Unit

UNIT 10.27 Extraction of Proteins from Formalin-Fixed, Paraffin-Embedded Tissue Using the Qproteome Extraction Technique and Preparation of Tryptic Peptides for Liquid Chromatography/Mass Spectrometry Analysis

  1. Thibault Geoui1,
  2. Henning Urlaub2,
  3. Uwe Plessmann2,
  4. Peter Porschewski1

Published Online: 1 APR 2010

DOI: 10.1002/0471142727.mb1027s90

Current Protocols in Molecular Biology

Current Protocols in Molecular Biology

How to Cite

Geoui, T., Urlaub, H., Plessmann, U. and Porschewski, P. 2010. Extraction of Proteins from Formalin-Fixed, Paraffin-Embedded Tissue Using the Qproteome Extraction Technique and Preparation of Tryptic Peptides for Liquid Chromatography/Mass Spectrometry Analysis. Current Protocols in Molecular Biology. 90:VI:10.27:10.27.1–10.27.12.

Author Information

  1. 1

    QIAGEN GmbH, Hilden, Germany

  2. 2

    Max Planck Institute for Biophysical Chemistry, Göttingen, Germany

Publication History

  1. Published Online: 1 APR 2010
  2. Published Print: APR 2010

Abstract

This unit provides a robust, reliable, and easy-to-use kit-based method for extraction of intact, non-degraded proteins from formalin-fixed, paraffin-embedded (FFPE) tissue, and their subsequent use for analysis by liquid chromatography/mass spectrometry (LC/MS). After deparaffinization, proteins are extracted from unstained sections of FFPE rat liver tissue. After a simple cleanup step using organic extraction, the sample is transferred into a buffer optimized for trypsin digestion of the extracted proteins. Subsequently, LC/MS is used to identify the proteins that gave rise to the tryptic peptides. Comparing formalin-fixed and frozen tissues, good correlation is observed in the mass spectrometric pattern attributable to the tryptic peptides and number of identified proteins. Since FFPE tissues are generally available in clinical practice, this method can be used to analyze biomarkers in different pathological situations (e.g., healthy vs. diseased). The method can also be used for protein extraction from fresh-frozen tissue. Curr. Protoc. Mol. Biol. 90:10.27.1-10.27.12. © 2010 by John Wiley & Sons, Inc.

Keywords:

  • formalin-fixed tissue;
  • mass spectrometry;
  • proteomics;
  • protein extraction