Simplified high-sensitivity sequencing of a major histocompatibility complex class i-associated immunoreactive peptide using matrix-assisted laser desorption/ionization mass spectrometry

Amina S. Woods, Alex Y.C. Huang, Robert J. Cotter, Gary R. Pasternack, Drew M. Pardoll, Elizabeth M. Jaffee

Research output: Contribution to journalArticle

Abstract

Cytotoxic T cells (CTL) are known to recognize small peptide fragments of cytoplasmic proteins bound to major histocompatibility complex (MHC) class I molecules on cell surfaces. Recent work indicates that tumor antigens are processed and presented in a manner similar to viral antigens. Identification of the peptides recognized by tumor-specific CTL would provide valuable information about their parent proteins, as well as allowing for the development of recombinant antigen-specific tumor vaccines. While highly represented MHC-bound peptides have been routinely purified by reversed-phase HPLC for Edman degradation sequencing, identification and sequencing of infrequent peptides that represent the biologically relevant targets of tumor-specific CTL have proved elusive. We have combined matrix-assisted laser desorption/ionization mass spectrometry with on-slide exopeptidase digestion to successfully identify and directly sequence a model tumor-specific peptide antigen derived from an integrated viral gene. The enhanced sensitivity of this technique (femtomolar range) allows for the sequencing of specific MHC-bound peptides derived from as few as 1 × 109 cells.

Original languageEnglish (US)
Pages (from-to)15-25
Number of pages11
JournalAnalytical biochemistry
Volume226
Issue number1
DOIs
StatePublished - Mar 1995

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Molecular Biology
  • Cell Biology

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