Determination of estradiol by biotin-avidin-amplified electrochemical enzyme immunoassay

A simple, sensitive biotin-avidin-amplified electrochemical enzyme-linked immunosorbent assay (ELISA) for the determination of estradiol (E-2) was proposed in this paper. The complex of biotinylated anti-E-2 antibody and horseradish peroxidase-labeled avidin (HRP-avidin) were regarded as a probe in this system. The activity of labeled enzyme was measured with electrochemical methods using o-phenylenediamine as substrate. Coupled with the plate-coated antigen, indirect ELISA format using E-2-ovalbumin, the electrochemical detection was performed for E-2 with the detection limit of 21.0 pg/ml, and the linear range of determination of 50.0-500.0 pg/ml. The proposed method has been used for the determination of E-2 in river water with satisfactory results. Compared with the traditionally spectrophotometric ELISA detection, this method shows greatly heightened sensitivity. The limit of detection improved by about two orders of magnitude, which is very suitable for the conditions with extremely low concentration of analyte or very small volumes of sample present.