Quantitative protein expression analysis of CLL B cells from mutated and unmutated IgVH subgroups using acid-cleavable isotope-coded affinity tag reagents

David R. Barnidge, Diane F. Jelinek, David C. Muddiman, Neu E. Kay

Research output: Contribution to journalArticlepeer-review

15 Scopus citations

Abstract

Relative protein expression levels were compared in leukemic B cells from two patients with chronic lymphocytic leukemia (CLL) having either mutated (M-CLL) or unmutated (UM-CLL) immunoglobulin variable heavy chain genes (IgVH). Cells were separated into cytosol and membrane protein fractions then labeled with acid-cleavable ICAT reagents (cICAT). Labeled proteins were digested with trypsin then subjected to SCX and affinity chromatography followed by LC-ESI-MS/MS analysis on a linear ion trap mass spectrometer. A total of 9 proteins from the cytosol fraction and 4 from the membrane fraction showed a 3-fold or greater difference between M-CLL and UM-CLL and a subset of these were examined by Western blot where results concurred with cICAT abundance ratios. The abundance of one of the proteins in particular, the mitochondrial membrane protein cytochrome c oxidase subunit COX G was examined in 6 M-CLL and 6 UM-CLL patients using western blot and results showed significantly greater levels (P < 0.001) in M-CLL patients vs UM-CLL patients. These results demonstrate that stable isotope labeling and mass spectrometry can complement 2D gel electrophoresis and gene microarray technologies for identifying putative and perhaps unique prognostic markers in CLL.

Original languageEnglish (US)
Pages (from-to)1310-1317
Number of pages8
JournalJournal of Proteome Research
Volume4
Issue number4
DOIs
StatePublished - Jul 2005

Keywords

  • B cell
  • CLL
  • Linear ion trap mass spectrometer
  • M-CLL
  • Quantitative comparison
  • UM-CLL
  • cICAT

ASJC Scopus subject areas

  • Biochemistry
  • Chemistry(all)

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