The DetectX^® Glutathione (GSH) Fluorescent Detection Kit utilizes a proprietary non-fluorescent molecule, ThioStar^®, that covalently binds to the free thiol group on GSH to yield a highly fluorescent product. After mixing the sample or standard with ThioStar^® and incubating at room temperature for 15 minutes, the GSH-generated signal is read at 510 nm in a fluorescent plate reader with excitation at 390 nm. Addition of a reaction mixture converts all the GSSG into free GSH, which then reacts with the excess ThioStar^® during a second 15 minute incubation, yielding the signal related to Total GSH content.

Glutathione (GSH) is the highest concentration non-protein thiol in mammalian cells and is present in concentrations of 0.5 to 10 mM. GSH plays a key role in many biological processes, including the synthesis of proteins and DNA, the transport of amino acids, and the protection of cells against oxidation. Harmful hydrogen peroxide cellular levels are minimized by the enzyme glutathione peroxidase (GP) using GSH as a reductant. The oxidized GSH dimer, GSSG, is formed from GSH and peroxide by the GP reaction. An important role of GSSG in the NFkB activating signal cascade is suggested by the fact that the potent NFkB inducer TPA increases intracellular GSSG levels and GSSG/GSH ratios.