Rapid Staining of Proteins in Polyacrylamide Gels with Nile Red

  • Joan-Ramon Daban
  • Salvador Bartolomé
  • Antonio Bermúdez
Part of the Springer Protocols Handbooks book series (SPH)


Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) is one of the most powerful methods for protein analysis (1,2). Unfortunately, the typical procedures for the detection of protein bands after SDS-PAGE, using the visible dye Coomassie blue and silver staining, have several time-consuming steps and require the fixation of proteins in the gel. This chapter describes a rapid and very simple method for protein staining in SDS gels developed in our laboratory (3,4). The method is based on the fluorescent properties of the hydrophobic dye Nile red (9-diethylamino-5H-benzo[α]phenoxazine-5-one; see Fig. 1), and allows the detection of <0.1 µg of unfixed protein per band about 6 min after the electrophoretic separation. Furthermore, it has been shown elsewhere (5) that, in contrast to the current staining methods, Nile red staining does not preclude the direct electroblotting of protein bands, and does not interfere with further sequencing and immunodetection analysis.


Critical Micelle Concentration Sodium Sulfite Photographic Negative Polaroid Film Intense Agitation 
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  1. 1.
    Andrews, A. T. (1986) Electrophoresis. Theory, Techniques, and Biochemical and Clinical Applications. Oxford University Press, Oxford, UK.Google Scholar
  2. 2.
    Zewert, T. E. and Harrington, M. G. (1993) Protein electrophoresis. Curr. Opin. Biotechnol. 4, 3–8.PubMedCrossRefGoogle Scholar
  3. 3.
    Daban, J.-R., Samsó, M., and Bartolomé, S. (1991) Use of Nile red as a fluorescent probe for the study of the hydrophobic properties of protein-sodium dodecyl sulfate complexes in solution. Anal. Biochem. 199, 162–168.PubMedCrossRefGoogle Scholar
  4. 4.
    Daban, J.-R., Bartolomé, S., and Samsó, M. (1991) Use of the hydrophobic probe Nile red for the fluorescent staining of protein bands in sodium dodecyl sulfate-polyacrylamide gels. Anal. Biochem. 199, 169–174.PubMedCrossRefGoogle Scholar
  5. 5.
    Bermúdez, A., Daban, J.-R., Garcia, J. R., and Mendez, E. (1994) Direct blotting, sequencing and immunodetection of proteins after five-minute staining of SDS and SDS-treated IEF gels with Nile red. BioTechniques 16, 621–624.PubMedGoogle Scholar
  6. 6.
    Greenspan, P. and Fowler, S. D. (1985) Spectrofluorometric studies of the lipid probe Nile red. J. Lipid Res. 26, 781–789.PubMedGoogle Scholar
  7. 7.
    Sackett, D. L. and Wolff, J. (1987) Nile red as a polarity-sensitive fluorescent probe of hydrophobic protein surfaces. Anal. Biochem. 167, 228–234.PubMedCrossRefGoogle Scholar
  8. 8.
    Samsó, M., Daban, J.-R., Hansen, S., and Jones, G. R. (1995) Evidence for sodium dodecyl sulfate/protein complexes adopting a necklace structure. Eur. J. Biochem. 232, 818–824.PubMedCrossRefGoogle Scholar

Copyright information

© Humana Press Inc., Totowa, NJ 1996

Authors and Affiliations

  • Joan-Ramon Daban
    • 1
  • Salvador Bartolomé
    • 1
  • Antonio Bermúdez
    • 1
  1. 1.Department de Bioquímica i Biologia MolecularUniversitat Autònoma de BarcelonaBellaterra (Barcelona)Spain

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