【作者】 李树香； 王欣怡； 刘敬； 周亚； 徐静； 邹剑平；

【Author】 LI Shu-xiang1,WANG Xin-yi1,LIU Jing1,ZHOU Ya1,XU Jing1,ZOU Jian-ping2 (1.National Vaccine and Serum Institute,Beijing 100024,China; 2.Beijing Jianping Jiuxing Biomedical Science and Technology Co.,Ltd.,Beijing 100085,China)

【机构】 北京生物制品研究所 第三研究室； 北京健平九星生物医药科技有限公司；

【摘要】 目的:研制血管内皮生长因子检测试剂盒(酶标法),用于类风湿性关节炎的临床诊断。方法:以～株抗血管内皮生长因子特异性单克隆抗体作为捕获抗体,另一株单抗作为检测抗体,棋盘滴定确定抗体使用浓度。制备VEGF165参考品,对样品稀释液、封闭液等实验条件进行优化。对试剂盒进行分析性能评估确定其质控标准。通过临床试验验证其在类风湿性关节炎检测中的有效性。结果:棋盘滴定确定捕获抗体浓度为10μg/mL,酶标抗体1;20 000应用,冻干VECF165参考品含量720～880 pg/mL,变异系数<10%。5%BSA作为封闭液和样品稀释液,试剂盒线性范围0～400 pg/mL,最低检出量0 pg/mL,不同浓度样本批内精密度9.83%～13.73%,不同时间批内精密度5.47%～14.08%,批间精密度7.88%,准确性94%～105%,特异性良好,效期末产品各项指标符合标准。确定了正常人VEGF参考范围:<142.2 pg/mL,作为判断类风湿性关节炎活动期指标灵敏度为70.5%,特异性80%。结论:成功研制了血管内皮生长因子检测试剂盒,完成临床试验评价。更多还原

【Abstract】 Objective:To develop a vascular endothelial growth factor assay kit for clinical diagnosis of rheumatoid arthritis.Methods:We choose two anti-vascular endothelial growth factor monoclonal antibodies as capture antibody and detective antibody respectively and determined the concentration of antibodies used by checkboard titration.To prepare freeze-dried VEGF165 reference material.The sample diluention,blocking solution and other experimental conditions were optimized.To determine the quality control standards by performance evaluation.Verify the effectiveness for RA detection through clinical trials.Results:The concentration of capture antibody and detect antibody were indentified as 10μg/mL and 1:20 000 respectively.The content of freeze-dried reference material is 720～880 pg/mL.Coefficient of variation is less than 10%.The sample diluent and block solution was 5%BSA.The linear range is 0～400 pg/mL.The minimum detectable concentration is 0 pg/mL.The intra precision of different concentration samples is 9.8%～13.7%.The intra precision of different detection times is 5.4%～14%.The inter precision is 7.8%.The precision performance is 94%～105%.Not found cross reaction with other cytokines.The Indicators of effective product at the end meet quality control standards.Normal reference values of VEGF:<142.2 pg/mL.As an indicator of active rheumatoid arthritis,sensitivity is 70.5%.Specificity is 80%.Conclusion:The vascular endothelial growth factor assay kit was successfully prepared. Evaluation of clinical trials completed.更多还原