title: Identification of target and function specific antibodies for effective drug delivery.
abstract: Phage antibody technology is a powerful approach for generating human antibodies to virtually any target antigen. For many therapeutic applications, it is useful to generate antibodies that bind to cell-surface receptors in a manner where binding results in internalization of the antibody. This allows use of the antibody to deliver toxic payloads intracellularly to achieve a therapeutic effect. Here we describe how phage antibody libraries can be directly selected on tumor cell lines to generate antibodies binding cell-surface receptors and which are rapidly internalized upon binding. Protocols are provided showing how to (1) directly select internalizing antibodies from phage antibody libraries; (2) screen phage antibodies in a high-throughput flow cytometry assay for binding to the tumor cell line used for selection; (3) identify the antigen bound by the phage antibody using immunoprecipitation and mass spectrometry; and (4) verify and quantitate such that phage antibodies are internalized.
authors.affiliation: University of California, San Francisco, CA, USA.
authors.forename: Yu
authors.initials: Y
authors.lastname: Zhou
authors.forename: James D
authors.initials: JD
authors.lastname: Marks
mesh_terms: D000912:Antibodies, Neoplasm / Q000276:immunology*; D000918:Antibody Specificity; D000951:Antigens, Neoplasm / Q000737:chemistry / Q000276:immunology; D001681:Biological Assay; D045744:Cell Line, Tumor; D016503:Drug Delivery Systems / Q000379:methods*; D004705:Endocytosis; D006801:Humans; D007135:Immunoglobulin Variable Region / Q000276:immunology; D013058:Mass Spectrometry; D019151:Peptide Library
chemical_list: D000912:Antibodies, Neoplasm; D000951:Antigens, Neoplasm; D007135:Immunoglobulin Variable Region; D019151:Peptide Library