title: Comparison of particulate 3,3',5,5'-tetramethylbenzidine and 3,3'-diaminobenzidine as chromogenic substrates for immunoblot.
abstract: In horseradish peroxidase (EC: 1.11.1.7)-dependent immunoblot assays, particulate 3,3',5,5'-tetramethylbenzidine (TMB) is shown to be a more efficient immunoblot substrate than the standard substrate 3,3'-diaminobenzidine (DAB), because TMB is easily prepared, stable, and less carcinogenic than is DAB. Assays of antibody in a serially diluted human immunodeficiency virus (HIV) control serum (CDC reference CAT# VS2151) have the same sensitivity limits with both DAB and TMB (1:312,500). Complete, working substrate solutions of H2O2/TMB/enhancer and of H2O2/DAB were stored at room temperatures and at 48 degrees C respectively. Periodic tests showed the TMB substrate system to be functional after four weeks at 48 degrees C and after eight weeks at room temperature, while the DAB system was functional after one week at 48 degrees C and after four weeks at room temperature. The stability, safety, and convenience of the commercially available TMB kits make this substrate ideal for immunoblot tests.
authors.affiliation: Centers for Disease Control.
authors.forename: J A
authors.initials: JA
authors.lastname: Brand
authors.forename: V C
authors.initials: VC
authors.lastname: Tsang
authors.forename: W
authors.initials: W
authors.lastname: Zhou
authors.forename: S B
authors.initials: SB
authors.lastname: Shukla
mesh_terms: D015100:3,3'-Diaminobenzidine / Q000494:pharmacology; D001560:Benzidines / Q000494:pharmacology; D011232:Chemical Precipitation; D002863:Chromogenic Compounds / Q000494:pharmacology; D006678:HIV / Q000187:drug effects / Q000276:immunology*; D015483:HIV Antibodies / Q000276:immunology; D015488:HIV Antigens / Q000302:isolation & purification*; D006801:Humans; D015151:Immunoblotting / Q000379:methods*; D011933:Reagent Kits, Diagnostic
chemical_list: D001560:Benzidines; D002863:Chromogenic Compounds; D015483:HIV Antibodies; D015488:HIV Antigens; D011933:Reagent Kits, Diagnostic; D015100:3,3'-Diaminobenzidine; C021758:3,3',5,5'-tetramethylbenzidine