strange double band in electrophoretic pattern - (Jan/28/2009 )

I used a 5% separating with a 3% stacking and run for 90 min in total at 50 mA (constant). Gel was fixed in acetic acid/ethanol solution, stained by Coomassie Blue R-250. most of bands even markers (1st left lane)appear as two bands which seem from one protein. I'm very confused with what reason lead to band split.

Thank your for your time.

Maybe a problem with your buffer are you using SDS PAGE classical or MES buffers ?

pesji on Jan 29 2009, 09:52 AM said:

I use classical SDS-PAGE buffer, do you mean it has a problem on buffer pH? I checked when I made them at room temperature, Stock stacking and separating solution are pH 6.8 and 8.8 respectively. Maybe I need to make a double check.