Studies On Gene Engineered Vaccine Of The Anti-idiotype MAb Against Vibrio Alginolyticus

Fish disease played the key role on restricting the development of aquaculture industry. Various attempts, including chemical, antibiotic and attenuated or inactivated vaccine, had been used to treat and prevent bacterial infectious diseases in farmed fish. The chemicals may have residual effect that could endanger fish health. Using antibiotics on fish were easy to generate resistant strain of bacteria. Safety of attenuated or inactivated vaccine for fish, environment and consumers were difficult to control. So people seek for the more safe and economical methods to treat and prevent bacterial infectious fish diseases.The novel vaccines were occurred. They include synthetic peptide vaccine, nucleic acid vaccine, anti-idiotype vaccine and so on. But these novel vaccines had some shortcoming on safety and economy. Anti-idiotype antibody vaccines have advantages over conventional vaccines. It had good immunogenicity and reliability but expensiveness. Acheaper vaccine that had higher affinity and stronger penetration was hoped to construced by recombinant DNA technology. So we choose the scFv of the anti-idiotype mAb against vibrio alginolyticus. The scFv was composed of antibody heavy and light chain variable domains joined with a synthetic linker peptide. The scFv shows the more beneficial and economical compared with parental antibody. Therefore, using recombinant DNA technology to develop the scFv of the anti-idiotype mAb against vibrio alginolyticus should be considered as the more economical and practical way.Fish disease has caused heavy economic losses in the aquaculture industry. Vibrio alginolyticus was one of the pathogens that infected fresh and marine fish. We had obtained the hybridoma cell 2F4 secreting anti-idiotype monoclonal antibody (mAb) against vibrio alginolyticus. Fish immunized with this anti-idiotype mAb showed effective protection. It had good immunogenicity and reliability. So it was applied in the aquaculture industry.To construct of gene engineered vaccine of the anti-idiotype mAb against vibrio alginolyticus, in this work total RNA was extracted from hybridoma cell 2F4 secreting anti-idiotype mAb against vibrio alginolyticus and VH and Vl gene fragment was amplified by RT-PCR. The scFv was arranged in Vh -Linker- Vl orientation, being joined together with 15-amino-acids (Gly4Ser)3 linker. The amplified scFv gene fragment was approximately 750bp. This 747bp DNA fragment encoded for a protein fragment of 249 amino acid residues. The VH gene and VL gene feagments of this sequence included three complementarity-determining regions (CDRs), four framework regions (FRs) and two characteristic cysteines residues, respectively. This 747bp DNA fragment contained a stong antigenic epitope and this antigenic epitope was successfully expressed as a fusion protein in bacteria using prokaryotic expression plasmid. The...