Application of a monoclonal antibody specific for the ORF92 capsid protein of Cyprinid herpesvirus 2

Cyprinid herpesvirus 2 (CyHV-2) infection is a recently discovered viral disease of farmed silver crucian carp (Carassius auratus gibelio) in mainland China that has resulted in massive fish mortality and appears to be spreading cross the world. Various detection methods based on nucleic acids have been developed, but effective immunological diagnostic method has not been reported for CyHV-2. In this study, recombinant ORF92 protein was used as a capture antigen to identify cells and tissues infected with CyHV-2 by immunological methods. Firstly, the open reading frame of CyHV-2 ORF92 was cloned into the PGEX-4T-3 vector and expressed in Escherichia coli. Purified recombinant ORF92 protein was then used as an antigen to prepare monoclonal antibodies, and an efficient hybridoma cell line was selected by dot-blot assay. The resulting mAb-187 was applied to detect CyHV-2 in infected Ryukin goldfish fin (RyuF-2) cells and fish tissues by western blotting and immunofluorescence assays. The monoclonal antibody could specifically identify CyHV-2 in infected RyuF-2 cells and gill, kidney, and spleen tissues, suggesting it could be applied for CyHV-2 detection in crucian carp.