Antigenic peptides of the epizootic hematopoietic necrosis virus

The epizootic hematopoietic necrosis virus (EHNV) is a strain of the Iridovirus genus, which includes viruses seriously affecting native and aquacultured fish and amphibians. Despite its growing importance as a threat to fish farming, very little information is available on the biochemical and immunological nature of this virus. To identify and characterize the main antigenic determinants of EHNV, a panel of murine monoclonal antibodies was produced upon parenteral inoculation with live virus. A total of 124 primary hybridoma cultures from two fusions was found to produce antibodies reacting with EHNV by ELISA, but no neutralizing monoclonal antibody was detected. Twenty hybridoma cultures were randomly chosen for further study, and the antibodies secreted were analyzed by Western blotting, radioimmunoprecipitation, and immunostaining of infected cells. Only three MAbs immunoprecipitated the 50-kDa EHNV major capsid protein (MCP) from infected cell lysates, but they did not stain this protein in Western blotting. Eight and five further MAbs recognized peptides of approximately 15 and 18 kDa, respectively. Four antibodies could not be mapped into any viral protein, although they specifically immunostained virus-infected cells and reacted with purified EHNV virions by ELISA. These latter MAbs and the three antibodies directed at the MCP are likely to recognize conformation-dependent epitopes on the virus capsid proteins.