犬细小病毒样颗粒的制备及免疫原性分析

Construction and immune effect evaluation of canine parvovirus like particles

犬细小病毒病是由犬细小病毒(canine parvovirus,CPV)引起犬的一种急性、高度传染性和致死性的病毒性疾病。该病主要通过接种疫苗预防,没有特效治疗方法。为了制备犬细小病毒样颗粒(virus like particles,VLPs),本研究通过杆状病毒表达系统表达CPV VP2蛋白。将表达后的蛋白经过蔗糖密度梯度离心法纯化后,负染色处理置于透射电子显微镜下观察。结果显示VP2蛋白组装成直径25 nm左右的六边形或圆形样VLPs。血凝试验表明,该VLPs可以凝集猪红细胞,血凝价为27。进一步将该VLPs经弗氏佐剂乳化后免疫小鼠,血凝抑制试验结果显示VLPs可诱导小鼠产生211的血凝抑制抗体。因此,通过杆状病毒表达系统表达VP2蛋白,VP2蛋白可在细胞内组装成与病毒粒子类似的VLPs,该VLPs可以诱导小鼠产生血凝抑制抗体,为CPV VLPs疫苗的制备奠定了基础。

Canine parvovirus disease is an acute,highly contagious,and fatal viral disease caused by Canine parvovirus(CPV) in dogs.The disease is mainly prevented by vaccination and there is no specific treatment method.In order to produce canine parvovirus like particles(VLPs),the CPV VP2 protein was expressed using a baculovirus expression system.The expressed proteins were subsequently purified through sucrose density gradient centrifugation and negatively stained for observation under transmission electron microscopy.The results showed that the VP2 protein assembled into hexagonal or circular VLPs with a diameter of approximately 25 nm.The results of hemagglutination test demonstrated that the VLPs were capable of agglutinating pig red blood cells,with a hemagglutination value of 27.Furthermore,the VLPs were emulsified with Freund's adjuvant and administered to mice as an immunization strategy.The hemagglutination inhibition test results revealed that the VLPs elicited the production of 211 hemagglutination inhibition antibodies in mice.Hence,VP2 protein can be expressed through baculovirus expression system to assemble into VLPs,which is similar to CPV virion.Generally,CPV VLPs produced through baculovirus expression system can elicit the production of hemagglutination inhibitory antibodies in mice,thereby providing a basis for the development of CPV VLPs vaccines.