Expression of GP5 gene of PRRSV CH-1a strain and development of monoclonal antibodies against GP5

A fragment encoding the first 26 amino acids of glycoprotein 5(GP5) of PRRSV strain CH-1a was deleted,the truncated fragment was cloned into a prokaryotic expression vector pGEX-6p-1,and a fusion-expressed protein rtGP5 of 42ku was obtained in E.coli.The protein rtGP5 showed a strong reaction to the PRRSV-positive sera in Western-blotting analysis.BALB/c mice were immunized intrape-(ritoneally) with purified rtGP5 protein.Murine myeloma cells were fused with the splenocytes of the immunized mice after the third immunization.An indirect ELISA coated with rtGP5 and GST was used to screen hybridomas for production of specific antibody in hybridoma culture fluid.15 hybridomas clones producing-steadily MAbs were obtained after 3 cycles of cloning.All the 15 MAbs showed strong reactivity in IFA on the PRRSV infected Marc-145 cells.All the MAbs belong to IgG1 isotype,and the light chains of the antibodies are κ chain,which were isotyped by ELISA using a commercial kit.The MAbs against PRRSV GP5 protein could be used for further analysis of the structure and function of PRRSV,and identification of epitopes on GP5 which may be helpful for the design of new vaccines.