stehen, |
| wird es keine |
| Tel.:+49.351.463.5465 Gerechtigkeit geben."|
| |
|krieger at rcs.urz.tu-dresden.de H.Achternbusch |
\ krieger at mibm.ruf.uni-freiburg.de /
~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~
--
/~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~\
| Jens Krieger "Solange die Alpen noch stehen, |
| wird es keine |
| Tel.:+49.351.463.5465 Gerechtigkeit geben."|
| |
|krieger at rcs.urz.tu-dresden.de H.Achternbusch |
\ krieger at mibm.ruf.uni-freiburg.de /
~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~