A novel CE-based
enzyme immunoassay (CE-EIA) method was developed in
o-aminophenol (OAP)-H(2)O(2)-horseradish
peroxidase (HRP) system and applied to benign
liver disease and
hyperthyroidism research in the clinical practical field. In the presented method, after the
enzyme immunoreaction, the HRP-labeled antibody or HRP-labeled
antigen catalyzed the
enzyme substrate OAP and H(2)O(2). The product of the enzymatic catalysis reaction
2-aminophenoxazine-3-one (AP) was determined using electrochemical detection on a Pt
electrode at the outlet of the reaction capillary. Factors influencing the performance, including running
buffer concentration, separation, and detection voltage, were investigated to the optimum conditions. Noncompetitive and competitive models were utilized to detect
alpha-fetoprotein (AFP) and
thyroxine (T(4)) in human sera, respectively. The linear ranges and the detection limits (S/N = 3) were from 1.5 to 66.6 ng/mL and 0.48 ng/mL for AFP, and from 1.7 to 260.0 ng/mL and 1.0 ng/mL for T(4). The results of this method were linear proportional to those of spectrophotometric ELISA method, giving a good prospect for a new clinical diagnostic instrument.