A 2-D liquid-phase chromatography for proteomic analysis in plant tissues

Two-dimensional liquid chromatography based on a high-performance chromatofocusing in the first dimension followed by high-resolution reversed-phase chromatography in the second dimension can be used as a complementary approach to protein separation with two-dimensional gel electrophoresis. In this work, Arabidopsis thaliana proteins obtained from different tissue extracts were resolved by using a new automated system, ProteomeLab PF 2D commercialized by Beckman Coulter (Fullerton, CA, USA). In particular, protein patterns obtained after two different extraction procedures (MgSO4 and urea buffer) were compared. Reproducibility of the protein patterns was also confirmed in different injections of the same sample and in the comparative analyses of some proteins by MALDI-TOF/MS. Computer analysis of the chromatograms revealed that with this two-dimensional liquid phase technique, hundreds of “virtual bands” can be identified and compared in crude plant protein lysates.