Sensitive and nonenzymatic measurement of hydrogen peroxide in biological systems

The increasing demand in detecting H(2)O(2) under various experimental conditions is only partly fulfilled by most conventional peroxidase-based assays. This article describes a sensitive and nonenzymatic H(2)O(2) assay that is based on the chemiluminescence reaction of luminol with hypochlorite. It allows the determination of H(2)O(2) down to nanomolar concentrations. Actual H(2)O(2) concentrations rather than a turnover of H(2)O(2) can be determined in monolayer cultures, perfusates, suspensions of intact cells, organelles, and crude homogenates. One of the strengths of this assay is that it may be used to assess fast enzyme kinetics (catalase, glutathione peroxidase, oxidases) at very low H(2)O(2) concentrations. Its use together with a glucose oxidase/catalase system appears to be a powerful tool in studying signal functions of H(2)O(2) in various biological systems on a quantitative basis. Several applications are discussed in detail to demonstrate the technical requirements and analytical potentials.