HISTOCHEMICAL DEMONSTRATION OF SUCCINIC DEHYDROGENASE ACTIVITY IN TISSUE SECTIONS BY A MODIFIED TECHNIQUE

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Abstract
Previous methods for demonstrating succinic dehydrogenase activity histochemically involve ditetrazolium or neotetrazolium as indicator dyes in conjunction with aerobic or anaerobic incubation of tissue sections. Using the anaerobic technique of Padykula, it appeared that enhancement of staining in the absence of O2 might be due to an alleviation of H competition by the cytochrome system. This hypothesis was tested by adding NaCN to the incubating medium (which contained neotetrazolium as an indicator dye) at concns. capable of poisoning cytochrome oxidase. Tissue sections incubated in this manner showed similar and in certain cases greater amts. of enzyme activity than previously reported by others. Another possible explanation for increased deposition of diformazan, is cyanide trapping of any oxalacetate which may be formed during processing or incubation of tissue sections. The effect of cyanide on this potent inhibitor of succinic dehydrogenase activity was clearly demonstrated. In addition, the use of a more alkaline 0.1 [image] phosphate buffer, pH 8.2 tended to increase staining. Thus, the confirmation of results by Brodie and Gots who found that the reduction of a tetrazolium indicator dye by yeast diaphorase was 23% greater at pH 8.3 than at pH 7.5. The specificity of the staining reaction in the histochemical demonstration of succinic dehydrogenase under these modified conditions was maintained in view of the fact that no staining occurred when the substrate Na succinate was omitted from the incubating medium.