Application

3,3′,5,5′-Tetramethylbenzidine (TMB) is a noncarcinogenic substitute (Ames test negative) for benzidine suitable as peroxidase substrate for use in ELISA procedures. The substrate produces a soluble end product that is pale blue in color and can be read spectrophotometrically at 370 or 620-650 nm. The TMB reaction may be stopped with 2 M H₂SO₄ (resulting in a yellow color), and read at 450 nm. A sensitive and specific reagent for the detection of blood, assay of hemoglobin, assay of peroxidases.

3,3′,5,5′-Tetramethylbenzidine has been used as a substrate for peroxidase in:enzyme-immunoassay (EIA)indirect competitive enzyme-linked immunosorbent assay (ELISA)sandwich enzyme-linked immunosorbent assay