Establishment of an indirect ELISA method for bovine respiratory syncytial virus rG protein
10.16303/j.cnki.1005-4545.2025.09.08
- VernacularTitle:牛呼吸道合胞体病毒重组G蛋白间接ELISA方法的建立
- Author:
Hong LI
1
;
Guanxin HOU
;
Chihuan LI
;
Siping ZHU
;
Chao REN
;
Xintong ZHU
;
Xiaochen LIU
;
Yulai DONG
;
Qiumei SHI
;
Zhiqiang ZHANG
Author Information
1. 河北科技师范学院动物科技学院河北省预防兽医重点实验室,河北秦皇岛0660042
- Publication Type:Journal Article
- Keywords:
bovine respiratory syncytial virus;
rG protein;
indirect ELISA
- From:
Chinese Journal of Veterinary Science
2025;45(9):1878-1887
- CountryChina
- Language:Chinese
-
Abstract:
In order to establish a serological method for the detection of bovine respiratory syncytial virus,the prokaryotic expression of four proteins of BRSV,G,F,P,and M was carried out,and the most suitable coating antigen was screened to establish an indirect ELISA detection method.The results showed that the four recombinant proteins of BRSV,rG,rF,rP and rM were successfully expressed.The results of checkerboard screening showed that the P/N value of rG protein was the largest,which was determined to be the best coating antigen established by indirect ELISA meth-od.The optimal reaction conditions for indirect ELISA were as follows:the mass concentration of rG protein coating was 1 mg/L,37℃ for 2 h;3％BSA 37℃ block for 1 h;Serum was diluted 1∶50 and incubated at 37℃ for 1h;Secondary antibody 1∶5 000 dilution,37℃ for 30min;The color development conditions of the substrate were 37℃ for 15 min;Thirty negative sera were selected,and the cut-off value was determined to be 0.63 by the established indirect ELISA method.The re-sults of the specificity test showed that the indirect ELISA method established in this test only recognized BRSV-positive serum,and did not react with IBRV,BCoV,and BPIV3-positive serum.The results of repeatability test showed that the method had good repeatability,and the coefficient of variation within and between batches was less than 10％.The results of the sensitivity test showed that the BRSV-positive serum was still positive when diluted to 1∶8 192.The indirect ELISA method established in this experiment was used to detect 100 clinical serum samples at the same time,and the total coincidence rate of the two reached 90.48％,the positive coincidence rate was 93.42％,and the negative coincidence rate was 82.75％.The indirect ELISA established in this test can be used for the detection of bovine respiratory syncytial virus in clinical practice.
