Preparation of monoclonal antibody against bovine viral diarrhea virus and estab-lishment of double antibody sandwich ELISA method
10.16303/j.cnki.1005-4545.2025.11.06
- VernacularTitle:牛病毒性腹泻病毒单克隆抗体的制备及双抗体夹心ELISA方法的建立
- Author:
Qianyue MA
1
;
Jiaxuan LI
1
;
Yanping JIANG
1
;
Wen CUI
1
;
Xinyuan QIAO
1
;
Changcheng ZHU
1
;
Shize HAO
1
Author Information
1. 东北农业大学动物医学学院黑龙江省动物疾病防控技术与制剂创制重点实验室,哈尔滨 150030
- Publication Type:Journal Article
- Keywords:
bovine viral diarrhea virus;
monoclonal antibody;
double antibody sandwich ELISA
- From:
Chinese Journal of Veterinary Science
2025;45(11):2343-2350
- CountryChina
- Language:Chinese
-
Abstract:
The purpose of this study was to prepare high affinity monoclonal antibodies(mAbs)a-gainst bovine viral diarrhea virus(BVDV)and establish a double antibody sandwich ELISA detec-tion method.BVDV was purified by differential ultracentrifugation and used to immunize BALB/c mice.Hybridoma cells were prepared by fusing spleen cells from the immunized mice with SP2/0 cells.Positive cells were screened by indirect ELISA.A double-antibody sandwich ELISA method for detecting BVDV was developed using monoclonal antibody 4D11 as the capture antibody and HRP-labeled monoclonal antibody 3F3 as the detection antibody.The results of the ELISA and the determination of the variable region gene sequence of monoclonal antibodies indicated that the two monoclonal antibodies recognize different antigenic epitopes.Specificity tests showed that two monoclonal antibodies specifically recognize BVDV and did not cross-react with other bovine viru-ses associated with diarrhea.Indirect immunofluorescence assay and Western blot assay demonstra-ted that both mAbs exhibited strong reactivity with BVDV.The double antibody sandwich ELISA detection method established in this study had good specificity.The sensitivity test revealed that the method could detect a minimum virus amount of 3.1 × 104 TCID50.The reproducibility test showed that the inter-batch coefficient of variation(Cv)was between 2.47％and 7.44％,and the intra-batch Cv was between 1.71％and 9.89％,indicating good reproducibility.The establishment of this method provides an effective technical tool for the rapid diagnosis and prevention and con-trol of BVDV.
