Ottawa Institute of Systems Biology, Faculty of Medicine, University of Ottawa, 451 Smyth Road, Ottawa, Ontario, Canada.
Talanta. 2010 Feb 15;80(4):1526-31. doi: 10.1016/j.talanta.2009.07.018. Epub 2009 Jul 14.
We developed a new method consisting of the proteomic reactor coupled with step pH fractionation for the analysis of low-abundance proteins from minute amount of sample. These new reactors were implemented using both SAX and SCX materials. The pH fractions from the SAX reactor provided higher peptide and protein identification than SCX reactor and conventional solution digestion. Interestingly, the physical characteristics (pI, molecular weight, missed cleavage site and grand average hydrophobicity (GRAVY) index, and number of acid and basic amino acid) of the peptides obtained from the SAX and SCX proteomic reactors are drastically different. Furthermore, nearly half of the peptides observed from the pH fractionations from the SAX reactor are of low abundance while only 22% low-abundance proteins are observed with conventional in-solution digestion following 2D LC-MS/MS analysis.
我们开发了一种新方法，该方法由蛋白质组学反应器与分步 pH 分级相结合，用于从小量样本中分析低丰度蛋白质。这些新的反应器都使用 SAX 和 SCX 材料实现。与 SCX 反应器和常规溶液消化相比，来自 SAX 反应器的 pH 级分提供了更高的肽和蛋白质鉴定率。有趣的是，从 SAX 和 SCX 蛋白质组学反应器获得的肽的物理特性（等电点、分子量、缺失切割位点和平均疏水性（GRAVY）指数、酸性和碱性氨基酸的数量）有很大的不同。此外，从 SAX 反应器的 pH 分级中观察到的近一半肽是低丰度的，而只有 22%的低丰度蛋白质是通过传统的 2D LC-MS/MS 分析后在溶液中消化观察到的。
