THE STAINING OF ACIDIC PROTEINS ON POLYACRYLAMIDE GELS - ENHANCED SENSITIVITY AND STABILITY OF STAINS-ALL STAINING IN COMBINATION WITH SILVER-NITRATE

A number of acidic proteins, such as those found in bone and dentin, a re poorly resolved on acrylamide gels using Coomassie blue or silver n itrate staining. The cationic dye Stains-all allows visualization and identification of these proteins due to their differential staining: h ighly acidic proteins stain blue and intact proteoglycans stain purple , whereas less acidic proteins stain pink. However, the use of Stains- all is limited due to relatively poor staining sensitivity and lack of stability to light. A procedure which addresses these deficiencies ha s been developed utilizing established protocols for Stains-all staini ng followed by silver nitrate incubation and development. In this way, phosphoproteins such as osteopontin, bone sialoprotein, dentin phosph ophoryn, and other acidic glycoproteins are visualized at higher sensi tivity (greater than fivefold) and staining stability than normally ac hieved with just Stains-all. The protocol stains a greater variety of proteins than a combined alcian blue/silver staining procedure previou sly described. Utilizing the Stains-all/silver protocol, porcine bone osteopontin, a protein not visualized by standard silver staining, can be observed in amounts as little as 0.25 ng on polyacrylamide gels. F urthermore, densitometric scans demonstrate that the staining intensit y is proportional to osteopontin amount and can be used for quantifica tion over a range from 0.25 to 50 ng. (C) 1997 Academic Press.