Initial development and validation of a novel extraction method for quantitative mining of the formalin-fixed, paraffin-embedded tissue proteome for biomarker investigations

Nirmalan, NJ; Hughes, C; Peng, J; McKenna, T; Langridge, J; Cairns, DA; Harnden, P; Selby, PJ; Banks, RE

Abstract

Annotated formalin-fixed, paraffin-embedded
(FFPE) tissue archives constitute a valuable resource for
retrospective biomarker discovery. However, proteomic
exploration of archival tissue is impeded by extensive
formalin-induced covalent cross-linking. Robust methodology
enabling proteomic profiling of archival resources
is urgently needed. Recent work is beginning to support
the feasibility of biomarker discovery in archival tissues,
but further developments in extraction methods which are
compatible with quantitative approaches are urgently
needed. We report a cost-effective extraction methodology
permitting quantitative proteomic analyses of small
amounts of FFPE tissue for biomarker investigation. This
surfactant/heat-based approach results in effective and
reproducible protein extraction in FFPE tissue blocks. In
combination with a liquid chromatography-mass spectrometry-
based label-free quantitative proteomics methodology,
the protocol enables the robust representative
and quantitative analyses of the archival proteome.
Preliminary validation studies in renal cancer tissues have
identified typically 250-300 proteins per 500 ng of tissue
with 1D LC-MS/MS with comparable extraction in FFPE
and fresh frozen tissue blocks and preservation of tumor/
normal differential expression patterns (205 proteins, r )
0.682; p < 10-15). The initial methodology presented here
provides a quantitative approach for assessing the potential
suitability of the vast FFPE tissue archives as an
alternate resource for biomarker discovery and will allow
exploration of methods to increase depth of coverage and
investigate the impact of preanalytical factors.

Citation

Nirmalan, N., Hughes, C., Peng, J., McKenna, T., Langridge, J., Cairns, D., …Banks, R. (2011). Initial development and validation of a novel extraction method for quantitative mining of the formalin-fixed, paraffin-embedded tissue proteome for biomarker investigations. Journal of Proteome Research, 10(2), 896-906. https://doi.org/10.1021/pr100812d

• Journal Article Type: Article
• Publication Date: Jan 1, 2011
• Deposit Date: Dec 20, 2011
• Journal: Journal of Proteome Research
• Print ISSN: 1535-3893
• Publisher: American Chemical Society
• Peer Reviewed: Peer Reviewed
• Volume: 10
• Issue: 2
• Pages: 896-906
• DOI: https://doi.org/10.1021/pr100812d
• Publisher URL: http://dx.doi.org/10.1021/pr100812d