A picomolar spectrophotometric assay for superoxide dismutase

During the aerobic xanthine oxidase reaction, O₂^? is produced and accumulates to a steady state determined by a balance between the rate of production of this radical and its rate of dismutation. Addition of ferricytochrome c then results in a biphasic reduction, the very rapid phase of which reflects reaction of the accumulated O₂^?, while the slower phase corresponds to the continuing production of this radical. Superoxide dismutase suppresses the accumulation of O₂^? during the xanthine oxidase reaction and thus diminishes the burst of reduction seen upon addition of ferricytochrome c. This effect has been utilized, at pH 10.2, as the basis of an assay that permits measurement of picomolar levels of superoxide dismutase. The theory and practice of this ultrasensitive assay are described.