Determination of estradiol in human serum using magnetic particles-based chemiluminescence immunoassay.
A magnetic particles (MPs)-based chemiluminescence immunoassay (CLIA) with high sensitivity, specificity, and reproducibility was proposed for the evaluation of estradiol (E(2)) in human sera. The MPs coated with secondary antibody were used as dispersed solid phase for the immunoassay, and the horseradish peroxidase (HRP)-luminol-H(2)O(2) chemiluminescent system with high sensitivity was chosen as the detection system. The method showed specific recognition to E(2), without cross-reaction for the major steroids, including estrone (E(1)), estriol (E(3)), dihydrotestosterone (DHT), androstenedione, and testosterone (T), which was commonly found in human serum. The addition of sodium trichloracetate (Na-TCA) in the enzyme buffer as a blocking agent contributed to the realization of direct analysis of E(2) in human serum without extraction. Besides, the effects of several physicochemical parameters, including the dilution ratios of E(2)-6-HRP conjugate and anti-E(2) polyclonal antibody, immunoreaction time, chemiluminescent (CL) substrate volume, volume of MPs, and CL reaction time, were studied and optimized. The proposed method had a detection limit of 2.51pgmL(-1) with a larger working range of 15-1000pgmL(-1). The inter-assay and intra-assay coefficient of variation (CV) were both less than 15%. The average recoveries of three different spiked concentration samples were 93.3, 106 and 101%, respectively. The method has been successfully applied to the determination of E(2) in 105 human sera and showed a good correlation compared with the commercial radioimmunoassay (RIA) kit with a correlative coefficient of 0.9892. This method has exhibited great potential in the fabrication of diagnostic kit and could be used in the clinical analysis of E(2) in human serum.
Xin
Tian-Bing
TB
College of Chemistry and Environmental Science, Hebei University, Baoding 071002, China.
Liang
Shu-Xuan
SX
Wang
Xu
X
Li
Haifang
H
Lin
Jin-Ming
JM
D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D004958:Estradiol / Q000097:blood* / Q000276:immunology; D006801:Humans; D008163:Luminescent Measurements / Q000379:methods*; D008280:Magnetics; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
D004958:Estradiol