The use of prontosil for the visualization of carbonic anhydrase bands in polyacrylamide gels. Application to the carbonic anhydrase isozymes of erythrocytes and white skeletal muscle of the rabbit.
Prontosil, a carbonic anhydrase inhibitor of orange-red colour, is used to visualize carbonic anhydrase bands during isoelectric focusing in polyacrylamide gels. 5-60 ng of the sulfonamide Prontosil are added to the 100-200 microliters samples before application to the gels. Bound Prontosil moves into the gel together with carbonic anhydrase and stains the enzyme bands formed there, while unbound Prontosil remains on top of the gels. The method is specific, no proteins other than carbonic anhydrase were observed to be stained, and it requires no special equipment. It was applied to chloroform/ethanol extracts of erythrolysates and white muscle homogenates from rabbits. Densitometric evaluation of the Prontosil-stained bands obtained with these extracts showed that rabbit red cells contain roughly equal amounts of carbonic anhydrase isoenzymes B and C while in rabbit white skeletal muscle isoenzyme C is predominant and little B enzyme occurs. These results confirm previous findings obtained by affinity chromatography of erythrolysates and muscle homogenates.
D000818:Animals; D001391:Azo Compounds; D002256:Carbonic Anhydrases / Q000032:analysis*; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods; D004912:Erythrocytes / Q000201:enzymology*; D006801:Humans; D007527:Isoenzymes / Q000032:analysis; D009132:Muscles / Q000201:enzymology*; D011817:Rabbits; D013379:Substrate Specificity; D010128:p-Aminoazobenzene / Q000031:analogs & derivatives; D044482:Benzeneacetamides
D001391:Azo Compounds; D002256:Carbonic Anhydrases; D007527:Isoenzymes; D010128:p-Aminoazobenzene; C003359:sulfamidochrysoidine; D044482:Benzeneacetamides