摘要
目的建立血清促红细胞生成素(EPO)的ELISA试剂盒及一种检测血清EPO的双抗体夹心ELISA方法,并对其临床检测方法学进行研究。方法EPO抗体用缓冲液稀释,150μL/孔滴加到96孔板,用以包被。以正常人血清和临床血清为标本,用ELISA法观察其灵敏度、回收率、线性试验、稳定性。结果最佳包被抗体浓度为1∶600,最佳抗原工作浓度为1∶100,酶标抗体工作浓度为1∶5000。标准曲线的回归方程为:y=0.017x+0.3002,r=0.9710,P<0.05。灵敏度为0.46U/L,高、低浓度样品平均回收率分别为106.74%、104.78%。血清标本20、80U/L批内变异分别为3.044%、7.964%,批间变异分别为1.379%、12.915%。结论双抗体夹心ELISA法检测血清EPO,其敏感性、特异性、准确度均良好,为临床快速、准确、方便检测EPO提供了实验依据。
Objective To develop a double antibody sandwich ELISA method for detecting erythropoietin （EPO）, establish ELISA kit of the serum EPO, then initially study the research methodology of its clinical detection. Methods Coat the 96-microwell plate with EPO antibody was diluted with buffer, the each well of the 96-microwell plate should be dropped with 150μL antibody per well. The test sensitivity,recovery,linearity, and stability using ELISA method,with serum from healthy individuals and patients as clinical samples. Results The optimal antibody concentration was 1：600, the best working concentration of antigen was 1：100,and the best working concentration of enzyme labeled antibody was 1：5 000. The standard protein curve regression equation was y=0.017x＋0.300 2,r=0.971 0, P〈0.05. The sensitivity was 0.46 U/L,and the mean recoveries of samples with high and low concentrations were 106.74%, 104.78% respectively. The coefficient of variation of intra-assay ranged were 3.044% ,7.964% and the coefficient of variation of inter-assay ranged were 1.379% ,12.915% ,the serum sample was 20,80 U/L. Conclusion Double-antibody sandwich ELISA assay of serum EPO has good sensitivity ,specificity,accuracy ,and provides the experimental evidence for quick, accurate, and convenient clinical EPO detection.
出处
《实用临床医学（江西）》
CAS
2009年第4期1-4,共4页
Practical Clinical Medicine
基金
天津市科技型中小企业技术创新专项资金资助(08ZXCXSH03300)
