摘要
研究制备了快速检测新冠病毒(severe acute respiratory syndrome coronavirus 2,SARS-CoV-2)及其表面刺突糖蛋白受体结合域(receptor-binding domain,RBD)的ELISA试剂盒.该试剂盒由一对RBD单克隆抗体作为捕获抗体与检测抗体,其中检测抗体进行辣根过氧化物酶(HRP)偶联,通过方阵滴定法确定双抗夹心ELISA的最适工作条件.另外,对检测试剂盒进行线性、特异性、准确度、精密度等进行了验证.结果表明,双抗夹心ELISA方法中捕获抗体和酶标检测抗体的效价分别为1∶160000和1∶320000,试剂盒的定量检测下限为31 ng/mL.经验证,组装的试剂盒精密度为0.71%~1.59%,平均准确度为96.53%,与其他的同种属灭活病毒无交叉反应.试剂盒各项参数均符合《中国药典》(三部)2020版标准,表明该试剂盒适用于新冠病毒以及RBD抗原的快速检测.
In this study,an ELISA kit for rapid detection of severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)and its surface spike glycoprotein receptor binding domain(RBD)was prepared.The kit consists of a pair of RBD monoclonal antibodies as capture antibodies and detection antibodies which were coupled to horseradish peroxidase(HRP),and determines by square titration to the optimal operating conditions for dual antisandwich ELISA.In addition,the linearity,specificity,accuracy and precision of the detection kit were verified.The results showed that the titers of the captured antibody and the microplate detection antibody in the double-antisandwich ELISA method were 1:160000 and 1:320000 respectively,and the lower limit of quantitative detection of the kit was 31 ng/mL.Furthmore,the kits have a precision of 0.71%~1.59%and an average accuracy of 96.53%,also no cross-reactivity with other inactivated viruses of the same species.All parameters of the kit meet the 2020 standards of the Chinese Pharmacopoeia(PartⅢ),indicating that the kit is suitable for the rapid detection of SARS-CoV-2 and RBD antigen.
作者
周立娟
井申荣
苑荣亮
简建升
陈伟
邵聪文
ZHOU Li-juan;JING Shen-rong;YUAN Rong-liang;JIAN Jian-sheng;CHEN Wei;SHAO Cong-wen(College of Medicine,Kunming University of Science and Technology,Kunming 650500,Yunnan,China;Zhejiang Tianyuan Biological Pharmaceutical Co.,Ltd.,Hangzhou 311116,Zhejiang,China)
出处
《云南大学学报（自然科学版）》
CAS
CSCD
北大核心
2022年第4期870-876,共7页
Journal of Yunnan University(Natural Sciences Edition)
基金
国家自然科学基金(81860357)。
关键词
抗体效价
双抗夹心法
新型冠状病毒
捕获抗体
检测抗体
antibody titer
double antibody sandwich method
new coronavirus
capture antibody
detection antibody
