Subject Area物理化学
Enrichment of Phosphopeptides by a Novel SAX Capillary Monolithic Column for Sensitive MALDI-TOF MS Analysis
Dong MM(董铭铭); Wu MH(吴明火); Wang FJ(王方军); Qin HQ(秦洪强); Han GH(韩广辉); Dong J(董靖); Wu RA(吴仁安); Ye ML(叶明亮); Liu Z(刘震); Zou HF(邹汉法)
Conference NameThe 24th International Symposium on Micro-scale Bioseparation,
Conference Date2009-10-19
Conference Place中国
Funding Organization大连化学物理研究所
AbstractProtein phosphorylation is one of the most biologically relevant and ubiquitous post-translational modifications. MALDI-TOF MS is a powerful tool for the analysis of protein phosphorylation by detection of phosphopeptides in phosphoprotein digest. However, if protein digest is directly submitted to MALDI-TOF MS analysis, the sensitivity for detection of phosphopeptides is very poor due to the ion suppression by nonphosphorylated peptides. So the isolation and enrichment of phosphopeptides prior to MS analysis is crucial for protein phosphorylation analysis. Here, a SAX organic-silica hybrid capillary monolith was prepared to enrich phosphopeptides from protein digest for sensitive MALDI-TOF MS analysis. The SAX monolithic column was prepared by a novel “one-pot” process by concurrently using organic monomers and alkoxysilanes. As the optimal loading and elution buffers for SAX chromatography of phosphopeptides are basic and acidic solutions, respectively, the protein digests without any pretreatment could be loaded onto the SAX column under basic pH conditions, after removing non-phosphopeptides by washing, the bound phosphopeptides were eluted with 5% formic acid directly onto a MALDI target and analyzed by MALDI-TOF MS. Above approach significantly reduced the sample loss and simplified the analytical procedures. Because of the excellent MS detection compatible procedure and the nanoscale SAX column, the detection limit of 50 amol for detection of phosphopeptides from β-casein digest was achieved.
Document Type会议论文
Corresponding AuthorZou HF(邹汉法)
Recommended Citation
GB/T 7714
Dong MM,Wu MH,Wang FJ,et al. Enrichment of Phosphopeptides by a Novel SAX Capillary Monolithic Column for Sensitive MALDI-TOF MS Analysis[C],2009:259/1.
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