Skip to main content

Two-Dimensional Difference Gel Electrophoresis

  • Protocol
  • First Online:
Two-Dimensional Electrophoresis Protocols

Part of the book series: Methods in Molecular Biology ((MIMB,volume 519))

Summary

The introduction of two-dimensional fluorescent difference gel electrophoresis has enabled the extensive screening of differential protein expression levels with higher confidence and greater sensitivity than using the classical two-dimensional electrophoresis (2DE) approach. Using this technology, multiple protein samples can be labeled with up to three different fluorescent dyes. These labeled protein samples are mixed and applied on the same 2DE gel, subsequently scanned and analyzed by specialized software tools. The possibility to run two or more protein samples on a single gel, as well as the introduction of an internal standard on each gel drastically reduces the gel-to-gel variability and thus results in higher levels of certainty with regard to the differential character of the expressed proteins.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 129.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 169.00
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  1. Liang, P., and Pardee, A. B. (1992) Differential display of eukaryotic messenger RNA by means of the polymerase chain reaction. Science 257, 967–971.

    Article  PubMed  CAS  Google Scholar 

  2. Diatchenko, L., Lau, Y. F., Campbell, A. P., Chenchik, A., Moqadam, F., Huang, B., et al (1996) Suppression subtractive hybridization: a method for generating differentially regulated or tissue-specific cDNA probes and libraries. Proc. Natl. Acad. Sci. U S A 93, 6025–6030.

    Article  PubMed  CAS  Google Scholar 

  3. Schena, M., Shalon, D., Davis, R. W., and Brown, P. O. (1995) Quantitative monitoring of gene expression patterns with a complementary DNA microarray. Science 270, 467–470.

    Article  PubMed  CAS  Google Scholar 

  4. Anderson, L., and Seilhamer, J. (1997) A comparison of selected mRNA and protein abundances in human liver. Electrophoresis 18, 533–537.

    Article  PubMed  CAS  Google Scholar 

  5. Ünlü, M., Morgan, M. E., and Minden, J. S. (1997) Difference gel electrophoresis: a single gel method for detecting changes in protein extracts. Electrophoresis 18, 2071–2077.

    Article  PubMed  Google Scholar 

  6. Tonge, R., Shaw, J., Middleton, B., Rowlinson, R., Rayner, S., Young, J., et al (2001) Validation and development of fluorescence two-dimensional differential gel electrophoresis proteomics technology. Proteomics 1, 377–396.

    Article  PubMed  CAS  Google Scholar 

  7. Alban, A., David, S. O., Bjorkesten, L., Andersson, C., Sloge, E., Lewis, S., and Currie, I.(2003) A novel experimental design for comparative two-dimensional gel analysis: Two-dimensional difference gel electrophoresis incorporating a pooled internal standard. Proteomics 3, 36–44.

    Article  PubMed  CAS  Google Scholar 

  8. Van den Bergh, G., and Arckens, L. (2004) Fluorescent two-dimensional difference gel electrophoresis unveils the potential of gel-based proteomics. Curr. Opin. Biotechnol. 15, 38–43.

    Article  PubMed  CAS  Google Scholar 

  9. Shaw, J., Rowlinson, R., Nickson, J., Stone, T., Sweet, A., Williams, K., and Tonge, R. (2003) Evaluation of saturation labelling two-dimensional difference gel electrophoresis fluorescent dyes. Proteomics 3, 1181–1195.

    Article  PubMed  CAS  Google Scholar 

  10. Qu, Y., Moons, L., and Vandesande, F. (1997) Determination of serotonin, catecholamines and their metabolites by direct injection of supernatants from chicken brain tissue homogenate using liquid chromatography with electrochemical detection. J. Chromatogr. B Biomed. Sci. Appl. 704, 351–358.

    Article  PubMed  CAS  Google Scholar 

  11. Görg, A., Obermaier, C., Boguth, G., and Weiss, W. (1999) Recent developments in two-dimensional gel electrophoresis with immobilized pH gradients: wide pH gradients up to pH 12, longer separation distances and simplified procedures. Electrophoresis 20, 712–717.

    Article  PubMed  Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Gert Van den Bergh .

Editor information

Editors and Affiliations

Rights and permissions

Reprints and permissions

Copyright information

© 2009 Humana Press, a part of Springer Science+Business Media, LLC

About this protocol

Cite this protocol

Bergh, G. (2009). Two-Dimensional Difference Gel Electrophoresis. In: Tyther, R., Sheehan, D. (eds) Two-Dimensional Electrophoresis Protocols. Methods in Molecular Biology, vol 519. Humana Press. https://doi.org/10.1007/978-1-59745-281-6_29

Download citation

  • DOI: https://doi.org/10.1007/978-1-59745-281-6_29

  • Published:

  • Publisher Name: Humana Press

  • Print ISBN: 978-1-58829-937-6

  • Online ISBN: 978-1-59745-281-6

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics