乙肝表面抗原DNA疫苗初免—蛋白疫苗加强免疫策略增强免疫反应的蛋白质组学分析
发布时间：2018-10-12 13:19
【摘要】：DNA初免-蛋白质加强的免疫策略已经广泛应用以增强这两种疫苗的免疫原性,但是这种免疫增效的机制目前还不清楚。我们以pVAX1质粒为载体构建了表达乙肝表面抗原的真核表达质粒pVAX-S,用pVAX-S和重组乙肝表面抗原以C57BL/6小鼠为实验动物实施了DDD、DDS、SSS和SSD4种不同的免疫策略。在第一次免疫后第3周、7周、11周、15周、19周和23周用酶联免疫吸附法检测了这些小鼠的总抗体滴度,第11周用混合淋巴细胞反应检测了不同免疫组小鼠的T细胞免疫记忆强度和树突状细胞刺激T细胞增殖的能力,第23周用RocheAssay Medium分光法按效应细胞/靶细胞1:50的比例检测了CTL活性。我们观察到DDS组小鼠拥有最强的免疫记忆,无论抗体滴度、CTL活性、T细胞免疫记忆强度还是树突状细胞激活T细胞的能力都比其它免疫组以及空白对照组的小鼠更强。我们进一步进行了比较血清蛋白质组学的研究,以期能阐明这种免疫增效作用的机制。以非免疫组小鼠作参照胶,我们发现DDS组小鼠有5个蛋白上调、17个蛋白下调;而SSS组小鼠则有7个蛋白上调、10个蛋白下调。在DDS组中唯一出现的3个蛋白质periplakin、F-box protein 30和calpain的生理功能都和免疫反应有一定的关系。这3个蛋白可以作为潜在的成功免疫的新型分子标记物,也可能成为疫苗作用机制的新研究目标。在我们的一个在研项目中,我们用猪带绦虫囊尾蚴抗原膜联蛋白B1 DNA疫苗和蛋白质疫苗对小鼠进行免疫后分析其血清蛋白质组,初步得到6个特异性表达蛋白的质谱结果,这6个蛋白与乙肝表面抗原DDS/SSS免疫组小鼠血清中鉴定的6个特异性表达蛋白无一相同,说明这3个从DDS免疫组中发现的特异性表达蛋白具有抗原特异性。
[Abstract]:DNA immune-protein enhanced immunization strategy has been widely used to enhance the immunogenicity of the two vaccines, but the mechanism of this immune synergism is not clear. The eukaryotic expression plasmid pVAX-S, expressing hepatitis B surface antigen was constructed by using pVAX1 plasmid as vector. The different immunological strategies of DDD,DDS,SSS and SSD4 were carried out by using pVAX-S and C57BL/6 mice as experimental animals. The total antibody titers of the mice were detected by enzyme-linked immunosorbent assay (Elisa) at week 3, week 7, week 11, week 15, week 19 and week 23 after the first immunization. At the 11th week, the T cell immune memory intensity and the ability of dendritic cells to stimulate T cell proliferation were measured by mixed lymphocyte reaction in mice of different immune groups. The activity of CTL was measured by RocheAssay Medium spectrophotometry at 1:50 of effector cell / target cell at the 23rd week. We observed that DDS mice had the strongest immune memory, whether antibody titer, CTL activity, T cell immune memory intensity or the ability of dendritic cells to activate T cells were stronger than those of other immune groups and blank control groups. We further studied the comparison of serum proteomics in order to elucidate the mechanism of this immune synergism. Using non-immunized mice as reference glue, we found that 5 proteins were up-regulated and 17 proteins down-regulated in DDS group, while 7 proteins were up-regulated and 10 proteins down-regulated in SSS group. The physiological functions of periplakin,F-box protein 30 and calpain, the only three proteins in DDS group, were related to the immune response to some extent. These three proteins can be used as potential novel molecular markers for successful immunization, and may also become new research targets for the mechanism of vaccine action. In one of our research projects, we immunized mice with cysticercus cellulosae antigen integrin B1 DNA vaccine and protein vaccine and analyzed their serum proteome, and obtained the mass spectrometry results of six specific expressed proteins. These six proteins were not the same as those identified in the serum of mice immunized with hepatitis B surface antigen (DDS/SSS), indicating that the three specific expressed proteins from DDS immunized group had antigen-specificity.
【学位授予单位】：第二军医大学
【学位级别】：硕士
【学位授予年份】：2006
【分类号】：R392
本文编号：2266272
[Abstract]:DNA immune-protein enhanced immunization strategy has been widely used to enhance the immunogenicity of the two vaccines, but the mechanism of this immune synergism is not clear. The eukaryotic expression plasmid pVAX-S, expressing hepatitis B surface antigen was constructed by using pVAX1 plasmid as vector. The different immunological strategies of DDD,DDS,SSS and SSD4 were carried out by using pVAX-S and C57BL/6 mice as experimental animals. The total antibody titers of the mice were detected by enzyme-linked immunosorbent assay (Elisa) at week 3, week 7, week 11, week 15, week 19 and week 23 after the first immunization. At the 11th week, the T cell immune memory intensity and the ability of dendritic cells to stimulate T cell proliferation were measured by mixed lymphocyte reaction in mice of different immune groups. The activity of CTL was measured by RocheAssay Medium spectrophotometry at 1:50 of effector cell / target cell at the 23rd week. We observed that DDS mice had the strongest immune memory, whether antibody titer, CTL activity, T cell immune memory intensity or the ability of dendritic cells to activate T cells were stronger than those of other immune groups and blank control groups. We further studied the comparison of serum proteomics in order to elucidate the mechanism of this immune synergism. Using non-immunized mice as reference glue, we found that 5 proteins were up-regulated and 17 proteins down-regulated in DDS group, while 7 proteins were up-regulated and 10 proteins down-regulated in SSS group. The physiological functions of periplakin,F-box protein 30 and calpain, the only three proteins in DDS group, were related to the immune response to some extent. These three proteins can be used as potential novel molecular markers for successful immunization, and may also become new research targets for the mechanism of vaccine action. In one of our research projects, we immunized mice with cysticercus cellulosae antigen integrin B1 DNA vaccine and protein vaccine and analyzed their serum proteome, and obtained the mass spectrometry results of six specific expressed proteins. These six proteins were not the same as those identified in the serum of mice immunized with hepatitis B surface antigen (DDS/SSS), indicating that the three specific expressed proteins from DDS immunized group had antigen-specificity.
【学位授予单位】：第二军医大学
【学位级别】：硕士
【学位授予年份】：2006
【分类号】：R392
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相关期刊论文 前2条
1 李林,吴家睿,李伯良;蛋白质组学的产生及其重要意义[J];生命科学;1999年02期
2 李宁,许红韬;蛋白质组研究的现状与展望[J];生物技术通讯;2000年04期
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