【摘要】 目的构建口蹄疫病毒(FMDV)多价DNA疫苗,并检测其免疫原性。方法以复合多表位表达盒OAAT及AsiaⅠ型FMDV的P1-2A-3C基因为基础,构建FMDV多价DNA疫苗pIRES-OAAT-P1-2A-3C,并用间接免疫荧光(IFA)方法检测目的蛋白在HeLa细胞中的表达。进一步进行小鼠免疫试验,并应用ELISA法检测小鼠血清抗体,ELISPOT检测小鼠脾淋巴单细胞IFN-γ的分泌水平,流式细胞术检测脾T淋巴细胞亚群数量,淋巴细胞转化试验检测特异性淋巴细胞增殖水平。结果所构建的FMDV多价DNA疫苗在HeLa细胞中获得了正确表达。免疫小鼠后,血清特异性抗体水平、脾淋巴单细胞IFN-γ的分泌、脾T淋巴细胞亚群CD4+和CD8+的数量及特异性淋巴细胞增殖水平均显著提高。结论已成功构建了FMDV多价DNA疫苗,并诱导小鼠产生了特异性的细胞免疫和体液免疫应答。更多还原

【Abstract】 Objective To construct polyvalent foot-and-mouth disease virus(FMDV) DNA vaccine and determine its immunogenicity. Methods Polyvalent FMDV DNA vaccine pIRES-OAAT-P1-2A-3C was constructed based on complex poly-epitope expression cassette OAAT and P1-2A-3C gene of FMDV type Asia Ⅰand transfected to HeLa cells. Identify the expressed target protein by IFA. Immunize mice with pIRES-OAAT-P1-2A-3C, and determine the serum antibody by ELISA, the secretion of IFN-γ by splenic lymphocytes by ELISPOST, the number of splenic T lymphocyte subgroups by flow cytometry, and the proliferation level of specific lymphocytes by lymphocyte transformation test. Results The constructed polyvalent FMDV DNA vaccine was correctly expressed in HeLa cells. The serum specific antibody level, secretion level of IFN-γ by splenic lymphocytes, the number of splenic T lymphocyte subgroups and the proliferation level of specific lymphocytes of mice after immunization with the DNA vaccine increased significantly. Conclusion Polyvalent FMDV DNA vaccine was successfully constructed and induced specific cellular and humoral immune responses in mice.更多还原