The RPIII Fusion Protein Of Eg95-1,Eg95-2,Eg95-3 That The Short Peptide Epitope Of Echinococcus Granulosusinduce To Induce The Immune Response Characteristics Of Mice

Objective: To find the vaccine candidate against hydatidosis,this study aims at the potential function of the recombinant protein rP? of Eg95-1?Eg95-2 and Eg95-3 in Echinococcus granulosus.In order to discuss the immuno protection and its mechanisms of the recombinant protein rP? of Eg95-1?Eg95-2 and Eg95-3,we will observe the characteristic of the recombinant protein rP? of Eg95-1?Eg95-2 and Eg95-3 vaccine candidate factor through the modol of mice which immunized with the recombinant protein rP? of Eg95-1,Eg95-2 and Eg95-3 and infected by Echinococcus granulosus.Method: Our research group have already used bioinformatics technology to analyse and obtain the information of the T-B union epitope of Eg95 and using phage display technology to identify and screen.To constructed M13KE/Eg95-1,M13KE/Eg95-2 and M13KE/Eg95-3 plasmid,then expressed and purified the recombinant protein rP? of Eg95-1?Eg95-2 and Eg95-3.The first part of the experiment: the mice were randomly divided into four groups.Respectively using the recombinant protein rP? of Eg95-1,Eg95-2 and Eg95-3 to immune the femal Balb/C mice,the forth group is the control group.Then we used ELISA to detect the level of antibodies(IgG and IgE)in mice serum each group in week 0,week 2,week 4,week 6,week 8,week 10 and to detect the level of cytokines(IL-2?IL-4?IL-10?TNF-? and IFN-?)in week 0 and week 10.We examed The proliferation rates of spleen lymphocytes with different stimulation by MTT in week 10.The second part of the experiment : the mice were randomly divided into four groups.Respectively we used the recombinant protein rP? of Eg95-1,Eg95-2 and Eg95-3 to immune the femal Balb/C mice,the fourth group is the control group.Then all the mice challenged intraperitoneally with E.g protoscoleces in week 8,executed in week 32.We Used ELISA to detect the level of antibodies(IgG and IgE)and cytokines(IL-2?IL-4?IL-10,TNF-? and IFN-?)in mice serum each group in week 0,week 2,week 4,week 6,week 8,week 10.We examed the proliferation rates of spleen lymphocytes with different stimulation by CCK in week 32.Result: To expressed and Purified the recombinant protein rP? of Eg95-1?Eg95-2 and Eg95-3 successfully.The first part of the experiment: the recombinant protein rP ? of Eg95-1,Eg95-2 and Eg95-3 immune the mice respectively,start from the 4th weeks,the levels of antibodies(Ig G and IgE)are increased,and the level of IgE peaked in week 8,gradually falled in week 10,but the level still higher than week 0 with statistical significance(P < 0.05).The levels of cytokines(IL-2?IL-4?IL-10?TNF-? and IFN-?)of Eg95-1 group,Eg95-2 group,Eg95-3 in week 10 obvious higher than in week 0(P < 0.05).The levels of cytokines(IL-2?IL-4?IL-10?TNF-? and IFN-?)of the control group have no significant change.Spleen lymphocytes of Eg95-1 group,Eg95-2 group,Eg95-3 group and control group all can be stimulated induced to proliferate by ConA,and Eg95-1 group,Eg95-2 group and Eg95-3 group can also be stimulated induced to proliferate by Eg95 antigen,and in week 10,three experimental groups were statistically significant(P < 0.05).The second part of the experiment: the levels of antibodies(IgG and Ig E)and cytokines(IL-2?IL-4?IL-10?TNF-? and IFN-?)of Eg95-1 group,Eg95-2 group,Eg95-3 groups in week 8,week 10,week 32 compared with week 0 were all increased,with statistical significance(P < 0.05).The levels of cytokines(IL-2,TNF-? and IFN-?)of Eg95-1 group,Eg95-2 group,Eg95-3 group peaked in week 10,gradually falled in week 32,but the levels still higher than week 0 with statistical significance(P < 0.05).The levels of cytokines(IL-2,TNF-? and IFN-?)of control group peaked in week 10,gradually falled in week 32,compared with week 0 were statistically significant(P < 0.05).The levels of cytokines(IL-4 and IL-10)of Eg95-1 group,Eg95-2 group,Eg95-3 group and control group are on the rise,the week 8,week 10,week 32 all have statistical significance.After Eg95 antigen stimulated,spleen lymphocytes of Eg95-1 group,Eg95-2 group,Eg95-3 groups and control group were all markedly proliferation,statistically significant compared with no irritant.Conclusion: the recombinant protein rP? of Eg95-1?Eg95-2 and Eg95-3 to induce mainly IgG immune mice humoral immune response.the recombinant protein rP? of Eg95-1?Eg95-2 and Eg95-3 can be induced by immune mice produce humoral and cellular immune response in mice.the recombinant protein rP? of Eg95-1?Eg95-2 and Eg95-3 can induce the secondary infected mice to produce strong protection,and can stimulate the host humoral immune response and cellular immune response to play a role of immune protection.the recombinant protein rP? of Eg95-1?Eg95-2 and Eg95-3 can be used as final host canine vaccine candidate molecules,as we later connects effective antigen epitope to develop the MAP for studying the prevention of Echinococcus granulosus stronger,lay the foundation for more effective vaccine.