Staining of the Ca2+-binding proteins, calsequestrin, calmodulin, troponin C, and S-100, with the cationic carbocyanine dye "Stains-all".
The Ca2+-binding proteins, calsequestrin, calmodulin, troponin C, and S-100, have all been shown to stain dark blue or purple with the cationic carbocyanine dye "Stains-all", while most proteins stain red or pink. Stains-all staining of these Ca2+-binding proteins was 50-100% more intense than Coomassie blue staining, and the blue stained calsequestrin and troponin C could be detected in muscle extracts. Densitometric scans of Stains-all-stained gels revealed that interaction of the dye with Ca2+-binding proteins changed the absorption spectrum of the dye. The dye-protein complex absorbed maximally at 615 nm, reflecting a state that results from the binding of individual dye molecules at anionic sites. Stains-all also interacted with undenatured Ca2+-binding proteins in aqueous solution forming a complex absorbing maximally at 600 nm. A simple assay for Ca2+-binding proteins has been developed using this property. These results suggest that the interaction of the dye with anionic sites within these Ca2+-binding proteins produces the dye-protein complex which absorbs at 600-615 nm. Stains-all staining of these Ca2+-binding proteins will be useful in their identification and purification from various tissues. It might also be a valuable tool in the identification of potential Ca2+-binding proteins.
D000818:Animals; D002135:Calcium-Binding Proteins / Q000032:analysis*; D002147:Calmodulin / Q000032:analysis; D002155:Calsequestrin / Q000032:analysis; D004591:Electrophoresis, Polyacrylamide Gel; D011817:Rabbits; D009418:S100 Proteins / Q000032:analysis; D012519:Sarcoplasmic Reticulum / Q000032:analysis; D013194:Staining and Labeling; D014336:Troponin / Q000032:analysis; D019209:Troponin C
D002135:Calcium-Binding Proteins; D002147:Calmodulin; D002155:Calsequestrin; D009418:S100 Proteins; D014336:Troponin; D019209:Troponin C