title: Sample Preparation for Mass Spectrometry-Based Proteomics; from Proteomes to Peptides.
abstract: Mass spectrometry (MS) has become the predominant technology to analyze proteins due to it ability to identify and characterize proteins and their modifications with high sensitivity and selectivity (Aebersold and Mann, Nature 422(6928):198-207, 2003; Han et al., Curr Opin Chem Biol 12(5):483-490, 2008). While mass spectrometry instruments have improved rapidly over the past couple of decades, mass spectrometry results have remained largely dependent on sample preparation and quality. Sample ionization and mass measurements are susceptible to a wide variety of interferences, including buffers, salts, polymers, and detergents. These contaminants also impair MS system performance, often requiring time consuming maintenance or costly repairs to restore function. The goal of this chapter is to describe the rationale, considerations, and general techniques used to prepare samples for proteomic mass spectrometry analysis.
authors.affiliation: Thermo Fisher Scientific, Rockford, IL, USA. john.rogers@thermofisher.com.
authors.forename: John C
authors.initials: JC
authors.lastname: Rogers
authors.affiliation: Thermo Fisher Scientific, Rockford, IL, USA.
authors.forename: Ryan D
authors.initials: RD
authors.lastname: Bomgarden
mesh_terms: D002458:Cell Fractionation; D002853:Chromatography, Liquid; D006367:HeLa Cells; D057166:High-Throughput Screening Assays; D006801:Humans; D010455:Peptides / Q000302:isolation & purification*; D011506:Proteins / Q000302:isolation & purification*; D059748:Proteolysis; D020543:Proteome; D040901:Proteomics / Q000379:methods*; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D010455:Peptides; D011506:Proteins; D020543:Proteome
keywords: Digestion; Digestion comparison; Filter-assisted sample preparation (FASP); In-gel digestion; In-solution digestion; Lysis; Protein chromatography; Protein depletion or enrichment; Protein extraction