A fluorometric method for determination of C3b inactivator.
A simple method for the determination of C3b inactivator in human plasma was investigated. Fluorescent-labeled C3, which was prepared by treatment with methylamine and a fluorescent thiol reagent, N-(dimethylamino-4-methylcoumarinyl)-maleimide, was used as the substrate. Diluted human plasma is incubated with fluorescent C3, reduced with 2-mercaptoethanol, and subjected to sodium dodecyl sulfate polyacrylamide gel electrophoresis. The fluorescent cleavage product is then determined by scanning the gels with a fluorospectrophotometer. This method is simple and sensitive enough to determine C3b inactivator activity in 1 microliter of human plasma.
D003180:Complement C3b Inactivator Proteins / Q000032:analysis*; D004228:Dithionitrobenzoic Acid; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008301:Maleimides / Q000138:chemical synthesis; D013050:Spectrometry, Fluorescence / Q000379:methods; D013439:Sulfhydryl Reagents; D013997:Time Factors
D003180:Complement C3b Inactivator Proteins; D008301:Maleimides; D013439:Sulfhydryl Reagents; C019347:N-(7-dimethylamino-4-methylcoumarinyl)maleimide; D004228:Dithionitrobenzoic Acid