Armenta Jenny M, Gu Binghe, Thulin Craig D, Lee Milton L
Department of Chemistry and Biochemistry, Brigham Young University, Provo, UT 84602-5700, USA.
J Chromatogr A. 2007 Apr 27;1148(1):115-22. doi: 10.1016/j.chroma.2007.02.089. Epub 2007 Mar 2.
A butyl methacrylate-co-ethylene dimethacrylate (BuMA-co-EDMA) monolith was synthesized by UV initiated polymerization at the inlet end of a 75 microm I.D. fused silica capillary that had been previously coated with a protein compatible polymer, poly(vinyl)alcohol. The monolith was used for on-line preconcentration of proteins followed by capillary electrophoresis (CE) separation. For the analysis of standard proteins (cytochrome c, lysozyme and trypsinogen A) this system proved reproducible. The run-to-run %RSD values for migration time and corrected peak area were less than 5%, which is typical of CE. As measured by frontal analysis using lysozyme as solute, saturation of a 1cm monolith was reached after loading 48 ng of protein. Finally, the BuMA-co-EDMA monolithic preconcentrator was coupled to a protein G monolithic column via a zero dead volume union. The coupled system was used for on-line removal of IgG, preconcentration of standard proteins and CE separation. This system could be a valuable sample preparation tool for the analysis of low abundance proteins in complex samples such as human serum, in which high abundance proteins, e.g., human serum albumin (HSA) and immunoglobulin G (IgG), hinder identification and quantification of low abundance proteins.
通过紫外引发聚合反应，在一根内径为75微米、预先涂覆了与蛋白质兼容的聚合物聚乙烯醇的熔融石英毛细管的入口端合成了甲基丙烯酸丁酯-二甲基丙烯酸乙烯酯共聚物（BuMA-co-EDMA）整体柱。该整体柱用于蛋白质的在线预浓缩，随后进行毛细管电泳（CE）分离。对于标准蛋白质（细胞色素c、溶菌酶和胰蛋白酶原A）的分析，该系统证明具有可重复性。迁移时间和校正峰面积的批间相对标准偏差（%RSD）值小于5%，这是毛细管电泳的典型值。以溶菌酶为溶质通过前沿分析测定，加载48 ng蛋白质后，1厘米的整体柱达到饱和。最后，通过零死体积连接器将BuMA-co-EDMA整体预浓缩器与蛋白G整体柱连接。该耦合系统用于在线去除IgG、标准蛋白质的预浓缩和毛细管电泳分离。该系统可能是一种有价值的样品制备工具，用于分析复杂样品（如人血清）中的低丰度蛋白质，其中高丰度蛋白质（如人血清白蛋白（HSA）和免疫球蛋白G（IgG））会阻碍低丰度蛋白质的鉴定和定量。
