title: A rapid screening and production method using a novel mammalian cell display to isolate human monoclonal antibodies.
abstract: Antibody display methods are increasingly being used to produce human monoclonal antibodies for disease therapy. Rapid screening and isolation of specific human antibody genes are valuable for producing human monoclonal antibodies showing high specificity and affinity. In this report, we describe a novel mammalian cell display method in which whole human IgG is displayed on the cell surface of CHO cells. Cells expressing antigen-specific human monoclonal IgGs with high affinity on the cell surface after normal folding and posttranscriptional modification were screened using a cell sorter. The membrane-type IgG-expressing CHO cells were then converted to IgG-secreting cells by transfection with a plasmid coding Cre recombinase. This mammalian cell display method was applied to in vitro affinity maturation of monoclonal C9 IgG specific to the human high-affinity IgE receptor (FcεRIα). The CDR3 of the C9 heavy chain variable region gene was randomly mutated and inserted into pcDNA5FRT/IgG. A C9 IgG (CDRH3r)-expressing CHO cell display library consisting of 1.1×10(6) independent clones was constructed. IgG-displaying cells showing high reactivity to FcεRIα antigen were screened by the cell sorter, resulting in the establishment of a CHO cell line producing with higher reactivity than the parent C9 IgG.
authors.affiliation: Department of Bioscience and Biotechnology, Faculty of Agriculture, Kyushu University, 6-10-1 Hakozaki, Higashi-ku, Fukuoka 812-8581, Japan; Japan Society for the Promotion of Science, 8 Ichibancho, Chiyoda-ku, Tokyo 102-8472, Japan.
authors.forename: Kosuke
authors.initials: K
authors.lastname: Tomimatsu
authors.forename: Shin-ei
authors.initials: SE
authors.lastname: Matsumoto
authors.forename: Hayato
authors.initials: H
authors.lastname: Tanaka
authors.forename: Makiko
authors.initials: M
authors.lastname: Yamashita
authors.forename: Hidekazu
authors.initials: H
authors.lastname: Nakanishi
authors.forename: Kiichiro
authors.initials: K
authors.lastname: Teruya
authors.forename: Saiko
authors.initials: S
authors.lastname: Kazuno
authors.forename: Tomoya
authors.initials: T
authors.lastname: Kinjo
authors.forename: Takeki
authors.initials: T
authors.lastname: Hamasaki
authors.forename: Ken-ichi
authors.initials: K
authors.lastname: Kusumoto
authors.forename: Shigeru
authors.initials: S
authors.lastname: Kabayama
authors.forename: Yoshinori
authors.initials: Y
authors.lastname: Katakura
authors.forename: Sanetaka
authors.initials: S
authors.lastname: Shirahata
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis* / Q000302:isolation & purification*; D000921:Antibody-Producing Cells / Q000378:metabolism; D016466:CHO Cells; D002462:Cell Membrane / Q000378:metabolism; D061505:Cell Surface Display Techniques / Q000379:methods*; D006224:Cricetinae; D003412:Cricetulus; D005785:Gene Conversion; D006801:Humans; D007074:Immunoglobulin G / Q000378:metabolism; D008969:Molecular Sequence Data; D019151:Peptide Library; D017455:Receptors, IgE / Q000737:chemistry / Q000378:metabolism; D011995:Recombination, Genetic / Q000235:genetics; D019076:Transgenes
chemical_list: D000911:Antibodies, Monoclonal; C543355:FcepsilonRI alpha-chain, human; D007074:Immunoglobulin G; D019151:Peptide Library; D017455:Receptors, IgE
keywords: CHO cells; Cre recombinase; Human high affinity IgE receptor; Human monoclonal antibody; In vitro maturation; Mammalian cell display