Immune Protection Of CSFV DNA Vaccine And Enhancing Its Immune Efficiency

Immune Protection of CSFV DNA Vaccine and Enhancing Its Immune efficiencyImmune response against classical swine fever virus (CSFV) was elicited after immunization of pig with CSFV E2 gene eukaryotic expression plasmid pcDSW. The immunized pigs could be protected from the challenge of a lethal dose of CSFV virulent strain, Shimen.Mice were immunized with plasmids, pIRSTIL-2 and pIRSTIL-3, expressing E2 gene and murine IL-2 or IL-3, respectively. The results showed that IL-2 and IL-3 could obviously enhance the immune response level of CSFV DNA vaccine, alone or cooperatively. The cDNAs of porcine IL-2 and Il-6 were obtained from blood by RT-PCR and cloned into the pGEM-T Vector. A eukaryotic expression plasmid pIRE2PIL2, expressing the CSFV E2 gene and porcine IL-2, was contructed and was inoculated into piglets. The piglets immunized with both pIRE2PIL2 and pIRE2, which express the CSFV E2 gene only, were obtained a comparatively protection level after challenge with CSFV virulent strain, Shimen. Some enhancing immune effect was demostrated by that the amounts of white blood cells(WBC) of piglets immunized with pIRE2PIL2 didn't decreased after challenge but that of piglets immunized with pIRE2 decreased obviously.Two hybridoma secreting monoclonal antibodies(McAb) against the epitopes on CSFV E2 were achieved by injection of Balb/c mice with pIRE2 plasmid prepared in DNA vaccine formula (resulting in McAbs GE and GB). These two McAbs were further characterized by their reactivities, specificities and neutralizing abilities vs. CSFV. The results showed that: a. Both McAbs could reacted in ELISA and Western-blot detection with both prokaryoteexpressed E2 protein and the antigens prepared from tissue culture of CSFV virulent strain Shimen. b. Both McAbs could neutralize the infection of rabbits with attenuated CSFV vaccine lapinized virus or c-strain, c. In indirect ELISA both McAbs could reacted with 9 field strains of CSFV. GB could crossly reacted with bovine viral diarrhea virus (BVDV), while GE could not. indicating GB probably recognizes a common antigenic epitope of pestvirus.Two more experiments relating to the above programme have been done:A. 9.5kb cDNA fragment of the 12.3kb genome of CSFV avirulent strain. HCLV or cstrain. was obtained by RT-PCR from the spleen or lymph nodes of rabbit infected with HCLV.B. A 18mer of peptide (ACDCRGDCFCGPKKKRKV, named RNLS), which contains a motif of RGD and a nuclear localization signal (NLS) of SV40 large T-antigen. was synthized. The primary experiment showed that RNLS could enhence the transfection level of DNA obviously...