Prokaryotic expression and binding activity analysis of nanoantibodies against vac-cinia virus protein D8
10.16303/j.cnki.1005-4545.2024.08.20
- VernacularTitle:痘苗病毒蛋白D8纳米抗体的原核表达及结合活性分析
- Author:
Jingjing TIAN
1
;
Xiaolan YANG
;
Tao LI
;
Hui WANG
;
Ruiwen FAN
Author Information
1. 山西农业大学动物医学学院,山西 太谷 030801
- Keywords:
vaccinia virus;
nanoantibody;
SUMO;
binding activity
- From:
Chinese Journal of Veterinary Science
2024;44(8):1728-1734
- CountryChina
- Language:Chinese
-
Abstract:
This study aims to obtain nanoantibody of vaccinia virus protein D8 by SUMO tag and analyze its binding activity with antigen.The sequence was designed and synthesized to construct the prokaryotic expression plasmid pKMD-SUMO-D8,which was transformed into Escherichia coli BL21(DE3)receptor cells and induced to express.Anti-SUMO-D8 with His and SUMO tags were purified,and then all tags were removed by SUMO protease to further obtain Anti-D8.Final-ly,the binding activity was detected and analyzed by indirect ELISA and IFA.The results showed that the prokaryotic expression plasmid pKMD-SUMO-D8 and recombinant strains were construc-ted.Under the condition of 30 ℃,the expression of Anti-SUMO-D8 was best when induced by IPTG with 0.1 mmol/L for 4 h.High purity Anti-SUMO-D8 and Anti-D8 nanoantibodies were ob-tained.Indirect ELISA results showed that both Anti-SUMO-D8 and Anti-D8 had binding activity with antigen.Under the same conditions,the binding activity of Anti-D8 to vaccinia virus and E8L was higher than that of Anti-SUMO-D8.Indirect IFA results showed that 48 h after infection with vaccinia virus,some nuclei of the experimental group showed green fluorescence under fluorescence microscope.Nanoantibodies of vaccinia virus protein D8 could be efficiently prepared by SUMO tag,and the prepared nanoantibodies had binding activity.This study provides new idea for the preparation of nanoantibody,and also lays a foundation for further study of the biological function of vaccinia virus protein D8 nano-antibody.
