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A novel enzyme-linked immunosorbent assay using a mixture of human native and recombinant proteinase-3 significantly improves the diagnostic potential for antineutrophil cytoplasmic antibody-associated vasculitis
  1. J Damoiseaux1,
  2. C Dähnrich2,
  3. A Rosemann2,
  4. C Probst2,
  5. L Komorowski2,
  6. C A Stegeman3,
  7. K Egerer4,
  8. F Hiepe4,
  9. P van Paassen1,
  10. W Stöcker2,
  11. W Schlumberger2,
  12. J W Cohen Tervaert1
  1. 1
    Department of Clinical and Experimental Immunology, University Hospital Maastricht, The Netherlands
  2. 2
    EUROIMMUN AG, Lübeck, Germany
  3. 3
    Department of Nephrology, University Medical Center Groningen, The Netherlands
  4. 4
    Department for Rheumatology and Clinical Immunology, Charité University of Medicine Berlin, Germany
  1. J Damoiseaux, Department of Clinical and Experimental Immunology, University Hospital Maastricht, PO Box 5800, 6202 AZ Maastricht, The Netherlands; jan.damoiseaux{at}mumc.nl

Abstract

Background: Antineutrophil cytoplasmic antibodies (ANCA) with a C-ANCA or P-ANCA pattern are detected in ANCA-associated vasculitis (AAV). While in most patients with AAV a C-ANCA pattern is due to reactivity with proteinase-3 (PR3)-ANCA, some C-ANCA-positive sera do not react with PR3.

Objective: The development and evaluation of a direct enzyme-linked immunosorbent assay (ELISA) for PR3-ANCA with increased sensitivity.

Methods: A mixture of human native (hn) and human recombinant (hr) PR3 was used as antigen coating. The resulting ELISA (anti-PR3-hn-hr) was compared with ELISAs using directly coated hn-PR3 or hr-PR3, as well as with a hn-PR3 capture ELISA. Assay characteristics were determined in patients with AAV (n = 248), with special attention for those patients with C-ANCA (n = 132), as well as disease controls (n = 585) and healthy controls (n = 429). Additionally, for prediction of relapses serial samples of 46 patients with PR3-AAV were analysed.

Results: At a predefined specificity of 99% both ELISAs containing hr-PR3 revealed a substantial increase in sensitivity. For the prediction of relapses by rises in PR3-ANCA titres the capture ELISA was most optimal (odds ratio 12.5). With an odds ratio of 8.9 the novel anti-PR3-hn-hr ELISA was second best.

Conclusions: Owing to the very high sensitivity of the novel anti-PR3-hn-hr ELISA for the detection of PR3-ANCA in C-ANCA-positive samples of patients with AAV this assay has an excellent diagnostic performance. This feature is combined with a good predictability of clinical relapses in patients with PR3-AAV. These characteristics challenge the dogma that, for detection of PR3-ANCA, capture ELISAs are superior for diagnosis and follow-up.

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Footnotes

  • Competing interests: CD, AR, CP and LK all are employees of EUROIMMUN AG, Lübeck Germany. W Schlumberger and W. Stöcker are board members of EUROIMMUN AG. The other authors have declared no conflict of interest.

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