摘要
目的构建柯萨奇病毒B组 3型 (CoxsackievirusesgroupB3,CVB3)VP1基因的真核表达重组质粒pcDNA3/VP1,并观察它对小鼠的免疫保护作用。方法从CVB3感染细胞中扩增出VP1片段 ,构建真核表达重组质粒 pcDNA3/VP1,经大量扩增纯化后 ,肌内注射免疫BALB/c小鼠 ,每次免疫后的第 6天 ,断尾取血检测CVB3中和抗体 ;3次免疫后用致死量 (10 0 0TCID50 )的CVB3感染小鼠 ,观察 pcDNA/VP1对小鼠的免疫保护作用。 结果pcDNA3/VP1重组质粒免疫小鼠后 ,能诱导机体产生中和抗体 ,各次免疫后抗体滴度分别为 <1∶5 ,1∶5 .7和1∶34.8;用致死量CVB3感染小鼠 ,pcDNA3/VP1重组质粒免疫组、pcDNA3质粒对照组及生理盐水对照组生存率分别为 30 %、2 0 %及 15 % ,3组小鼠生存率无明显差异 (P >0 .0 5 )。结论 pcDNA3/VP1重组质粒在小鼠体内能够表达并能诱导机体产生中和抗体 ,抗体滴度随免疫次数增加 。
ObjectiveTo construct the eukaryotic expression plasmid pcDNA3/VP1 of Coxsackieviruses group B3(CVB3) and to observe its protective effect on mice.MethodsThe VP1 segment was amplified from CVB3 infected HeLa cells with RT PCR and insert into eukaryotic expression plasmid pcDNA3. E.Coli DH 5α was transformed and pcDNA3/VP1 DNA was prepared and purified. BALB/c mice were immunized with this DNA vaccine by tibialis anterio muscle injecting. Sera were harvested from immune mice after every immunization and stored for neutralization antibody titer. The mice were infected with 1 000 TCID 50 CVB3 after three times of inoculation and the pretective effects of pcDNA3/VP1 were observed.ResultsNeutralizing antibodies of immune mice were produced after immunization with eukaryotic expression plasmid pcDNA3/VP1. The titers of antibody were < 1∶5,1∶ 5.7 and 1∶ 34.8 , respectively. The survival rate of pcDNA3/VP1 immunizated group, pcDNA3 immunizated control group, Saline control group was 30 %? 20 % and 15 %, respectively. The survival rate of the three groups had no significant difference(P> 0.05 ).ConclusionThe eukaryotic expression plasmid pcDNA3/VP1 could express in mice and induce mice to produce neutralizing antibodies. The titer of antibody was increased along with the immunization, but the level of antibody was low and there was no evident protective effect in mice immuned with pcDNA3/VP1 to lethal dose infection of CVB3.
出处
《河北医科大学学报》
CAS
2002年第3期133-136,共4页
Journal of Hebei Medical University
