Wu Kang, Liu Peng, Guo Bingnan, Zhang Yinsheng, Zhao Lixiang, Yang Jicheng, Liu Haiyan
Laboratory of Cellular and Molecular Tumor Immunology, Cyrus Tang Hematology Center, Jiangsu Institute of Hematology, The First Affiliated Hospital, Soochow University, Suzhou, PR China.
Hybridoma (Larchmt). 2010 Feb;29(1):31-6. doi: 10.1089/hyb.2009.0065.
Recombinant human myxovirus resistance protein A (MxA) was successfully expressed by an Escherichia coli expression system. After immunization and cell fusion, a mouse hybridoma (3C2) producing MAbs to MxA was established. Hybridoma 3C2 was further characterized using indirect ELISA, Western blot analysis, immunofluorescent staining, and immunoprecipitation. The ELISA results showed that the titer of 3C2 was between 1:6400 and 1:12800 in ascitic fluids. The isotype of the monoclonal antibody was tested to be IgG1kappa. 3C2 can also specifically recognize human MxA protein in various formats by Western blot analysis, immunofluorescent staining, and immunoprecipitation assay. We further demonstrated that 3C2 could be used to detected MxA expression induce by type I interferon in A549 cell line and human peripheral blood mononuclear cells by Western blot in a dose-dependent manner.
重组人黏液病毒抗性蛋白A（MxA）通过大肠杆菌表达系统成功表达。免疫和细胞融合后，建立了产生抗MxA单克隆抗体的小鼠杂交瘤（3C2）。使用间接ELISA、蛋白质免疫印迹分析、免疫荧光染色和免疫沉淀对杂交瘤3C2进行了进一步表征。ELISA结果显示，腹水液中3C2的效价在1:6400至1:12800之间。检测到单克隆抗体的同种型为IgG1κ。通过蛋白质免疫印迹分析、免疫荧光染色和免疫沉淀试验，3C2还可以特异性识别各种形式的人MxA蛋白。我们进一步证明，3C2可用于通过蛋白质免疫印迹以剂量依赖性方式检测I型干扰素在A549细胞系和人外周血单个核细胞中诱导的MxA表达。
