Immune Reaction And Protective Efficacy In Mice Of Oral Recombinant Lactococcus Lactis Mixed Antigens Of Helicobacter Pylori

ObjectivesIn present study, we aimed to induce and express the recombinant vaccine and oral gavage BALB/c mice for a month, then use H.pylori international standard strains11637challenge and test their colonization rate. To evaculate immune response and immune protection effect produced by different vaccineMethodl.Recombinant vaccine NZ3900/pNZ8110-ureB, NZ3900/pNZ8110-lpp20, NZ3900/pNZ8110-omp22-hpaA,NZ3900/pNZ8149-ureB,NZ3900/pNZ8110-LysM-hpaA,NZ3900/pNZ8149-SPusp45-ureB were cultured overnight and induced by40ng/ml nisin. SDS-PAGE identified recombinant immune protein quantity while Western-blot identified immnological specificity2. TB1/pMAL-c2X-ureB, TB1/pMAL-c2X-hpaA,TB1/pMAL-c2X-lpp20, TB1/pMAL-c2X-omp22were cultured overnight and induced by0.3mmol/L IPTG Crusing by ultrasound, purifying by Amylose affinity chromato-graphy, and determ-inating protein concentration by BCA3.The BALB/c mice were randomly divided into9groups, one time pre week for four times. Two week after the last immunization, half of the mice were sacrificed and collected their serum、succus gastricus.Rest of them were attacked with H.pylori, and then they were sacrificed two weeks after challenged, and the stomach was collected for semi-quantitative analysis based on urease test. The immune protective effect of different recombinant L.lactis was evaluate4. Statistical analysis of dataThe data was inputted to computer and the analysis was performed on SPSS17.0.The quantitative variable was analyzed by independent one-way ANOVA. The difference between two groups was analyzed by Bonferroni test.Results1.SDS-PAGE gel photo showed that the hybridization band of recombinant proteins of Lpp20, UreB, Omp22-HpaA, LysM-HpaA induced by Nisin were displayed at19kd,66kd,53kd,29kd,respectively, and Western-blot showed the recombinant proteins had good immunogenicity and immune reactivity.2. TB1/pMAL-c2X-ureB, TB1/pMAL-c2X-hpaA,TB1/pMAL-c2X-lpp20, TB1/pMAL-c2X-omp22were induced by IPTG and purified by Amylose affinity chromatography.3.Systemic antibody responses induced by recombinant vaccine were higher than pNZ3900/8149group, pNZ3900/8110group and PBS group, While the group of NZ3900/pNZ8110-LysM-hpaA were the highest. After H.pylori challenged, rapid urease test showed all of the recombinant vaccine had a protective effects.Conclusions1.Successfully obtained the high purity recombinant H.pylori antigen of Ureb, HpaA, Lpp20, Omp222. All of the oral gavage recombinant vaccine have immunogenicity.3.It discovered that NZ3900/pNZ8110-LysM-hpaA has the best immunorea-ction.