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目的:研究并比较结核分枝杆菌免疫保护性抗原DNA(Ag85A和ESAT-6)疫苗联合免疫,BCG免疫以及联合DNA疫苗初免—BCG加强免疫等不同的免疫策略,诱导免疫应答效果观察。方法:健康雌性BALB/c小鼠24只,随机分成PBS阴性对照组,DNA初免—BCG异源加强组,DNA(Ag85A和ESAT-6)初免DNA同源加强组和BCG阳性对照组,共进行3次免疫,初免2次,最后1次加强,间隔2周1次。PBS组3次均注射PBS溶液;DNA/BCG组以质粒DNA免疫2次,最后1次以BCG加强免疫;DNA/DNA组3次均以质粒DNA进行免疫;BCG组则注射PBS溶液2次后以BCG免疫。末次免疫后4、6、8周分别分离血清测定总IgG水平,同时分离小鼠脾细胞,体外经TB-PPD刺激后进行淋巴细胞增殖实验(XTT法)并测定脾细胞培养上清中IFN-γ和IL-4水平。结果:DNA/BCG、DNA/DNA、BCG组体外经TB-PPD刺激后均检测到特异性IgG抗体产生,3组平均效价为1∶120、1∶160、1∶80,DNA/DNA组的抗体效价高于另外2组;小鼠脾细胞体外经TB-PPD刺激后,均能产生特异性淋巴细胞增殖并诱生较强的IFN-γ反应,其中DNA/BCG组IFN-γ的分泌水平高于DNA/DNA组和BCG组(P<0.05)。结论:联合DNA疫苗初免—BCG加强的免疫策略能在小鼠体内诱导较强的特异性细胞免疫反应,产生高水平的IFN-γ。
OBJECTIVE: To study and compare the immune response induced by immunomodulatory antigen DNA of Mycobacterium tuberculosis (Ag85A and ESAT-6), BCG immunization and DNA vaccine prime-BCG booster immunization. Methods: Twenty-four healthy female BALB / c mice were randomly divided into PBS-negative control group, DNA prime-BCG heterozygote group, DNA (Ag85A and ESAT-6) DNA prime-enhancing group and BCG positive control group A total of 3 immunizations, the initial 2 times, the last one to strengthen, 1 interval 2 weeks. The PBS / PBS group was injected with PBS three times, the DNA / BCG group was immunized twice with plasmid DNA and the last one with BCG. The DNA / DNA group was immunized with plasmid DNA three times, and the BCG group was injected with PBS solution twice Immunized with BCG. Serum total IgG levels were determined at 4, 6 and 8 weeks after the last immunization, and splenocytes were isolated at the same time. The proliferation of spleen cells was stimulated by TB-PPD in vitro (XTT method) and IFN- γ and IL-4 levels. RESULTS: Specific IgG antibody production was detected in all DNA / BCG, DNA / DNA and BCG groups after TB-PPD stimulation. The average titers of the three groups were 1: 120, 1: 160, 1: Of the antibody titers were higher than the other two groups. The spleen cells stimulated by TB-PPD in vitro could produce specific lymphocyte proliferation and induced a strong IFN-γ response, in which DNA / BCG group IFN-γ The secretion level was higher than DNA / DNA group and BCG group (P <0.05). CONCLUSION: The combination of DNA vaccine prime-BCG boosted immunization strategy can induce strong specific cellular immune responses in mice and produce high levels of IFN-γ.