摘要
[目的]制备一种人抗PD-L1抗体,建立其酶联免疫吸附分析(ELISA)的检测方法。[方法]将表达抗体重链和轻链的质粒转染到HEK-293细胞制备抗体,并建立ELISA方法对抗体进行检测。[结果]间接ELISA法的最佳抗原包被浓度为0.25μg/mL,抗PD-L1抗体标准品的起始浓度为0.25μg/mL,最适封闭液浓度为2%BSA,最适封闭时间为2 h,二抗的最佳稀释度为1∶4 000。细胞上清中的抗PD-L1抗体样品1的滴度为125,浓度66.66 ng/mL,灵敏度为6.3 ng/mL;样品2的滴度为125,浓度为81.8 ng/mL,灵敏度为5.9 ng/mL。[结论]建立了一种人抗PD-L1单克隆抗体的间接ELISA检测方法,经对样品1和样品2的批内批间重复性试验统计分析,变异系数均<10%,该ELISA法适用于该抗体的检测。
[Objective] A human anti-PD-L1 antibody was prepared and its enzyme-linked immunosorbent assay(ELISA) was established.[Methods]The plasmids expressing the heavy chain and light chain of anti-PD-L1 antibody were transfected into HEK-293 cells to prepare the antibody,and the ELISA method was established to detect the antibody.[Results] Indirect ELISA optimal antigen coating concentration was 0.25 μg/mL; the initial concentration of anti-PD-L1 antibody standard was 0.25 μg/mL; the optimal blocking concentration was 2% BSA; the optimal blocking time was 2 h; the optima dilution of the secondary antibody was 1∶ 4 000.The titer of the anti-PD-L1 antibody sample 1 in the cell supernatant was 125,the concentration was 66.66 ng/mL,the sensitivity was 6.3 ng/mL; the titer of sample 2 was 125,the concentration was 81.8 ng/mL,the sensitivity was 5.9 ng/mL from batch to batch of the two samples.[Conclusion] An indirect ELISA method for the establishment of a human anti-PD-L1 monoclonal antibody was established.By repeatability test of the inter-intra Samples 1 and 2 batches of statistical analysis,coefficients of variation were 〈 10%,the ELISA method is suitable for the detection of this antibody.
出处
《生物技术》
北大核心
2017年第6期552-556,共5页
Biotechnology
基金
吉林省科技计划发展项目("持留性结核杆菌肽类抑制剂药物"
No.20150101226JC)
关键词
抗程序性死亡配体-1抗体
酶联免疫吸附分析
检测
灵敏度
anti-procedure death ligand-1-antibody
enzyme-linked immunosorbent assay
detection
sensitivity