Assessment of the 2-D Gel-Based Proteomics Application of Clinically Archived Formalin-Fixed Paraffin Embedded Tissues
- 260 Downloads
Hospital tissue repositories possess a vast and valuable supply of disease samples with matched retrospective clinical information. Detection and characterization of disease biomarkers in formalin-fixed paraffin-embedded (FFPE) tissues will greatly aid the understanding of the diseases mechanisms and help in the development of diagnostic and prognostic markers. In this study, the possibility of using full-length proteins extracted from clinically archived FFPE tissues in two-dimensional (2-D) gel-based proteomics was evaluated. The evaluation was done based on two types of tumor tissues (breast and prostate) and two extraction protocols. The comparison of the 2-D patterns of FFPE extracts obtained by two extraction protocols with the matching frozen tissue extracts showed that only 7–10 % of proteins from frozen tissues can be matched to proteins from FFPE tissues. Most of the spots in the 2-D FFPE’s maps had pl 4–6, while the percentages of proteins with pl above 6 were 3–5 times lower in comparison to the fresh/frozen tissue. Despite the three-fold lower number of the detected spots in FFPE maps compared to matched fresh/frozen maps, 67–78 % of protein spots in FFPE could not be matched to the corresponding spots in the fresh/frozen tissue maps indicating irreversible protein modifications. In conclusion, the inability to completely reverse the cross-linked complexes and overcome protein fragmentation with the present day FFPE extraction methods stands in the way of effective use of these samples in 2-D gel based proteomics studies.
KeywordsFormalin-fixed paraffin-embedded (FFPE) Proteomics Two-dimensional polyacrylamide gel electrophoresis (2-D PAGE) Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) Protein extraction
- 2-D DIGE
Two-dimensional difference in-gel electrophoresis
- 2-D PAGE
Two-dimensional polyacrylamide gel electrophoresis
Sodium dodecyl sulfate
Sodium dodecyl sulfate polyacrylamide gel electrophoresis
Many thanks to Dr. Katerina Kubelka-Sabit and Dr. Vanja Filipovski from Clinical Hospital “Sistina”, Skopje, for providing the fresh and FFPE tissues used in this study.
The study has been approved by the Ethics Committee of the Macedonian Academy of Sciences and Arts.
Conflict of interest
The authors have declared no conflict of interests.
- 21.Nirmalan NJ, Hughes C, Peng J, McKenna T, Langridge J, Cairns DA, Harnden P, Selby PJ, Banks RE (2011) Initial development and validation of a novel extraction method for quantitative mining of the formalin-fixed, paraffin-embedded tissue proteome for biomarker investigations. J Proteome Res 10(2):896–906CrossRefGoogle Scholar
- 28.Sprung RW Jr, Brock JW, Tanksley JP, Li M, Washington MK, Slebos RJ, Liebler DC (2009) Equivalence of protein inventories obtained from formalin-fixed paraffin-embedded and frozen tissue in multidimensional liquid chromatography-tandem mass spectrometry shotgun proteomic analysis. Mol Cell Proteomics 8(8):1988–1998CrossRefGoogle Scholar