title: Phage display revisited: Epitope mapping of a monoclonal antibody directed against Neisseria meningitidis adhesin A using the PROFILER technology.
abstract: There is a strong need for rapid and reliable epitope mapping methods that can keep pace with the isolation of increasingly larger numbers of mAbs. We describe here the identification of a conformational epitope using Phage-based Representation OF ImmunoLigand Epitope Repertoire (PROFILER), a recently developed high-throughput method based on deep sequencing of antigen-specific lambda phage-displayed libraries. A novel bactericidal monoclonal antibody (mAb 9F11) raised against Neisseria meningitidis adhesin A (NadA), an important component of the Bexsero(®) anti-meningococcal vaccine, was used to evaluate the technique in comparison with other epitope mapping methods. The PROFILER technology readily identified NadA fragments that were capable of fully recapitulating the reactivity of the entire antigen against mAb 9F11. Further analysis of these fragments using mutagenesis and hydrogen-deuterium exchange mass-spectrometry allowed us to identify the binding site of mAb 9F11 (A250-D274) and an adjoining sequence (V275-H312) that was also required for the full functional reconstitution of the epitope. These data suggest that, by virtue of its ability to detect a great variety of immunoreactive antigen fragments in phage-displayed libraries, the PROFILER technology can rapidly and reliably identify epitope-containing regions and provide, in addition, useful clues for the functional characterization of conformational mAb epitopes.
authors.affiliation: a Scylla Biotech Srl , Messina , Italy.
authors.forename: Veronica Lanza
authors.initials: VL
authors.lastname: Cariccio
authors.affiliation: a Scylla Biotech Srl , Messina , Italy.
authors.forename: Maria
authors.initials: M
authors.lastname: Domina
authors.affiliation: b Department of Human Pathology , University of Messina , Messina , Italy.
authors.forename: Salvatore
authors.initials: S
authors.lastname: Benfatto
authors.affiliation: c Department Clinical and Experimental Medicine , University of Messina , Messina , Italy.
authors.forename: Mario
authors.initials: M
authors.lastname: Venza
authors.affiliation: c Department Clinical and Experimental Medicine , University of Messina , Messina , Italy.
authors.forename: Isabella
authors.initials: I
authors.lastname: Venza
authors.affiliation: d GSK Vaccines , Siena , Italy.
authors.forename: Agnese
authors.initials: A
authors.lastname: Faleri
authors.affiliation: d GSK Vaccines , Siena , Italy.|e Department of Life Sciences , University of Siena , Siena , Italy.
authors.forename: Marco
authors.initials: M
authors.lastname: Bruttini
authors.affiliation: d GSK Vaccines , Siena , Italy.
authors.forename: Erika
authors.initials: E
authors.lastname: Bartolini
authors.affiliation: d GSK Vaccines , Siena , Italy.
authors.forename: Marzia Monica
authors.initials: MM
authors.lastname: Giuliani
authors.affiliation: d GSK Vaccines , Siena , Italy.
authors.forename: Laura
authors.initials: L
authors.lastname: Santini
authors.affiliation: d GSK Vaccines , Siena , Italy.
authors.forename: Brunella
authors.initials: B
authors.lastname: Brunelli
authors.affiliation: d GSK Vaccines , Siena , Italy.
authors.forename: Nathalie
authors.initials: N
authors.lastname: Norais
authors.affiliation: d GSK Vaccines , Siena , Italy.
authors.forename: Erica
authors.initials: E
authors.lastname: Borgogni
authors.affiliation: b Department of Human Pathology , University of Messina , Messina , Italy.
authors.forename: Angelina
authors.initials: A
authors.lastname: Midiri
authors.affiliation: b Department of Human Pathology , University of Messina , Messina , Italy.
authors.forename: Roberta
authors.initials: R
authors.lastname: Galbo
authors.affiliation: b Department of Human Pathology , University of Messina , Messina , Italy.
authors.forename: Letizia
authors.initials: L
authors.lastname: Romeo
authors.affiliation: b Department of Human Pathology , University of Messina , Messina , Italy.
authors.forename: Carmelo
authors.initials: C
authors.lastname: Biondo
authors.affiliation: d GSK Vaccines , Siena , Italy.
authors.forename: Vega
authors.initials: V
authors.lastname: Masignani
authors.affiliation: c Department Clinical and Experimental Medicine , University of Messina , Messina , Italy.|f Charybdis Vaccines Srl , Messina , Italy.
authors.forename: Giuseppe
authors.initials: G
authors.lastname: Teti
authors.affiliation: g Department of Biosciences and Territory , University of Molise , Pesche , Isernia , Italy.
authors.forename: Franco
authors.initials: F
authors.lastname: Felici
authors.affiliation: a Scylla Biotech Srl , Messina , Italy.|b Department of Human Pathology , University of Messina , Messina , Italy.
authors.forename: Concetta
authors.initials: C
authors.lastname: Beninati
mesh_terms: D018829:Adhesins, Bacterial / Q000276:immunology*; D000818:Animals; D000907:Antibodies, Bacterial / Q000276:immunology*; D000911:Antibodies, Monoclonal / Q000276:immunology*; D001428:Bacterial Vaccines / Q000276:immunology; D061505:Cell Surface Display Techniques / Q000379:methods*; D018604:Epitope Mapping / Q000379:methods*; D022401:Meningococcal Vaccines; D009345:Neisseria meningitidis / Q000276:immunology*; D010446:Peptide Fragments / Q000276:immunology
chemical_list: D018829:Adhesins, Bacterial; D000907:Antibodies, Bacterial; D000911:Antibodies, Monoclonal; D001428:Bacterial Vaccines; D022401:Meningococcal Vaccines; D010446:Peptide Fragments
keywords: Deep sequencing Phage-based Representation OF ImmunoLigand Epitope Repertoire (PROFILER); NadA; Neisseria meningitidis; epitope mapping; phage display; vaccines