Capabilities Using 2-D DIGE in Proteomics Research

Abstract

The use of two-dimensional gel electrophoresis for differential analysis in proteomics was revolutionized by the introduction of 2-D fluorescence difference gel electrophoresis (2-D DIGE). This fluorescence-based technique allows the use of multiplexed samples and an internal standard that virtually eliminates gel-to-gel variability, resulting in increased confidence that differences found between samples are due to real induced changes, rather than inherent biological variation or experimental variability. 2-D DIGE has quickly become the "gold standard" for gel-based proteomics for studying tissues, as well as cell culture and bodily fluids such as serum or plasma. This chapter will describe the basic 2-D DIGE technique using minimal labeling, image acquisition using high-quality fluorescence scanners, and software capable of analyzing the multiplexed images and normalizing the data using the internal standard.

Proteomic Dissection of Seed Germination and Seedling Establishment in Brassica napus

“…By introduction of fluorescent cyanine dyes and an internal standard, the ability of 2-D DIGE to separate and resolve complex proteomic patterns was greatly improved (Van den Bergh and Arckens, 2004 ; Rozanas and Loyland, 2008 ). Although, 2-D DIGE has not totally overcome the shortfalls of gel-based proteomics technology, there is still not a method that can completely replace 2-D DIGE to simultaneously separate and display several thousand proteins from complex samples.…”