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Optimizing the Difference Gel Electrophoresis (DIGE) Technology

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Clinical Proteomics

Part of the book series: Methods in Molecular Biology™ ((MIMB,volume 428))

Summary

Difference gel electrophoresis (DIGE) technology has been used to provide a powerful quantitative component to proteomics experiments involving 2D gel electrophoresis. DIGE combines spectrally resolvable fluorescent dyes (Cy2, Cy3, and Cy5) with sample multiplexing for low technical variation, and uses an internal standard methodology to analyze replicate samples from multiple experimental conditions with unsurpassed statistical confidence for 2D gel-based differential display proteomics. DIGE experiments can facilely accommodate sufficient independent (biological) replicate samples to control for the large interpersonal variation expected from clinical samples. The use of multivariate statistical analyses can then be used to assess the global variation in a complex set of independent samples, filtering out the noise from technical variation and normal biological variation thereby focusing on the underlying variation that can describe different disease states. This chapter focuses on the design and implementation of the DIGE methodology employing the use of a pooled-sample internal standard in conjunction with the minimal CyDye chemistry. Notes are also provided for the use of the alternative saturation labeling chemistry.

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Author information

Authors and Affiliations

  1. Proteomics Laboratory, Mass Spectrometry Research Center, Department of Biochemistry, Vanderbilt University School of Medicine, Nashville, TN

    David B Friedman

  2. Cambridge Centre for Proteomics, Department of Biochemistry, University of Cambridge, UK

    Kathryn S Lilley

Authors
  1. David B Friedman
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  2. Kathryn S Lilley
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Editors and Affiliations

  1. Biomedical Research Foundation, Academy of Athens, Athens, Greece

    Antonia Vlahou

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© 2008 Humana Press, a part of Springer Science+Business Media, LLC

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Friedman, D.B., Lilley, K.S. (2008). Optimizing the Difference Gel Electrophoresis (DIGE) Technology. In: Vlahou, A. (eds) Clinical Proteomics. Methods in Molecular Biology™, vol 428. Humana Press. https://doi.org/10.1007/978-1-59745-117-8_6

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  • DOI: https://doi.org/10.1007/978-1-59745-117-8_6

  • Publisher Name: Humana Press

  • Print ISBN: 978-1-58829-837-9

  • Online ISBN: 978-1-59745-117-8

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