Nesbitt Chandra A, Lo Josephine T M, Yeung Ken K C
Department of Chemistry, The University of Western Ontario, London, Ont., N6A 5B7 Canada.
J Chromatogr A. 2005 May 6;1073(1-2):175-80. doi: 10.1016/j.chroma.2004.09.081.
An effective protein preconcentration technique specifically designed for microliter-volume samples is presented. The preconcentration is based on the capturing of protein in its isoelectric point (pI) within an applied electric field, using a pH junction created by a discontinuous buffer system. The buffers were chosen to selectively preconcentrate proteins of neutral pI, myoglobin in this case, while removing other proteins with acidic or basic pIs. For the suppression of electro-osmotic flow (EOF) and protein adsorption, the capillary inner wall was modified with a zwitterionic phospholipid bilayer coating. A preconcentration factor of up to 1700 was obtained for a 1 microg/mL solution of myoglobin. The preconcentration was completed in approximately 20 min. Several sample introduction conditions were presented to accommodate sample volume from one to a few hundreds of microliters. The final volume of the preconcentrated sample band was estimated to be approximately 5 nL.
本文介绍了一种专门为微升体积样品设计的有效蛋白质预浓缩技术。该预浓缩基于在施加电场中利用不连续缓冲系统产生的pH交界捕获处于其等电点（pI）的蛋白质。选择这些缓冲液以选择性地预浓缩中性pI的蛋白质，在此情况下为肌红蛋白，同时去除具有酸性或碱性pI的其他蛋白质。为了抑制电渗流（EOF）和蛋白质吸附，用两性离子磷脂双层涂层对毛细管内壁进行了修饰。对于1μg/mL的肌红蛋白溶液，获得了高达1700的预浓缩因子。预浓缩在约20分钟内完成。提出了几种进样条件，以适应1至数百微升的样品体积。预浓缩样品带的最终体积估计约为5 nL。
