A simple colorimetric method for the measurement of hydrogen peroxide produced by cells in culture.
A simple, rapid and inexpensive method for the measurement of hydrogen peroxide (H2O2) produced by cells in culture is described. The assay is based on the horseradish peroxidase (HRPO)-mediated oxidation of phenol red by H2O2 which results in the formation of a compound demonstrating increased absorbance at 610 nm. A linear relationship between absorbance at 610 nm and concentration of H2O2 was found in the 1--60 microM (1--60 nmoles/ml) range. Due to the non-toxic character of phenol red and HRPO, the assay permits measurement of H2O2 production and release by macrophages for time intervals of 5--60 min under regular tissue culture conditions. Using this assay, the ability of a number of agents to induce H2O2 release by guinea pig peritoneal macrophages was demonstrated. These agents were: phorbol myristate acetate (PMA), opsonized zymosan, concanavalin A (Con A), wheat germ agglutinin (WGA), N-formyl-methionyl-leucyl-phenylalanine (FMLP) and A23187.
D000818:Animals; D000001:Calcimycin / Q000494:pharmacology; D002478:Cells, Cultured; D003124:Colorimetry / Q000379:methods*; D003208:Concanavalin A / Q000494:pharmacology; D006168:Guinea Pigs; D006861:Hydrogen Peroxide / Q000378:metabolism*; D037102:Lectins / Q000494:pharmacology; D008264:Macrophages / Q000378:metabolism*; D009239:N-Formylmethionine / Q000031:analogs & derivatives / Q000494:pharmacology; D009240:N-Formylmethionine Leucyl-Phenylalanine; D009842:Oligopeptides / Q000494:pharmacology; D010637:Phenolsulfonphthalein; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D015054:Zymosan / Q000494:pharmacology
D037102:Lectins; D009842:Oligopeptides; D003208:Concanavalin A; D000001:Calcimycin; D009239:N-Formylmethionine; D009240:N-Formylmethionine Leucyl-Phenylalanine; D015054:Zymosan; D006861:Hydrogen Peroxide; D010637:Phenolsulfonphthalein; D013755:Tetradecanoylphorbol Acetate