A discontinuous method for catalase determination at 'near physiological' concentrations of H2O2 and its application to the study of H2O2 fluxes within cells.
We describe a discontinuous method for the measurement of catalase using the Ferrous Oxidation in Xylenol orange (FOX) assay. Samples containing catalase are incubated with H2O2 for varying time intervals prior to rapid mixing of aliquots of the incubation mixtures with FOX reagent, which measures residual H2O2. Absorbance is then read at 560 nm after 30-min incubation at room temperature. Decay of H2O2 is proportional to catalase activity in the original sample. An adaptation of the method involves exposure of intact cells to H2O2-generating systems in the presence of aminotriazole. The extent of catalase inactivation allows estimates of total H2O2 accumulation within the cell.
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Department of Medicine, University College London Medical School, UK.
Wolff
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D000640:Amitrole / Q000494:pharmacology; D000818:Animals; D002374:Catalase / Q000032:analysis* / Q000037:antagonists & inhibitors / Q000097:blood; D002417:Cattle; D004789:Enzyme Activation / Q000187:drug effects; D004912:Erythrocytes / Q000201:enzymology; D005296:Ferrous Compounds / Q000378:metabolism; D005456:Fluorescent Dyes; D006801:Humans; D006861:Hydrogen Peroxide / Q000097:blood / Q000378:metabolism*; D007700:Kinetics; D008099:Liver / Q000201:enzymology; D010084:Oxidation-Reduction; D010636:Phenols; D051381:Rats; D015203:Reproducibility of Results; D013454:Sulfoxides; D014992:Xylenes
D005296:Ferrous Compounds; D005456:Fluorescent Dyes; D010636:Phenols; D013454:Sulfoxides; D014992:Xylenes; D006861:Hydrogen Peroxide; D002374:Catalase; C016833:xylenol orange; D000640:Amitrole