Zhang Jiajia, Sun Ziyan, Sun Shaohua, Zhang Kaili, Deng Dafu, He Ping, Zhang Pingping, Xia Nengwen, Jiang Sen, Zheng Wanglong, Meurens Francois, Zhu Jianzhong
College Veterinary Medicine, Yangzhou University, China; Joint International Research Laboratory of Agriculture and Agri-Product Safety, Yangzhou University, China; Comparative Medicine Research Institute, Yangzhou University, China; Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou University, Yangzhou 225009, China.
Swine and Poultry Infectious Diseases Research Center, Faculty of Veterinary Medicine, University of Montreal, Saint-Hyacinthe, QC J2S 2M2, Canada; Department of Veterinary Microbiology and Immunology, Western College of Veterinary Medicine, University of Saskatchewan, Saskatoon, SK S7N 5E2, Canada.
Vet Microbiol. 2025 Apr;303:110437. doi: 10.1016/j.vetmic.2025.110437. Epub 2025 Feb 19.
The African Swine Fever Virus (ASFV) poses a significant threat to the global pig farming industry. The major icosahedral capsid protein p72 plays a key role in the assembly of virus particles and infection process. As one major antigen of ASFV, p72 has been widely utilized as a marker for diagnosing infection. In this study, five p72 specific monoclonal antibodies (mAbs) were generated through the immunization of mice followed by cell fusion. Among the five hybridomas, clones 1B7, 2F3, 5D3, and 5D4 recognized a novel epitope of FHDMVGHHILGACH, while clone 1D7 recognized a new epitope, GPLLCNIHDL. Both linear epitopes were found to be highly conserved across all genotypes I and II ASFV isolates, with the first located at the pseudo hexagonal base of the p72 trimer and the latter situated at the FG insertion loop. The antigenic epitopes were capable of competing with the binding of corresponding mAbs in p72 protein based ELISA, and peptide based ELISA effectively detected ASFV antibodies in clinical samples. Furthermore, we developed and optimized a sandwich ELISA using the mAb 2F3 for efficient detection of ASFV p72 antigen. Our study not only provides valuable tools for assessing p72 function assay, but also lays the foundation for serological diagnosis, prevention and control of ASF.
非洲猪瘟病毒（ASFV）对全球养猪业构成重大威胁。主要的二十面体衣壳蛋白p72在病毒粒子组装和感染过程中起关键作用。作为ASFV的一种主要抗原，p72已被广泛用作诊断感染的标志物。在本研究中，通过免疫小鼠然后进行细胞融合，产生了五种p72特异性单克隆抗体（mAb）。在这五个杂交瘤中，克隆1B7、2F3、5D3和5D4识别出一个新表位FHDMVGHHILGACH，而克隆1D7识别出一个新表位GPLLCNIHDL。发现这两个线性表位在所有I型和II型ASFV分离株中高度保守，第一个位于p72三聚体的假六边形底部，第二个位于FG插入环。这些抗原表位能够在基于p72蛋白的ELISA中与相应mAb的结合竞争，并且基于肽的ELISA能够有效检测临床样本中的ASFV抗体。此外，我们开发并优化了一种使用mAb 2F3的夹心ELISA，用于高效检测ASFV p72抗原。我们的研究不仅为评估p72功能测定提供了有价值的工具，也为非洲猪瘟的血清学诊断、预防和控制奠定了基础。
