Cai Jiaxi, Li Zishu, Wang Yu, Fang Shuren, Fang Xiaohan, Xue Xianghong
Department of Viral Infectious Diseases of Special Animals, Institute of Special Animal and Plant Sciences, Chinese Academy of Agricultural Sciences, Changchun, China; Jilin Provincial Key Laboratory of Special Economic Animal Molecular Biology, Institute of Special Animal and Plant Sciences, Chinese Academy of Agricultural Sciences, Changchun, China.
Department of Viral Infectious Diseases of Special Animals, Institute of Special Animal and Plant Sciences, Chinese Academy of Agricultural Sciences, Changchun, China; Jilin Provincial Key Laboratory of Special Economic Animal Molecular Biology, Institute of Special Animal and Plant Sciences, Chinese Academy of Agricultural Sciences, Changchun, China.
J Virol Methods. 2025 Apr;333:115098. doi: 10.1016/j.jviromet.2024.115098. Epub 2024 Dec 9.
Hundreds of millions of the domestic dogs worldwide are routinely inoculated with the modified live vaccines for canine distemper virus (CDV) every year. However, the corresponding serological diagnostic and detections are always lacking, thus, there is an urgent demand to establish its unique diagnostic technologies to produce high-quality antigenic biomolecules. In the present study, the ectodomain (et) of CDV hemagglutinin (H) protein was firstly expressed in a soluble and secreted forms by an Expi293F transient transfection system based on its antigenic secondary structure analysis. The yields of CDV H(et) protein was 2.6 g/L with purity over 95 % in supernatant of Expi293F cells. The CDV H(et) protein elicited comparative antibodies levels to the CDV virions in rabbit by ELISA and neutralization test. The purified polyclonal antibodies of immunized with CDV H(et) protein recognized both wild-type and vaccine CDV strains. More importantly, the purified polyclonal antibodies of CDV H(et) protein revealed significantly higher viral neutralizing activity than those from CDV-3 virions, which highlighted that the critical role of CDV H protein to elicit viral specific and protective neutralizing antibodies. Taken together, the CDV H(et) protein produced in mammalian expression systems was high-quality and good immunogenicity, and would be with great potential to serve as a serological diagnostic antigen or a novel CDV subunit vaccine in future.
全球每年有数亿只家养犬常规接种犬瘟热病毒（CDV）的减毒活疫苗。然而，相应的血清学诊断和检测方法一直缺乏，因此，迫切需要建立其独特的诊断技术以生产高质量的抗原生物分子。在本研究中，基于CDV血凝素（H）蛋白的抗原二级结构分析，首先通过Expi293F瞬时转染系统以可溶性和分泌形式表达其胞外域（et）。Expi293F细胞上清中CDV H（et）蛋白的产量为2.6 g/L，纯度超过95%。通过ELISA和中和试验，CDV H（et）蛋白在兔体内诱导出与CDV病毒粒子相当的抗体水平。用CDV H（et）蛋白免疫纯化的多克隆抗体可识别野生型和疫苗CDV毒株。更重要的是，CDV H（et）蛋白纯化的多克隆抗体显示出比CDV - 3病毒粒子纯化的多克隆抗体显著更高的病毒中和活性，这突出了CDV H蛋白在诱导病毒特异性和保护性中和抗体中的关键作用。综上所述，在哺乳动物表达系统中产生的CDV H（et）蛋白质量高、免疫原性好，未来作为血清学诊断抗原或新型CDV亚单位疫苗具有巨大潜力。
