Protein Extraction from Formalin-fixed, Paraffin-embedded Tissue Sections: Quality Evaluation by Mass Spectrometry

Shan-Rong Shi ¹, Cheng Liu ¹, Brian M. Balgley ¹, Cheng Lee ¹ and Clive R. Taylor ^1*

¹ Department of Pathology, University of Southern California Keck School of Medicine, Los Angeles, California (S-RS,CL,CRT), and Calibrant Biosystems, Gaithersburg, Maryland (BMB,CL)

^* To whom correspondence should be addressed. E-mail: taylor{at}pathfinder.hsc.usc.edu .

Submitted on October 4, 2005
Accepted on 27 November 2005

	Abstract

A satisfactory protocol of protein extraction has been established based on the heat-induced antigen retrieval (AR) technique that has been widely applied in immunohistochemistry for archival formalin-fixed, paraffin-embedded (FFPE) tissue sections since early 1990s. Based upon the basic principle of AR, an initial serial experiment to identify an optimal protocol of heat-induced protein extraction was carried out using FFPE mouse tissues. Based on these experiments the optimal protocol for extraction of proteins was then performed on an archival FFPE tissue of human renal carcinoma. Five 10 µm-FFPE sections were boiled in a retrieval solution of Tris-HCl containing 2% sodium dodecyl sulfate (SDS) for 20 min, followed by incubation at 60°C for 2 hr. Fresh tissue taken from the same case of renal carcinoma was processed for extraction of proteins by a conventional method using RIPA solution, to compare the efficiency of protein extraction from FFPE tissue sections with extraction from fresh tissue. In addition, further sections of the same FFPE sample were processed by the same procedure without heating treatment, as a control. Evaluation of the quality of protein extracted from FFPE tissue was done using gel electrophoresis and mass spectrometry, showing most identified proteins extracted from FFPE tissue sections were overlapped with those extracted from fresh tissue.

Key Words: protein extraction, antigen retrieval, formalin-fixed, paraffin-embedded tissue, mass spectrometry

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