title: Masked selection: a straightforward and flexible approach for the selection of binders against specific epitopes and differentially expressed proteins by phage display.
abstract: Phage display is a well-established procedure to isolate binders against a wide variety of antigens that can be performed on purified antigens, but also on intact cells. As selection steps are performed in vitro, it is possible to focus the outcome of the selection on relevant epitopes by performing some additional steps, such as depletion or competitive elutions. However in practice, the efficiency of these steps is often limited and can lead to inconsistent results. We have designed a new selection method named masked selection, based on the blockade of unwanted epitopes to favor the targeting of relevant ones. We demonstrate the efficiency and flexibility of this method by selecting single-domain antibodies against a specific portion of a fusion protein, by selecting binders against several members of the seven transmembrane receptor family using transfected HEK cells, or by selecting binders against unknown breast cancer markers not expressed on normal samples. The relevance of this approach for antibody-based therapies was further validated by the identification of four of these markers, Epithelial cell adhesion molecule, Transferrin receptor 1, Metastasis cell adhesion molecule, and Sushi containing domain 2, using immunoprecipitation and mass spectrometry. This new phage display strategy can be applied to any type of antibody fragments or alternative scaffolds, and is especially suited for the rapid discovery and identification of cell surface markers.
authors.affiliation: Inserm,U1068, CRCM, Marseille, F-13009, France;
authors.forename: Klervi
authors.initials: K
authors.lastname: Even-Desrumeaux
authors.forename: Damien
authors.initials: D
authors.lastname: Nevoltris
authors.forename: Marie Noelle
authors.initials: MN
authors.lastname: Lavaut
authors.forename: Karima
authors.initials: K
authors.lastname: Alim
authors.forename: Jean-Paul
authors.initials: JP
authors.lastname: Borg
authors.forename: Stéphane
authors.initials: S
authors.lastname: Audebert
authors.forename: Brigitte
authors.initials: B
authors.lastname: Kerfelec
authors.forename: Daniel
authors.initials: D
authors.lastname: Baty
authors.forename: Patrick
authors.initials: P
authors.lastname: Chames
mesh_terms: D000941:Antigens / Q000276:immunology / Q000378:metabolism; D001943:Breast Neoplasms / Q000276:immunology / Q000378:metabolism; D045744:Cell Line, Tumor; D061505:Cell Surface Display Techniques; D018604:Epitope Mapping / Q000379:methods*; D000939:Epitopes / Q000378:metabolism*; D005260:Female; D057809:HEK293 Cells; D019073:HT29 Cells; D006367:HeLa Cells; D006801:Humans; D047468:Immunoprecipitation; D019169:Jurkat Cells; D061986:MCF-7 Cells; D011485:Protein Binding; D011506:Proteins / Q000276:immunology / Q000378:metabolism*; D040901:Proteomics / Q000379:methods
chemical_list: D000941:Antigens; D000939:Epitopes; D011506:Proteins