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Two‐dimensional separations of peptides and proteins by comprehensive liquid chromatography‐capillary electrophoresis
Author(s) -
Larmann John P.,
Lemmo Anthony V.,
Moore Alvin W.,
Jorgenson James W.
Publication year - 1993
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/elps.1150140169
Subject(s) - chromatography , capillary electrophoresis , chemistry , dimension (graph theory) , resolution (logic) , instrumentation (computer programming) , reversed phase chromatography , two dimensional chromatography , sample (material) , electrophoresis , analytical chemistry (journal) , high performance liquid chromatography , computer science , artificial intelligence , mathematics , pure mathematics , operating system
Analysis of biological samples is problematic because of their complex composition. Reversed phase liquid chromatography (RPLC), size exclusion chromatography (SEC), and, more recently, capillary zone electrophoresis (CZE) are routinely used for the analysis of these samples, but are eventually limited because they are one‐dimensional (1‐D) methods. As sample complexity increases, the separation efficiency necessary to resolve a large number of sample components in one dimension becomes prohibitively high. A solution to this problem has been to use a two‐dimensional (2‐D) approach. Each dimension in a 2‐D separation relies on a different separating mechanism. By expanding the separation into two dimensions, sample components unresolved in the first dimension can often be separated in the second. This circumvents the requirement for extremely high efficiencies in either dimension. Two‐dimensional slab gel electrophoresis has been used successfully in this area, but a more instrumental approach is desired. In this paper we describe three coupled‐column approaches to 2‐D separations. First, microcolumn SEC‐CZE is explored as a means of 2‐D protein analysis. Next, RPLC‐CZE is investigated for analysis of peptides in tryptic maps. Finally, RPLC is coupled with fast CZE (FCZE), a unique form of CZE analysis, for fast 2‐D analysis of peptides. Details of the instrumentation used in these 2‐D systems will be presented along with the results of some typical 2‐D analyses.