Description
Stop and stabilize TMB microwell substrate reactions.
Stop Solution for TMB Substrates (STOPT) is a proprietary acidic formulation used to stop TMB microwell substrate reactions. It is suitable for all endpoint ELISAs using a tetramethylbenzidine (TMB) substrate reaction for color development.
TMB substrates are oxidized by peroxidase enzymes to yield a soluble blue-green reaction product. In endpoint assays, the reaction can be stopped by adding equal volumes of Stop Solution for TMB Substrates. Addition of STOPT changes the chromagen color from blue-green to yellow, where it can be read at 450 nm, and concurrently stabilizes the yellow TMB product for one hour. Stopping the reaction will increase the sample absorbance value up to 3-fold. To avoid overdeveloping the TMB substrate reaction, the blue-green reaction product should be periodically monitored on an ELISA plate reader using 620-650 nm absorbance filter settings. When OD values reach approximately 0.7 units, the reaction should be stopped with STOPT.
For best results, the absorbance should be monitored and read before values exceed 2.5 OD units. If the yellow reaction product yields OD values above 2.5 units, it is recommended to dilute the antibodies/conjugates or shorten the incubation period.
Stop Solution for TMB Substrates is ready to use at 1X. Stop the reaction by adding an equal volume of STOPT to the substrate in every well of the plate