Extraction and characterization of adenovirus proteins from sodium dodecylsulfate polyacrylamide gel electrophoresis by matrix-assisted laser desorption/ionization mass spectrometry

J Am Soc Mass Spectrom. 2000 Apr;11(4):356-61. doi: 10.1016/s1044-0305(00)00101-x.

Abstract

A new methodology for the extraction and characterization of proteins from Coomassie-stained sodium dodecylsulfate polyacrylamide gel electrophoresis using matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) has been described. The utility of this methodology was demonstrated in the characterization of adenovirus proteins. The key steps in the extraction and destaining process involve washing the excised band with a combination of solvents that include 10% acetic acid, acetonitrile, methanol, and formic acid:water:isopropanol mixture. By using this procedure, we determined adenovirus proteins with molecular weights ranging from 10,000 to 110,000 Da by MALDI-MS, obtaining a detection limit of approximately 6 pmol. Parallel experiments were successfully carried out to analyze adenovirus proteins from Cu-stained gels. It was observed that increase in laser intensity resulted in significant improvements in the quality of MALDI mass spectra for the analysis of inefficiently destained proteins from Cu-stained gels.

MeSH terms

  • Adenoviridae / chemistry*
  • Coloring Agents
  • Copper
  • Electrophoresis, Polyacrylamide Gel
  • Molecular Weight
  • Rosaniline Dyes
  • Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
  • Viral Proteins / chemistry*
  • Viral Proteins / isolation & purification

Substances

  • Coloring Agents
  • Rosaniline Dyes
  • Viral Proteins
  • Copper
  • coomassie Brilliant Blue