Quantitative Profiling of the Human Substantia Nigra Proteome from Laser-capture Microdissected FFPE Tissue.

Molecular & Cellular Proteomics : MCP
Eva GriesserAngus I Lamond

Abstract

Laser-capture microdissection (LCM) allows the visualization and isolation of morphologically distinct subpopulations of cells from heterogeneous tissue specimens. In combination with formalin-fixed and paraffin-embedded (FFPE) tissue it provides a powerful tool for retrospective and clinically relevant studies of tissue proteins in a healthy and diseased context. We first optimized the protocol for efficient LCM analysis of FFPE tissue specimens. The use of SDS containing extraction buffer in combination with the single-pot solid-phase-enhanced sample preparation (SP3) digest method gave the best results regarding protein yield and protein/peptide identifications. Microdissected FFPE human substantia nigra tissue samples (∼3,000 cells) were then analyzed, using tandem mass tag (TMT) labeling and LC-MS/MS, resulting in the quantification of >5,600 protein groups. Nigral proteins were classified and analyzed by abundance, showing an enrichment of extracellular exosome and neuron-specific gene ontology (GO) terms among the higher abundance proteins. Comparison of microdissected samples with intact tissue sections, using a label-free shotgun approach, revealed an enrichment of neuronal cell type markers, such as tyrosine hydroxyla...Continue Reading

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Citations

Jun 25, 2020·Proteomics·Ferdinand von Eggeling, Franziska Hoffmann
Aug 5, 2020·Annual Review of Cell and Developmental Biology·Pawan Kishor Singh, Miratul M K Muqit

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Methods Mentioned

BETA
Fusion Tribrid
acetylation
laser-capture microdissection
laser-capture proteomics

Software Mentioned

MaxQuant
DataShop
Perseus
Xcalibur
DAVID
Excel
Instant Clue
Volcano Plot

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