Evaluation of two-dimensional differential gel electrophoresis for proteomic expression analysis of a model breast cancer cell system

The technique of fluorescent two-dimensional (2D) differ-ence gel electrophoresis for differential protein expres-sion analysis has been evaluated using a model breast cancer cell system of ErbB-2 overexpression. Labeling of paired cell lysate samples with N-hydroxy succinimidyl ester-derivatives of fluorescent Cy3 and Cy5 dyes for sep-aration on the same 2D gel enabled quantitative, sensitive, and reproducible differential expression analysis of the cell lines. SyproRuby staining was shown to be a highly sensitive and 2D difference gel electrophoresis-compati-ble method for post-electrophoretic visualization of pro-teins, which could then be picked and identified by matrix-assisted laser-desorption ionization mass spectroscopy. Indeed, from these experiments, we have identified mul-tiple proteins that are likely to be involved in ErbB-2