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On‐Tissue Hydrogel‐Mediated Nondestructive Proteomic Characterization: Application to fr/fr and FFPE Tissues and Insights for Quantitative Proteomics Using a Case of Cardiac Myxoma
Author(s) -
Taverna Domenico,
Mignogna Chiara,
Santise Gianluca,
Gaspari Marco,
Cuda Giovanni
Publication year - 2019
Publication title -
proteomics – clinical applications
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.948
H-Index - 54
eISSN - 1862-8354
pISSN - 1862-8346
DOI - 10.1002/prca.201700167
Subject(s) - quantitative proteomics , proteomics , chemistry , trypsin , proteolytic enzymes , multiplex , microbiology and biotechnology , biomedical engineering , pathology , biology , biochemistry , bioinformatics , medicine , enzyme , gene
Purpose The application of a methodology for quantitative protein analysis from formalin‐fixed and paraffin‐embedded (FFPE) tissue by using hydrogels. Miniaturized polymeric gels are placed onto histologically defined tissue regions in order to perform localized digestion for bottom‐up proteomics. Hydrogel‐extracted peptides are then labeled with tandem mass tags (TMT) reagents for relative protein quantification. A cardiac myxoma biopsy is used. Experimental design Multiple hydrogels, incorporating the proteolytic enzyme trypsin, are placed on serial tissue sections, and processed for digestion and TMT derivatization. SCX fractionation before LC‐MS/MS analysis and bioinformatics analysis are carried out. Results Two histologically different areas on both FFPE and frozen sections of the same cardiac myxoma biopsy are compared. In total, 1949 (FFPE) and 2491 (frozen) proteins are identified, with a total overlap of 56%. The quantitative comparison highlighted 15 (FFPE) and 138 (frozen) differentially expressed proteins between myxoma regions. Conclusion The methodology successfully detects numerous protein signals from FFPE and frozen specimens and is able to differentiate between tissue regions. A fast and reliable tissue preparation for quantitative protein analysis by minimum sample manipulation is developed. This offers an option for on‐tissue proteomics analysis while preserving the inherent spatial information on the tissue.