【摘要】 目的　研究联合运用狂犬病毒糖蛋白基因重组腺病毒与同一抗原核酸疫苗对小鼠的免疫效果。方法　构建狂犬病毒糖蛋白 (GP)的真核表达质粒pcDNA3.1 CVS N2cGP作为核酸疫苗 ,瞬时转染COS 7细胞 ,间接免疫荧光试验检测pcDNA3.1 CVS N2cGP的表达 ;以基因枪方法进行初次免疫接种和首次加强免疫 ,以表达同一抗原的复制缺陷型重组腺病毒通过鼻腔接种进行第二次加强免疫 ;ELISA试验检测血清狂犬病毒特异性IgG抗体 ,快速荧光灶抑制试验 (RFFIT)检测狂犬病毒中和抗体。结果　间接免疫荧光试验表明pcDNA3.1 CVS N2cGP能有效地表达GP于转染的细胞膜上 ,ELISA试验表明小鼠接受基因枪核酸免疫后 ,仅诱导产生低水平的特异性抗体 ,而用重组腺病毒进行加强免疫后 ,特异性抗体水平显著提高。结论　联合运用狂犬病毒糖蛋白基因重组腺病毒与同种抗原核酸疫苗可克服它们单独使用时各自的缺点 ,能有效地诱导小鼠产生抗狂犬病毒特异免疫。更多还原

【Abstract】 Objective To investigate the immune response of mice primed by DNA particle bombing and boosted by immunization with replication-defective adenoviral recombinant bearing rabies glycoprotein gene. Methods The full-length glycoprotein gene of rabies virus CVS-N2c was inserted into plasmid pcDNA3.1. COS-7 cells were transfected with the recombinant plasmid pcDNA3.1/CVS-N2c GP and the expression of GP was determined by indirect immune fluorescent test. Mice were immunized with a heterogeneous prime-booster regime: First, DNA particle bombing vaccination was conducted twice by helium gas drived gene gun at a interval of two weeks. Then, replication defective adenoviral recombinant was used for booster at fifteenth weeks. The level of rabies virus specific antibody was measured by ELISA and the neutralizing antibody was titrated by a rapid fluorescent focus inhibition test (RFFIT). Results Indirect immune fluorescent test indicated that the glycoprotein was successfully expressed on the surface of COS-7 cells. Low level titre of specific antibody was induced by DNA immunization. RFFIT results showed that the neutralizing antibody reached 0.66 IU/ml after secondary DNA immunization and dramatic rising of neutralizing antibody was observed when the mice was boosted with 2×10 6 PFU recombinant replication defective adenovirus, the mean titre reached 90 IU/ml at seventeenth weeks. Conclusion Combined immunization of DNA vaccine and replication-defective recombinant adenovirus bearing rabies CVS-N2c glycoprotein gene induced potential immune response against rabies virus.更多还原