Iodonitrotetrazolium (chloride)

Iodonitrotetrazolium chloride (INT, CAS: 146-68-9) is a colorless tetrazolium-based chromogenic reagent commonly used as a chromogenic electron acceptor for the colorimetric determination of dehydrogenase activity. In enzyme-coupled redox reactions, INT accepts electrons from reduced cofactors through an electron transfer mediator and is reduced to water-soluble red formazan with the absorption maximum at approximately 490 nm, allowing enzyme activity to be quantified spectrophotometrically.

In lactate dehydrogenase (LDH) assays, LDH released into the culture supernatant catalyzes the oxidation of L-lactate to pyruvate with the concomitant reduction of NAD+ to NADH. In the presence of the electron-coupling reagent 1-methoxy-5-methylphenazinium methyl sulfate (MPMS), electrons generated from NADH are transferred to INT, reducing the tetrazolium salt to formazan. The absorbance of the formazan is directly proportional to the LDH activity present in the sample.

Reference:

[1]. Kaja S, Payne AJ, Naumchuk Y, Koulen P. Quantification of Lactate Dehydrogenase for Cell Viability Testing Using Cell Lines and Primary Cultured Astrocytes. Curr Protoc Toxicol. 2017 May 2;72:2.26.1-2.26.10. doi: 10.1002/cptx.21. PMID: 28463416; PMCID: PMC5501254.

[2]. Rayamajhi M, Zhang Y, Miao EA. Detection of pyroptosis by measuring released lactate dehydrogenase activity. Methods Mol Biol. 2013;1040:85-90. doi: 10.1007/978-1-62703-523-1_7. PMID: 23852598; PMCID: PMC3756820.