ImmunoTek™ TMB (3,3',5,5'-Tetramethylbenzidine) Stable Liquid Substrate for HRPO, 1.25mM, Soluble

RT

Molecular Biology Grade

ImmunoTek™ TMB (3, 3', 5, 5'-Tetramethylbenzidine) Stable Liquid Substrate for HRPO, 1.25mM, Soluble

Tetramethylbenzidine (TMB) is an excellent substrate for detecting horseradish peroxidase (HRP) labeled probes. TMB produces a soluble end product which is blue in color that is quantitated at 650nm, providing an ultrasensitive quantitative substrate system.

TMB has been shown to be a safe-sensitive substrate for the assay of horseradish peroxidase (HRP). Initially, in the presence of HRP and hydrogen peroxide, a one-electron oxidation product is formed.

This compound, a cation free radical, is blue in color wth an adsorption maximum at 653nm. Further reaction with HRP/H2O2 or acidification of the radical with acid yields the diimine terminal oxidation product adsorbing light at 450 nm. The extinction coefficient of the radical (E653 nm = 5.9x10e4 mol-1 cm-1) provide a remarkably sensitive system for the assay of HRP and HRP labeled probes.

TMB is a single component reagent stable at room temperature and not sensitive to normal laboratory light. It is optimized with respect to TMB and hydrogen peroxide concentrations and yields a linear response with the concentrations of HRP usually employed in immunologic assays.

Application:
Kinetic and Endpoint Assays of Horseradish Peroxidase

Method:
After reacting with a HRP labeled probe, tissue sections or blots are incubated in the TMB solution. A one electron oxidation occurs resulting in the formation of a stable blue precipitate at the sites of HRP activity.

Features:
• Single component system
• Room temperature stable
• Exhibits low background levels
• Safe to use - no weighing of hazardous powders
• Save preparation time - use right out of the bottle
• Ultra-low sensitivity
• Minimal lot-to-lot variation

Reagent Provided:
Tetramethylbenzidine Solution:
Contains TMB, 1.25mM/L in an acetate buffer with proprietary stabilizers and precipitating agents.

Storage and Stability:
Stable for 12 months at RT. To extend the stability of product, refrigerate at 4°C.