| Objective:The accurate quantification of serum E2 has high guiding significance for women’s physiological and pathological conditions.This study is based on the principle of LICA and hopes to establish a competitive mode of LICA detection system for quantification of E2 concentration in serum.And make an evaluation of analysis performance and clinical of this detection system.Methods:1.Preliminary establishment of a light-initiated chemiluminescent assay for serum E2:Based on LICA,the receptor beads were labeled with monoclonal anti-human estradiol antibodies,and estradiol or estriol was labeled with biotin,then the clinical samples were added for incubation,after that the streptavidin-coated donor beads were added,which constituted the detection environment.In this study,the best response mode was selected firstly,then the better competitive antigens were selected,finally the optimal concentration of receptor beads and biotinylated antigens was determined by checkerboard titration.Thus,a method based on LICA of serum E2 was established.2.Evaluation of the performance of LICA in measuring E2:On the basis of conditional optimization,we draw the standard curve of the fitting equation.The analytical performance of the initially established method was evaluated.The main indicators included sensitivity,precision,accuracy,and specificity.133 clinical samples were selected and analyzed with IMMULITE 2000 analyzer and VIDAS analyzer to verify the accuracy of the detection system.In this study,Graph Pad Prism7 software was used for mathematical model of dose-response curve and related statistical analysis.Results:1.The initial establishment of a light-initiated chemiluminescent assay in determining serum E2:We optimized the reaction mode,the type of competitive antigen and the optimal dilution of receptor beads and biotin-labeled antigen.Finally,the best response mode was determined as one-step method,the optimal competitive antigen was biotin-labeled estriol antigen,monoclonal anti-human estradiol antibody labeled receptor microspheres,and the optimal dilution of biotin-labeled estriol antigen degrees are 1:200 and 1:18×10~4 respectively.2.Performance characteristics of LICA in determining E2:The expected detection range of E2 was 20-5000 pg/m L.The analytical and functional sensitivities were 7.16 and 13.7 pg/m L,respectively.The intra-and interassay coefficients of variation were both below 15%,and the recovery rate ranged from 97.5%to 106.8%.The interference rates ranged from-3.6%to 5.4%and met detection requirements for E2 in hyperbilirubinemia,hemolysis,and lipemia in clinical samples.In addition,the cross-reactivity rates between E2 and structural analogs and some reproductive hormones varied from 1.9%to 10.6%which showed that LICA is highly specific for E2.Moreover,our results showed high accordance with the IMMULITE 2000(y=0.6695x+47.92,r~2=0.843)and VIDAS systems(y=1.099x-821.5,r~2=0.9392).Conclusion:This study successfully established a competitive mode of LICA for quantitation of serum estradiol.The detection system is a homogeneous detection method,with good detection performance,high sensitivity,strong specificity,good repeatability,and strong anti-interference ability,and the small amount of clinical samples is good for patients to seek medical treatment and can be used for quantitatively detect the concentration of estradiol in serum,and compared with some traditional methods and common clinical methods,the detection system has simple steps,no separation and washing steps,which greatly shortens the reaction time,moreover the cost is low,and it is easy to automate.It has a good application prospect in the detection of estradiol,which is helpful for the development of estradiol diagnostic kit. |
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