Lucigenin caused a concentration-dependent increase in superoxide production by xanthine oxidase plus xanthine. This was seen, in terms of superoxide dismutase-inhibitable reduction of cytochrome c; in spite of the ability of univalently reduced lucigenin to directly reduce cytochrome c. It follows that in the absence of this interference, by the cytochrome, an even greater increase in superoxide production mediated by lucigenin would have been observed. Clearly lucigenin luminescence should not be relied upon as a method for measurement of, or even for detection of, superoxide.