Detection of hemoprotein peroxidase activity on polyvinylidene difluoride membrane.
The feasibility of detecting hemoproteins after electroblotting was examined. Hemoproteins were subjected to lithium dodecyl sulfate-polyacrylamide gel electrophoresis (LDS-PAGE) and then electroblotted and peroxidase activity was detected with 3,3',5,5'-tetramethylbenzidine The sensitivity and specificity of tetramethylbenzidine staining of LDS-PAGE gels was retained when proteins were electroblotted. Subsequent staining of the membrane with Coomassie blue R-250 revealed a protein pattern similar to that in the polyacrylamide gel. Thus electroblotting of hemoproteins does not affect resolution of the electrophoretic pattern and heme-associated peroxidase activity. Additionally, the ability to stain hemoproteins on polyvinylidene difluoride membranes offers the advantage of utilizing the same membrane for further biochemical and immunological characterizations.
Dutta
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Department of Biochemistry, University of California, Riverside 92521.
Henry
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HL
D000818:Animals; D001560:Benzidines; D002645:Chickens; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods; D006418:Heme / Q000032:analysis; D006420:Hemeproteins / Q000032:analysis*; D008297:Male; D008567:Membranes, Artificial; D008970:Molecular Weight; D010544:Peroxidases / Q000378:metabolism*; D011145:Polyvinyls; D012394:Rosaniline Dyes; D012967:Sodium Dodecyl Sulfate; D005465:Fluorocarbon Polymers
D001560:Benzidines; D006418:Heme; D006420:Hemeproteins; D008567:Membranes, Artificial; D010544:Peroxidases; D011145:Polyvinyls; D012394:Rosaniline Dyes; D012967:Sodium Dodecyl Sulfate; C024865:polyvinylidene fluoride; D005465:Fluorocarbon Polymers; C021758:3,3',5,5'-tetramethylbenzidine; C024757:Coomassie brilliant blue R; C028913:dodecyl sulfate