College of Veterinary Medicine, Yangzhou University, Yangzhou 225009, China.
Joint International Research Laboratory of Agriculture and Agri-Product Safety, The Ministry of Education of China, Yangzhou University, Yangzhou 225009, China.
Int J Mol Sci. 2023 Sep 11;24(18):13934. doi: 10.3390/ijms241813934.
Porcine deltacoronavirus (PDCoV) is an emerging virus that poses a significant threat to the global swine industry. Its membrane (M) protein is crucial for virion assembly and virus-host interactions. We selected the hydrophilic region of M protein for prokaryotic expression, purification, and recombinant protein production. Utilizing hybridoma technology, we prepared the monoclonal antibody (mAb) 24-A6 against M protein. The mAb 24-A6 was shown to be suitable for use in immunofluorescence assays, western blotting, and immunoprecipitation, with specificity for PDCoV and no cross-reactivity with other five porcine viruses. The M protein was observed to be expressed as early as 3 h after PDCoV infection, increasing its expression over the duration of infection. Notably, the antigenic epitope of the M protein identified as SPESRL recognized by mAb 24-A6 was found within a conserved structural domain (SWWSFNPETNNL) of the coronavirus M protein, indicating a crucial overlap between a functionally important viral assembly region and a region recognized by the immune system. Our findings provide valuable insights into mAb 24-A6 targeting the antigenic epitope of M protein and may contribute to the development of diagnostic tools for PDCoV infection and fundamental research into the function of PDCoV M protein.
猪德尔塔冠状病毒（PDCoV）是一种新兴病毒，对全球养猪业构成重大威胁。其膜（M）蛋白对于病毒粒子组装和病毒-宿主相互作用至关重要。我们选择 M 蛋白的亲水区域进行原核表达、纯化和重组蛋白生产。利用杂交瘤技术，我们制备了针对 M 蛋白的单克隆抗体（mAb）24-A6。mAb 24-A6 适合用于免疫荧光分析、western blot 和免疫沉淀，特异性针对 PDCoV，与其他五种猪病毒无交叉反应。PDCoV 感染后 3 小时即可观察到 M 蛋白的表达，随着感染时间的延长，其表达逐渐增加。值得注意的是，mAb 24-A6 识别的 M 蛋白抗原表位 SPESRL 位于冠状病毒 M 蛋白的保守结构域（SWWSFNPETNNL）内，表明在一个功能重要的病毒组装区域和一个被免疫系统识别的区域之间存在关键重叠。我们的研究结果为 mAb 24-A6 针对 M 蛋白抗原表位提供了有价值的见解，并可能有助于开发用于 PDCoV 感染的诊断工具和对 PDCoV M 蛋白功能的基础研究。
