Fu Yuanfang, Li Dong, Cao Yimei, Zhou Peng, Li Kun, Zhao Zhixun, Li Pinghua, Bai Xingwen, Bao Huifang, Wang Songtai, Zhao Lixia, Wang Xiaoxiao, Liu Zaixin, Sun Pu, Lu Zengjun
State Key Laboratory of Veterinary Etiological Biology, College of Veterinary Medicine, Lanzhou University, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, National Foot-and-Mouth Disease Reference Laboratory, Lanzhou 730000, PR China.
Lanzhou Animal Health Supervision, Lanzhou 730000, PR China.
J Virol Methods. 2023 Apr;314:114676. doi: 10.1016/j.jviromet.2023.114676. Epub 2023 Jan 18.
Foot-and-mouth disease (FMD) is a highly contagious and economically devastating disease of cloven-hoofed animals. Vaccination and surveillance against non-structure protein (NSP) are the most efficacious and cost-effective strategy to control this disease. Therefore, vaccine purity control is vital for successful prevention. Currently, vaccine purity is tested by an in-vivo test that recommended in the World Organization for Animal Health (WOAH), but it is time consuming and costly. Herein, we develop a double-antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA) for quantitative detection of residual NSPs in inactivated FMD virus (FMDV) vaccines. In this assay, the monoclonal antibody 3A24 was selected as capture antibody and biotinylated 3B4B1 (Biotin-3B4B1) as detection antibody. A standard curve was developed using the NSP 3AB concentration versus OD value with the linear range of concentration of 2.5-160 ng/mL. The lowest limit of detection was 2.5 ng/mL. In addition, we determined 2.5 ng/mL of NSP as an acceptable threshold value of FMD vaccine purity using a dose-response experiment in cattle. The DAS-ELISA combined with the threshold value of FMD vaccine purity could provide a quick and simple tool for evaluation the antigenic purity of FMD vaccine during the manufacturing process.
口蹄疫（FMD）是一种偶蹄动物的高度传染性且对经济有严重破坏的疾病。针对非结构蛋白（NSP）的疫苗接种和监测是控制该疾病最有效且具成本效益的策略。因此，疫苗纯度控制对于成功预防至关重要。目前，疫苗纯度通过世界动物卫生组织（WOAH）推荐的体内试验进行检测，但该方法耗时且成本高。在此，我们开发了一种双抗体夹心酶联免疫吸附测定法（DAS - ELISA），用于定量检测灭活口蹄疫病毒（FMDV）疫苗中的残留NSP。在该测定法中，选择单克隆抗体3A24作为捕获抗体，生物素化的3B4B1（生物素 - 3B4B1）作为检测抗体。利用NSP 3AB浓度与OD值绘制标准曲线，浓度线性范围为2.5 - 160 ng/mL。最低检测限为2.5 ng/mL。此外，我们通过在牛身上进行的剂量反应实验确定2.5 ng/mL的NSP为口蹄疫疫苗纯度的可接受阈值。结合口蹄疫疫苗纯度阈值的DAS - ELISA可为评估口蹄疫疫苗生产过程中的抗原纯度提供一种快速简便的工具。
