Epitope mapping of anti-PR3 antibodies using chimeric human/mouse PR3 recombinant proteins.
Autoantibodies against proteinase 3 (PR3) and myeloperoxidase (MPO) (ANCA = anti-neutrophil cytoplasmic antibodies) are used as diagnostic tools for patients with small vessel vasculitis. ANCA are detected by different assays, but the correlation between the results of these assays is generally poor. The overall aim of the study was to provide a framework for the future development of new assays with an increased diagnostic yield. In order to express discrete epitopes of human PR3 (hPR3), the nonantigenic molecules murine PR3 (mPR3) and human leucocyte elastase (HLE) were used as a framework. We constructed recombinant chimeric vectors and were able to produce 6 hPR3/mPR3 proteins and 3 hPR3/HLE proteins. Anti-PR3 monoclonal antibodies differed in their binding pattern to the chimeras, but no distinct binding region could be identified for any monoclonal antibody. The recombinant hPR3/mPR3 were also tested in ELISA with sera from patients with Wegener's granulomatosis with renal involvement. The results show that patients have antibodies to different constructs, indicating that the patients vary in their antibody repertoire from the beginning of the disease, and that patients may have antibodies from a broad range of clones early in the course of the disease. Recombinant hPR3/mPR3 chimeric proteins have a potential to be used as antigens in future ANCA assays.
Selga
D
D
Department of Nephrology, Lund University Hospital, Lund, Sweden. Diana.Selga@njur.lu.se
Segelmark
M
M
Wieslander
J
J
Gunnarsson
L
L
Hellmark
T
T
D000595:Amino Acid Sequence; D000818:Animals; D019268:Antibodies, Antineutrophil Cytoplasmic / Q000097:blood* / Q000276:immunology; D000911:Antibodies, Monoclonal / Q000276:immunology; D001323:Autoantibodies / Q000276:immunology; D002478:Cells, Cultured; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D018604:Epitope Mapping / Q000379:methods*; D005822:Genetic Vectors; D014890:Granulomatosis with Polyangiitis / Q000175:diagnosis* / Q000276:immunology; D006801:Humans; D019272:Leukocyte Elastase / Q000235:genetics; D051379:Mice; D008969:Molecular Sequence Data; D053146:Myeloblastin; D009195:Peroxidase / Q000276:immunology; D011993:Recombinant Fusion Proteins / Q000276:immunology; D011994:Recombinant Proteins / Q000276:immunology; D012697:Serine Endopeptidases / Q000235:genetics / Q000276:immunology*; D014162:Transfection
D019268:Antibodies, Antineutrophil Cytoplasmic; D000911:Antibodies, Monoclonal; D001323:Autoantibodies; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D009195:Peroxidase; D012697:Serine Endopeptidases; D019272:Leukocyte Elastase; D053146:Myeloblastin