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MDPF Staining of Proteins on Western Blots

  • F. Javier Alba
  • Joan-Ramon Daban
Protocol
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Part of the Springer Protocols Handbooks book series (SPH)

Abstract

We describe a method for the detection of total protein patterns on polyvinylidene difluoride (PVDF) membranes using the fluorogenic dye 2-methoxy-2,4-diphenyl3(2H)-furanone (MDPF) (1). This method is based on the fluorescent properties of this dye (2). As can be seen in Fig. 1, MDPF (A) and the hydrolysis product (C) are nonfluorescent; only the adduct B formed with the proteins is fluorescent. This makes unnecessary the destaining of the PVDF membrane after protein labeling. The whole process of staining with MDPF is completed in about 20 min. Wet membranes are translucent, allowing the visualization of MDPF labeled protein bands by transillumination with UV light (see Fig. 2 A). Electrophoretic bands containing less than 10 ng of protein transferred to PVDF membranes can be detected after the reaction with MDPF.

Keywords

PVDF Membrane Borate Buffer Sodium Borate Transfer Buffer Electrophoretic Band 
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References

  1. 1.
    Alba, F. J. and Daban, J.-R. (1998) Rapid fluorescent monitoring of total protein patterns on sodium dodecyl sulfate-polyacrylamide gels and Western blots before immunodetection and sequencing. Electrophoresis, 19, 2407–2411.PubMedCrossRefGoogle Scholar
  2. 2.
    Weigele, M., DeBernardo, S., Leimgruber, W., Cleeland, R., and Grunberg, E. (1973) Fluorescent labeling of proteins. A new methodology. Biochem. Biophys. Res. Commun. 54, 899–906.PubMedCrossRefGoogle Scholar
  3. 3.
    Alba, F. J. and Daban, J.-R. (1997) Nonenzymatic chemiluminescent detection and quantitaion of total protein on Western and slot blots allowing subsequent immunodetection and sequencing. Electrophoresis 18, 1960–1966.PubMedCrossRefGoogle Scholar

Copyright information

© Humana Press Inc., Totowa, NJ 2002

Authors and Affiliations

  • F. Javier Alba
    • 1
  • Joan-Ramon Daban
    • 1
  1. 1.Departament de Bioquímica i Biologia MolecularUniversität Autónoma de BarcelonaBellaterra (Barcelona)Spain

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