The bromophenol dye front traveled all the way to the bottom of the gel but the spots are seen mostly on top.. Even the molecular marker on the right looks like it traveled out of the gel. I tried checking for possible reasons that might have caused this rare anomaly but I could not find any that fits the problem that I have encountered. Thanks.
It might be a problem in the lower gel buffer. You might do new SDS-PAGE gel buffers.
1. What is the concentration of your agarose low melting to seal de strip?. I am using agarose 1% in distilled water with bromophenol blue and it works well.
2. Also, what voltage are you using and how long?.
Dear Dominic
What is %T of 2nd dimention gel?
It s also caused by a too low SDS concentration. Therefore not all proteins (especially the high MW proteins) are saturated with SDS.
Also, it seems that the gels have buble and you must the gasess the gels
It's a 12.5% gel. And the agarose sealing solution is 1% with bromophenol blue.. Another thing that I also observed was that the agarose sealing solution actually expanded out of the gels from the top of the plates after the run was completed. Could this be due to heat during the run or maybe even impurities in the protein sample??
What kind of 2-DE system are you using? I would say you might have a leakage problem between the anode/cathode buffer.
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