Thermo Scientific Pierce DAB Substrate Kit enables the use of 3,3'-diaminobenzidine (DAB) in Western blotting and The role of tissue staining methods in performing chromogenic detection of horseradish peroxidase (HRP) activity.

DAB (3,3'-diaminobenzidine hydrochloride; MW = 214.1) reacts with HRP in the presence of peroxide, where the peroxidase-conjugated antibody binds to the sample An insoluble brown product is produced. The brown precipitate is insoluble in alcohol and other organic solvents, making it an excellent substrate for immunohistochemical staining that requires the use of traditional counterstains and mounting media. Pierce DAB Substrate Kit includes 10X DAB solution and stable peroxide substrate buffer. Crystalline DAB (powder) is also available.

DAB Substrate Features:

•HRP Substrate—for detection of horseradish peroxidase activity on solid media, including nitrocellulose and PVDF membranes and fixed tissue samples
• Chromogenesis—can be visualized without the use of special equipment; DAB reaction produces brown bands or Spots
• Packaging Options—Choose powders for custom formulations with buffers and peroxides, or select complete kits of pre-made solutions for immediate use

Pierce DAB Substrate Kit Features:

•Easy to use—Dilute 10X stock solution to 1X
•Safer in supplied stable peroxide buffer —Pre-prepared DAB solutions can be used with this chemical to minimize safety concerns
• Complete—Includes convenient stable hydrogen peroxide buffer solution (Product No. 34062)
• Stable—Store refrigerated at 4°C for at least One year; no need for frozen storage

DAB has been used in a variety of applications including Western blotting, immunohistochemistry techniques, and electron microscopy. A number of enhancement methods have been reported for DAB involving different buffer conditions, addition of metal ions, and post-processing applications (Refs 1-7; see also Pierce Metal Enhanced DAB Substrate Kit, Product number 34065). One of the disadvantages of DAB compared to other substrates is the need for immediate use of the working solution, since it starts the reaction process when hydrogen peroxide is added. This results in an increase in the background color of the substrate. However, the brown band produced by the DAB reaction does not fade as quickly as the color products of other precipitated HRP substrates.

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DAB substrate powder (3,3 '-Diaminobenzidine hydrochloride)