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

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 i n the characterization of adenovirus proteins. The key steps in the extract ion and destaining process involve washing the excised band with a combinat ion of solvents that include 10% acetic acid, acetonitrile, methanol, and f ormic acid:water: isopropanol mixture. By using this procedure, we determin ed adenovirus proteins with molecular weights ranging from 10,000 to 110,00 0 Da by MALDI-MS, obtaining a detection limit of approximately 6 pmol. Para llel experiments were successfully carried out to analyze adenovirus protei ns from Cu-stained gels. It was observed that increase in laser intensity r esulted in significant improvements in the quality of MALDI mass spectra fo r the analysis of inefficiently destained proteins from Cu-stained gels. (J Am Soc Mass Spectrom 2000, 11, 356-361) (C) 2000 American Society for Mass Spectrometry.