title: Determining protein subcellular localization in mammalian cell culture with biochemical fractionation and iTRAQ 8-plex quantification.
abstract: Protein subcellular localization is a fundamental feature of posttranslational functional regulation. Traditional microscopy based approaches to study protein localization are typically of limited throughput, and dependent on the availability of antibodies with high specificity and sensitivity, or fluorescent fusion proteins. In this chapter we describe how Localization of Organelle Proteins by Isotope Tagging (LOPIT), a mass spectrometry based workflow coupling biochemical fractionation and iTRAQ™ 8-plex quantification, can be applied for the high-throughput characterization of protein localization in a mammalian cell culture line.
authors.affiliation: Department of Genetics, University of Cambridge, Downing Street, Cambridge, CB2 3EH, UK, ac587@cam.ac.uk.
authors.forename: Andy
authors.initials: A
authors.lastname: Christoforou
authors.forename: Alfonso Martinez
authors.initials: AM
authors.lastname: Arias
authors.forename: Kathryn S
authors.initials: KS
authors.lastname: Lilley
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D002853:Chromatography, Liquid; D008322:Mammals; D011506:Proteins / Q000378:metabolism*; D013347:Subcellular Fractions / Q000378:metabolism*; D053719:Tandem Mass Spectrometry
chemical_list: D011506:Proteins