中兽医医药杂志2025，Vol.44Issue(5)：35-40,6.DOI:10.13823/j.cnki.jtcvm.2025.064
多房棘球绦虫TSP5单克隆抗体的制备与鉴定
Preparation and identification of monoclonal antibodies against TSP5 of Echinococcus multilocularis
摘要
Abstract
The objective was to prepare and identify a monoclonal antibody(mAb)against Tetraspanin 5(TSP5)of Echinococcus multilocularis,and evaluate its potential application in the diagnosis and immunotherapy of echinococcosis.The prokaryotic expression vector pET-28a-TSP5 was constructed by gene synthesis.Recombinant TSP5 protein was expressed and purified.Balb/c mice were immunized with the protein,and hybridoma cells were screened by hybridoma technology.Positive clones were subcloned by limiting dilution.Antibody titer was detected by indirect ELISA,and binding specificity to native TSP5 in hydatid tissues was analyzed by immunofluorescence.The variable region sequences of the antibody were sequenced and analyzed.The results showed that the pET-28a-TSP5 vector was successfully constructed,and a recombinant TSP5 protein of approximately 8.6 kDa(corresponding to the extracellular domain)was expressed and purified.A hybridoma cell line named 3B7G12,stably secreting anti-TSP5 mAb,was obtained.The antibody titer reached 1∶243 000 by ELISA.Immunofluorescence results showed that the mAb specifically recognized TSP5 protein on the surface of protoscoleces and cyst walls in E.multilocularis tissues.Sequencing of the variable regions revealed well-defined CDR sequences with high potential affinity.A high-titer and highly specific anti-TSP5 monoclonal antibody was successfully prepared,which accurately localized TSP5 protein in E.multilocularis tissues.This antibody provides a new tool for molecular diagnosis of alveolar echinococcosis and lays an experimental foundation for the development of CAR-T therapy and antibody-based drugs.关键词
多房棘球绦虫/Tetraspanin 5/单克隆抗体/包虫病/CAR-T免疫治疗Key words
Echinococcus multilocularis/Tetraspanin 5/monoclonal antibody/echinococcosis/CAR-T immunotherapy分类
农业科技引用本文复制引用
罗琦,程永康,陈思思,常岭迪,李子华..多房棘球绦虫TSP5单克隆抗体的制备与鉴定[J].中兽医医药杂志,2025,44(5):35-40,6.基金项目
宁夏回族自治区重点研发计划项目(2022BSB03092) （2022BSB03092）
