摘要
以重组A型口蹄疫病毒(FMDV)为抗原免疫BALB/c小鼠,应用杂交瘤技术制备单克隆抗体,并通过间接ELISA、有限稀释法及Western-blotting等试验进行筛选及鉴定。结果显示,获得了1株能够持续稳定分泌抗FMDV 3D蛋白单克隆抗体的杂交瘤细胞株(命名为5G2);经鉴定染色体数介于102～106条之间,与预期结果相符;单克隆抗体的稳定性检测表明其细胞培养上清抗体效价为1∶1 280,腹水抗体效价为1∶25 600,说明该株单克隆抗体的性质比较稳定;单克隆抗体特异性试验结果显示,获得的腹水单抗与3D蛋白、重组A型FMDV、A型FMDV、Asia1型FMDV及O型FMDV均可发生特异性反应,而与猪水疱病病毒及水疱性口炎病毒不发生反应。该非结构蛋白3D单克隆抗体的成功制备为进一步利用该单抗开发诊断试剂和研究病毒基因功能提供了技术手段。
Monoclonal antibody（McAb） was prepared by hybridoma technique after BALB/c mice were immunized with recombinant serotype A foot-and-mouth disease virus（FMDV）,and indirect ELISA was used for screening positive cell clones. The positive clone was subcloned to get monoclone cell lines by limi- ting dilution method and Western-blot was used for identification. Eventually, a hybridoma cell line（SG2） which secreted stably monoclonal antibodies against 3D protein of FMDV was obtained. The chromosome number ranged between 102 and 106 ,which was consistent as expected. Tests for stability of McAb showed that the antibody titer of the cell culture supernatant was 1 ： 1 280,while the antibody titer of aseites was 1 ： 25 600,indicating that secretion antibody ability was quite stable. The specificity test results showed that McAb could react specifically with the 3D protein, recombinant serotype A FMDV, serotype A FM- DV,serotype Asia1 FMDV and serotype O FMDV,which could not react with swine vesicular disease virus and vesicular stomatitis virus. The monoclonal antibody against the nonstructural protein 3D was successfully prepared,providing a tool for establishment of the method for diagnosing FMDV and for studies on function analysis of the 3D protein.
出处
《中国兽医科学》
CAS
CSCD
北大核心
2013年第5期488-493,共6页
Chinese Veterinary Science
基金
国家高技术研究发展计划(863)项目(2011AA10A211-1)
甘肃省高层次人才科技创新创业扶持行动项目(1013JHTA008)
国家农业产业体系项目(CARS-39)
关键词
口蹄疫病毒
3D蛋白
单克隆抗体
foot-and-mouth disease virus
3D protein
monoclonal antibody
