猪流行性腹泻病毒单克隆抗体8A3的CHO细胞瞬时表达与稳转细胞池构建  

Transient Expression of Porcine Epidemic Diarrhea Virus Monoclonal Antibody 8A3 in CHO Cells and Construction of Stable Transduced Cell Pool

作　　者：周丽璇 孙玉洁 张云静 谭菲菲 黄柏成 田克恭 ZHOU Li-xuan;SUN Yu-jie;ZHANG Yun-jing;TAN Fei-fei;HUANG Bai-cheng;TIAN Ke-gong(College of Veterinary Medicine,Henan Agricultural University,Zhengzhou,Henan,450002,China;National Research Center for Veterinary Medicine,Luoyang,Henan,471003,China)

机构地区：[1]河南农业大学动物医学院,河南郑州450002 [2]国家兽用药品工程技术研究中心,河南洛阳471003

出　　处：《动物医学进展》2023年第5期1-6,共6页Progress In Veterinary Medicine

基　　金：郑洛新自创区创新引领型产业集群专项(201200211200)。

摘　　要：为实现猪流行性腹泻病毒(Porcine epidemic diarrhea virus,PEDV)单克隆抗体(monoclonal antibody,mAb)的体外高效表达,首先利用基因工程技术获得了PEDV鼠源mAb 8A3的轻链和重链序列,然后构建真核表达载体pCHO1.0-8A3-H-L,再利用CHO细胞对mAb 8A3进行瞬时表达,最后经稳定转染以获得稳定表达8A3的CHO细胞池。结果显示,CHO细胞表达的mAb 8A3在免疫印迹和间接免疫荧光分析中均可特异性识别PEDV,且具有正确的IgG抗体构型;且CHO细胞表达的mAb 8A3可以中和G2a、G2b和G1基因型PEDV。实现了PEDV mAb 8A3的CHO瞬时转染表达,并获得了稳定表达8A3的CHO细胞池,为PEDV mAb稳定表达细胞系的筛选奠定了基础,同时为PEDV的诊断和治疗提供了新型基因工程抗体材料。In order to achieve a high expression level of porcine epidemic diarrhea virus(PEDV)monoclonal antibody(mAb)in vitro,firstly,the light chain and heavy chain sequences of PEDV high neutralizing activity mAb 8A3 were obtained by genetic engineering technology,and then the antibody eukaryotic expression vector pCHO1.0-8A3-H-L was constructed,after that,the CHO transient expression system was used to express mAb 8A3.Finally,the CHO cell pool stably expressing 8A3 was obtained by stable transfection.The results showed that the CHO-expressed mAb 8A3 could recognize PEDV in both immunoblotting and indirect immunofluorescence analysis,and showed the correct IgG antibody configuration.In addition,CHO-expressed mAb 8A3 can neutralize PEDV of G2a,G2b and G1 genotypes.In conclusion,this study achieved the CHO transient transfection and expression of PEDV high neutralizing activity mAb 8A3,and constructed a stably transfected CHO cell pool,which provided a basis for the construction of PEDV mAb stable cell lines,and also provides a new genetically engineered antibody material for the diagnosis and treatment studies of PEDV.

关 键 词：猪流行性腹泻病毒 单克隆抗体 中国仓鼠肾细胞 瞬时表达 稳定表达 

分 类 号：S852.651[农业科学—基础兽医学] S858.28[农业科学—兽医学]