Tau蛋白外显子特异性单抗制备及初步应用  

Preparation and Application of Tau Exon-Specific Monoclonal Antibodies

作　　者：刘恋 陈嘉 胡超 夏影 杨微 王琳 石琦[2] 董小平[2,3] 陈志宝 陈操[2,3] LIU Lian;CHEN Jia;HU Chao;XIA Ying;YANG Wei;WANG Lin;SHI Qi;DONG Xiaoping;CHEN Zhibao(College of Life Science and Technology,Heilongjiang Bayi Agricultural University,Daqing 163000,China;National Institute for Viral Disease Control and Prevention,Chinese Center for Disease Control and Prevention,State Key Laboratory for Infectious Disease Prevention and Control y Beijing 102206,China;Center for Biosafety Mega-Science,Chinese Academy of Sciences,Wuhan 430071,China)

机构地区：[1]黑龙江八一农垦大学生命科学技术学院,大庆163000 [2]中国疾病预防控制中心病毒病预防控制所,传染病预防控制国家重点实验室,北京102206 [3]中国科学院生物安全大科学研究中心,武汉430071

出　　处：《病毒学报》2020年第2期214-222,共9页Chinese Journal of Virology

基　　金：中国医学科学院中央级公益性科研院所基本科研业务费专项资金(项目号:2018RC330004),题目:朊病毒感染中tau蛋白病变特点研究;国家自然科学基金(项目号:81772197,81401670,81630062),题目:朊病毒感染致脑组织微环境钙离子稳态变化及相关蛋白作用机制研究,题目:朊病毒病中tau蛋白异构体分布和磷酸化特点的研究,题目:朊病毒持续感染机制的研究;国家重点研发计划(项目号:2018YFC1200305),题目:BSL-4实验室关键设备性能及风险评估与发展战略研究;重点实验室课题(项目号:2019SKLID401,2016SKLID603),题目:宿主天然免疫在朊病毒清除和神经损伤中作用的研究,题目:朊病毒复制和转化过程中形态变化探索研究;黑龙江省自然科学基金(项目号:C2018044),题目:VCP蛋白在朊病毒病中的变化及对朊病毒复制影响的研究;黑龙江八一农垦大学研究生创新科研项目(项目号:YJSCX2018-Y66),题目:朊病毒感染神经细胞时tau蛋白异构体内外分泌能力差异研究。

摘　　要：目前由于缺失特异性的商品化抗体,不同tau异构体在中枢神经系统疾病中,尤其在朊病毒病中的变化鲜有报道。本研究利用杂交瘤技术制备了抗tau蛋白外显子2和10的单克隆抗体,同时对制备抗体的灵敏度、特异性、免疫反应亲和力、种属反应性等方面进行了评价,随后对制备的抗体应用范围进行了摸索。结果显示,制备的四株单抗特异性高、灵敏度好,尤其可以识别小鼠脑组织中的tau蛋白。制备的四株单抗可用于间接细胞免疫荧光实验和多种感染神经细胞和感染小鼠模型脑组织的蛋白免疫印迹和免疫组织化学实验。综上,本研究制备了四株tau外显子特异性单抗,上述单抗具有较广的应用范围,将成为神经退行性疾病中tau蛋白检测的有力工具。Changes of different tau isoforms in central nervous system diseases,especially in prion diseases have rarely been reported due to a lack of specific antibodies available commercially. In the present study,four monoclonal antibodies(mAbs) against exon 2 and exon 10 of tau proteins were prepared respectively with hybridoma technique. The prepared monoclonal antibodies were evaluated in terms of sensitivity,specificity,affinity in the immune response,and species reactivity. Subsequently,the application range of the prepared antibody was explored. Results showed that the prepared monoclonal antibodies had high specificity and sensitivity. Also,they could recognize tau protein in mouse brain specifically. Furthermore,the four prepared monoclonal antibodies could be used in cell immunofluorescence,Western Blot and immunohistochemistry experiments with various prion infected nerve cells and brain tissues from prion infected mouse models. Taken together,four tau exon-specific monoclonal antibodies were prepared and had a wide application range. These antibodies could become powerful tools for tau analyses in neurodegenerative diseases.

关 键 词：TAU蛋白 外显子2 外显子10 单克隆抗体 朊病毒病 

分 类 号：R373.9[医药卫生—病原生物学] R511[医药卫生—基础医学]