Streamlined Protocol for Deep Proteomic Profiling of FAC-sorted Cells and Its Application to Freshly Isolated Murine Immune Cells.

Molecular & Cellular Proteomics : MCP
Samuel A MyersSteven A Carr

Abstract

Proteomic profiling describes the molecular landscape of proteins in cells immediately available to sense, transduce, and enact the appropriate responses to extracellular queues. Transcriptional profiling has proven invaluable to our understanding of cellular responses; however, insights may be lost as mounting evidence suggests transcript levels only moderately correlate with protein levels in steady state cells. Mass spectrometry-based quantitative proteomics is a well-suited and widely used analytical tool for studying global protein abundances. Typical proteomic workflows are often limited by the amount of sample input that is required for deep and quantitative proteome profiling. This is especially true if the cells of interest need to be purified by fluorescence-activated cell sorting (FACS) and one wants to avoid ex vivo culturing. To address this need, we developed an easy to implement, streamlined workflow that enables quantitative proteome profiling from roughly 2 μg of protein input per experimental condition. Utilizing a combination of facile cell collection from cell sorting, solid-state isobaric labeling and multiplexing of peptides, and small-scale fractionation, we profiled the proteomes of 12 freshly isolated, ...Continue Reading

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Citations

Sep 27, 2019·Journal of Leukocyte Biology·Arjun SukumaranJennifer Geddes-McAlister
Mar 19, 2020·Molecular & Cellular Proteomics : MCP·Anne-Claude GingrasAlma L Burlingame
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Jan 17, 2021·Molecular & Cellular Proteomics : MCP·Anne-Claude GingrasAlma L Burlingame
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