Human acidic mammalian chitinase (AMCase); molecular cloning, protein expression, and production of monoclonal antibody

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Event Abstract Back to Event Human acidic mammalian chitinase (AMCase); molecular cloning, protein expression, and production of monoclonal antibody Kanokwan Lowhalidanon1, Piyanat Meekrathok1, Sunisa Thongsom1, Wipa Suginta1 and Panida Khunkaewka1* 1 Suranaree University of Technology, Biochemistry-Electrochemistry Research Unit, Thailand Human acidic mammalian chitinase (AMCase), is a 50 kDa protein containing a 30 kDa N-terminal catalytic domain that can hydrolyze chitin. This enzyme belongs to family 18 glycosyl hydrolases. High abundant of this enzyme was found in gastrointestinal tract. In lung, interleukin-13 was proposed to play a key role as primary effector that induces secretion of AMCase by airway epithelial cells and macrophages. The secreted AMCase activates elevation of monocyte chemoattractant protein 1 (MCP-1) and eotaxin-1, which induce the recruitment of eosinophils, T cells, and macrophages to site of inflammation in lung epithelial cells. Nonetheless, the mechanisms underlying AMCase mediated cells recruitment and its surface receptor have not been identified. Present study, the gene encoding AMCase was isolated from the human cDNA, cloned into pQE-tri vector, and the recombinant protein was expressed in E. coli M15. The affinity-purified AMCase was further used as immunogen for monoclonal antibody (mAb) production by standard hybridroma technique. Two specific mAbs against AMCase were successfully raised and named as 4G1 and 6E5. Both monoclones belong to IgG1 isotype. Western blot analysis showed that the mAbs strongly reacted with a corresponding protein with molecular weight of about 50 kDa in human acute monocytic leukemia cell line, THP1, and peripheral blood mononuclear cells. In conclusion, the AMCase was cloned and expressed in E. Coli system. Two specific mAbs to AMCase were raised and specifically reacted to human AMCase. The generated mAbs will be used for further functional analysis of the AMCase in regulation of the immune system. Acknowledgements This work was supported by the National Research Council of Thailand and Suranaree University of Technology, Thailand. References Louise E. Donnelly and Peter J. Barnes. (2004). Acidic mammalian chitinase – a potential target for asthma therapy. TRENDS in Pharmacological Science Vol.25. Keywords: AMCase, Molecular cloning, protein expression, Monoclonal antibody, Asthma Conference: 15th International Congress of Immunology (ICI), Milan, Italy, 22 Aug - 27 Aug, 2013. Presentation Type: Abstract Topic: Immune-mediated disease pathogenesis Citation: Lowhalidanon K, Meekrathok P, Thongsom S, Suginta W and Khunkaewka P (2013). Human acidic mammalian chitinase (AMCase); molecular cloning, protein expression, and production of monoclonal antibody. Front. Immunol. Conference Abstract: 15th International Congress of Immunology (ICI). doi: 10.3389/conf.fimmu.2013.02.00189 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: 11 Mar 2013; Published Online: 22 Aug 2013. * Correspondence: Dr. Panida Khunkaewka, Suranaree University of Technology, Biochemistry-Electrochemistry Research Unit, Nakhonratchasima, 30000, Thailand, kpanida@sut.ac.th 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 Kanokwan Lowhalidanon Piyanat Meekrathok Sunisa Thongsom Wipa Suginta Panida Khunkaewka Google Kanokwan Lowhalidanon Piyanat Meekrathok Sunisa Thongsom Wipa Suginta Panida Khunkaewka Google Scholar Kanokwan Lowhalidanon Piyanat Meekrathok Sunisa Thongsom Wipa Suginta Panida Khunkaewka PubMed Kanokwan Lowhalidanon Piyanat Meekrathok Sunisa Thongsom Wipa Suginta Panida Khunkaewka 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.