Development and characterization of two monoclonal antibodies against grouper iridovirus 55L and 97L proteins

Grouper iridovirus (GIV) is one of the most important viral pathogens in grouper, particularly at the fry and fingerling stages. The study of GIV pathogenicity has been hampered by the lack of proper immunological reagents to study the expression and function of viral proteins in the infected cells. In this study, two mouse monoclonal antibodies (mAbs) against GIV 55L and 97L proteins were produced. Enzyme-linked immunosorbent assay (ELISA) and Western blotting were used to screen these hybridomas, resulting in the identification of two high-affinity mAbs named GIV55L-mAb-2 and GIV97L-mAb-3, respectively. Both mAbs belong to the IgG1 isotype and were effective in detecting their respective target viral protein. Reverse-transcription polymerase chain reaction (RT-PCR) and Western blot analyses of GIV-infected GK cells revealed that GIV 97L is an immediate early gene, whereas GIV 55L a late one. The localization of 55L and 97L in GIV-infected cells was further characterized by immunofluorescence microscopy with the mAbs. The 55L protein mainly aggregated in the cytoplasm while 97L distributed in both the nucleus and cytoplasm of the infected cells. These studies demonstrate the validity of the two mAbs as immunodiagnostic and research reagents.