protein spreading in gel - (Nov/27/2008 )
Has anyone had a problem that protein are laterally spreading in the SDS-PAGE gel? Or does anyone know a cure for the problem? My bands are sharp but the width of the bands are two times the sample well so that I cannot run samples in adjacent cells.
I have heard that too high salt concentration can cause this problem so after extracting the proteins I have desalted the samples with PD-10 desalting columns and elueted them to 0.05 M Tris-HCl, pH 8.0. It wont help. After eluation I have precipitated the samples with 80% acetone and resuspended to Laemmli sample buffer. Usually I need to add 2 ul of 1M Tris-HCl, pH 8.0 to samples before loading to gel as otherwise proteins form vertical streaks and diffuse bands in the gel. Does anyone know why?
Can you give us a picture of this spreading? Would be helpful