AN OSMIOPHILIC DISTYRYL DITETRAZOLIUM SALT (DS-NBT) FOR ULTRASTRUCTURAL DEHYDROGENASE ACTIVITY

The preparation of a new osmiophilic ditetrazolium salt (DS-NBT) containing two styryl groups, but retaining the essential structural features of nitro-BT, is given. DS-NBT may be used to demonstrate succinic dehydrogenase activity (SDH) faster than with nitro-BT and fully as rapidly as with TC-NBT. DS-NBT is more stable than TC-NBT and it is more reliably and more readily osmicated than TC-NBT. The ultrastructural sites of SDH and NADH 2 diaphorase activity are similar to those shown for cytochrome oxidase activity with diaminobenzidine in rat heart. The sites of lactic dehydrogenase activity are found in the sarcoplasmic reticulum and mitochondria of rat skeletal muscle fixed in formaldehyde for 3 hr. Tissue penetration of the doubly charged ditetrazolium salt (DS-NBT) was facilitated by 40-sec sonication and by 5% dimethyl sulfoxide. Illustrative examples and controls are included.