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目的:构建双亚型(H5/H7)流感病毒DNA疫苗,并检测其免疫原性。方法:构建共表达流感病毒H5亚型血凝素(HA)和H7亚型HA的DNA疫苗,采用间接免疫荧光法(IFA)检测目的蛋白在幼仓鼠肾细胞(BHKcell)中的表达。分为pVAX1-H5-H7、pVAX1-H5、pVAX1-H7和pVAX1空质粒对照组4组(每组15只)进行小鼠免疫实验,并应用ELISA法检测小鼠血清抗体,流式细胞术检测小鼠脾T淋巴细胞亚群数量。结果:所构建的双亚型(H5/H7)流感病毒DNA疫苗在BHK细胞中的表达产物可激发荧光抗体产生绿色免疫荧光,且对照pVAX1空载体转染BHK细胞无免疫荧光产生。使用双亚型(H5/H7)流感病毒DNA疫苗免疫小鼠后,小鼠产生了针对H5和H7抗原的血清特异性抗体,流式细胞术检测免疫组小鼠脾T淋巴细胞亚群CD4+和CD8+的数量显著高于对照组(P<0.05)。结论:成功构建的双亚型(H5/H7)流感病毒DNA疫苗可以诱导小鼠产生特异性的细胞免疫和体液免疫应答。
Objective: To construct a double subtype (H5 / H7) influenza virus DNA vaccine and test its immunogenicity. Methods: DNA vaccine co-expressing H5 subtype hemagglutinin (HA) and H7 subtype HA of influenza virus was constructed and the expression of the target protein in baby hamster kidney cells (BHKcell) was detected by indirect immunofluorescence assay (IFA). Immunized mice were divided into 4 groups (n = 15) with pVAX1-H5-H7, pVAX1-H5, pVAX1-H7 and pVAX1 empty plasmid control group. Serum antibodies were detected by ELISA. Flow cytometry Mouse spleen T lymphocyte subsets. Results: The expressed product of the H5 / H7 influenza virus DNA vaccine in BHK cells stimulated the production of green immunofluorescence by fluorescent antibody, and no immunofluorescence was detected in BHK cells transfected with pVAX1 empty vector. After immunization of mice with double subtype (H5 / H7) influenza virus DNA vaccine, mice developed serum-specific antibodies against H5 and H7 antigens. The splenic T lymphocyte subsets CD4 + and The number of CD8 + was significantly higher than that of the control group (P <0.05). Conclusion: The successfully constructed double subtype (H5 / H7) influenza virus DNA vaccine can induce mice to produce specific cellular and humoral immune responses.