柯萨奇病毒B1/B3型二价VP1基因疫苗的构建及其免疫试验  

A bivalent VP1 gene vaccine against Coxsackie virus B1/B3

作　　者：吴晓羽[1] 赵铁强[1] 田野[1] 沈景霞[1] 孟繁超[1] 刘雅君[2] 王乐民[2] 李晓光[2] 钟照华[3] 李呼伦[3] 

机构地区：[1]哈尔滨医科大学第一临床医学院心内科,150001 [2]哈尔滨市第一医院心内科 [3]哈尔滨医科大学微生物教研室

出　　处：《中华医学杂志》2001年第8期480-484,共5页National Medical Journal of China

基　　金：国家自然科学基金资助项目 (30 0 70 30 1)

摘　　要：目的　构建柯萨奇病毒 (CV)B1/B3型二价VP1基因免疫质粒 ,并探讨其免疫原性。方法　通过逆转录聚合酶链反应 (RT PCR)技术扩增CVB1的主要中和抗原VP1基因 ,将分别由CMV和SV40启动子控制的CVB1和CVB3的VP1基因反向串联插入到真核表达载体pCR3 Uni中。将该质粒转染鼠NIH3T3细胞 ,于转染后 2 4、48、96h进行免疫荧光及RT PCR检测。应用该重组质粒免疫BALB/c小鼠 ,于免疫前、免疫后 2、4、6周分别断尾采血 ,用ELISA方法检测CVB特异性抗体。结果　获得了含有CVB1和CVB3的VP1基因免疫质粒 ,RT PCR及免疫荧光检测结果表明该质粒在NIH3T3细胞获得瞬时表达 ,小鼠免疫后 4、6周均有特异性抗体产生。结论　本研究构建的CVB1/B3型二价VP1基因疫苗能诱导小鼠产生CVB特异性抗体 ,可以作为候选基因疫苗。Objective To construct a bivalent VP1 gene vaccine against Coxsackie virus B1 and B3 (CVB1 and CVB3) and to test its immunogenicity in mice. Methods The VP1 gene of CVB1 was amplified by RT PCR. The VP1 genes of CVB1 and of CVB3 under the control of cytomegalovirus(CMV)promoter and SV40 promoter respectively were inserted oppositely into the pCR3 Uni, a eukaryotic expression vector. The resultant bivalent gene vaccine plasmid was used to inoculate 3 week old BALB/c mice. Immunofluorescence test and RT PCR were conducted 24, 48, and 96 hours after the transfection. Sera of the tested animals were sampled at 0, 2, 4, and 6 weeks after the inoculation and detected for CVB specific antibodies by ELISA. Results A bivalent gene vaccine plasmid of CVB1 and CVB3 was obtained. Samples of 4 and 6 weeks after the inoculation were shown to be CVB antibody positive, while those of 0 and 2 weeks after the inoculaiton shown negative. Conclusion The gene vaccine thus constructed induces CVB speicific antibodies in mice, and can serve as gene vaccine candidate for human beings.

关 键 词：柯萨奇病毒 基因疫苗 免疫试验 病毒性心肌炎 

分 类 号：R512.5[医药卫生—内科学]