中国兽医科学2024，Vol.54Issue(9)：1195-1201,7.DOI:10.16656/j.issn.1673-4696.2024.0145
不同信号肽对猪瘟E0蛋白在HEK-293F细胞中分泌表达的影响及E0-ELISA方法的建立
Effect of different signal peptides on secretion expression of CSFV E0 protein in HEK-293F cells and establishment of E0-ELISA method
摘要
Abstract
To establish an indirect ELISA method using CSFV E0 protein as a coating antigen,we con-structed recombinant expression plasmids of CSFV E0 fused with 5 commonly used signal peptides and their endogenous signal peptides.Subsequently,these plasmids were transiently transfected into HEK-293F cells for expression.Western-blot results revealed the expression of E0 protein mediated by different signal peptides in the cell culture supernatant.Among them,E0 protein mediated by human in-sulin(Insulin)signal peptide showed the highest secretion level.The purified E0 protein was used as the coating antigen to establish an indirect ELISA method.173 swine serum samples were concurrently tested using the E0-ELISA and IDEXX ELISA kit.The results demonstrated a concordance rate of 92.49％between E0-ELISA and IDEXX ELISA.Compared to the virus neutralization test(VNT),the concordance rate between E0-ELISA and VNT was 95.38％,while IDEXX ELISA showed a concordance rate of 91.33％with VNT.These results indicate that E0-ELISA is more sensitive than IDEXX ELISA.In this study,we achieved ef-ficient glycosylation-mediated expression of classical swine fever virus(CSFV)E0 protein in HEK-293F cells using different signal peptides.We successfully established the E0-ELISA method using this E0 protein as the coating antigen,providing a valuable technical tool for the clinical differentiation of CSFV field infection.关键词
E0蛋白/信号肽/猪瘟病毒/HEK-293F/间接ELISAKey words
E0 protein/signal peptide/classical swine fever virus/HEK-293F/indirect ELISA分类
农业科技引用本文复制引用
方琪,罗烨,梁彤彤,张珊玲,冯泽中,谢诗晴,郑金,尹德明,余兴龙..不同信号肽对猪瘟E0蛋白在HEK-293F细胞中分泌表达的影响及E0-ELISA方法的建立[J].中国兽医科学,2024,54(9):1195-1201,7.基金项目
湖南省重点领域研发计划项目(2022NK2049) （2022NK2049）
