| Human chorionic gonadotropin(h CG), consisting of α and β subunits combined via the binding between the ionic and hydrophobic bonds, is secreted by the syncytiotrophoblast cells. As produced in human placenta and luteal cells, h CG not only plays important role in implantation and embryo nutrition[1-4], but also possesses the diagnostic value for the gynecological inflammation and chorionic disease[5,6]. This study mainly focused on the investigation and development by using the chemiluminescence method for β-HCG detection, beside this, evaluation of the performance was also involved.Methods:From paring and screening among 62 β-sited antibodies provided by Yimeinuo company. Investigation on the components and study about the reaction system of the kit: according to the principle of the optimal reaction of antigen and antibody, the product was determined by the coated mode, sample dilution, enzyme conjugate, reaction mode, and the calibration system was established. Study on the analysis and reference interval of the kit: establish the linear range, repeatability, accuracy, limit of detection, specificity and stability index of the kit. 1302 sample from nondiseased subjects were chose for investigation, and we set 99% of the results as credible, by using SPSS to calculate the referenced value. According to the results got from two hospitals, to get the clinical accuracy of the kit was verified by the examination of different pathological population.Results:We chose one pair optimal antibodies as the raw materials, and we’ve also set up the quality standards. Investigation on the components and study about the reaction system of the kit, Coating: the coating buffer we used was 0.2M PB, PH 6.5. Enzyme combination: PH7.5, 0.01 M Tris-HCl, 50% FBS. Sample diluent:PH7.5, 0.01 M TAPS, 3% BSA. Calibrator: PH7.5, 0.01 M PBS, 3% BSA, the international referenced materials’ number was 77364. Study about the reaction system: two steps, incubated for 60 minutes and 30 minutes, and then washed for 5 times. 50μL substrate A and 50μL substrate B were separately added, and the light values were determined in 5 minutes. The recommended specimen type was serum. By turning the light values for logarithmic transformation, the calibration curve was then created according to four-parameter model. Study on the analysis and reference interval of the kit: linear range is from 1 to 1000 m IU/m L, within-run precision, between-run precision and within-batch precision were all less than 10%; when analyzed LH(500m IU/m L), FSH(500m IU/m L), TSH(500μIU/m L), all the results didn’t beyond 1.0m IU/m L; and the interference ratio caused by 50mg/d L bilirubin, 150mg/d L hemoglobin and 6000mg/d L triglycerides were all less than 10% respectively; the LOD(limit of detection) was 1.0 m IU/m L; the dilution recovery ranged from 85% to 150%; we tested the accelerated stability, in-use stability and real-time stability of the whole kit, and it showed that the kit kept stable. The referenced value was set 5.0 m IU/m L. According to the results got from two hospitals: the People’s Hospital(110 samples) and the Third Affiliated Hospital of the College of Chinese medicine(236 samples), both the positive and negative and total coincidence of the frontier hospital were 100%. And as for the latter hospital, its positive coincidence was 99.01% while the negative coincidence was 100%, its total coincidence was 99.03%. There was no significant difference between these two hospitals, and their results were in good linearity. Compared to the control reagents, the specific data of the two hospitals were summarized as follows, which suggested the two series of data were in good correlation.Conclusion:This study evaluated the performance of the quantitative detection method for β-HCG(chemiluminescence), among which involved the LOD, repeatability, specificity, dilution recovery, stability, the reference value and the clinical study. As compared with the international authorized similar kits that had already been on market, our kit possessed good clinical coincidence and better accuracy, and we’d also achieved their level as for the performance. |
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