ABSTRACT
Objective To generate the spaO-ompA fusion gene of Salmonella paratyphi A and its prokaryotic expression system,and to determine the immunoprotection of the recombinant expression product rSpaO-OmpA.Methods A flexible peptide sequence was used to link spaO and ompA genes and a prokaryotic expression system of spaO-ompA fusion gene was subsequently generated.SDS-PAGE and Bio-Rad Agarose Image Analyzer were applied to examine the expression as well as the yield of the target recombinant protein rSpaO-OmpA.The antigenicity and immunoreactivity of rSpaO-OmpA were determined using immunodiffusion test,Western Blot assay and micro-Widal's test.By a mouse infection model,the immunoprotection of rSpaO-OmpA against the lethal challenge of S.paratyphi A was determined.In the animal protective test,the recombinant expressed SpaO (rSpaO) and OmpA ( rOmpA ) were used as the controls.Results The generated spaO-ompA fusion gene had 100％ nucleotide and amino acid sequence identities compared to the single spaO or ompA gene.The constructed prokaryotic expression system IPTG E.coli BL21DE3pET42a-spaO-ompA expressed the recombinant protein rSpaO-OmpA.rSpaO-OmpA combined with the antiserum against wholecell of S.paratyphi A to present positive hybridization signal and induced specific antibody in the immunized rabbits.Immunization with 100 or 200 μg rSpaO-OmpA contributed 66.7％ (8/12) or 83.3％ (10/12) immunoprotective rates in mice when the animals were attacked with S.paratyphi A.The immunoprotective rates produced by rSpaO-OmpA were significantly higher than that of equal rSpaO or rOmpA( P＜0.05 ).The sera from rSpaO-OmpA immunized mice presented 1∶5-1∶40 agglutination titers to the H antigens of different S.paratyphi species,and 1∶1-1∶16 immunodiffusion titers to rSpaO,rOmpA and rSpaO-OmpA proteins,respectively.Conclusion The artificially fusion antigen,rSpaO-OmpA,has more powerful immunogenicity and immunoprotection that the equal rSpaO or rOmpA.
ABSTRACT
AIM: To investigate the changes on immunological function in antigen-sensitized mice injected by dermatophagoides farinaeliposomes (DF) subcutaneously. METHODS: BALB/c mice were sensitized with DF antigen injected subcutaneously. After injected by DF liposomes with 1, 3, and 9 ?g?g -1 dosage respectively, the total IgE, total IgG, DF specific IgE(sIgE), DF specific IgG (sIgG), IL-4, and IFN-? were detected by ELISA. RESULTS: The total IgE, sIgE, total IgG, and sIgG in the serum of sensitized mice increased, the level of IFN-? reduced and IL-4 increased. After injected subcutaneously by DF liposomes, the total IgE, sIgE and IL-4 in the serum of mice of all the treatment groups reduced, and IFN-? raised, and the total IgG, sIgG raised in the serum of mice administrated with small and medium dosage, but the low dosage was more obvious. CONCLUSION: DF liposomes have some effects on the immunological function in mite-sensitized mice in the low dosage.