title: Shotgun Proteomics Analysis of Estrogen Effects in the Uterus Using Two-Dimensional Liquid Chromatography and Tandem Mass Spectrometry.
abstract: Shotgun (gel-free) proteomics is a useful approach to perform identification and relative quantification of protein in complex mixtures such as tissue homogenates, biological fluids, cell lysates, and extracellular proteins. Incorporation of separative and analytical techniques such as two-dimensional liquid chromatography at nanoscale (2D-nanoLC) coupled to tandem mass spectrometry (MS/MS analysis) into the shotgun protocol provides an excellent strategy. This chapter describes the application of the shotgun proteomics protocol to evaluate the identity and expression analysis of proteins from rat uterus after estrogen (ethinylestradiol) treatment. The steps of the protocol involve sample preparation (digestion), 2D-nanoLC-MS/MS analysis, and shotgun proteomics analysis including bioinformatics tools for data conversion, organization, and interpretation.
authors.affiliation: Division of Basic Biomedical Sciences, Sanford School of Medicine, University of South Dakota, 414 E. Clark St., Vermillion, SD, 57069, USA. eduardo.callegari@usd.edu.
authors.forename: Eduardo A
authors.initials: EA
authors.lastname: Callegari
mesh_terms: D000818:Animals; D002853:Chromatography, Liquid; D019295:Computational Biology; D030562:Databases, Protein; D004967:Estrogens / Q000494:pharmacology*; D004997:Ethinyl Estradiol / Q000494:pharmacology*; D005260:Female; D036103:Nanotechnology; D011506:Proteins / Q000378:metabolism*; D040901:Proteomics / Q000379:methods*; D051381:Rats; D053719:Tandem Mass Spectrometry; D014599:Uterus / Q000187:drug effects* / Q000378:metabolism; D057188:Workflow
chemical_list: D004967:Estrogens; D011506:Proteins; D004997:Ethinyl Estradiol
keywords: Estrogen; Liquid chromatography; Mass spectrometry; Peptides; Proteins; Proteomics; Rat; Uterus; “Shotgun” proteomics