Enzyme immunoassay test system for interoperative control of the specificity of recombinant capsid protein of porcine circovirus type 2

Porcine circovirus infections, primarily caused by porcine circovirus type 2 (PCV2), are one of the most significant problems in global pig production. Vaccination is a key tool for control and prevention of porcine circovirus infections. Recombinant analogs of capsid protein (rCap) of porcine circovirus type 2 (PCV2) are key components of modern diagnostic test systems and subunit vaccines. The stability of its antigenic activity is a critical parameter determining the efficiency and reproducibility of the functional product. The aim of this study was to develop and test an enzymelinked immunosorbent assay system for interoperational monitoring of the specific activity of rCap protein. To obtain recombinant analogues of the PCV-2 capsid protein, the previously constructed recombinant E. coli BL21(DE3)pLysSORF2 producer strain (for prokaryotic analogue) and the stably expressing Chinese hamster ovary cell line CHO-K1/PCVORF2 (for eukaryotic analogue) were used; protein purification was performed according to previously developed protocols. Chromatographically purified IgG isolated from a pool of sera from PCV-2-seropositive pigs was used for sensitization of the solid phase (polystyrene plates); the specific peroxidase conjugate was obtained by the periodate method. The study resulted in the production of a stable peroxidase conjugate with a labeling density of 1.5:1. The developed test system enabled semiquantitative analysis of recombinant raw materials, demonstrating good reproducibility (coefficient of variation <7%). The test system was found to be universal and capable of assessing the activity of rCap variants obtained in both prokaryotic and eukaryotic expression systems, accurately recognizing their antigenic determinants. Thus, the implementation of the semiquantitative ELISA method in the production cycle will standardize the process of in-process quality control of biomass obtained using recombinant DNA technologies and guarantee stable antigenic characteristics of vaccines and diagnostic products.