Preparation and characterization of a monoclonal antibody against MCP of largemouth bass ranavirus and investigation of the virus's tissue tropism

Largemouth bass ranavirus (LMBV) is a highly contagious pathogen causing severe symptoms and significant mortality in freshwater fish. With no effective vaccine or treatment, rapid detection and control are critical for managing outbreaks. In this study, we report the development and characterization of monoclonal antibodies (mAbs) targeting the major capsid protein (MCP) of the LMBV. Furthermore, these mAbs, in conjunction with quantitative polymerase chain reaction (qPCR) techniques, were employed to investigate tissue tropism and distribution following LMBV infection in largemouth bass. The findings revealed that three hybridoma cell lines, designated as 45ED8, 9F5F2 and 20D7, consistently secreted anti-MCP mAbs. The 45ED8 and 9F5F2 mAbs were identified as IgG1 subtype antibodies, whereas 20D7 was classified as an IgG2b subtype antibody. All mAbs specifically detected LMBV in western blot (WB), immunofluorescence assay (IFA), and immunohistochemistry (IHC) without cross-reacting with other fish pathogens. One of the mAbs demonstrated significant neutralizing activity against LMBV, with a neutralization titer of 1:128. IHC analysis revealed positive signals in all tested samples from LMBV-infected fish, with particularly strong signals observed in the liver, spleen, and intestines. These findings are consistent with qPCR results. In light of the current scarcity of widely accessible reagents for LMBV, these specific mAbs hold potential for advancing research and establishing a foundation for novel diagnostic and therapeutic approaches. Furthermore, the observed tissue tropism enhances our understanding of the pathogenesis and host-virus interactions of LMBV, thereby informing future research endeavors. © 2025 Published by Elsevier B.V.