ABSTRACT
<p><b>OBJECTIVE</b>To establish and optimize the proteomic analysis of protoscoleces-specific antigens from Echinococcus granulosus. To provide a foundation for identifying specific antigens in the soluble proteins of E. granulosus protoscoleces for further research.</p><p><b>METHODS</b>Brood capsules were collected aseptically from fertile E. granulosus cysts from the livers of an infected patient. The fertile E. granulosus cysts were fractured, and protoscoleces were collected by centrifugation. The soluble proteins of protoscoleces were acquired using the 2D Quant kit according to the manufacturer's instructions. We employed two-dimensional electrophoresis (2-DE) combined with immunoblot assay (Western blot) to analyze the soluble components of E. granulosus protoscoleces antigens. The 2-DE and immunoblot maps obtained were analyzed with PDQuest 8.0 image analysis software.</p><p><b>RESULTS</b>About 233 soluble protein spots were identified with Coomassie-stained gels. Most of the proteins had a molecular weight of 16,000 Da to 117,000 Da, and an isoelectric point value of 3.0 to 10.0. 2-DE immunoblot was conducted and 57 specific antigen spots were observed, among which 23 spots were identified.</p><p><b>CONCLUSION</b>2-DE combined with Western blot is the key to successful proteomic analysis and presents a new possibility for searching the specific E. granulosus protoscoleces antigens.</p>
Subject(s)
Animals , Antigens, Helminth , Chemistry , Metabolism , Blotting, Western , Methods , Echinococcus granulosus , Classification , Metabolism , Electrophoresis, Gel, Two-Dimensional , Gene Expression Regulation , Helminth Proteins , Proteomics , MethodsABSTRACT
<p><b>OBJECTIVE</b>To investigate the protective immunity against Echinococcus granulosus in mice immunized with rEg14-3-3.</p><p><b>METHODS</b>ICR mice were subcutaneously immunized three times with rEg14-3-3, followed by the challenge with Echinococcus granulosus protoscoleces intraperitoneally and then sacrificed after six months of post-challenge to detect the proliferation of splenocytes by MTT assay, and to measure the secretion of IL-2, IL-4, IL-10, and IFN-γ by ELISA. The rate of reduced hydatid cyst and the levels of IgE, IgG and IgG subclasses in sera were examined.</p><p><b>RESULTS</b>Mice vaccinated with rEg14-3-3 and challenged with protoscoleces revealed significant protective immunity of 84.47%. ELISA analysis indicated that the immunized mice generated specific high levels of IgG and the prevailing isotypes of IgG were IgG1 and IgG2a. Splenocytes from mice immunized with rEg14-3-3 showed a significant proliferation response. The secretion of IFN-γ and IL-2 increased significantly in the vaccinated mice whereas there was no significant difference in IL-4 and IL-10 levels between vaccinated and control mice.</p><p><b>CONCLUSION</b>The results indicate that the rEg14-3-3 vaccine could induce a high level of protective immunity as a promising vaccine candidate to prevent cystic echinococcosis.</p>