3,3′–diaminobenzidine (DAB)–H2O2–HRP voltammetric enzyme-linked immunoassay for the detection of carcionembryonic antigen

A new voltammetric enzyme-linked immunoassay system of 3,3'-diaminobenzidine (DAB)-H2O2-horseradish peroxidase (HRP) has been presented and used for the sensitive detection of carcinoembryonic antigen (CEA) in human serum. In this proposed procedure, DAB was firstly used as the electroactive substrate in the HRP catalyzed oxidation reaction in the present of H2O2. The generated product produced a sensitive second-order derivative linear sweep voltammetric peak at potential of -0.62 V (vs. SCE) in Britton-Robinson (BR) buffer solution. The free HRP could be measured in a linear range from 2.5 x 10(-6)-2.5 x 10(-2) unit/ml and a detection limit of about 1.5 x 10(-6) unit/ml. Under the optimal experiment conditions, CEA could be detected in the linear range from 0.50 to 80 ng/ml with a detection limit of 0.5 ng/ml. The proposed electrochemical enzyme-linked immunosorbent assay method is simple, inexpensive, reproducible and sensitive, which shows promising for detecting CEA in the clinical diagnosis.