This study examines the S2 protein of the Nephropathogenic Infectious Bronchitis virus (NIBV), essential for the virus's adaptation and expanded tissue and host range. Using bioinformatics, its hydrophilicity, hydrophobicity, signal peptides, antigenic epitopes, secondary and tertiary structures were analyzed. The NIBV-S2 gene was amplified via RT-PCR, cloned into the pET-32a (+) vector to form the recombinant plasmid pET-SX9-S2, and transferred into BL21(DE3) cells. An anti-rHis-S2 polyclonal antibody was generated using the expressed rHis-S2 as the antigen. The indirect ELISA titer of this polyclonal antibody was 1:1024,000. This polyclonal antibody against NIBV-S2 effectively detects NIBV-S2 and is suitable for western bloting, immunofluorescence, and Co-IP experiments. Detection showed a marked rise in NIBV-S2 protein expression from 3 to 9 dpi, peaking at 9 dpi, followed by a decline from 9 to 15 dpi as the infection progressed. At 9 dpi, there was a notable downregulation of the ABCG2, a uric acid transporter, in the kidneys of NIBV-infected chickens compared to the control group. Furthermore, analyses employing molecular docking, immunofluorescence, co-immunoprecipitation (Co-IP), and molecular dynamics (MD) simulations have demonstrated that the NIBV-S2 protein possesses the ability to bind to ABCG2. This observation was further substantiated by in vitro inhibition experiments, which revealed that the expression level of the S protein decreased following the application of the ABCG2 inhibitor Ko143. In summary, NIBV utilizes its S2 protein to interact with ABCG2, thereby facilitating viral colonization in the kidneys, which may contribute to urate accumulation.

To Cite This Article: Liu Z, Shi Y, Wu M, Chen C, Zheng K, Qiu L, Li M, Gao X, Yin C and Guo X, 2026. Development and application of polyclonal antibody against s2 protein of nephropathogenic infectious bronchitis virus. Pak Vet J, 46(5): 1219-1230. http://dx.doi.org/10.29261/pakvetj/2026.109