Microproteomic analysis of 10,000 laser captured microdissected breast tumor cells using short-range sodium dodecyl sulfate-polyacrylamide gel electrophoresis and porous layer open tubular liquid chromatography tandem mass spectrometry.

ABSTRACT: Precise proteomic profiling of limited levels of disease tissue represents an extremely challenging task. Here, we present an effective and reproducible microproteomic workflow for sample sizes of only 10,000 cells that integrates selective sample procurement via laser capture microdissection (LCM), sample clean-up and protein level fractionation using short-range SDS-PAGE, followed by ultrasensitive LC-MS/MS analysis using a 10 μm i.d. porous layer open tubular (PLOT) column. With 10,000 LCM captured mouse hepatocytes for method development and performance assessment, only 10% of the in-gel digest, equivalent to ∼1000 cells, was needed per LC-MS/MS analysis. The optimized workflow was applied to the differential proteomic analysis of 10,000 LCM collected primary and metastatic breast canc

SUBMITTER: Thakur D 

PROVIDER: S-EPMC3205921 | biostudies-literature | 2011 Nov

REPOSITORIES: biostudies-literature