毛细管等电聚焦/反相液相色谱/质谱联用

Capillary isoelectric focusing/reversed phase liquid chromatography/mass spectrometry.

作者信息

Lee Cheng S, Balgley Brian M

机构信息

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

出版信息

Methods Mol Biol. 2009;492:233-40. doi: 10.1007/978-1-59745-493-3_13.

DOI:10.1007/978-1-59745-493-3_13

PMID:19241036

Abstract

The vast number of proteins present in the proteome of a typical organism requires that separations be performed on the mixture prior to introduction into a mass spectrometer for protein identification and quantification. An integrated protein separation platform, combining capillary isoelectric focusing (CIEF) with reversed phase liquid chromatography (RPLC), is described to provide high resolving power for the analysis of complex protein mixtures. Thus, the proteins are systematically resolved according to their differences in isoelectric point and hydrophobicity using combined CIEF/RPLC separations. A key feature of the CIEF-based multidimensional separation platform is the elimination of protein loss and dilution in an integrated platform while achieving comprehensive and ultrasensitive analysis of protein profiles within small cell populations or limited tissue samples.

摘要

典型生物体蛋白质组中存在的大量蛋白质要求在将混合物引入质谱仪进行蛋白质鉴定和定量之前，先对其进行分离。本文描述了一种将毛细管等电聚焦（CIEF）与反相液相色谱（RPLC）相结合的集成蛋白质分离平台，用于为复杂蛋白质混合物的分析提供高分辨率。因此，利用CIEF/RPLC组合分离技术，蛋白质根据其等电点和疏水性的差异被系统地分离。基于CIEF的多维分离平台的一个关键特性是在集成平台中消除蛋白质损失和稀释，同时实现对小细胞群体或有限组织样本内蛋白质谱的全面和超灵敏分析。