摘要
目的构建日本血吸虫铁蛋白proVAX/GMCSF-SjFerDNA疫苗,观察其诱导小鼠产生抗攻击感染的免疫保护性效果。方法小量制备pGMC/SjFer重组质粒DNA,PCR扩增GMCSF-SjFer基因,将此目的片段亚克隆到真核表达载体proVAX中,构建重组真核表达质粒proVAX/GMCSF-SjFer。雌性昆明鼠随机分为对照组、免疫组和空质粒组。空质粒组小鼠股四头肌注射proVAX,免疫组同法注射重组质粒proVAX/GMCSF-SjFer,对照组注射生理盐水。按0、2、6周设计接种3次。末次接种后第2周,感染血吸虫尾蚴(40±2)条/鼠。感染后第42d剖杀小鼠,灌注法冲虫,计数虫荷和克肝卵数。免疫前和免疫后2周留取小鼠血清,用ELISA法检测抗血吸虫成虫抗体水平。结果经PCR和双酶切鉴定,获得1条约为960bp的目的片段和2条酶切片段。免疫组小鼠的减虫率及减卵率分别为22.46%和29.40%,空质粒对照组分别为3.11%和7.79%,差异有统计学意义(P<0.05)。ELISA检测免疫组小鼠血清特异性IgG抗体水平增高。结论构建的DNA疫苗proVAX/GMCSF-SjFer能诱导小鼠产生一定水平的保护性免疫。
Objective To observe the protective immunity induced by the DNA vaccine co-expressing of GMCSF and SjFer of Schistosoma japonicum in mice.Methods The DNA of pGMC/SjFer was prepared.The gene of GMCSF-SjFer was amplified by PCR and digested with restrictive endonuclease,and directional cloned into the eukaryotic expression vector proVAX to construct the recombinant plasmid proVAX/GMCSF-SjFer.Female mice were randomly divided into control group,immunized group and plasmid proVAX group.Each mouse was injected in quadriceps femoris with plasmid proVAX（control group） and recombinant plasmid proVAX/GM-CSF-SjFer（immunized group）.Each mouse was immunized three times at 0,2,6 week,and challenged with 40±2 cercariae of S.japonicum by abdominal skin penetration 2 week post immunization.Then,the mice were sacrificed and perfused at 42 days later.The recovered worms and hepatic eggs were counted.Antibody level in the sera of mice before and two weeks after immunization was determined with ELISA.Results A 960 bp fragment was amplified by PCR,and the same fragment was obtained by restriction enzyme digestion with recombinant plasmids.Compared with the control group,the worm reduction rate was 22.46%,and its egg reduction rate was 29.40% in the immunized group.There was higher specific IgG in the serum of mice in the immunized group.Conclusion The ferritin DNA vaccine（proVAX/GMCSF-SjFer） could induce partial protective immunity against Schistosoma japonicum in mice.
出处
《中国病原生物学杂志》
CSCD
2008年第8期601-604,共4页
Journal of Pathogen Biology
基金
湖南省自然科学基金资助项目(No07JJ3031)
