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Piacenza d'Adige, Italy

Sinico R.A.,Clinical Immunology and Renal Unit | Radice A.,Microbiology Institute
Clinical and Experimental Rheumatology | Year: 2014

Antineutrophil cytoplasmic antibodies (ANCA) are considered the diagnostic biomarker of some necrotising vasculitis such as granulomatosis with polyangiitis (GPA), microscopic polyangiitis (MPA) and, to a lesser extent, eosinophilic granulomatosis with polyangiitis (EGPA). According to the current recommendations, combining indirect immunofluorescence and proteinase 3 (PR3) and myeloperoxidase (MPO) antigen specific immunometric assays, in the proper clinical setting, assures the best diagnostic specificity. When such conditions are satisfied, ANCA are detected in up to 90% of patients with active generalised GPA and MPA and in about 40% of patients with EGPA. Cytoplasmic ANCA (C-ANCA) with specificity for PR3 are usually found in patients with GPA whereas perinuclear ANCA (P-ANCA) in patients with MPA and EGPA. However, ANCA antigen specificity is more closely associated with disease phenotype and prognosis than clinical diagnosis. The clinical value of serial ANCA testing in monitoring disease activity is still debated. Recently, new promising developments in methodology and techniques (computer- based image analysis of imunofluorescence patterns, novel generation of PR3-/MPO-ANCA immunometric assays and multiplex technology) have been proposed but studies comparing the performances of the different assays are scarce. © Clinical and Experimental Rheumatology 2014.

Radice A.,Microbiology Institute | Bianchi L.,Nephrology and Clinical Immunology | Maggiore U.,University of Parma | Vaglio A.,University of Parma | Sinico R.A.,Nephrology and Clinical Immunology
Clinical Chemistry and Laboratory Medicine | Year: 2013

Background: PR3-ANCA, the serological marker of granulomatosis with polyangiitis (GPA), is usually detected by immunometric assays, with purified PR3 directly coated onto the solid-phase. Novel methods for PR3-ANCA detection have been developed to improve the performance of traditional PR3-ANCA specific assays, but little is known about their diagnostic performance in real-life clinical settings. This study aimed to compare the performance of nine different commercial PR3-ANCA specific assays, including traditional and newer ones, for the diagnosis of GPA. Methods: The evaluated assays for PR3-ANCA detection were representative of the first, second, and third generation tests (direct, capture and anchor assays, respectively). A third-generation assay employing both human and recombinant PR3 was also evaluated. The study population consisted of 55 GPA patients, 175 disease controls (representing most diseases in differential diagnosis with primary small-vessel vasculitis) including 52 with microscopic polyangiitis, and 20 healthy subjects. We performed the primary evaluation of test sensitivity using cut-off points which provided adequate and identical specificity for each test. Results: Although specificity and area under the ROC curve did not differ significantly between the different assays, substantial differences in sensitivity at 98%-specificity were found in some instances (p<0.001). Compared to first generation direct PR3-ANCA specific assays, some of the second and third generation tests increased the positive predictive value (PPV) for GPA diagnosis. Conclusions: Some of the newer PR3-ANCA specific assays have better PPV than traditional ones. © 2013 by Walter de Gruyter Berlin Boston.

Mahler M.,INOVA Diagnostics Inc. | Radice A.,Microbiology Institute | Yang W.,INOVA Diagnostics Inc. | Bentow C.,INOVA Diagnostics Inc. | And 3 more authors.
Clinica Chimica Acta | Year: 2012

Background: The detection of anti-proteinase 3 (PR3) and anti-myeloperoxidase (MPO) autoantibodies represents a serological hallmark in the diagnosis of small vessel vasculitis such as granulomatosis with polyangiitis (GPA) and microscopic polyangiitis (MPA). We evaluated novel chemiluminescence assays (CIAs) for PR3- and MPO-ANCA detection and investigated their utility for disease activity monitoring. Methods: Sera collected from GPA (n = 41) and MPA (n = 30) patients were tested by QUANTA Lite® PR-3 and MPO ELISAs (INOVA Diagnostics) and by the QUANTA Flash™ PR3 and MPO CIAs (INOVA). Precision and linearity were analyzed following reference guidelines. The recently launched reference sera for PR3-and MPO-ANCA (Centers of Disease Control and prevention, CDC) were used to establish international units for the new assays. Disease activity was determined using the Birmingham Vasculitis Activity Score. Results: The international standards for PR3-and MPO-ANCA yielded results of 403. CU and 332. CU in the novel CIAs, respectively. The linearity analysis showed linear regression values > 0.97 with slopes between 0.96 and 1.04. Total variation obtained from the precision study showed CV% of ≤ 7.4 for PR3-ANCA and ≤ 12.8 for MPO-ANCA. Good agreement (Spearman rho ≥ 0.89) was observed between CIA and ELISA. PR3-ANCA determined by CIA, but not by ELISA, was correlated with disease activity. No correlation was found for MPO-ANCA. Conclusion: The novel PR3- and MPO-ANCA CIAs show good precision, linearity and correlation to ELISA. In addition, PR3-ANCA by CIA show correlation with disease activity. © 2012 Elsevier B.V.

Radice A.,Microbiology Institute | Bianchi L.,Clinical Immunology and Renal Unit | Sinico R.A.,Clinical Immunology and Renal Unit
Autoimmunity Reviews | Year: 2013

Antineutrophil cytoplasmic antibodies (ANCA) are the serological hallmark of some idiopathic systemic vasculitides, such as granulomatosis with polyangiitis (GPA), microscopic polyangiitis (MPA) and, to a lesser extent, Churg-Strauss syndrome (CCS), the so-called ANCA-associated vasculitides (AAV).ANCA were first detected by immunofluorescence (IIFT), subsequently the target antigens myeloperoxidase (MPO) and proteinase 3 (PR3) were identified, allowing the development of the quantitative, antigen-specific assays. According to the guidelines, combining IIFT and PR3-ANCA/MPO-ANCA assures the optimal diagnostic specificity. Antigen specificity does not effectively differentiate among the different AAV, however C-ANCA/PR3-ANCA are mainly found in GPA, while P-ANCA/MPO-ANCA are more prevalent in MPA and CSS. Despite their diagnostic value, the performance of the widespread immunometric assays for ANCA testing is disappointing, particularly for the low sensitivity. In recent years, more "sensitive" assays have been developed, using the microplate as well as fully the automated technologies, with promising preliminary results. ANCA, may be detected in a number of pathological conditions other than small vessel vasculitis. However, in most of these non-vasculitic patients ANCA do not recognize MPO or PR3 as target antigens, but other granulocyte components, often multiple or unknown specificities. A positive ANCA result by itself is not diagnostic for AAV, clinical evidence and possibly histological confirmation are always required. On the other hand, a negative test result cannot completely rule out a diagnosis of AAV, as AAV without detectable ANCA exist. The appropriate use of ANCA testing strongly improves the diagnostic accuracy and clinical usefulness of the results. © 2012.

Damoiseaux J.,Maastricht University | Agmon-Levin N.,The Zabludowicz Center for Autoimmune Diseases | van Blerk M.,Scientific Institute of Public Health | Chopyak V.,Lviv National Medical University | And 10 more authors.
Clinical and Experimental Rheumatology | Year: 2014

Objective: One of the main goals of the European Autoimmunity Standardisation Initiative (EASI) is the harmonisation of test-algorithms for autoantibodies related to systemic autoimmune rheumatic diseases (SARD). Methods: A questionnaire was used to gather information on methodology, interpretation, and the algorithm for detection of anti-nuclear antibodies (ANA) in relation to their antigen-specificity. The questionnaire was sent to 1200 laboratories in 12 European countries. Results: The response rate was 47.2%. The results reveal not only apparent differences between countries, but also within countries. Conclusion: Awareness of these differences may as such already stimulate harmonisation, but the observed differences may also direct recommendations that may further contribute to achieving the EASI goal of harmonisation of autoimmune diagnostics for SARD. © Clinical and Experimental Rheumatology 2014.

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