Comparison of chromogens for the determination of horseradish peroxidase as a marker in enzyme immunoassay.
o-Phenylenediamine, 2,2'-azino-di(3-ethylbenzthiazoline sulphonic acid-6) (ABTS), o-dianisidine and 4-aminoantipyrine were compared as chromogens for the determination of horseradish peroxidase. Highest sensitivity in the determination of horseradish peroxidase-IgG conjugates in dissolved form was obtained with o-phenylenediamine. When these conjugates were used in a two-site binding enzyme immunoassay for hepatitis B surface antigen (HBsAg), the steepest calibration curve and the lowest detection limit were obtained when ABTS was used to determine the immune complexes bound to the solid phase. Non-ionic detergents, such as polyoxyethylene-sorbitol ester, retarded horseradish peroxidase inactivation, resulting in a chromogen-dependent activity rise of horseradish peroxidase. An optimised determination of horseradish peroxidase is reported, in which the sensitivity of the solid phase enzyme immunoassay is doubled by the use of o-dianisidine.
D000675:Ampyrone; D052160:Benzothiazoles; D002863:Chromogenic Compounds; D003962:Dianisidine; D005069:Evaluation Studies as Topic; D006735:Horseradish Peroxidase / Q000378:metabolism*; D007124:Immunoenzyme Techniques; D010544:Peroxidases / Q000378:metabolism*; D010655:Phenylenediamines; D013451:Sulfonic Acids
D052160:Benzothiazoles; D002863:Chromogenic Compounds; D010655:Phenylenediamines; D013451:Sulfonic Acids; D000675:Ampyrone; C002502:2,2'-azino-di-(3-ethylbenzothiazoline)-6-sulfonic acid; D006735:Horseradish Peroxidase; D010544:Peroxidases; D003962:Dianisidine