Coomassie brilliant blue R-250 as a highly sensitive pre-stain for immediate visualization of human serum proteins on polyacrylamide gel disc electrophoresis and raising monospecific antibodies.
We hereby describe electrophoretic analysis of normal human serum pre-stained with Coomassie Brilliant Blue R-250 (CBB). The proposed method was optimized by studying as many as 450 disc electrophoretic separations and 30 variables. The method when compared with the post-electrophoretic staining by Amido Black (AB) revealed that the pre-stained discs were intensely well defined and resolved within 2 hours with a transparent gel. Gels stained with AB, despite a prolonged destaining, showed a residual dye retention making identification of the faint components difficult. Protein eluted from the CBB pre-stained gels retained its purity and immunoreactivity and conjugates of the two prototype proteins namely the albumin and the transferrin produced high-titre monospecific antisera in immunized rabbits.
Saoji
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AM
Kulkarni
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Naghi
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D000580:Amido Black; D000911:Antibodies, Monoclonal / Q000096:biosynthesis*; D001798:Blood Proteins / Q000032:analysis* / Q000276:immunology; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D012394:Rosaniline Dyes; D013194:Staining and Labeling
D000911:Antibodies, Monoclonal; D001798:Blood Proteins; D012394:Rosaniline Dyes; C004692:coomassie Brilliant Blue; D000580:Amido Black