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对结核杆菌四联DNA疫苗的免疫应答和保护效果进行了评价。用结核分枝杆菌Ag85B,MPT-64,MPT-63和ESAT-6等4个分泌蛋白基因的表达载体首次免疫小鼠21d后,Ag85B抗体滴度达到1:200,二次免疫后为1:102400,三次免疫后为1:1288。MPT-64和MPT-63的抗体滴度在首次免疫21d后即高达1:25600,二、三次免疫后呈下降趋势。三次免疫21d后均未检测到ESAT-6的特异性抗体。用该DNA四联苗免疫小鼠后,4种蛋白都诱导脾脏细胞产生较高水平的细胞应答(产生了特异性γ-干扰素)。三次免疫后经静脉强毒攻击,四联苗免疫组小鼠肺脏的细菌数比对照空载体组减少了99.8％,保护水平显著高于BCG疫苗组、对肺组织的病理形态特征观察表明,空载体pJW4303的对照小鼠,肺部严重损伤,肺实质干酪样坏死,坏死结节占肺实质的50％～70％,而四联苗免疫的小鼠,肺组织结构正常,实质无肉芽肿,肺泡轮廓清晰。研究证实,Ag85B,MPT-64,MPT-63和ESAT-6四种分泌蛋白编码基因组成的四联DNA疫苗具有很高的免疫应答水平和保护效率。
The immune response and protective effect of Mycobacterium tuberculosis quadruple DNA vaccine were evaluated. The titer of Ag85B antibody reached 1: 200 after the first immunization with the expression vectors of four secreted protein genes of Mycobacterium tuberculosis Ag85B, MPT-64, MPT-63 and ESAT-6, 102400, 1: 1288 after three immunizations. Antibody titers of MPT-64 and MPT-63 were as high as 1: 25600 after 21 days of initial immunization, and declined after two or three immunizations. ESAT-6 specific antibodies were not detected after 21 days of three immunizations. After immunization of mice with this DNA quadriplex, all four proteins induced a higher level of cellular response in spleen cells (producing specific gamma interferon). After three immunizations, the number of bacteria in the lungs of mice immunized with quadruplex vaccine was 99.8% less than that of the control group, and the level of protection was significantly higher than that of the BCG vaccine group. The observation of the pathological features of the lung tissue showed that empty The control mice of the vector pJW4303 had severe lung injury and caseous necrosis of the lung. The necrotic nodules accounted for 50% to 70% of the lung parenchyma. However, the mice immunized with the quadruple vaccine had normal lung tissue structure and virtually no granulomas. Alveolar outline clear. The study confirmed that Ag85B, MPT-64, MPT-63 and ESAT-6 four secreted protein encoding gene composed of DNA vaccine has a high level of immune response and protection efficiency.