小鼠H-Y单克隆抗体ELISA检测方法的建立  

ELISA for Determination of Mouse H-Y Monoclonal Antibody

作　　者：杭苏琴[1] 刘为敏[1] 牛树理[1] 冯紫云[1] 

机构地区：[1]南京农业大学动物科技学院,江苏南京210095

出　　处：《中国兽医学报》2004年第6期547-549,共3页Chinese Journal of Veterinary Science

摘　　要：以纯系 BAL B/ c雄性小鼠脾细胞腹腔注射免疫同系雌性小鼠 9次 ,获得的抗血清经雌、雄鼠脾细胞吸收后用于精子细胞毒性试验 ,测得 H- Y抗血清效价为 1/ 16 0。选取免疫应答最好的雌鼠脾细胞与 SP2 / 0骨髓瘤细胞融合 ,用精子细胞毒性方法筛选效价较高的细胞株制备腹水 ,建立优化的 EL ISA反应条件。优化后的 EL ISA反应条件为 :抗原4℃过夜 ,加 H- Y抗血清 37℃反应 12 0 m in,加 HRP- Ig G 37℃反应 30 min,加 TMB 2 5℃ 30 min。优化后的 EL ISA与常规 EL ISA比较 ,可以明显降低阴性吸光值 ,检测灵敏度从 1/ 32 0上升为 1/ 12BALB/c female mice were immunized by intraperitoneal injections of syngenetic male spleen cells for nine times.Sperm cytotoxicity test was used to examine antisera and give a titer of 160.The splenocytes from mice producing high immune response were fused with SP2/0 cells in 50% PEG.Sperm cytotoxicity test and ELISA were used for selecting cell strains with high titer of monoclonal antibody.Optimal ELISA:antigen was immbolized to polystyrene plate well overnight at 4℃,washing three times and then add H-Y antiserum for 120 minutes at 37℃,washing again and add enzyme-labelled HRP- IgG for 30 minutes at 37℃,washing it five times and substrate TMB was added for 30 minutes.The sensibility of the ELISA was improved from 1∶320 to 1∶(1 280).

关 键 词：小鼠 H-Y单克隆抗体 ELISA检测方法 抗血清制备 

分 类 号：S852.5[农业科学—基础兽医学] S854.43[农业科学—兽医学]