title: Recombinant production and characterization of human anti-influenza virus monoclonal antibodies identified from hybridomas fused with human lymphocytes.
abstract: In previous studies, hybridomas producing human immunoglobulin G, the antibodies 5E4 and 5A7 against influenza A and B virus were established using a novel human lymphocyte fusion partner, SPYMEG. In the present study, we succeeded in achieving the recombinant production and secretion of 5E4 and 5A7 in Chinese hamster ovary cells. Our N-glycan analysis by intact-mass detection and liquid chromatography mass spectrometry showed that recombinant 5E4 and 5A7 have one N-glycan and the typical mammalian-type N-glycan structures similar to those in hybridomas. However, the glycan distribution was slightly different among these antibodies. The amount of high-mannose-type structures was under 10% of the total N-glycans of recombinant 5E4 and 5A7, compared to 20% of the 5E4 and 5A7 produced in hybridomas. The amount of galactosylated N-glycans was increased in recombinants. Approximately 80% of the N-glycans of all antibodies was fucosylated, and no sialylated N-glycan was found. Recombinant 5E4 and 5A7 neutralized pandemic influenza A virus specifically, and influenza B virus broadly, quite similar to the 5E4 and 5A7 produced in hybridomas, respectively. Here we demonstrated that recombinants of antibodies identified from hybridomas fused with SPYMEG have normal N-glycans and that their neutralizing activities bear comparison with those of the original antibodies.
authors.affiliation: International Center for Biotechnology, Osaka University, Yamada-oka 2-1, Suita, Osaka 565-0871, Japan; The Japan Science and Technology Agency/Japan International Cooperation Agency, Science and Technology Research Partnership for Sustainable Development, Tokyo, Japan.
authors.forename: Ryo
authors.initials: R
authors.lastname: Misaki
authors.affiliation: International Center for Biotechnology, Osaka University, Yamada-oka 2-1, Suita, Osaka 565-0871, Japan.
authors.forename: Natsuko
authors.initials: N
authors.lastname: Fukura
authors.affiliation: International Center for Biotechnology, Osaka University, Yamada-oka 2-1, Suita, Osaka 565-0871, Japan.
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Kajiura
authors.affiliation: The Japan Science and Technology Agency/Japan International Cooperation Agency, Science and Technology Research Partnership for Sustainable Development, Tokyo, Japan; Department of Virology, Research Institute for Microbial Diseases, Osaka University, Yamada-oka 3-1, Suita, Osaka 565-0871, Japan; Graduate School of Life and Environmental Sciences, Osaka Prefecture University, Izumisano, Osaka 598-8531, Japan.
authors.forename: Mayo
authors.initials: M
authors.lastname: Yasugi
authors.affiliation: The Japan Science and Technology Agency/Japan International Cooperation Agency, Science and Technology Research Partnership for Sustainable Development, Tokyo, Japan; Department of Virology, Research Institute for Microbial Diseases, Osaka University, Yamada-oka 3-1, Suita, Osaka 565-0871, Japan; Kanonji Institute, The Research Foundation for Microbial Diseases of Osaka University, Kanonji, Kagawa, Japan.
authors.forename: Ritsuko
authors.initials: R
authors.lastname: Kubota-Koketsu
authors.affiliation: The Japan Science and Technology Agency/Japan International Cooperation Agency, Science and Technology Research Partnership for Sustainable Development, Tokyo, Japan; Department of Virology, Research Institute for Microbial Diseases, Osaka University, Yamada-oka 3-1, Suita, Osaka 565-0871, Japan.
authors.forename: Tadahiro
authors.initials: T
authors.lastname: Sasaki
authors.affiliation: The Japan Science and Technology Agency/Japan International Cooperation Agency, Science and Technology Research Partnership for Sustainable Development, Tokyo, Japan; Ina Laboratory, Medical & Biological Laboratories Corporation, Ltd., Ina, Nagano, Japan.
authors.forename: Masatoshi
authors.initials: M
authors.lastname: Momota
authors.affiliation: The Japan Science and Technology Agency/Japan International Cooperation Agency, Science and Technology Research Partnership for Sustainable Development, Tokyo, Japan; Medical & Biological Laboratories Corporation, Ltd., Nagoya, Aichi, Japan.
authors.forename: Ken-Ichiro
authors.initials: K
authors.lastname: Ono
authors.affiliation: International Center for Biotechnology, Osaka University, Yamada-oka 2-1, Suita, Osaka 565-0871, Japan.
authors.forename: Takao
authors.initials: T
authors.lastname: Ohashi
authors.affiliation: The Japan Science and Technology Agency/Japan International Cooperation Agency, Science and Technology Research Partnership for Sustainable Development, Tokyo, Japan; Department of Virology, Research Institute for Microbial Diseases, Osaka University, Yamada-oka 3-1, Suita, Osaka 565-0871, Japan.
authors.forename: Kazuyoshi
authors.initials: K
authors.lastname: Ikuta
authors.affiliation: International Center for Biotechnology, Osaka University, Yamada-oka 2-1, Suita, Osaka 565-0871, Japan; The Japan Science and Technology Agency/Japan International Cooperation Agency, Science and Technology Research Partnership for Sustainable Development, Tokyo, Japan. Electronic address: fujiyama@icb.osaka-u.ac.jp.
authors.forename: Kazuhito
authors.initials: K
authors.lastname: Fujiyama
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis / Q000276:immunology*; D057134:Antibodies, Neutralizing / Q000096:biosynthesis / Q000276:immunology*; D000914:Antibodies, Viral / Q000097:blood / Q000276:immunology*; D002459:Cell Fusion / Q000379:methods; D006224:Cricetinae; D006801:Humans; D006825:Hybridomas / Q000276:immunology / Q000378:metabolism; D007074:Immunoglobulin G / Q000096:biosynthesis / Q000276:immunology*; D009980:Influenza A virus / Q000276:immunology*; D009981:Influenza B virus / Q000276:immunology*; D008214:Lymphocytes / Q000276:immunology / Q000378:metabolism
chemical_list: D000911:Antibodies, Monoclonal; D057134:Antibodies, Neutralizing; D000914:Antibodies, Viral; D007074:Immunoglobulin G
keywords: Chinese hamster ovary; Glycosylation; Human antibody production; SPYMEG