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[目的]构建卡氏肺孢子虫抗原p55基因片段DNA疫苗,并研究该疫苗的体外表达及在小鼠体内的免疫原性。[方法]构建真核表达质粒pcDNA3.1(+)-582,用免疫印渍技术检测目的蛋白的免疫原性。将雌性BALB/c小鼠分为3组,以肌肉注射方式分别注射重组表达质粒pcDNA3.1(+)-582(免疫组)、pcDNA3.1(+)及PBS(对照组)。ELISA法检测小鼠血清中特异性抗体水平,MTT法检测免疫小鼠的淋巴细胞增殖能力。[结果]成功构建了重组真核表达质粒pcDNA3.1(+)-582,免疫印渍技术证实目的蛋白与免疫后的小鼠阳性血清能发生特异性反应;ELISA结果显示免疫后pcDNA3.1(+)-582组小鼠抗体水平呈增高的趋势,pcDNA3.1(+)-582组小鼠淋巴细胞的增殖指数明显高于pcDNA3.1(+)组及PBS组。免疫组与对照组相比较差异显著,差异有统计学意义(P﹤0.05)。[结论]卡氏肺孢子虫抗原p55基因片段DNA疫苗构建成功,免疫小鼠后能诱导产生一定的体液免疫和细胞免疫。
[Objective] The aim of the study was to construct DNA vaccine of Pneumocystis carinii antigen p55 gene fragment and to study its in vitro expression and immunogenicity in mice. [Method] The eukaryotic expression plasmid pcDNA3.1 (+) - 582 was constructed and the immunogenicity of the target protein was detected by immunoblotting. Female BALB / c mice were divided into 3 groups and injected with recombinant plasmid pcDNA3.1 (+) - 582 (immunized group), pcDNA3.1 (+) and PBS (control group) respectively by intramuscular injection. The serum levels of specific antibodies were detected by ELISA and the proliferation of lymphocytes was detected by MTT assay. [Results] Recombinant eukaryotic expression plasmid pcDNA3.1 (+) - 582 was constructed successfully. The immunoprecipitation assay confirmed that the target protein reacted specifically with the immunized mouse sera. The results of ELISA showed that pcDNA3.1 (+ +) - 582 mice showed an increasing trend. The proliferation index of lymphocytes in pcDNA3.1 (+) - 582 group was significantly higher than that in pcDNA3.1 (+) group and PBS group. The difference between the immunized group and the control group was significant (P <0.05). [Conclusion] The DNA vaccine of Pneumocystis carinii antigen p55 gene fragment was successfully constructed. After immunization, the mice could induce certain humoral and cellular immunity.