大鼠血管内皮细胞分泌蛋白质组的质谱蛋白质组分析方法改进
Improved mass spectrometric proteomic profiling of the secretome of rat vascular endothelial cells.
作者信息
Pellitteri-Hahn M C, Warren M C, Didier D N, Winkler E L, Mirza S P, Greene A S, Olivier M
机构信息
National Center for Proteomics Research, Biotechnology and Bioengineering Center, Medical College of Wisconsin, Milwaukee, Wisconsin 53226, USA.
出版信息
J Proteome Res. 2006 Oct;5(10):2861-4. doi: 10.1021/pr060287k.
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.
摘要
体外培养细胞时血清白蛋白的污染会显著阻碍对细胞分泌蛋白的质谱分析。在此，我们报告一种针对大鼠血管内皮细胞的新型洗涤和培养技术，该技术可大幅降低这些细胞分泌蛋白组中细胞培养常用添加剂牛血清白蛋白（BSA）的浓度。细胞经过严格冲洗后，在无血清培养基中培养24小时，而不会明显阻碍细胞生长或活力。通过串联质谱（LC-MS/MS）鉴定，严格冲洗的细胞中BSA扫描的百分比（平均13.2%）显著低于中度冲洗或未冲洗的细胞处理组（分别平均为35.2%和45.2%）。此外，严格洗涤处理使得通过LC-MS/MS能够更可靠地鉴定大鼠内皮细胞分泌蛋白组中的更大部分。
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