识别呼吸道合胞病毒融合后F蛋白的单克隆抗体制备及应用

Preparation and Application of A Monoclonal Antibody Targeting the Postfusion F Protein of the Respiratory Syncytial Virus

人呼吸道合胞病毒(Human respiratory syncytial virus,HRSV)是引起5岁以下儿童急性下呼吸道感染的主要病原体之一,至今尚无安全有效的预防RSV感染的疫苗。融合(F)蛋白是目前RSV疫苗研发的主要抗原靶标,存在prefusion(pre⁃F)构象和postfusion(post⁃F)构象,而pre⁃F构象蛋白上含有诱导机体产生高效中和抗体的中和表位。因此,在靶向pre⁃F蛋白疫苗研发前期工作中,对于F蛋白两种构象形式相对含量的评估可以进一步指导RSV疫苗免疫原再设计。目前已报道的F蛋白抗体大多识别pre构象,本研究拟开发出一种特异性识别post⁃F蛋白的工具抗体,希望能够评估post⁃F蛋白的相对含量。通过RSV A2 post⁃F重组蛋白免疫小鼠获得了特异性识别post⁃F蛋白的鼠源抗体12A12,结果显示12A12对于post⁃F蛋白具有良好的广谱结合活性,并且对不同来源的post⁃F蛋白在不同方法学应用上都有一定的检测能力,表现出成为特异性检测post⁃F蛋白工具抗体的潜力。

The human respiratory syncytial virus(HRSV)is one of the main pathogens causing acute infections of the lower respiratory tract in children under 5 years of age.A safe and efficacious RSV vaccine is not available.Fusion(F)protein is present in two conformations:prefusion(pre⁃F)and postfusion(post⁃F),which is the main antigen target for RSV vaccine development.The pre⁃F protein conformation contains neutralizing epitopes inducing highly potent neutralizing antibodies.Therefore,in the preliminary research of development of vaccines using pre⁃F protein,evaluation of the relative content of the two conformational forms of F protein can provide guidance for the immunogen redesign of the RSV vaccine.Most F⁃protein antibodies of the RSV reported previously recognize the pre⁃F conformation.We intended to develop a“tool”antibody that specifically recognizes and can evaluate the relative content of post⁃F protein.The murine monoclonal antibody 12A12 was obtained by immunizing mice with RSV A2 post⁃F recombinant protein.Results showed that 12A12 had broad⁃spectrum binding activity for post⁃F protein and certain capabilities of detecting post⁃F from different sources in different methodological applications,indicating the potential to serve as a tool antibody to identify post⁃F protein.