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Preparation of a New Tetrazolium Salt Which Yields a Blue Pigment on Reduction and Its Use in the Demonstration of Enzymes in Normal and Neoplastic Tissues^*

Alexander M. Rutenburg, M.D., Ralph Gofstein, B.S. and Arnold M. Seligman, M.D.

(From the Kirstein Laboratory for Surgical Research, Beth Israel Hospital, Boston and the Department of Surgery, Harvard Medical School)

The preparation of a ditetrazolium salt (IV) is given. When reduced, this water-soluble, pale-yellow salt (BT) is converted to a blue, water-insoluble pigment (diformazan). The toxicity of BT in mice is greater than the toxicity of triphenyl tetrazolium chloride. Dehydrogenase activity was demonstrated mainly in liver and kidney in vivo and in vitro with BT. Results with blocks of fresh tissue differed from the results with homogenates. Specific dehydrogenase activity was demonstrated in extracts of homogenates by the addition of specific substrates such as sodium succinate, xanthine, sodium lactate, and sodium malate. Cysteine desulfurase was demonstrated with cysteine as substrate. Tumors of animals and man contained no succinic dehydrogenase and little cysteine desulfurase by this technique.

^* This investigation was aided by a research grant from the National Cancer Institute, National Institutes of Health, Public Health Service, (in part) by a research grant from the American Cancer Society (Massachusetts Division) and (in part) by an institutional grant to Harvard University from the American Cancer Society.

Kirstein fellow in Surgery, Beth Israel Hospital.

Received 10/10/49.

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