甲型H1N1流感病毒样颗粒疫苗的初步研究

Preliminary study on H1N1 virus-like particle vaccine

目的对甲型H1N1病毒样颗粒的免疫原性和保护效果进行初步研究,为研发不依赖鸡胚生产工艺的流感疫苗提供新的思路。方法使用昆虫-杆状病毒表达系统表达并纯化甲型H1N1流感病毒A/California/07/2009(H1N1)的病毒样颗粒;通过Western blot、血凝、单向免疫扩散,透射电镜等方法鉴定病毒样颗粒;使用A/California/07/2009(H1N1)裂解疫苗作为对照,腹腔注射免疫Balb/c小鼠,并使用A/Beijing/501/2009(H1N1)进行攻毒,评价病毒样颗粒疫苗的免疫原性及保护效果。结果经鉴定,纯化后的病毒样颗粒血凝滴度为1:512,HA含量为92.9μg/ml,颗粒大小在100 nm左右,二免10 d后IgG抗体效价7.9×103,血凝抑制实验测得血抑(Hemagglutination Inhibition,HI)效价384,对50LD50的A/Beijing/501/2009(H1N1)病毒的保护率达到100%。结论甲型H1N1流感病毒样颗粒的免疫原性显著高于裂解疫苗,为发展不依赖鸡胚生产的流感亚单位疫苗提供了技术保证。

The pandemic virus of swine-derived influenza A（H1N1） continues to cause infection worldwide,especially in younger age groups.The manufacture process of conventional influenza vaccines relies on the growth of embryonated chicken eggs,which proved labor intensive and slow.In this study,we produced 2009 H1N1 virus-like particle（VLP） comprised Hemagglutinin（HA）,Neuraminidase（NA） and Matrix 1（M1） protein of 2009 H1N1 influenza virus in insect cells by Bac-to-Bac baculovirus expression systems.Purified 2009 H1N1 VLP was characterized by hemagglutination assay,negative staining electron microscopy,single-radial-immunodiffusion assay and Western blot.The results showed that 2009 H1N1 VLP morphologically resembled influenza virions and exhibited biological characteristics of 2009 H1N1 influenza virus.4-6 weeks old Balb/c mice were vaccinated with purified 2009 H1N1 VLP and then challenged with 2009 H1N1 virus.Results of ELISA and hemagglutination inhibition assay showed that 2009 H1N1 VLP could obviously elicit strong immune responses than split vaccinated groups.In addition,2009 H1N1 VLP provides protective immunity in mice against the 2009 pandemic H1N1 influenza virus.All the results suggest that 2009 H1N1 VLP could be an excellent influenza vaccine that should be developed further.