title: Improved mass spectrometric proteomic profiling of the secretome of rat vascular endothelial cells.
abstract: Serum albumin contamination of cells cultured in vitro significantly impedes the mass spectrometric analysis of proteins secreted by the cells. Here we report a novel washing and culturing technique for rat vascular endothelial cells that considerably reduces the concentration of the commonly used additive for cell culture, bovine serum albumin (BSA), in the secretome of these cells. Cells are rinsed stringently and cultured for 24 h in serum-free media without appreciably impeding cell growth or viability. The percentage of BSA scans identified by tandem mass spectrometry (LC-MS/MS) in stringently rinsed cells (average 13.2%) was significantly lower than either the moderately rinsed or no rinse cell treatments (average 35.2% and 45.2% respectively). Furthermore, the stringent wash treatment allowed the confident identification of a larger portion of the secretome of rat endothelial cells by LC-MS/MS.
authors.affiliation: National Center for Proteomics Research, Biotechnology and Bioengineering Center, Medical College of Wisconsin, Milwaukee, Wisconsin 53226, USA.
authors.forename: M C
authors.initials: MC
authors.lastname: Pellitteri-Hahn
authors.forename: M C
authors.initials: MC
authors.lastname: Warren
authors.forename: D N
authors.initials: DN
authors.lastname: Didier
authors.forename: E L
authors.initials: EL
authors.lastname: Winkler
authors.forename: S P
authors.initials: SP
authors.lastname: Mirza
authors.forename: A S
authors.initials: AS
authors.lastname: Greene
authors.forename: M
authors.initials: M
authors.lastname: Olivier
mesh_terms: D000818:Animals; D002417:Cattle; D018929:Cell Culture Techniques; D002478:Cells, Cultured; D003470:Culture Media / Q000737:chemistry; D042783:Endothelial Cells / Q000737:chemistry* / Q000378:metabolism; D004730:Endothelium, Vascular / Q000166:cytology; D013058:Mass Spectrometry; D020543:Proteome / Q000032:analysis*; D040901:Proteomics / Q000379:methods*; D051381:Rats; D012710:Serum Albumin, Bovine / Q000032:analysis
chemical_list: D003470:Culture Media; D020543:Proteome; D012710:Serum Albumin, Bovine