Characterization of monoclonal antibodies against Muscovy duck reovirus sigmaB protein

The sigmaB protein of Muscovy duck reovirus (DRV), one of the major structural proteins, is able to induce neutralizing antibody in ducks, but the monoclonal antibody (MAb) against sigmaB protein has never been characterized.

Four hybridoma cell lines secreting anti-DRV sigmaB MAbs were obtained, designated 1E5, 2F7, 4E3 and 5D8. Immunoglobulin subclass tests differentiated them as IgG2b (1E5 and 4E3) and IgM (2F7 and 5D8). Dot blot and western blotting assays showed that MAbs reacted with His-sigmaB protein in a conformation-independent manner. Competitive binding assay indicated that the MAbs delineated two epitopes, A and B of sigmaB. Immunofluorescence assay indicated that the four MAbs could specifically bind to Vero cells infected with DRV and sigmaB was distributed diffusely in the cytoplasma of infected cells. MAbs had universal reactivity to all DRVs tested in an antigen-capture enzyme-linked immunosorbent assay.

Results of this research provide important information about the four monoclonal antibodies and therefore the MAbs may be useful candidate for the development of a MAb capture ELISA for rapid detection of DRVs. In addition, it showed that the sigmaB protein was located in the cytoplasma of infected cells by immunofluorescence assay with MAbs. Virus isolation and RT-PCR are reliable way for detection of DRV infection, but these procedures are laborious, time consuming, and requiring instruments. These obvious diagnosis problems highlight the ongoing demand of rapid, reproducible, and automatic methods for the sensitive detection of DRV.