羊布鲁氏菌强毒株16M感染小鼠巨噬细胞模型的建立

摘要

Macrophage was the strongest anti-body cells of phagocytosis, while, Brucella could multiply in the cell and could not be killed. Therefor,we established the infection model,for continuous research Brucella and it's the host cell,and the interaction of baceria cell and host cell surface-associated membrane protein,discovering the intracellular interaction mechanism for next step. Infection of mouse macrophage cell line RAW264.7 with Brucella melitensi virulent strain 16M were performed 4 h at a cell/bacteria ratio of 1 : 500. Brucella-infected marophages were examined by indirect immunofluorescence assay and transmission electron microscopy assay. The optimal concentration ratio of primary antibody and secondary antibody in indirect immunofluorescence assay were 1 : 80 and 1 : 80, and the membrane depression and the endocytosis bodies were observed by transmission electron microscope. Expriment reduced the environmental factors of the infection process, optimized concentration ratio of primary and secondary antibodies in indirect immunofluorescence assay,established infection model.%巨噬细胞是机体抗吞噬能力最强的细胞,但布鲁氏菌不但不能被巨噬细胞杀死,反而能在胞内大量繁殖,因此,本研究建立其感染模型,为下一步继续研究布鲁氏菌与其宿主细胞表面相关膜蛋白之间的作用和胞内寄生机制奠定基础.用羊布鲁氏菌强毒株16M感染巨噬细胞系264.7(细胞和细菌比例为1:500),感染时间为4h,建立布鲁氏菌感染巨噬细胞模型,做间接免疫荧光试验和透射电镜试验.间接免疫荧光试验中一抗与二抗最佳稀释度分别为1:80和1:80,电镜下观察到细菌侵入细胞时膜凹陷,形态发生变化,形成内吞小体.本试验减少感染过程所涉及的环境因素,优化了间接免疫荧光试验所需的一抗和二抗浓度比,成功建立感染模型.