DAB Substrate Kit

The Kit contains the following reagents to make an active stable substrate solution for use with the enzyme horseradish peroxidase:

1. 12 ml Stable DAB Solution (Store below -20^oC)
2. 6 ml Peroxide Solution (Store 2-8^oC)
3. 6 ml Buffer Stock Solution (Store 2-8^oC)

Store the DAB (25X) stock solution below 0^oC. The remaining reagents are stored at 2-8^oC.

DAB Substrate (3,3/-Diaminobenzidine) is the most commonly used substrate for demonstrating the presence of horseradish peroxidase in immunohistochemistry or immunoblotting. It produces a stable water/alcohol insoluble reddish-brown product. The slides may be dehydrated through alcohols to xylene and mounted with any commercially available mounting media.

1. To 5ml distilled water add 100µl (2 drops) of buffer stock solution.
2. Add 200µl (4 drops) of DAB stock solution.
3. Add 100µl (2 drops) of Hydrogen Peroxide solution. Mix well. (Working solution stable for at least 6 hours at room temperature.)

Application For use as an insoluble substrate for horseradish peroxidase. This substrate solution is suitable for use in immunoblotting and immunohistological staining procedures.

Caution: DAB is a possible carcinogen. Avoid contact by using gloves and mask when handling.

1. Nakane, P.K. and Pierce, G.B. (1967) J. Histochem. Cytochem. 14, 929
2. Barile, F. and Trombetta, L.D., (1982) J Histochem. 5:12
3. Rose, N.R., Friedman, H. and Fahey, J.L., eds. (1986) Manual of Clinical Laboratory Immunology, 2rd. ed. ASM, Washington, D.C.
4. Hsu, S. and Soban, E. (1982) J. Histochem. Cytochem. 30:1079