Direct sequencing of peptides transferred onto polyvinylidene difluoride membrane from sodium dodecyl sulfate–polyacrylamide gel electrophoresis by electroblotting

Abstract

Protein stained with coomassie brilliant blue (CBB) on polyacrylamide gels were digested with lysylendopeptidase in the presence of sodium dodecyl sulfate. The resulting peptides were then separated by sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS–PAGE) and electroblotted onto a polyvinylidene difluoride (PVDF) membrane. After CBB R-250 of the staining peptide bands on the PVDF-blotted they were loaded directly into the sequencer. Using PVDF membrane for peptide separation and sequencing is quite simple and in contrast to separation of peptides by reversed-phase liquid chromatography, it is not affected by sodium dodecyl sulfate.