Quantification of β-catenin signaling components in colon cancer cell lines, tissue sections, and microdissected tumor cells using reaction monitoring mass spectrometry

Yi Chen, Mike Gruidl, Elizabeth Remily-Wood, Richard Z. Liu, Steven Eschrich, Mark Lloyd, Aejaz Nasir, Marilyn M. Bui, Emina Huang, David Shibata, Timothy Yeatman, John M. Koomen

Research output: Contribution to journalArticlepeer-review

42 Scopus citations

Abstract

Reaction monitoring mass spectrometry has emerged as a powerful tool for targeted detection and quantification of proteins in clinical samples. Here, we report the use of gel electrophoresis for protein fractionation and liquid chromatography coupled to multiple reaction monitoring mass spectrometry (LC-MRM) screening for quantitative analysis of components from the Wnt/β-catenin signaling pathway, which contributes to colon tumor formation and progression. In silico tools are used to design LC-MRM screens for each target protein. Following successful peptide detection, stable isotope labeled peptides are synthesized and developed as internal standards. Then, the assays are implemented in colon cancer cell lines to achieve detection in minimal amounts of cells, compatible with direct translation to clinical specimens. Selected assays are compared with qualitative results from immunoblotting (Westerns) and translated to individual frozen colon tissue sections and laser capture microdissected tumor cells. This LC-MRM platform has been translated from in vitro models to clinical specimens, forming the basis for future experiments in patient assessment.

Original languageEnglish (US)
Pages (from-to)4215-4227
Number of pages13
JournalJournal of Proteome Research
Volume9
Issue number8
DOIs
StatePublished - Aug 6 2010
Externally publishedYes

Keywords

  • biomarker assessment
  • colon cancer
  • frozen tissue
  • laser capture microdissection
  • liquid chromatography coupled to multiple reaction monitoring
  • Wnt/β-catenin signaling

ASJC Scopus subject areas

  • Biochemistry
  • Chemistry(all)

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