Quantitative double-staining methods for tryptophyl and cysteinyl residues and protein in model films

The reliability of three new staining methods developed by the authors were examined with polyacrylamide gel films containing concanavalin A and ovalbumin. The methods were a 2-nitrophenylsulphenyl chloride technique for staining tryptophyl and cysteinyl residues together, a nitrophenylsulphenyl-2-mercaptoethanol method for tryptophyl residues only, and the combination of both these techniques with Coomassie Brilliant Blue for staining total protein. The relative contents of tryptophyl and cysteinyl residues of concanavalin A and ovalbumin in the films, calculated from absorbance values at 370 and 650 nm of double-stained films, were in good agreement with the theoretical values calculated from their amino acid compositions. Thus, these quantitative double-staining methods are reliable and should be useful for histochemical investigations of changes in the amount and composition of tissue proteins.