| Summary: | 碩士 === 國立臺灣科技大學 === 化學工程系 === 103 === A convenient method for fabricating a sensitive indirect sandwich type electrochemical ELISA for the detection of estradiol using a chitosan electrodeposited platinum (Pt) wire microelectrode was proposed. In this assay, anti-17β estradiol produced in goat was used as the capture antibody which was immobilized on the chitosan coated Pt wire microelectrode, anti-17β estradiol 6 antibody produced in mouse was used as the detection antibody, and goat anti-mouse IgG conjugated with alkaline phosphatase (AP) was used as the secondary antibody. 4-Aminophenyl phosphate (4-APP) was chosen as the AP substrate and the oxidation potential of the electroactive AP product, 4-amino phenyl (4-AP), on the Pt electrode was determined to be +0.14 V (vs. Ag/AgCl). The electrochemical ELISA was detected by constant potential amperometry at +0.14 V in the Tris buffer (pH 9.0). The sensitivity (limit of detection) of this assay was 0.27 pg/mL with a wide detection range from 0.27 pg/mL up to 100 ng/mL. The assay specificity evaluated by testing the cross-reactivity of the assay against progesterone and 17α-ethynylestradiol was found to be 0.033% and 3.35%, respectively. The calibration curve of this assay tested in serum was also established. In this study, a sensitive, inexpensive, disposable, and easy to be fabricated electrochemical ELISA for estradiol was demonstrated.
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