Protein extraction from formalin-fixed, paraffin-embedded tissue sections: quality evaluation by mass spectrometry.
Author(s): Shi SR, Liu C, Balgley BM, Lee C, Taylor CR
Publication: J Histochem Cytochem, 2006, Vol. 54, Page 739-43
PubMed ID: 16399996 PubMed Review Paper? No
Purpose of Paper
Conclusion of Paper
Studies
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Study Purpose
The purpose of this study was to compare protein banding patterns, observed via SDS-PAGE analysis, of fresh frozen and FFPE specimens subjected to antigen retrieval protocols differing in solution pH and temperature.
Summary of Findings:
SDS protein profiles were comparable among freshly frozen specimens and FFPE specimens subjected to heat-induced antigen retrieval in a Tris-HCl buffer containing 2% SDS, with comparable results observed at the two solution pHs investigated. However, antigen retrieval at room temperature failed to produce observable protein bands.
Biospecimens
Preservative Types
- Formalin
- Frozen
Diagnoses:
- Neoplastic - Carcinoma
Platform:
Analyte Technology Platform Protein 1D/2D gels Pre-analytical Factors:
Classification Pre-analytical Factor Value(s) Analyte Extraction and Purification Temperature of heat-induced retrieval 100 degrees C
Room temperature
Analyte Extraction and Purification Antigen retrieval Tris-HCl buffer with SDS, pH 7
Tris-HCl buffer with SDS, pH 9
Biospecimen Preservation Type of fixation/preservation Formalin (buffered)
Snap frozen
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Study Purpose
The purpose of this study was to compare protein profiles obtained via capillary isoelectric focusing (CIEF) nano-reverse-phase liquid chromatography (RPLC)/ mass spectrometry among freshly frozen and FFPE specimens subjected to antigen retrieval protocols differing in solution pH and temperature.
Summary of Findings:
The majority of proteins identified were common among the differentially preserved specimens and antigen retrieval methods investigated, with 84% and 72% of proteins identified in fresh specimens also observed in FFPE specimens that underwent heat-induced retrieval in a solution of pH 7 or 9, respectively. FFPE specimens subjected to retrieval at room temperature yielded fewer identifiable proteins, representing a commonality of 54% with fresh frozen specimens. Distribution of identified proteins according to their identified subcellular locale was similar among preservation and antigen retrieval methods.
Biospecimens
Preservative Types
- Formalin
- Frozen
Diagnoses:
- Neoplastic - Carcinoma
Platform:
Analyte Technology Platform Protein LC-MS or LC-MS/MS Protein Capillary electrophoresis-MS Pre-analytical Factors:
Classification Pre-analytical Factor Value(s) Biospecimen Preservation Type of fixation/preservation Formalin (buffered)
Snap frozen
Analyte Extraction and Purification Temperature of heat-induced retrieval 100 degrees C
Room temperature
Analyte Extraction and Purification Antigen retrieval Tris-HCl buffer with SDS, pH 7
Tris-HCl buffer with SDS, pH 9