Expression and characterization of canine distemper virus hemagglutinin protein in suspension mammalian cells.

IF 2.2 4区 医学 Q3 BIOCHEMICAL RESEARCH METHODS Journal of virological methods Pub Date : 2024-12-09 DOI:10.1016/j.jviromet.2024.115098
Jiaxi Cai, Zishu Li, Yu Wang, Shuren Fang, Xiaohan Fang, Xianghong Xue
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引用次数: 0

Abstract

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.

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犬瘟热病毒血凝素蛋白在悬浮哺乳动物细胞中的表达及特性研究。
全世界每年有数亿只家养狗常规接种犬瘟热病毒(CDV)改良活疫苗。然而,相应的血清学诊断和检测一直缺乏,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。本研究通过对CDV血凝素(H)蛋白的抗原二级结构分析,利用Expi293F瞬时转染系统,首次以可溶性和分泌形式表达了CDV血凝素(H)蛋白的外结构域(et)。在Expi293F细胞上清液中CDV H(et)蛋白的产量为2.6g/L,纯度在95%以上。CDV H(et)蛋白通过酶联免疫吸附和中和试验在家兔体内引起了对CDV病毒粒子的比较抗体水平。纯化的CDV H(et)蛋白免疫多克隆抗体对CDV野生型和疫苗型均有识别。更重要的是,纯化的CDV H(et)蛋白多克隆抗体显示出比CDV-3病毒粒子更高的病毒中和活性,这突出了CDV H蛋白在引发病毒特异性和保护性中和抗体方面的关键作用。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,成本低,在未来作为一种血清学诊断抗原或新型CDV亚单位疫苗具有很大的潜力。
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来源期刊
CiteScore
5.80
自引率
0.00%
发文量
209
审稿时长
41 days
期刊介绍: The Journal of Virological Methods focuses on original, high quality research papers that describe novel and comprehensively tested methods which enhance human, animal, plant, bacterial or environmental virology and prions research and discovery. The methods may include, but not limited to, the study of: Viral components and morphology- Virus isolation, propagation and development of viral vectors- Viral pathogenesis, oncogenesis, vaccines and antivirals- Virus replication, host-pathogen interactions and responses- Virus transmission, prevention, control and treatment- Viral metagenomics and virome- Virus ecology, adaption and evolution- Applied virology such as nanotechnology- Viral diagnosis with novelty and comprehensive evaluation. We seek articles, systematic reviews, meta-analyses and laboratory protocols that include comprehensive technical details with statistical confirmations that provide validations against current best practice, international standards or quality assurance programs and which advance knowledge in virology leading to improved medical, veterinary or agricultural practices and management.
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