TMB Substrate

3,3′,5,5′-Tetramethylbenzidine (TMB) is a chromogenic substrate suitable for use in ELISA procedures, which utilize horseradish peroxidase conjugates. This substrate produces a soluble end product that is blue in color and can be read spectrophotometrically at 370 or 655 nm. The reaction maybe stopped with acid, resulting in a yellow solution that is read at 450 nm. This product is supplied as a ready-to-use peroxidase substrate containing TMB in a mildly acidic buffer. Prior to reaction with peroxidase, the substrate should be a colorless to light bluish-green solution. The substrate system develops a blue reaction product when reacted with peroxidase in microwell applications (e.g. ELISA assays). For end-point assays, acid can be used to stop the reaction, to yield a yellow end product. Since this substrate produces a soluble reaction product, it is not recommended for histochemistry or blotting.