Eukaryotic expression and monoclonal antibody preparation of CP312R recombi-nant protein of African swine fever virus
To prepare monoclonal antibodies against the CP312R protein of African swine fever vi-rus(ASFV),BALB/c mice were immunized with eukaryotic expression of CP312R protein of Af-rican swine fever virus(ASFV),and the reactivity of the obtained monoclonal antibodies was iden-tified by indirect immunofluorescence assay(IFA).The vector pOET3-CP312R was constructed u-sing the baculovirus expression system(BES),and it was co-transfected with the baculovirus ge-nome flashBACTM ULTRA into Sf9 insect cells to obtain the recombinant baculovirus AcMNPV-CP312R.The virus expressed the recombinant ASFV CP312R protein in an intracellular soluble form.Identification by Western blot showed that the recombinant CP312R protein could react spe-cifically with ASFV positive serum.In addition,two monoclonal antibodies(McAb)targeting the recombinant CP312R protein were obtained.The titer of McAbs were detected by indirect ELISA,and was found to be no less than1∶1 024 000.IFA detection showed that monoclonal antibodies reacted specifically with ASFV.This study provides material reserves for the development of Afri-can swine fever subunit vaccines and the establishment of serological testing methods.
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