title: Monoclonal antibody binding-site diversity assessment with a cell-based clustering assay.
abstract: The diversity of a panel of antibodies that target a specific antigen can be established in various assay formats. In conventional epitope binning assays purified antibodies are tested in a pairwise manner: two antibodies that compete with each other for binding to an antigen are grouped into the same cluster or bin, while they are assigned to two different clusters when they do not compete. Here we present a high through put assay that enables grouping of crude hybridoma supernatants without a need for antibody purification. In addition, the assay does not require recombinant protein, because it is conducted on cells that express the antigen of interest. Hence, one can use the antibody-clustering assay for cell surface proteins that are not amenable to purification. Heavy chain variable region (VH) sequencing shows that VH composition within clusters is conserved. Finally, the assay is in good agreement with a conventional epitope binning assay with purified antigen.
authors.affiliation: Igenica Inc., 863 Mitten Road Suite 102, Burlingame, CA 94010, USA. Electronic address: sliaochan@igenica.com.
authors.forename: Sindy
authors.initials: S
authors.lastname: Liao-Chan
authors.affiliation: Igenica Inc., 863 Mitten Road Suite 102, Burlingame, CA 94010, USA. Electronic address: jzachwieja@igenica.com.
authors.forename: Joseph
authors.initials: J
authors.lastname: Zachwieja
authors.affiliation: Igenica Inc., 863 Mitten Road Suite 102, Burlingame, CA 94010, USA. Electronic address: sgomez@igenica.com.
authors.forename: Steven
authors.initials: S
authors.lastname: Gomez
authors.affiliation: Igenica Inc., 863 Mitten Road Suite 102, Burlingame, CA 94010, USA. Electronic address: dana.duey@gmail.com.
authors.forename: Dana
authors.initials: D
authors.lastname: Duey
authors.affiliation: Igenica Inc., 863 Mitten Road Suite 102, Burlingame, CA 94010, USA. Electronic address: lippy1@gmail.com.
authors.forename: John
authors.initials: J
authors.lastname: Lippincott
authors.affiliation: Igenica Inc., 863 Mitten Road Suite 102, Burlingame, CA 94010, USA. Electronic address: jtheunissen@igenica.com.
authors.forename: Jan-Willem
authors.initials: JW
authors.lastname: Theunissen
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000145:classification / Q000276:immunology*; D000941:Antigens / Q000276:immunology; D001666:Binding Sites, Antibody / Q000276:immunology*; D045744:Cell Line, Tumor; D016000:Cluster Analysis; D017077:Culture Media, Conditioned / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes / Q000145:classification / Q000276:immunology*; D006801:Humans; D006825:Hybridomas / Q000276:immunology / Q000378:metabolism; D007074:Immunoglobulin G / Q000276:immunology / Q000378:metabolism; D007135:Immunoglobulin Variable Region / Q000145:classification / Q000276:immunology; D051379:Mice; D057507:Mice, 129 Strain
chemical_list: D000911:Antibodies, Monoclonal; D000941:Antigens; D017077:Culture Media, Conditioned; D000939:Epitopes; D007074:Immunoglobulin G; D007135:Immunoglobulin Variable Region
keywords: Antibody sequencing; Biosensor; Clustering assay; Engineered cell lines; Epitope binning; Monoclonal antibody