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作 者:宁唤唤 任瑞 康健 白鹭 路延之 谢燕玲 张芳琳 柏银兰 NING Huanhuan;REN Rui;KANG Jian;BAI Lu;LU Yanzhi;XIE Yanling;ZHANG Fanglin;BAI Yinlan(Department of Microbiology and Pathogen Biology,School of Preclinical Medicine,Air Force Medical University,Shaanxi Province,Xi’an 710032,China;School of Life Sciences,Yan’an University,Shaanxi Province,Yan’an 716000,China)
机构地区:[1]空军军医大学基础医学院微生物与病原生物学教研室,陕西西安710032 [2]延安大学生命科学学院,陕西延安716000
出 处:《中国医药导报》2023年第5期20-23,共4页China Medical Herald
基 金:国家“十三五”重大传染病专项课题(2018ZX10302302-002-004);国家自然科学基金面上项目(81971560、81671638);陕西省重点研发计划项目(2022ZDLSF01-07、2019ZDLSF02-04)。
摘 要:目的制备结核分枝杆菌(Mtb)6 kD早期分泌靶抗原(ESAT-6)单克隆抗体(mAb)并分析其特异性。方法用亲和层析法纯化目的蛋白ESAT-6,并用该蛋白免疫小鼠。分离免疫小鼠的脾细胞并与骨髓瘤细胞融合,筛选分泌mAb的杂交瘤细胞系。制备腹水并纯化mAb。酶联免疫吸附试验法分析mAb亚类和相对亲和力,蛋白印迹法检测其特异性。结果制备了可分泌ESAT-6抗体的杂交瘤细胞系,获得了纯化的mAb。mAb亚型为IgG1,相对亲和力为3.9×10^(-5)。该mAb可特异性识别Mtb的ESAT-6。结论制备抗ESAT-6的mAb为ESAT-6用于结核病诊断与防治制剂及其机制研究奠定了基础。Objective To prepare a monoclonal antibody(mAb)against 6 kD early secretory antigenic target(ESAT-6)of Mycobacterium tuberculosis and to analyze its specificity.Methods ESAT-6 was purified by affinity chromatography and immunized mice with the protein.Splenocytes from ESAT-6 immunized mice were fused with myeloma cells,and hybridoma cell lines secreting mAb were screened.mAb was purified from ascites.The subclass and relative affinity of mAb were assayed by enzyme-linked immunosorbent assay.The specificity of the mAb was analyzed by Western blot.Results The hybridoma cell line secreting ESAT-6 antibody was obtained and mAb was purified.The subclass of ESAT-6 mAb was IgG1 with a relative affinity of 3.9×10^(-5).The mAb specifically recognizes ESAT-6 in Mtb.Conclusion The preparation of mAb against ESAT-6 lays a foundation for the application of ESAT-6 in the diagnosis and treatment of tuberculosis and its mechanism.
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