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目的:构建柯萨奇病毒B3(CVB3)VP1基因重组腺病毒Ad/sVP1-C3d3,并观察对小鼠的免疫效果。方法:利用AdEasy-1系统构建、包装重组腺病毒Ad/sVP1-C3d3。BALB/c小鼠随机分为Ad/sVP1-C3d3、Ad/VP1、Ad和PBS4组,肌肉注射免疫,共免疫2次,每次间隔16d。用ELISA法和微量中和试验法分别检测血清CVB3VP1IgG和中和抗体滴度;CCK-8法检测特异性CTL杀伤活性;用致死量CVB3攻击小鼠后,检测血中病毒滴度。结果:成功构建、包装了重组腺病毒Ad/sVP1-C3d3。免疫小鼠后,Ad/sVP1-C3d3组血清CVB3VP1IgG滴度和中和抗体水平明显高于Ad/VP1(P<0.01),CTL杀伤活性明显高于Ad和PBS对照组(P<0.01)及Ad/VP1组(P<0.05),血清病毒滴度低于Ad和PBS对照组(P<0.05)。结论:重组腺病毒Ad/sVP1-C3d3能明显提高小鼠的细胞和体液免疫应答并降低血液病毒载量。
Objective: To construct the recombinant adenovirus Ad / sVP1-C3d3 of Coxsackievirus B3 (CVB3) VP1 gene and observe its immune effect on mice. Methods: AdEasy-1 system was constructed and packaged recombinant adenovirus Ad / sVP1-C3d3. BALB / c mice were randomly divided into Ad / sVP1-C3d3, Ad / VP1, Ad and PBS4 groups, immunized intramuscularly and co-immunized twice with 16d intervals. Serum CVB3VP1 IgG and neutralizing antibody titers were detected by ELISA and micro-neutralization test respectively. CCK-8 method was used to detect the specific CTL killing activity. After the mice were challenged with the lethal dose of CVB3, the virus titers in the blood were measured. Results: The recombinant adenovirus Ad / sVP1-C3d3 was successfully constructed and packaged. After immunization, CVB3VP1IgG titer and neutralizing antibody level in serum of Ad / sVP1-C3d3 group were significantly higher than that of Ad / VP1 (P <0.01), CTL cytotoxicity was significantly higher than Ad and PBS control group (P <0.01) / VP1 group (P <0.05). The titer of serum virus was lower than Ad and PBS control group (P <0.05). Conclusion: The recombinant adenovirus Ad / sVP1-C3d3 can significantly improve the cellular and humoral immune response and reduce the blood viral load in mice.