人博卡病毒VP2病毒样颗粒的表达、纯化及鉴定

Expression, purification and identification of human bocavirus VP2 virus-like particles

目的通过杆状病毒-昆虫细胞表达体系，获得纯重组人博卡病毒（rmoV）病毒蛋白2（VP2）病毒样颗粒，为HBoV的血清学检测方法的建立及疫苗研究提供实验基础。方法采用含有HBoV VP2基因的重组杆状病毒感染草地夜蛾9（SF9）细胞，一定时间后收集细胞及上清，经氯化铯（CsCI）不连续密度梯度超速离心后，获得高纯度人博卡病毒VP2病毒样颗粒。SDS—PAGE鉴定目的蛋白的纯度和分子量，透射电镜观察病毒样颗粒结构，Western blotting鉴定蛋白的抗原性。结果SDS—PAGE结果显示该研究获得的重组HBoV VP2病毒样颗粒分子量约为60kD，无明显杂带；该研究获得的HBoV VP2病毒样颗粒能和鼠抗人博卡病毒VP2多克隆抗体结合；电镜下HBoV VP2病毒样颗粒为对称正二十面体结构，直径约为20—25 nm。结论该研究成功获得高纯度的重组HBoV VP2病毒样颗粒，可用于人博卡病毒感染的血清学及免疫学研究。

[Objectives] To obtain pure recombinant human bocavirus （HBoV） VP2 virus-like particlesthrough the baculovirus-insect cell expression system, and provide experimental basis for establishment ofHBoV serological detection method and vaccine research. [Methods] Firstly, we made Spodoptera Frugiperda9 （SF9） cells with recombinant baculovirus who contains HBoV VP2 gene was infected, then collected cellsand the supematant after a certain timethe cesium chloride （CsC1） discontinuous density gradient over-speedcentrifugal was used to purify bocavirus VP2 virus-like particles. SDS-PAGE was used to appraisal proteinpurity and molecular weight, transmission electron microscope observe virus-like particles structure, Westernblotting identified the antigenicity of protein. [Results] In this study, the results of SDS-PAGE shows thatthe molecular weight of HBoV VP2 virus-like particles were about 60 kD, no obvious impurity; the HBoVVP2 virus-like particles obtained in this study can blind to mice anti-HBoV VP2 polyclonal antibody; Underthe electron microscope, HBoV VP2 virus-like particles is ieosahedral symmetry and approximately 20 nm-25nm in diameter. [Conclusion] This study successfully obtained high purity HBoV VP2 virus-like particles,and it can be used to bocavirus serology and immunology research.