“A modified method of histochemical demonstration for cytochrome oxidase” has been reported previously. Further studies on new couplers of pyrazolone compounds and others were performed, and 1- (3′-chloro) phenyl-3-methyl-5 pyrazolone was found to be most useful drug as the coupler for the apparent demonstration of this enzyme.
It was found, however, in this method that the change of color and diffusion of dye were observed in the sections stained, as the pH value of the reaction mixture was changed variously. These phenomena were supposed to ascribe to the condition of the reaction of the reagents.
In the present paper, in order to elucidate the problem, the oxidated reaction products of these reagents by the chemical oxidating agent were measured under the various pH values and concentrations of phosphate buffer by spectrophotometry, and the results from them were compared with histochemical optimum.