集成毛细管等电聚焦/纳米反相液相色谱联用电喷雾电离质谱用于完整酵母蛋白质的表征。

Integrated capillary isoelectric focusing/nano-reversed phase liquid chromatography coupled with ESI-MS for characterization of intact yeast proteins.

作者信息

Wang Yueju, Balgley Brian M, Rudnick Paul A, Evans Erin L, DeVoe Don L, Lee Cheng S

机构信息

Department of Chemistry and Biochemistry, University of Maryland, College Park, Maryland 20742, USA.

出版信息

J Proteome Res. 2005 Jan-Feb;4(1):36-42. doi: 10.1021/pr049876l.

DOI:10.1021/pr049876l

PMID:15707355

Abstract

An integrated protein concentration/separation platform, combining capillary isoelectric focusing (CIEF) with nano-reversed phase liquid chromatography (nano-RPLC), is developed to provide significant protein concentration and high resolving power for the analysis of complex protein mixtures. Upon completion of protein focusing, the proteins are sequentially and hydrodynamically loaded into individual trap columns using a group of microinjection and microselection valves. Repeated pro-tein loadings and injections into trap columns are carried out automatically until the entire CIEF cap-illary content is sampled and fractionated. Each CIEF fraction "parked" in separate trap columns is further resolved using nano-RPLC, and the eluants are analyzed using electrospray ionization-mass spectrometry.

摘要

开发了一种集成蛋白质浓缩/分离平台，将毛细管等电聚焦（CIEF）与纳米反相液相色谱（nano-RPLC）相结合，为复杂蛋白质混合物的分析提供显著的蛋白质浓缩和高分辨率。蛋白质聚焦完成后，使用一组微注射和微选择阀将蛋白质依次通过流体动力学方式加载到各个捕集柱中。自动对捕集柱进行重复的蛋白质加载和进样，直到对整个CIEF毛细管内容物进行采样和分级分离。每个“停泊”在单独捕集柱中的CIEF级分再通过nano-RPLC进一步分离，洗脱液用电喷雾电离质谱进行分析。