title: In silico designing, cloning, and heterologous expression of novel chimeric human B lymphocyte CD20 extra loop.
abstract: Design and production of monoclonal antibody for the diagnosis and immunotherapy of non-Hodgkin lymphoma require a suitable CD20 antigen as an effective immunogen. In this study, a new chimeric human CD20 extra loop (hCD20EXL) protein was designed by bioinformatics tools and was expressed in Escherichia coli BL21 DE3. Amino acid sequences, protein structure, immunogenicity, and other physicochemical property of potential antigens were in silico analyzed. Antigenicity, codon optimization, and other predictions of designed protein were determined by bioinformatics tools. The designed protein was heterologously expressed in E. coli and verified by SDS-PAGE and Western blot. Immunogenicity of this antigen was tested in mice, and reactivity of the antibodies was evaluated using flow cytometry. Experimental analysis confirmed the in silico prediction of the designed chimeric hCD20 in this study. Therefore, based on these results, it is suggested that the new chimeric hCD20 antigen could be an appropriate immunogen for production of monoclonal antibody in immunotherapy purposes.
authors.affiliation: Molecular biology Research Center, Baqiyatallah University of Medical Sciences, P.O. Box: 19395-5478, Tehran, Iran.
authors.forename: Mahdi
authors.initials: M
authors.lastname: Fasihi-Ramandi
authors.affiliation: Applied Microbiology Research Center, Baqiyatallah University of Medical Sciences, Tehran, Iran.
authors.forename: Jafar
authors.initials: J
authors.lastname: Amani
authors.affiliation: Department of Agricultural Biotechnology, National Institute of Genetic Engineering and Biotechnology (NIGEB), Tehran, Iran.
authors.forename: Ali-Hatef
authors.initials: AH
authors.lastname: Salmanian
authors.affiliation: Immunology Department, Tarbiat Modares University, Tehran, Iran.
authors.forename: Seyed Mohammad
authors.initials: SM
authors.lastname: Moazzeni
authors.affiliation: Molecular biology Research Center, Baqiyatallah University of Medical Sciences, P.O. Box: 19395-5478, Tehran, Iran. kazahmadi@yahoo.com.
authors.forename: Kazem
authors.initials: K
authors.lastname: Ahmadi
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis* / Q000276:immunology; D018951:Antigens, CD20 / Q000276:immunology*; D001402:B-Lymphocytes / Q000276:immunology*; D003001:Cloning, Molecular; D019295:Computational Biology; D004926:Escherichia coli / Q000235:genetics; D005434:Flow Cytometry; D006801:Humans; D051379:Mice; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000276:immunology
chemical_list: D000911:Antibodies, Monoclonal; D018951:Antigens, CD20; D011993:Recombinant Fusion Proteins
keywords: Bioinformatics; CD20; Heterologous expression; Immunotherapy; Lymphoma