Wenying Zhang , Yulong Wang , Guodong Wang , Hangbo Yu , Mengmeng Huang , Yulong Zhang , Runhang Liu , Suyan Wang , Hongyu Cui , Yanping Zhang , Yuntong Chen , Yulong Gao , Xiaole Qi 

Viruses. 2025 Jun 20;17(7):871. doi: 10.3390/v17070871.

Abstract

Infectious bursal disease virus (IBDV) is one of the most important immunosuppressive viruses in poultry, causing the global spread of infectious bursal disease (IBD). It poses a significant threat to the healthy development of the poultry industry. Vaccination is an effective approach for controlling IBDV infection. Therefore, reliable immune monitoring for IBDV is critical for maintaining poultry health. The enzyme-linked immunosorbent assay (ELISA) is a common technique used to detect specific antibodies in clinical serum testing and for the serological evaluation of IBDV vaccines. Among the currently available and under development IBDV vaccines, IBD VP2 subunit-based vaccines account for a considerable proportion. These vaccines stimulate the production of antibodies that are specific only to VP2. However, most IBDV antibody ELISA kits approved for use have applied the whole virus as the coating antigen, which does not adequately meet the diverse requirements for IBDV detection across different conditions. This study utilized a prokaryotic expression system to express the VP2 protein of the IBDV epidemic strain, assembling it into virus-like particles to be used as coating antigens. This approach enabled the establishment of an indirect ELISA method for detecting IBDV VP2 antibody (VP2-ELISA). The optimal coated antigen concentration was determined to be 2.5 μg/mL, with overnight coating at 4 °C; sealing with 5% skim milk at 37 °C for 4 h; serum dilution at 1:500 with incubation at 37 °C for 30 min; secondary antibody dilution at 1:4000 with incubation at 37 °C for 40 min; and then incubation with the substrate solution 3,3',5,5'-tetramethylbenzidine at room temperature for 20 min. The criterion for interpreting the detection results was OD450nm ≥ 0.111 indicates IBDV antibody positivity, while OD450nm < 0.111 indicates negativity. The established VP2-ELISA can specifically detect IBDV-positive sera at the lowest serum dilution of 1:6400, with intra- and inter-batch coefficients of variation of<2%. this="" indicates="" that="" the="" vp2-elisa="" exhibits="" good="" specificity="">

Keywords: ELISA; IBDV; VLP; VP2; antibody detection.