Rapid Staining of Proteins in Polyacrylamide Gels with Nile Red

Abstract

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.