LDH Cytotoxicity Assay-Fluorescence

Lactate dehydrogenase (LDH) is a soluble cytosolic enzyme present in most eukaryotic cells. LDH releases into culture medium upon cell death due to damage of plasma membrane. The increase of LDH activity in culture supernatant is proportional to the number of lysed cells. The LDH Cytotoxicity Assay Kit, Fluorescence (LDH-FL) is designed to measure lactate dehydrogenase (LDH) activity through a fluorescence-based reaction. This assay utilizes a reaction cocktail containing lactate, NAD⁺, diaphorase, and resazurin. LDH catalyzes the conversion of NAD⁺ to NADH in the presence of L-lactate. Subsequently, diaphorase uses the NADH produced to reduce resazurin to resorufin, which can be detected via fluorescence with an excitation wavelength of 560 nm and an emission wavelength of 590 nm. The amount of resorufin formed is directly proportional to the LDH released into the culture medium, providing a measure of cytotoxicity. This kit offers higher sensitivity in a shorter timeframe and can detect lower concentrations of LDH activity compared to the colorimetric version.