Nicotinamide adenine dinucleotide tetrazole reductase (NADH-TR) is predominantly located in the lipid bilayer of type I and II muscle fibers， primarily within mitochondria and the sarcoplasmic reticulum. This enzyme is a collective term for three distinct enzymes: NADPH dehydrogenase， NAD(P)-dehydrogenase， and NADH dehydrogenase. As a key component of the electron transport chain in aerobic metabolism， its abundance reflects the utilization of the tricarboxylic acid cycle cytochrome system and other oxidative metabolic pathways.

The staining mechanism of this kit utilizes NADH and colorless tetrazole salts as substrates. NADH-TR in tissues oxidizes and reduces these substrates into insoluble blue-purple substances (formaldehyde)， which deposit in the mitochondria of myofibrils， causing mitochondrial staining to appear blue-purple while leaving actin and myosin on myofibrils unstained. NADH-TR can classify muscle fibers into Type I and Type II based on their oxidative capacity. Type I muscle fibers (slow-twitch fibers) contain higher oxidase content than Type II muscle fibers (fast-twitch fibers). Consequently， after NADH-TR staining， Type I muscle fibers exhibit deep blue-purple coloration， while Type II fibers show varying shades depending on their subtypes: Type IIa appears blue-purple， and Type IIb appears light blue-purple.