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Profiling Solid Tumor Heterogeneity by LCM and Biological MS of Fresh-Frozen Tissue Sections

  • Donald J. Johann
  • Sumana Mukherjee
  • DaRue A. Prieto
  • Timothy D. Veenstra
  • Josip BlonderEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 755)

Abstract

The heterogeneous nature of solid tumors represents a common problem in mass spectrometry (MS)-based analysis of fresh-frozen tissue specimens. Here, we describe a method that relies on synergy between laser capture microdissection (LCM) and MS for enhanced molecular profiling of solid tumors. This method involves dissection of homogeneous histologic cell types from thin fresh-frozen tissue sections via LCM, coupled with liquid chromatography (LC)-MS analysis. Such an approach enables an in-depth molecular profiling of captured cells. This is a bottom-up proteomic approach, where proteins are identified through peptide sequencing and matching against a specific proteomic database. Sample losses are minimized, since lysis, solubilization, and digestion are carried out directly on LCM caps in buffered methanol using a single tube, thus reducing sample loss between these steps. The rationale for the LCM-MS coupling is that once the optimal method parameters are established for a solid tumor of interest, homogeneous histologic tumor/tissue cells (i.e., tumor proper, stroma, etc.) can be effectively studied for potential biomarkers, drug targets, pathway analysis, as well as enhanced understanding of the pathological process under study.

Key words

Thin fresh-frozen tissue sections Laser capture microdissection Liquid chromatography-mass spectrometry Solid tumor heterogeneity Biomarker Cancer 

Notes

Acknowledgments

This project was funded in whole or in part with federal funds from the National Cancer Institute, National Institutes of Health, under contract HSN261200800001E. The content of this publication does not necessarily reflect the views or policies of the Department of Health and Human Services, nor does the mention of trade names, commercial products, or organizations implies endorsement by the US Government.

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Copyright information

© Springer Science+Business Media, LLC 2011

Authors and Affiliations

  • Donald J. Johann
    • 2
  • Sumana Mukherjee
    • 2
  • DaRue A. Prieto
    • 1
  • Timothy D. Veenstra
    • 1
  • Josip Blonder
    • 1
    Email author
  1. 1.Laboratory of Proteomics and Analytical TechnologiesSAIC-Frederick, Inc., National Cancer Institute at FrederickFrederickUSA
  2. 2.Medical Oncology Branch, Center for Cancer ResearchNational Cancer Institute, National Institutes of HealthBethesdaUSA

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