Hao Yaxing, Li Zhirong, Wang Yifei, Liu Xiaobing, Ye Lilin
Institute of Immunology, Third Military Medical University, Gaotanyan, Shapingba District, Chongqing, 400038, China.
The First Affiliated Hospital, Biomedical Translational Research Institute and, Guangdong Province Key Laboratory of Molecular Immunology and Antibody Engineering, Jinan University, Guangzhou, 510632, China.
Methods Mol Biol. 2018;1707:15-38. doi: 10.1007/978-1-4939-7474-0_2.
B cell responses play a central role in humoral immunity, which protects an individual from invading pathogens by antigen-specific antibodies. Understanding the basic principles of the B cell responses during viral infection is of substantial importance for anti-viral vaccine development. In inbred mice, lymphocytic choriomeningitis virus (LCMV) infection elicits robust and typical T cell-dependent B cell responses, including germinal center reaction, memory B cell formation, and a long-lived plasma cell pool in bone marrow. Therefore, this system represents an ideal model to investigate anti-viral B cell responses. In this protocol, we describe how to propagate and quantify LCMV and successfully establish an acute LCMV infection in mice. This protocol also provides three different techniques to analyze B cell responses specific to an acute LCMV infection: the identification of germinal center (GC) B cells and follicular helper CD4 T (T) cells from the spleens and lymph nodes via flow cytometry, titration of LCMV-specific IgG in the serum after LCMV infection using an enzyme-linked immunosorbent assay (ELISA) analysis, and detection of LCMV-IgG secreted plasma cells from bone marrow with an enzyme-linked immunospot (ELISPOT) assay.
B细胞反应在体液免疫中起核心作用,体液免疫通过抗原特异性抗体保护个体免受入侵病原体的侵害。了解病毒感染期间B细胞反应的基本原理对于抗病毒疫苗的开发至关重要。在近交系小鼠中,淋巴细胞性脉络丛脑膜炎病毒(LCMV)感染引发强大且典型的T细胞依赖性B细胞反应,包括生发中心反应、记忆B细胞形成以及骨髓中长寿浆细胞库的形成。因此,该系统是研究抗病毒B细胞反应的理想模型。在本方案中,我们描述了如何繁殖和定量LCMV,并在小鼠中成功建立急性LCMV感染。本方案还提供了三种不同的技术来分析针对急性LCMV感染的B细胞反应:通过流式细胞术从脾脏和淋巴结中鉴定生发中心(GC)B细胞和滤泡辅助性CD4 T(Th)细胞;使用酶联免疫吸附测定(ELISA)分析在LCMV感染后血清中LCMV特异性IgG的滴定;以及用酶联免疫斑点(ELISPOT)测定法检测骨髓中分泌LCMV-IgG的浆细胞。