Gao Zichen, Hu Jianing, Cai Yiqin, Liu Ye, Yin Guihu, Guo Xinyu, Wang Ruiying, Zhong Meng, Liu Qingtao, Feng Xiuli
Key Laboratory of Animal Microbiology of China's Ministry of Agriculture, College of Veterinary Medicine, Nanjing Agricultural University, Nanjing 210095, China.
MOE Joint International Research Laboratory of Animal Health and Food Safety, College of Veterinary Medicine, Nanjing Agricultural University, Nanjing 210095, China.
Viruses. 2025 Mar 24;17(4):464. doi: 10.3390/v17040464.
Avian infectious bronchitis is caused by the avian infectious bronchitis virus (IBV), which poses a significant threat to the poultry industry and public health. The S1 protein of IBV plays a crucial role in the process of the virus invading host cells. To investigate the significant antigenic targets within the S1 protein, in this study, the truncated S1 sequence of the IBV M41 strain was cloned with approximately 660 bp and expressed. After purification and renaturation, the recombinant S1 protein was immunized into BALB/c mice. Then, following fusion with lymphocytes and SP2/0 cells, the indirect ELISA and Western blotting techniques were employed to screen hybridoma cell lines secreting monoclonal antibodies (mAbs) targeting the S1 protein. Antigenic epitopes of the mAbs were identified using truncated S1 fragments and peptide scanning. The results indicated that three hybridoma cell lines stably secreting S1 protein-specific mAbs (2A10, 4E9, and 5E12) were screened. The heavy chains of the three mAbs were IgG1, and all three mAbs contained kappa light chains. The identified minimal B-cell epitopes were RVSAMK and FYNLTV. Homology analysis showed these both epitopes were conserved across IBV subtypes and located on the S1 protein surface. The conserved β-sheet epitope RVSAMK and the surface-exposed, flexible loop epitope FYNLTV serve as ideal targets for broad-spectrum diagnostics and early infection detection, respectively. These epitopes provide unique structural advantages for antibody binding, enabling the design of multivalent epitope vaccines or the development of immunomodulatory drugs. They offer novel biomaterials and targets for antibody-based drug development and rapid detection methods for avian infectious bronchitis virus (IBV), holding significant potential for the prevention and control of IBV.
禽传染性支气管炎由禽传染性支气管炎病毒（IBV）引起，对家禽业和公共卫生构成重大威胁。IBV的S1蛋白在病毒侵入宿主细胞的过程中起关键作用。为了研究S1蛋白内的重要抗原靶点，本研究克隆了IBV M41株约660 bp的截短S1序列并进行表达。纯化复性后，将重组S1蛋白免疫BALB/c小鼠。然后，将淋巴细胞与SP2/0细胞融合后，采用间接ELISA和Western印迹技术筛选分泌靶向S1蛋白单克隆抗体（mAb）的杂交瘤细胞系。使用截短的S1片段和肽扫描鉴定mAb的抗原表位。结果表明，筛选出了3株稳定分泌S1蛋白特异性mAb（2A10、4E9和5E12）的杂交瘤细胞系。这3株mAb的重链均为IgG1，且均含有κ轻链。鉴定出的最小B细胞表位为RVSAMK和FYNLTV。同源性分析表明，这两个表位在IBV各亚型中均保守，且位于S1蛋白表面。保守的β-折叠表位RVSAMK和表面暴露的柔性环表位FYNLTV分别作为广谱诊断和早期感染检测的理想靶点。这些表位为抗体结合提供了独特的结构优势，有助于设计多价表位疫苗或开发免疫调节药物。它们为基于抗体的药物开发和禽传染性支气管炎病毒（IBV）快速检测方法提供了新型生物材料和靶点，在IBV的防控方面具有巨大潜力。
