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Related Experiment Videos

Assessing detection methods for gel-based proteomic analyses.

Lauren R Harris1, Matthew A Churchward, R Hussain Butt

  • 1Department of Physiology & Biophysics, Hotchkiss Brain Institute, Faculty of Medicine, University of Calgary, AB, T2N 4N1, Canada.

Journal of Proteome Research
|March 21, 2007
PubMed
Summary
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Sensitive protein detection in 2D gel electrophoresis is crucial. Sypro Ruby and Coomassie Blue offer high-sensitivity alternatives for protein staining, with Sypro Ruby showing superior stability.

Area of Science:

  • Proteomics
  • Biochemistry
  • Analytical Chemistry

Background:

  • Sensitive protein detection is critical for 2D gel electrophoresis (2DE) proteomic analyses.
  • Current limitations in 2DE often stem from protein detection sensitivity rather than resolution.
  • Fluorescent protein stains have enhanced in-gel detection sensitivity and protein quantification accuracy.

Purpose of the Study:

  • To evaluate the quality and cost-effectiveness of five commercial fluorescent protein stains.
  • To compare these stains against Sypro Ruby (SR), a commonly used stain.
  • To assess colloidal Coomassie Blue as a high-sensitivity alternative.

Main Methods:

  • Comparative analysis of five fluorescent stains: Krypton, Deep Purple, Rubeo, Flamingo, and Sypro Ruby (SR).

Related Experiment Videos

  • Evaluation of protein spot detection numbers and quantification accuracy.
  • Assessment of fluorophore stability, including repeated stain solution use.
  • Comparison of colloidal Coomassie Blue with SR using infrared fluorescence imaging.

    • Main Results:

    • All tested fluorescent stains demonstrated statistically comparable protein spot detection numbers.
    • Sypro Ruby exhibited superior fluorophore stability, allowing for repeated use of staining solutions.
    • Colloidal Coomassie Blue, when detected via infrared fluorescence imaging, was comparable to Sypro Ruby.
    • Detection sensitivity, not protein resolution, remains a key factor in 2DE.

    Conclusions:

    • Sypro Ruby and colloidal Coomassie Blue are high-sensitivity protein stains for 2DE.
    • The choice between Sypro Ruby and Coomassie Blue depends on available equipment and budget.
    • Both stains offer excellent alternatives for sensitive in-gel protein detection in proteomic studies.