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Novel methods for proteomics analysis of formalin-fixed, paraffin-embedded tissues (FFPE), and their application for biomarker discovery

Tanca, Alessandro (2010) Novel methods for proteomics analysis of formalin-fixed, paraffin-embedded tissues (FFPE), and their application for biomarker discovery. Doctoral Thesis.

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Abstract

A wealth of information on proteins involved in many aspects of disease is encased within formalin-fixed paraffin-embedded (FFPE) tissue repositories stored in hospitals worldwide. However, FFPE protein extracts, as described to date, often exhibit a low pattern complexity, and a poor suitability for downstream gel-based proteomic techniques. Thus, an optimised method for extraction of full-length proteins from FFPE tissues was developed. The results obtained analysing FFPE muscle, liver, and thyroid extracts, by GeLC-MS/MS, western immunoblotting, protein arrays, and ELISA, are presented and discussed. Moreover, 2D-PAGE-MS and 2D-DIGE-MS analyses of proteins extracted from fixed skeletal muscle and liver tissues are reported. Finally, the application of 2D-DIGE-MS and GeLC-MS/MS for differential proteomic investigation of FFPE diseased samples was pursued. First, a proteomic comparison between sheep pathological liver samples was carried out, and several stress biomarkers were detected. Subsequently, a thorough biomarker discovery study was conducted, analysing human lung neuroendocrine cancer tissues. GeLC-MS/MS analysis of 3 typical carcinoid and 3 small cell lung carcinoma cases led to the identification of over 400 unique proteins per disease class. According to statistical analysis, a panel of over 30 differentially expressed putative biomarkers is presented. In addition, also 2D-DIGE-MS data clearly support biomarkers identification.

Item Type:Doctoral Thesis
ID Code:3900
Publisher:Universita' degli studi di Sassari
Uncontrolled Keywords:FFPE, proteomics, 2D-DIGE, biomarker discovery
Subjects:Area 05 - Scienze biologiche > BIO/10 Biochimica
Area 06 - Scienze mediche > MED/07 Microbiologia e microbiologia clinica
Area 05 - Scienze biologiche > BIO/12 Biochimica clinica e biologia molecolare clinica
Divisions:001 Università di Sassari > 01 Dipartimenti > Scienze fisiologiche, biochimiche e cellulari
Cicli, scuole e corsi:Ciclo 22 > Scuole > Scienze biomolecolari e biotecnologiche > Biotecnologie molecolari e cellulari
Deposited On:18 May 2010 12:53

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