Production and identification of monoclonal antibody against PDCoV N protein
10.16303/j.cnki.1005-4545.2024.12.03
- VernacularTitle:猪δ冠状病毒N蛋白单克隆抗体的制备及鉴定
- Author:
Suxian LUO
1
;
Nannan GUO
;
Houjun HE
;
Deping SONG
;
Dongyan HUANG
;
Yuxin TANG
;
Yu YE
Author Information
1. 江西农业大学动物科学技术学院,江西南昌 330045
- Publication Type:Journal Article
- Keywords:
PDCoV;
nucleocaspid protein;
monoclonal antibodies
- From:
Chinese Journal of Veterinary Science
2024;44(12):2521-2525
- CountryChina
- Language:Chinese
-
Abstract:
Porcine deltacoronavirus(PDCoV)is one of the important pathogens associated with swine viral diarrhea,which results in enormous economic losses in the pig industry.Among the known encoding proteins,the nucleocapsid(N)protein is relatively conserved and serves as the dominant antigen of coronaviruses,making it an ideal candidate for the early and accurate diagnosis of infection.In this study,the PDCoV N protein was induced and expressed in a prokaryotic system using IPTG treatment at low temperatures.Six-week-old BALB/c mice were immunized with the recombinant PDCoV N protein purified by nickel column.Isolated spleen cells were harvested and fused with SP2/0 cells once the serum titers reached 1∶10 000,as determined by indirect ELISA.The positive monoclonal hybridoma with the highest titer of secretory antibodies were screened af-ter three rounds of limited dilution.Ultimately,indirect immunofluorescent assay(IFA)and West-ern blot were employed to confirm the characterization of the antibodies,and the subtypes of the heavy and light chains of the antibodies were determined.One strain of hybridoma against the PD-CoV N protein,designated as 2G12,was obtained,with a titer approaching 1∶40 960.The IFA as-say showed PDCoV was specifically bound to 2G12.Furthermore,the Western blot assay further showed PDCoV in the supernatant of infected cells or recombinant PDCoV N protein,reacted well with 2G12.The subtype of MAbs was determined as IgG1 with the light chains being κ.The MAbs generated against the PDCoV N protein in this study provide valuable support for the development of a diagnostic kit for PDCoV and play an important role in the functional research of the PDCoV N protein.
