Generation of antibodies by genetic immunization

描述：（申请人提供）基因免疫提供了主要的 与经典的蛋白质/肽免疫相比， 抗原特异性抗体。抗原跟随基因的体内表达 转移导致适当的修改和折叠，从而呈现 天然表位对于许多抗原候选物，没有纯化的蛋白可用 并且肽合成并不总是可行的。为了有效地使 通过遗传免疫方法产生有用的抗体，两种技术 需要克服的障碍，首先，需要能够筛选 抗体在遗传免疫小鼠血清中的存在，而不 抗原的可用性。这个问题可以通过表示 抗原从质粒DNA表达载体在体外或体内 高效基因转移。表达抗原的细胞可用于 免疫组织化学、流式细胞术或Western印迹分析，以鉴定 抗原特异性抗体。第二，高效、简单、经济， 需要体内基因转移来诱导体液免疫应答。米鲁斯 血管内质粒DNA基因转移方法满足这些要求， 可以有效地诱导针对表达的转基因的抗体应答。 在这个I期应用中，我们建议确定哪个血管内基因 转移方法产生最有效的体液免疫反应，针对 不同种类的表达抗原。我们还将优化蛋白质印迹 通过在不同细胞系中表达抗原的抗体筛选方法， 在肝细胞中。在第二阶段研究中，我们将进一步优化遗传学， 通过包括细胞因子基因的免疫方法， 时间表，并测试不同的表达载体和小鼠品系。的 将通过检测血清池来改进筛选方法（以限制 测定的初始数量），以及测试其他测定（免疫组织化学、流式细胞术、免疫荧光和免疫荧光）。 流式细胞术，ELISA）。这项技术将使Mirus能够提供全面的 抗体生成服务，为研究市场鉴定新型抗体 以及治疗目标，并引入基因免疫， 抗体筛选试剂盒。 拟议商业应用：不可用

DESCRIPTION: (Provided by Applicant) Genetic immunization provides major advantages over classic protein/peptide immunization for the generation of antigen-specific antibodies. In vivo expression of the antigen following gene transfer results in proper modification and folding, and thus presentation of natural epitopes. For many antigen candidates, no purified protein is available and peptide synthesis is not always feasible. To efficiently enable the generation of useful antibodies via genetic immunization methods, two technical hurdles need to be overcome, First, one needs to be able to screen for the presence of the antibodies in the sera of genetically immunized mice, without the availability of antigen. This problem can be solved by expressing the antigen from plasmid DNA expression vectors after in vitro or in vivo high-efficiency gene transfer. Cells expressing the antigen can be used in immunohistochemistry, flow cytometry, or Western blotting analysis to identify antigen-specific antibodies. Second, efficient, simple, and cost effective in vivo gene transfer is required to induce a humoral immune response. Mirus' intravascular plasmid DNA gene transfer methods fulfill these requirements and can efficiently induce antibody responses against expressed transgenes. In this Phase I application, we propose to determine which intravascular gene transfer method result in the most efficient humoral immune response against different classes of expressed antigens. We will also optimize Western blotting antibody screening methods by expressing antigens in different cell lines and in liver cells. During the Phase II studies, we will further optimize genetic immunization methods by including cytokine genes, refining the immunization schedule, and testing different expression vectors and mouse strains. The screening methods will be improved by testing pooling of sera (to limit the initial number of assays), and testing other assays (immunohistochemistry, flow cytometry, ELISA). This technology will allow Mirus to provide a comprehensive antibody generation service, identify novel antibodies for the research market as well as for therapeutic targets, and introduce genetic immunization and antibody screening kits. PROPOSED COMMERCIAL APPLICATION: NOT AVAILABLE