Generationof the polyclonal antibody against Zaire Ebola virus GP1 protein and development of indirect ELISA for antibody detection
10.16303/j.cnki.1005-4545.2025.06.12
- VernacularTitle:扎伊尔型埃博拉病毒GP1蛋白的多克隆抗体制备及间接ELISA检测方法的建立
- Author:
Xiao WU
1
;
Mengyao ZHANG
1
;
Hailun LI
1
;
Pei HUANG
1
;
Haili ZHANG
1
;
Xiaolei LIU
1
;
Hualei WANG
1
;
Yuanyuan LI
1
Author Information
1. 吉林大学动物医学学院人畜共患传染病重症诊治全国重点实验室,吉林长春 130062
- Publication Type:Journal Article
- Keywords:
Zaire Ebola virus;
prokaryotic expression;
GP protein;
polyclonal antibody;
indirect ELISA
- From:
Chinese Journal of Veterinary Science
2025;45(6):1194-1201
- CountryChina
- Language:Chinese
-
Abstract:
To establish an indirect enzyme linked immunosorbent assay(ELISA)method for the detection of Zaire Ebola virus(ZEBOV)specific antibodies,the full-length of ZEBOV GP1 gene was amplified by PCR and cloned into pET-30a(+)vector to generate the pET-30a(+)-GP1 plasmid.After expressed in the E.coli expression system,the purified GP1 protein was used as coating antigen to establish the indirect ELISA method for detection of ZEBOV antibody.The con-ditions including concentration of coating antigen and serum dilution were determined by chess-board titration.Specificity,sensitivity,and reproducibility of the established ELISA detection meth-od were evaluated.GP1 protein was successfully prepared by prokaryotic expression,and was used as the coatingantigen for indirect ELISA.By optimizing the reaction conditions,the optimal concen-tration of the coating antigen was determined to be 0.5 g/L;the optimal dilution of serum was cal-culated to be 1∶3 200;the optimal dilution of enzyme-labeled secondary antibody was measured to be 1∶20 000.The established method exhibited excellent specificity,sensitivity,and reproducibili-ty.In the present study,the GP1 protein was successfully expressed in the E.coli expression sys-tem and the high purity GP1 protein was used as the coating protein to establish an indirect ELISA assay for ZEBOV antibody.This method is highly specific,sensitive,and reproducible,which provides technical support for the fur-ther study of the biological function of GP1 and the detection of ZEBOV antibody in serum.
