Electroblotting of individual polypeptides from SDS/polyacrylamide gels for direct sequence analysis

Eur J Biochem. 1987 Nov 16;169(1):9-12. doi: 10.1111/j.1432-1033.1987.tb13573.x.

Abstract

A scheme for electroblotting of individual unstained protein bands from SDS/polyacrylamide gels and subsequent amino acid sequence analysis is described. Principal features are: detection of the polypeptide bands by visualization with KCl; electroblotting of excised gel pieces that correspond to the protein bands only; blotting onto polybrene-pretreated glass-fiber filter discs (12 mm diameter) placed in an electrophoretic concentrator. A high yield over all steps from gel application through electrophoresis, blotting, gas-phase sequencer degradation, and phenylthiohydantoin analysis is obtained with several different types of polypeptide (combined average yield over all steps 20%, spread 10-50%). Background is low and samples can be stored under vacuum for long periods after blotting.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence*
  • Electrophoresis, Polyacrylamide Gel*
  • Peptides*
  • Potassium Chloride

Substances

  • Peptides
  • Potassium Chloride