Preparation of Proteins and Peptides for Mass Spectrometry Analysis in a Bottom-up Proteomics Workflow

Overview

Journal Curr Protoc Mol Biol

Specialty Molecular Biology

Date 2009 Oct 10

PMID 19816929

Citations 118

Authors

Rebekah L Gundry

Melanie Y White

Christopher I Murray

Lesley A Kane

Qin Fu

Brian A Stanley

Jennifer E Van Eyk

Affiliations

Soon will be listed here.

Abstract

This unit outlines the steps required to prepare a sample for MS analysis following protein separation or enrichment by gel electrophoresis, liquid chromatography, and affinity capture within the context of a bottom-up proteomics workflow in which the protein is first broken up into peptides, either by chemical or enzymatic digestion, prior to MS analysis. Also included are protocols for enrichment at the peptide level, including phosphopeptide enrichment and reversed-phase chromatography for sample purification immediately prior to MS analysis. Finally, there is a discussion regarding the types of MS technologies commonly used to analyze proteomics samples, as well as important parameters that should be considered when analyzing the MS data to ensure stringent and robust protein identifications and characterization.

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