Recovering the proteome of archived biomedical specimens
Master of Science
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While several proteomic studies have undertaken the challenge of analyzing formalin fixed paraffin embedded (FFPE) tissue, few have developed optimized protocols for studying FFPE human brain tissue. This study will examine if an improved SDS free proteomic method applied to FFPE human brain tissue will produce a greater number of identified proteins then assess optimized methods to produce reliable, robust and reproducible results from this tissue. A modified guanidinium hydrochloride based extraction method was employed and the extracted proteins were analyzed using LC-MS/MS, for shotgun proteomics. The FFPE human brain tissue samples were acquired from archived specimens at the Griffith University School of Anatomy. By analyzing tissue samples from human FFPE brain, we evaluated conditions used for tissue lysis, digestion, and the effects of the fixation time and storage duration on the number of proteins obtained during shotgun proteomic analysis on FFPE tissue. Comparing the identified proteins and experimental procedures to the existing FFPE tissue studies in the literature, the limitations due to protein quality were identified, optimized processes proposed, and future studies including multiomic analysis of the human brain, were outlined.