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目的 构建高效表达结核分枝杆菌(Mycobacterium tuberculosis,MTB) ESAT6、rv3407、RpfB三个显性抗原基因(3 antigens,3Ag) 的重组人5型腺病毒(Adenovirus 5,Ad5) 载体疫苗,在小鼠模型中初步完成药效学研究.方法 用AdEasyTM XL腺病毒载体系统构建重组质粒pAd5-3Ag,经Pac I线性化后转染AD293细胞获得表达3Ag融合蛋白的Ad5-TPA-3Ag; 采用Western blot检测腺病毒载体中融合蛋白的表达; 将纯化的病毒颗粒以肌肉注射和滴鼻两种方式免疫小鼠,采用ELISA检测血清中抗体效价,对病毒的免疫剂量进行优化.结果 含3Ag基因的重组腺病毒载体构建成功.包装的病毒Ad5-TPA-3Ag以8 × 107 IFU的剂量免疫小鼠42d时各抗原的效价最高,ESAT6肌肉注射和滴鼻的最高效价分别为4. 54和4. 87; rv3407肌肉注射和滴鼻的最高效价分别为3. 41和3. 60; RpfB肌肉注射和滴鼻的最高效价为3. 63和2. 42.结论Ad5-TPA-3Ag在小鼠模型中可以诱导良好的免疫应答,可能成为一种新型的针对MTB潜伏感染的候选疫苗.“,”Objective To construct a novel recombinant human adenovirus 5 (Ad5) vectored vaccine against the triple antigenic Mycobacterium tuberculosis transgenes,ESAT6,rv3407 and RpfB (3Ag),and to evaluate its preliminary pharmacodynamical effects in murine models. Methods The Ad5 construction system of AdEasyTM XL was used to construct the plasmid pAd5-3Ag. After linearization by restriction enzyme Pac I,the pAd5-3Ag was transfected in AD293 cells to produce recombinant vector Ad5-TPA-3Ag against 3Ag for Western blot analysis. Purified Ad5-TPA-3Ag vectors were used to immunize murine models through intramuscular (IM) or intranasal (IN) routes,for testing serum antibody titers by ELISA,and for optimizing vector dosages. Results A viral vector plasmid containing 3Ag transgenes was successfully constructed. At 42 days after the packaged Ad5-TPA-3Ag was used to immunize murine models with a dosage of 8 × 107 IFU,anybody titers against each antigen were highest; the immune response was rapid and stable through the IM route with this dosage. The highest titer of antibody against ESAT6 was 4. 54 for the IM route and 4. 87 for the IN route; against rv3407,3. 41 and 3. 60; and against RpfB,3. 63 and 2. 42. Conclusions The Ad5-TPA-3Ag vaccine induced a good immune response in murine models; this vaccine may become a novel candidate vaccine against latent MTB infection.