The most convenient and chemically simple agent—diamide—is capable of producing a rapid diminution of the tripeptide thiol—glutathione (GSH)—within erythrocytes. An oxidant probe changes the oxidation state of the system; in the present case, diamide is an oxidant probe for thiols and changes the oxidation state of the thiols. Diamide perturbs the thiol status of a system. In most cases, the system can return to its original state by reduction. Information about the role of thiols in the biochemical, biophysical, and physiological economy of a biological system can be gained by the treatment of a system with an oxidant probe for thiols. The reaction of diamide with thiols can be followed spectrophotometrically between 300 and 325 nm and yields a second-order rate constant. The reduction of the diazene forms the diazane dicarboxylic acid bis(N,N-dimethylamide), a hydrazide that does not absorb down to 230 nm. The reaction of thiols with diazenecarbonyl derivatives—such as diamide—occurs in two observable stages, with thiolate anions (RS−) as the reactive species. The reaction proceeds via addition and displacement steps. In the case of GSH, the GS− anion adds to the diazene double bond to form a sulfenylhydrazine, which, in a second step, reacts with a second GS− anion at sulfur to yield a disulfide and a hydrazine.