3,3′,5,5′-Tetramethylbenzidine or TMB is a chromogenic substrate used in staining procedures in immunohistochemistry as well as being a visualising reagent used in enzyme-linked immunosorbent assays (ELISA).

[1] TMB is a white solid that forms a pale blue-green liquid in solution with ethyl acetate.

The resulting one-electron oxidation product is a diimine-diamine complex, which causes the solution to take on a blue colour,[2] and this colour change can be read on a spectrophotometer at the wavelengths of 370 and 650 nm.

The reaction can be halted by addition of acid or another stop reagent.

Using sulfuric acid turns TMB yellow, with a peak absorbance of 450 nm.