Expression Of The Truncated E2 Protein Of Classical Swine Fever Virus In Escherichi Coli And Preparation Of A Monoclonal Antibody Against E2 Protein

The envelope glycoprotein E2 of classical swine fever virus (CSFV) is responsible for the elicitation of neutralizing antibodies. It is often used in developing new type vaccines, clinical diagnostic reagents and studying immunopathological mechanisms of the virus.A tuncated E2 gene encoding the major antigenic regions of E2 protein was amplified by RT-PCR from the genomic RNA of CSFV C-strain and cloned into pPROEX-HTb expression vector, resulting in pPROEX-tE2. The truncated E2 protein (tE2) was expressed with high-level in pPROEX-tE2-transformed Escherichi coli cells after induction by IPTG, as demonstrated by SDS-PAGE analysis. The recombinant protein could be recognized by CSFV antisera in either Western blotting or ELISA.The tE2 protein purified on a Ni-chelating HisTrap affinity column was used to immunize BALB/c mice, of which spleen cells were fused with SP2/0 cells with PEG3250 after 3 immunizations. A hybridoma cell line stably secreting monoclonal antibody (McAb) directed against the tE2 protein was selected by ELISA. The McAb was proved to be CSFV-specific and recognize a linear epitope on the E2 protein of CSFV C-strain or Shimen strain. The McAb was identified to be IgG1 subtype and kappa light chain.In summary, the truncated E2 protein of CSFV was expressed in E. coli and a hybridoma cell line secreting McAb directed against CSFV E2 protein was developed, which can be used to develop diagnostic assays and study the structure-function of CSFV E2 protein.