Recently I am facing a problem with my SDS-PAGE gels (12% or 15%) which I never faced before. After the run when I blot the gel and stain the blot with Ponceu, I can see that my proteins have run only half or 2/3rd the distance and have stopped there. But the marker gets separated well till the end. If I stain the gel, there also I can see proteins distinctly stopping in between. I prepared fresh cell lysis buffer, sample loading buffer and fresh running buffer but the problem persists. Can anyone suggest whats wrong and how to get rid of the problem?
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#2
mdfenko
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Posted 03 March 2009 - 11:33 AM
have you also prepared fresh gel buffers (not just the electrode buffer)?
is there any difference between the lysis buffer that you use now and when the samples ran well?
is there any difference between the lysis buffer that you use now and when the samples ran well?
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#3
NI2009
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Posted 03 March 2009 - 11:39 PM
mdfenko, on Mar 3 2009, 11:33 AM, said:
have you also prepared fresh gel buffers (not just the electrode buffer)?
is there any difference between the lysis buffer that you use now and when the samples ran well?
is there any difference between the lysis buffer that you use now and when the samples ran well?
Yes, the Tris for the gels is also prepared recently. The pH was also checked before the use. I'll make them again anyway and see.
The composition of the lysis buffer is the same as before.
