title: Phage display for site-specific immunization and characterization of high-risk human papillomavirus specific E7 monoclonal antibodies.
abstract: The paper describes a method to use filamentous phage to display specific regions of proteins for immunization in order to direct the immune response towards a pre-defined region of the protein. The method called site-specific immunization (SSI) was evaluated using the E7 protein of oncogenic (high-risk) human papillomavirus (HPV) type 16 as a model system. This protein consists of sequence blocks also present in other viral and cellular proteins and in the corresponding protein of low-risk HPVs. A fragment of the HPV16 E7 oncoprotein specific for a group of high-risk viruses was identified by sequence comparison and displayed on filamentous phages in fusion with the major phage coat protein VIII. The recombinant phages triggered an immune response in mice against the full-length HPV16 E7 protein. Fusion of B-lymphocytes from the immunized animals with myeloma cells resulted in three hybridomas producing monoclonal antibodies (MAbs) with reactivity against the endogenous E7 protein. The specificity of the MAbs for the HPV16 E7 protein in cancer cell lines was confirmed by Western blot analyses and immunocytochemistry. The epitope of each MAb was roughly mapped by determining the reactivity against overlapping E7 fragments displayed on phage particles. The mimotopes of the MAbs were further determined by biopanning against a randomized peptide library displayed on phage and found to be unique for a sub-set of high-risk HPV E7 proteins. The combination of different phage display techniques for immunization and epitope mapping was efficient for generation and characterization of highly specific MAbs.
authors.affiliation: Fujirebio Diagnostics AB, Majnabbe Terminal, SE-414 55 Gothenburg, Sweden.
authors.forename: Maria
authors.initials: M
authors.lastname: Lidqvist
authors.forename: Olle
authors.initials: O
authors.lastname: Nilsson
authors.forename: Jan
authors.initials: J
authors.lastname: Holmgren
authors.forename: Christina
authors.initials: C
authors.lastname: Hall
authors.forename: Christian
authors.initials: C
authors.lastname: Fermér
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000918:Antibody Specificity; D018604:Epitope Mapping; D005260:Female; D007114:Immunization; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D009856:Oncogene Proteins, Viral / Q000276:immunology*; D050725:Papillomavirus E7 Proteins; D053918:Papillomavirus Vaccines / Q000276:immunology*; D019151:Peptide Library
chemical_list: D000911:Antibodies, Monoclonal; D009856:Oncogene Proteins, Viral; D050725:Papillomavirus E7 Proteins; D053918:Papillomavirus Vaccines; D019151:Peptide Library; C059731:oncogene protein E7, Human papillomavirus type 16