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The Basics of 2D DIGE

  • Phil BeckettEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 854)

Abstract

The technique of two-dimensional (2D) gel electrophoresis is a powerful tool for separating complex mixtures of proteins, but since its inception in the mid 1970s, it acquired the stigma of being a very difficult application to master and was generally used to its best effect by experts. The introduction of commercially available immobilized pH gradients in the early 1990s provided enhanced reproducibility and easier protocols, leading to a pronounced increase in popularity of the technique. However gel-to-gel variation was still difficult to control without the use of technical replicates. In the mid 1990s (at the same time as the birth of “proteomics”), the concept of multiplexing fluorescently labeled proteins for 2D gel separation was realized by Jon Minden’s group and has led to the ability to design experiments to virtually eliminate gel-to-gel variation, resulting in biological replicates being used for statistical analysis with the ability to detect very small changes in relative protein abundance. This technology is referred to as 2D difference gel electrophoresis (2D DIGE).

Key words

Two-dimensional gel electrophoresis 2D DIGE CyDye Multiplexing Difference gel electrophoresis 

Notes

Acknowledgments

Many thanks to Rita Marouga, Viola Ruddat, and Chris Rozanas (GE Healthcare) for their critical review and feedback.

Trademarks

CyDye™, Cy™, and Deep Purple™ are trademarks of GE Healthcare Ltd., a General Electric company.

Coomassie® is a registered trademark of ICI plc.

SYPRO® is registered trademark of Molecular Probes, Inc.

Sigma-Aldrich® is a registered trademark of Sigma Chemical Co.

2D Fluorescence Difference Gel Electrophoresis (Ettan DIGE) technology is covered by US Patent Numbers 6,043,025, 6,127,134, 6,426,190 and foreign equivalents and exclusively licensed from Carnegie Mellon University by GE Healthcare, Ltd., a General Electric company.

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Copyright information

© Springer Science+Business Media, LLC 2012

Authors and Affiliations

  1. 1.GE Healthcare Bio-SciencesPiscatawayUSA

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