A MICROSPECTROPHOTOMETRIC QUANTIFICATION OF CYTOCHROME OXIDASE ACTIVITY BY MEANS OF COLOR-MODIFIED DIAMINOBENZIDINE REACTION

概要

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A new microspectrophotometric method for quantifying cytochrome oxidase activity has been developed based on a color-modified diaminobenzidine (DAB) reaction. The reaction consists of pretreating tissue sections or cell smears with 10 mM cobalt acetate dissolved in 0.1 M tris-HCI buffer of pH 7.4 for 10 min at 37℃, washing and then treating with the DAB reagent for cytochrome oxidase for 15 min at 37℃. Mitochondria turn reddish blue in the stained sections or smears. This reaction is enzymatic and conforms to Michaelis-Menten's formula. The microspectrophotometric quantification is effected by scanning the selected cytoplasmic area at 575 nm or by measuring the cytoplasmic plug at two wavelengths, 575 and 665 nm. As an example, the monomodal distribution in cytochrome oxidase activity was demonstrated among isolated rat liver cells by the two wavelength method.
日本組織細胞化学会の論文