基于新型p22和p30双蛋白组合的间接ELISA法检测非洲猪瘟病毒抗体
Novel p22 and p30 dual-proteins combination based indirect ELISA for detecting antibodies against African swine fever virus.
作者信息
Li Jianda, Jiao Jian, Liu Na, Ren Sufang, Zeng Hao, Peng Jun, Zhang Yuyu, Guo Lihui, Liu Fei, Lv Tingting, Chen Zhi, Sun Wenbo, Hrabchenko Nataliia, Yu Jiang, Wu Jiaqiang
机构信息
Shandong Key Laboratory of Animal Disease Control and Breeding, Institute of Animal Science and Veterinary Medicine, Shandong Academy of Agricultural Sciences, Jinan, China.
School of Veterinary Medicine, Qingdao Agricultural University, Qingdao, China.
出版信息
Front Vet Sci. 2023 Feb 10;10:1093440. doi: 10.3389/fvets.2023.1093440. eCollection 2023.
INTRODUCTION
African swine fever virus (ASFV) infection is one of the most complex and fatal hemorrhagic viral diseases, causing a devastating loss to the swine industry. Since no effective vaccine is available, prevention and control of ASFV heavily depends on early diagnostic detection.
METHODS
In this study, a novel indirect ELISA was established for detecting antibodies against ASFV using dual-proteins, p22 and p30. Recombinants p22 and p30 were expressed and purified from vector system by recombined plasmids pET-KP177R and pET-CP204L. p22 and p30 were mixed as antigens for developing the indirect ELISA.
RESULTS
Through optimizing coating concentrations of p30 and p22, coating ratio (p30: p22 = 1:3), and serum dilution (as 1:600), the established ELISA performed higher specificity, sensitivity, and repeatability against ASFV-positive serum. Furthermore, 184 clinical serum samples from suspected diseased pigs were verified the established ELISA in clinical diagnosis. The results showed that compared with two commercial ELISA kits, the established ELISA possessed higher sensitivity and almost uniform coincidence rate.
CONCLUSION
The novel indirect ELISA based on dual-proteins p30 and p22 performed a valuable role in diagnostic detection of ASFV, providing a broad insight into serological diagnostic methods of ASFV.
引言
非洲猪瘟病毒（ASFV）感染是最复杂且致命的出血性病毒病之一，给养猪业带来了毁灭性损失。由于尚无有效疫苗，ASFV的预防和控制在很大程度上依赖于早期诊断检测。
方法
在本研究中，利用双蛋白p22和p30建立了一种新型间接ELISA法来检测抗ASFV抗体。通过重组质粒pET-KP177R和pET-CP204L从载体系统中表达并纯化重组体p22和p30。将p22和p30混合作为抗原以开发间接ELISA。
结果
通过优化p30和p22的包被浓度、包被比例（p30:p22 = 1:3）以及血清稀释度（为1:600），所建立的ELISA对ASFV阳性血清具有更高的特异性、敏感性和重复性。此外，对来自疑似患病猪的184份临床血清样本在临床诊断中验证了所建立的ELISA。结果表明，与两种商业ELISA试剂盒相比，所建立的ELISA具有更高的敏感性和几乎一致的符合率。
结论
基于双蛋白p30和p22的新型间接ELISA在ASFV的诊断检测中发挥了重要作用，为ASFV的血清学诊断方法提供了广阔的视野。
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