Event Abstract Back to Event A hybrid protein designed with epitopes from B. tropicalis and D. pteronyssinus allergens showed reduced IgE binding and allergenic activity Dalgys Martínez1, Marlon Múnera1, Jose Cantillo1, Luis Caraballo1, 2 and Leonardo Puerta1, 2* 1 University of Cartagena, Institute for Immunological Research, Colombia 2 Foundation for the Development of Medical and Biological Sciences (Fundemeb), Colombia Background: Allergens of the domestic mites D. pteronyssinnus and B. tropicalis are the main risk factor for asthma and allergic diseases in atopic individuals in tropical and subtropical regions of the word. Allergen-specific immunotherapy (AIT) with natural allergen extracts is the only treatment able to change the natural course of allergic diseases in most patients that follow the protocol of vaccination. In the last years recombinant allergens with different modifications have showed promising immunological properties for the development of vaccines for a more effective and safe AIT. Methods: Using an “in silico” approach we designed a single molecule containing epitopes of allergens from B. tropicalis (Blo t 5, Blo t 8 and Blo t 10) and D. pteronyssinus (Der p 1, Der p 2 and Der p 8), which we denominated MAVAC-BD-2.The nucleotide sequence encoding for the designed molecule was synthetized and cloned into an expression vector for expression in Escherichia coli. The recombinant protein was obtained in liquid culture induced by IPTG and purified by affinity chromatography. The IgE and IgG reactivity of MAVAC-BD-2 was determined by ELISA in 100 sera from mite allergic patients and 100 sera from non-allergic individuals. Basophil activation test was assessed in 9 mite-allergic patients by upregulation of CD203c expression as detected by flow cytometry. Basophils were stimulated with MAVAC-BD-2, mite extracts and the recombinants Blo t 5 and Der p 2 at concentration of 10 ug/ml. ELISA inhibition assays were performed using pool samples from allergic patients sensitized to domestic mites. Signed informed consent was obtained for all the procedures. Results: The protein was obtained in the induced culture in inclusion bodies; after treatment with denaturing buffer and refolding process the antibody reactivity was demonstrated by ELISA. Frequency of IgE binding to MAVAC-BD-2 in sera from mite allergy patients sensitized to B. tropicalis and D. pteronyssinus was 49%. IgE binding was also detected in 15% of sera from non-allergic individuals. IgG reactivity has a mean optical density of 1.3 in the allergic group vs. 1.09 in the non-allergic group (p <0.03). Sera from mite allergic subjects showed a lower IgE antibody reactivity to MAVAC-BD-2 compared to IgE antibody reactivity to B. tropicalis extract (p < 0.001), and to D. pteronyssinus (p < 0.001).The basophil activation test with the hybrid protein was significantly lower than those obtained with mite whole extract (p < 0.001) and single recombinant allergens Der p 2 and Blo t 5 (p < 0.001). By adsorption of the sera pool with MAVAC-BD-2, the IgG reactivity against the B. tropicalis and D. pteronyssinus whole extracts was inhibited by 41% and 30% respectively, indicating that some epitopes from of native allergen are represented in the hybrid molecule. Conclusion: A hybrid protein containing epitopes from two mite species was obtained. The lower IgE reactivity and reduce basophil activation compared to those induced by whole mite extract and purified allergens suggest that this molecule could be useful for the development of a mite allergen vaccine. Acknowledgements This project was founded by Colciencias and University of Cartagena, Colombia. (Grant 588-2013). Keywords: Mite allergy, Recombinant Proteins, IgE, Immunotherapy, Blomia tropicalis, Dermatophagoides pteronyssinus Conference: IMMUNOCOLOMBIA2015 - 11th Congress of the Latin American Association of Immunology - 10o. Congreso de la Asociación Colombiana de Alergia, Asma e Inmunología, Medellin, Colombia, 13 Oct - 16 Oct, 2015. Presentation Type: Poster Presentation Topic: Immunotherapy Citation: Martínez D, Múnera M, Cantillo J, Caraballo L and Puerta L (2015). A hybrid protein designed with epitopes from B. tropicalis and D. pteronyssinus allergens showed reduced IgE binding and allergenic activity. Front. Immunol. Conference Abstract: IMMUNOCOLOMBIA2015 - 11th Congress of the Latin American Association of Immunology - 10o. Congreso de la Asociación Colombiana de Alergia, Asma e Inmunología. doi: 10.3389/conf.fimmu.2015.05.00078 Copyright: The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers. They are made available through the Frontiers publishing platform as a service to conference organizers and presenters. The copyright in the individual abstracts is owned by the author of each abstract or his/her employer unless otherwise stated. Each abstract, as well as the collection of abstracts, are published under a Creative Commons CC-BY 4.0 (attribution) licence (https://creativecommons.org/licenses/by/4.0/) and may thus be reproduced, translated, adapted and be the subject of derivative works provided the authors and Frontiers are attributed. For Frontiers’ terms and conditions please see https://www.frontiersin.org/legal/terms-and-conditions. Received: 27 Apr 2015; Published Online: 14 Sep 2015. * Correspondence: Prof. Leonardo Puerta, University of Cartagena, Institute for Immunological Research, Cartagena, Bolivar, 0012, Colombia, lpuertal1@unicartagena.edu.co Login Required This action requires you to be registered with Frontiers and logged in. To register or login click here. Abstract Info Abstract The Authors in Frontiers Dalgys Martínez Marlon Múnera Jose Cantillo Luis Caraballo Leonardo Puerta Google Dalgys Martínez Marlon Múnera Jose Cantillo Luis Caraballo Leonardo Puerta Google Scholar Dalgys Martínez Marlon Múnera Jose Cantillo Luis Caraballo Leonardo Puerta PubMed Dalgys Martínez Marlon Múnera Jose Cantillo Luis Caraballo Leonardo Puerta Related Article in Frontiers Google Scholar PubMed Abstract Close Back to top Javascript is disabled. Please enable Javascript in your browser settings in order to see all the content on this page.