Based On The Human Plasma Proteome Of Immune Affinity Separation Technology

Human blood is a large repository of proteins, containing thousands of distinct proteins secreted, shed, or lost from cells, tissues and organs throughout the body. Plasma proteins often serve as indicators of disease and are a rich source for biomarker discovery. So there has been significant interest in comprehensively characterizing the protein content of plasma. However, plasma proteins have unique characteristic that the 12 or greater orders of magnitude in protein concentration limit the ability of mass spectrometry to effectively monitor the low abundant species. The intrinsic large dynamic range of plasma proteins makes the proteomic analysis very challenging. Hence, how to deplete the high abundant proteins and enrich the low abundant proteins (i.e., eliminate the interference of high abundant proteins to low abundant proteins) is crucial and a highlight in plasma proteome research. At present the immunoaffinity depletion technology for the most abundant proteins is accepted and applied widely by the researchers. But as the high abundant proteins are transporters or binders, immunoaffinty depletion may cause some proteins lost, which is the issue that researchers generally pay attention to and our research mainly focus on. In addition, the reason for the bound nonspecifically of the proteins to the column are explored and analyzed. Based on the immunoaffinity separation technology, not immunoaffinity depletion, the more comprehensive human plasma proteome study fulfilled, which approved the new strategy for large scale plasma proteome analysis.In the present study, two different immunoaffinity fractionation columns (Agilent, MARS; Genway, Seppro) for the top-6 or the top-12 proteins in plasma were investigated and both the proteins in column-bound ( MARS-BF and SepproTM-BF ) and flow-through fractions (MARS-FF and SepproTM-FF) were subsequently analyzed. A two-dimensional peptide separation strategy, utilizing chromatographic separation techniques, combined with tandem mass spectrometry (MS/MS) was employed for proteomic analysis of the four fractions. Using the established HUPO PPP criteria, a total of 2401 unique plasma proteins were identified. When more stringent criteria, based on searching against the reversed database, were implemented,...