Establishment and preliminary application of indirect ELISA method for detection of bovine parainfluenza virus type 3 based on HN protein
10.16303/j.cnki.1005-4545.2025.03.01
- VernacularTitle:基于HN蛋白的牛副流感病毒3型间接ELISA检测方法的建立及初步应用
- Author:
Hong LI
1
;
Rui AN
;
Chihuan LI
;
Siping ZHU
;
Yulai DONG
;
Tonglei WU
;
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 066004
- Publication Type:Journal Article
- Keywords:
bovine parainfluenza virus type 3(BPIV3);
HN protein;
indirect ELISA;
prokaryotic ex-pression
- From:
Chinese Journal of Veterinary Science
2025;45(3):397-403
- CountryChina
- Language:Chinese
-
Abstract:
In order to establish a serological method for the detection of bovine parainfluenza virus type 3(BPIV3),the prokaryotic expression and purification of BPIV3 HN,NP,F,and P proteins were carried out,and the optimal protein-coated antigen was screened,and an indirect ELISA de-tection method was established.The results showed that the four recombinant proteins of BPIV3,rHN,rNP,rF,and rP were expressed,and the checkerboard titration results showed that rHN pro-tein had the highest P/N value as the coating protein,so it was used for the subsequent method es-tablishment.The optimal reaction conditions for indirect ELISA were found to be:the mass con-centration of the antigen coating was 0.5 mg/L,37 ℃ 1.5 h,5％skim milk,overnight blocking at 4 ℃,serum dilution at 1∶50,incubation at 37 ℃ 1 h,secondary antibody dilution at 1∶10 000 and incubation at 37℃ 0.5 h,substrate reaction conditions were 37℃ for 12 min.The results of speci-ficity experiments showed that the established method could specifically identify BPIV3 antibody-positive serum with a sensitivity of 1∶800,and the coefficient of variation in the detection of intra-and inter-assay repeatability was less than 10％,and the overall coincidence rate of the same batch of samples detected with the SVANOVIR kit was 92.22％.This method was used to detect 192 se-rum samples in Hebei Province,and the positive rate of BPIV3 antibody in serum was 66.15％.The indirect ELISA detection method of BP1V3 antibody constructed in this study is suitable for large-scale clinical serological investigations,and provides valuable data support for the research and de-velopment of BPIV3 antigen and antibody detection kits in China.
