Preparation of monoclonal antibody against UL14 protein of pseudorabies virus and identification of its epitope
10.16303/j.cnki.1005-4545.2025.11.15
- VernacularTitle:伪狂犬病病毒UL14蛋白单克隆抗体制备及亚细胞定位
- Author:
Wenyan ZHANG
1
;
Da LIU
;
Peng PENG
;
Yan YAN
;
Weiren DONG
;
Jiyong ZHOU
Author Information
1. 浙江大学动物科学学院,浙江 杭州 310058;农业农村部动物病毒学重点实验室,浙江 杭州 3100
- Publication Type:Journal Article
- Keywords:
pseudorabies virus;
Interstitial protein UL14;
monoclonal antibody;
subcellular location
- From:
Chinese Journal of Veterinary Science
2025;45(11):2411-2419
- CountryChina
- Language:Chinese
-
Abstract:
The interstitial protein UL14 of pseudorabies virus(PRV)constitutes a crucial compo-nent for viral replication and virulence invasion.It can interact with other viral proteins to complete the infection of the host,rendering it one of the potentially important antiviral targets.In this stud-y,the PRV DX strain isolated in the laboratory was used as the parental strain.The recombinant UL14 protein was expressed and purified through the prokaryotic expression system.BALB/c mice were immunized with this protein as an antigen.After three rounds of subcloning screening,three hybridoma cell lines against the UL14 protein were obtained,named 1B4,1B5,and 1F2,respective-ly.The immunoreactivity of the antibodies was detected by immunofluorescence assay(IFA).The results showed that the antibody titers of the 1B4 and 1B5 strains were not lower than 1∶3 200,and that of the 1F2 strain was not lower than 1∶2 000.The results of western blot(WB)detection showed that the antibody titers of the three strains were all not lower than 1∶8 000.The results of indirect ELISA detection showed that the antibody titers in ascites were all not lower than 112 800.The results of the identification of the antigenic epitopes of the UL14 protein showed that the antigenic epitope recognized by 1B4 was 1 MFASDRRERRVRLAEAFQRE20,and the antigenic epitopes recognized by 1B5 and 1F2 were speculated to be 33GRADKKNPEFVRAFMAAKQAR53.After PRV infected susceptible cells,the temporal expression of the UL14 protein was detected by IF A using the prepared monoclonal antibodies.It was found that the temporal expression of UL14 was similar to that of gC,and the result of RT-qPCR of UL14 gene was consistent with IF A,indi-cating that UL14 might be a late gene during the PRV replication.The subcellular localization of UL14 after virus infection was detected by IFA,and it was found that the UL14 protein could be transferred from the cytoplasm to the nucleus and the expression of UL14 protein increased with the extension of infection time.This study provides a good research tool for further investigating on the function of the PRV UL14 protein and might contribute to the further study of the PRV replication mechanism mediated by the PRV UL14 protein.
