Combining isoelectric point-based fractionation, liquid chromatography and mass spectrometry to improve peptide detection and protein identification

Stephanie M. Cologna, William K. Russell, Peniel J. Lim, Gyula Vigh, David H. Russell

Research output: Contribution to journalArticle

12 Scopus citations

Abstract

The off-line coupling of an isoelectric trapping device termed membrane separated wells for isoelectric focusing and trapping (MSWIFT) to mass spectrometry-based proteomic studies is described. The MSWIFT is a high capacity, high-throughput, mass spectrometry-compatible isoelectric trapping device that provides isoelectric point (pI)-based separations of complex mixtures of peptides. In MSWIFT, separation and analyte trapping are achieved by migrating the peptide ions through membranes having fixed pH values until the peptide pI is bracketed by the pH values of adjacent membranes. The pH values of the membranes can be tuned, thus affording a high degree of experimental flexibility. Specific advantages of using MSWIFT for sample prefractionation include: (1) small sample volumes (~200 μL), (2) customized membranes over a large pH range, (3) flexibility in the number of desired fractions, (4) membrane compatibility with a variety of solvents systems, and (5) resulting fractions do not require sample cleanup before MS analysis. Here, we demonstrate the utility of MSWIFT for mass spectrometry-based detection of peptides in improving dynamic range and the reduction of ion suppression effects for high-throughput separations of tryptic peptides.

Original languageEnglish (US)
Pages (from-to)1612-1619
Number of pages8
JournalJournal of the American Society for Mass Spectrometry
Volume21
Issue number9
DOIs
StatePublished - Sep 1 2010
Externally publishedYes

ASJC Scopus subject areas

  • Structural Biology
  • Spectroscopy

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