Wang Ben, Xu Jinfeng, Zhang Hui, Lian Shizhen, Duan Yichang, Zhang Hongling, Hou Wei, Yin Baishuang, Zhu Yanzhu
College of Animal Science and Technology, Jilin Agriculture Science and Technology University, Jilin City, China.
Institute of Special Animal and Plant Sciences of Chinese Academy of Agricultural Sciences, Changchun, China.
Front Vet Sci. 2024 Aug 27;11:1440124. doi: 10.3389/fvets.2024.1440124. eCollection 2024.
Canine adenovirus (CAdV) had a high prevalence in fox populations and induced fox encephalitis. No ELISA kits specifically for CAdV-1 antigen had been commercialized for foxes in China. It is crucial to develop a rapid and accurate ELISA method for detecting of CAdV-1. The monoclonal antibodies (mAbs, IgG1A) and HRP-labeled polyclonal antibodies (pAbs) were used to establish the ELISA method in this experiment. The results showed that the optimal concentration and coating time for the mAbs (IgG1A) were 2.15 μg/mL and overnight at 4°C, respectively. The dilution ratio of the HRP-labeled pAbs was 1:2000. Five percent skimmed milk was selected as the blocking agent. The optimal incubation times for blocking, CAdV-1, and HRP-labeled pAbs were all 1 h. The cut-off value for negative rectal swab was determined to be 0.366 ± 0.032. The maximum dilution ratio was 100 TCID/mL. The ELISA method was positive to CAdV-1, and that was negative to CAdV-2, Canine Parvovirus () and Canine Distempervirus (). The ELISA method showed good repeatability, sensitivity, and specificity. Compared with RT-PCR, the sensitivity, specificity, and coincidence rates of the ELISA method were 93.75, 90.9, and 92.86%, respectively. These results indicate that the established ELISA method can be used for the large-scale screening and epidemiology surveillance of CAdV-1 in foxes.
犬腺病毒（CAdV）在狐狸种群中具有高流行率，并引发狐狸脑炎。在中国，尚无专门用于检测狐狸CAdV - 1抗原的ELISA试剂盒商业化。开发一种快速准确检测CAdV - 1的ELISA方法至关重要。本实验使用单克隆抗体（mAbs，IgG1A）和辣根过氧化物酶（HRP）标记的多克隆抗体（pAbs）建立ELISA方法。结果表明，mAbs（IgG1A）的最佳浓度和包被时间分别为2.15μg/mL和4℃过夜。HRP标记的pAbs的稀释比例为1:2000。选择5%脱脂牛奶作为封闭剂。封闭、CAdV - 1和HRP标记的pAbs的最佳孵育时间均为1小时。阴性直肠拭子的临界值确定为0.366±0.032。最大稀释比例为100 TCID/mL。该ELISA方法对CAdV - 1呈阳性，对CAdV - 2、犬细小病毒（）和犬瘟热病毒（）呈阴性。该ELISA方法具有良好的重复性、敏感性和特异性。与RT - PCR相比，ELISA方法的敏感性、特异性和符合率分别为93.75%、90.9%和92.86%。这些结果表明，所建立的ELISA方法可用于狐狸中CAdV - 1的大规模筛查和流行病学监测。
