人偏肺病毒DNA疫苗的构建和小鼠免疫反应的研究  

Construction of human metapneumovirus DNA vaccine and study on its immune response in mice

作　　者：刘文培[1] 郑丽舒[1] 段招军[1] 谢志萍[1] 张骞[1] 张万菊[1] 侯云德[1] 

机构地区：[1]中国疾病预防控制中心病毒病预防控制所病毒基因工程国家重点实验室,北京100052

出　　处：《中华实验和临床病毒学杂志》2009年第2期100-102,共3页Chinese Journal of Experimental and Clinical Virology

摘　　要：目的构建人偏肺病毒（hMPV）DNA疫苗，小鼠免疫后评价其细胞和体液免疫水平。方法利用PCR方法，从hMPV的cDNA中扩增融合蛋白FATM（缺失跨膜区）基因和基质蛋白M基因，构建DNA疫苗pcDNA3．1His．FATM和pcDNA3．1His—M，瞬时转染后用WesternBlot和间接免疫荧光方法检测F、M蛋白表达。疫苗肌内注射免疫小鼠，ELISA和ELISPOT方法分别检测血清IgG抗体和小鼠脾细胞CTL水平。结果WesternBlot和间接免疫荧光（IFA）方法证明构建的疫苗可表达FATM和M蛋白。pcDNA3．1His—FATM单独免疫小鼠，血清抗体滴度为1：44；与pcDNA3．1His-M联合免疫后，血清抗体滴度为1：64。ELISPOT检测证明，联合免疫组小鼠脾细胞产生IFN．7的效应CD8＋T细胞数为42±8．9，高于单独免疫组32±7．4的水平。结论DNA疫苗pcDNA3．1His—FATM可以诱导产生特异性的体液和细胞免疫，与pcDNA3．1His-M联合免疫，可以提高免疫水平。Objective To construct human metapneumovirns （hMPV） DNA vaccines and evaluate the cellular and humoral immune response in mice. Methods Fusion protein FATM （without transmembrane domain） gene and M gene of hMPV were amplified from eDNA by PCR, then DNA vaccines pcDNA3.1His-FATM and pcDNA3.1His-M were constructed to verify the expression of F and M protein by Western blotting and indirect immunofluorescent assay （IFA） respectively. Serum IgG and spleen cell CTL were detected with ELISA and ELISPOT assay after the BALB/c mice were immunized intramuscularly with the vaccines. Results The candidate DNA vaccines could express FATM and M protein as detected with Western blotting and IFA. The IgG antibody titers of mice was 1：44 when immunized with pcDNA3.1His-FATM, but could increase to 1：64 when coimmunized with pcDNA3.1His-M. ELISPOT assay demonstrated that IFN-7-secreting effector T cells reached 42 ± 8.9 in co-immunization group, higher than single vaccine pcDNA3.1His-FATM group （32 ± 7.4）. Conclusion DNA vaccine pcDNA3.1His-FATM could induce specific cellular and humoral immune responses, and the immune response could increase when co-immunization with pcDNA3.1His-M was carried out.

关 键 词：变性肺病毒 疫苗 DNA 免疫 细胞 抗体生成 

分 类 号：R3[医药卫生—基础医学]