title: Human autoimmune anti-proteinase 3 scFv from a phage display library.
abstract: This is the first study describing recombinant human antibody fragments directed to the autoantigen proteinase 3 (PR3) from an immune B cell source. Detection of these autoantibodies has proven valid for the diagnosis and monitoring of Wegener's granulomatosis. The described antibody fragment (scFv) was isolated from a phage display library prepared from the IgG-positive splenic lymphocytes of a patient with systemic autoimmunity. The cloning strategy was designed to maintain the diversity of the antibody variable gene repertoire, and sequencing of several variable genes demonstrated that all major heavy and light chain families were represented. We found an over-representation of particular heavy chain variable domains in splenic lymphocytes which differ from the ones frequently found in peripheral blood lymphocytes. It was possible to obtain specific scFv to PR3 after a single round of selection and the binding could be inhibited by the patients' sera. Although the antibody fragments in the splenic repertoire were found to be highly mutated, it was interesting to find that the selected scFv showed only limited somatic mutation. Furthermore, we could demonstrate that the removal of the mutations had no effect on binding specificity.
authors.affiliation: Department of Transfusion Medicine, University of Cambridge, UK.
authors.forename: R
authors.initials: R
authors.lastname: Finnern
authors.forename: E
authors.initials: E
authors.lastname: Pedrollo
authors.forename: I
authors.initials: I
authors.lastname: Fisch
authors.forename: J
authors.initials: J
authors.lastname: Wieslander
authors.forename: J D
authors.initials: JD
authors.lastname: Marks
authors.forename: C M
authors.initials: CM
authors.lastname: Lockwood
authors.forename: W H
authors.initials: WH
authors.lastname: Ouwehand
mesh_terms: D000595:Amino Acid Sequence; D001324:Autoantigens / Q000276:immunology*; D015551:Autoimmunity / Q000235:genetics; D001435:Bacteriophages / Q000235:genetics*; D001483:Base Sequence; D015723:Gene Library; D005803:Genes, Immunoglobulin; D014890:Granulomatosis with Polyangiitis / Q000276:immunology; D006801:Humans; D007128:Immunoglobulin Fragments / Q000032:analysis*; D007143:Immunoglobulin Heavy Chains / Q000235:genetics; D007135:Immunoglobulin Variable Region / Q000235:genetics; D008297:Male; D008875:Middle Aged; D008969:Molecular Sequence Data; D053146:Myeloblastin; D012697:Serine Endopeptidases / Q000276:immunology*
chemical_list: D001324:Autoantigens; D007128:Immunoglobulin Fragments; D007143:Immunoglobulin Heavy Chains; D007135:Immunoglobulin Variable Region; D012697:Serine Endopeptidases; D053146:Myeloblastin