2M Sulphuric Acid

Sulphuric acid plays a major role in ELISA experiments as a Stop solution where 3,3’,5,5’ - tetramethylbenzidine (TMB) acts as a substrate. Sulphuric acid plays a major role in ELISA experiments as a Stop solution where 3,3’,5,5’ - tetramethylbenzidine (TMB) acts as a substrate. Stop solution should fulfil two major requirements: 1) to terminate the reaction by effectively inhibiting the enzymatic activity of HRP, 2) to stabilize the oxidized products of the chromogenic or fluorogenic substrates. In the case of TMB, the stop solution, Sulphuric acid, not only inhibits color development but also converts the blue oxidation product of TMB to a yellow derivative which has an absorbance maxima at 450mn.