Improvement of indirect immunofluorescence technique to detect antineutrophil cytoplasmic antibodies and its impact on test positivity rate

The indirect immunofluorescence (IIF) technique for antineutrophil cytoplasmic antibodies (ANCA) detection is subject to substantial differences across laboratories. This study aimed to assess the impact of improvements in the IIF-ANCA technique on the positivity rate of ANCA tests. A cross-sectiona...

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Bibliographic Details
Published in:Brazilian Journal of Medical and Biological Research 2023, Vol.56 (1)
Main Authors: Zarur, E.B, Rodrigues, S.H, Ambrogini, O., Jr, Ferraz, M.L.G, Mastroianni- Kirsztajn, G, Andrade, L.E.C, de Souza, A.W.S
Format: Report
Language:English
Subjects:
Autoantibodies
Chronic active hepatitis
Diagnosis
Fluorescent antibody technique
Health aspects
Immunofluorescence
Measurement
Methods
Ulcerative colitis
Vasculitis
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Summary:The indirect immunofluorescence (IIF) technique for antineutrophil cytoplasmic antibodies (ANCA) detection is subject to substantial differences across laboratories. This study aimed to assess the impact of improvements in the IIF-ANCA technique on the positivity rate of ANCA tests. A cross-sectional study was performed with serum samples from patients with ANCAassociated vasculitis (AAV), autoimmune hepatitis (AIH), and ulcerative colitis (UC). A paired analysis was performed for IIFANCA results using the traditional method and a modified protocol after a series of specific adjustments in the technique based on the protocol of IIF-ANCA test performed at a nation-wide private laboratory in Brazil. ANCA specificity was assessed by ELISA for anti-proteinase 3 (PR3) and anti-myeloperoxidase (MPO) antibodies. Sixty-one patients were evaluated. The positivity rate of IIF-ANCA tests at disease presentation performed at the University reference laboratory was 32.3% in AAV, AIH, and UC patients, whereas the positivity rates of IIF-ANCA and ELISA tests in other laboratories were 75.0 and 72.7%, respectively. After modifications in the IIF-ANCA technique, there was a significant increase in the positivity rate (14.8 vs 34.3%; P=0.0002) and in median titers [1/40 (1/30-1/160) vs 1/80 (1/40-1/80); P=0.0003] in AAV, AIH, and UC patients. UC had the highest increment in positive results from 5.3 to 36.8%. There was poor agreement between MPO- or PR3-ANCA and both IIF-ANCA techniques. In conclusion, modifications in the IIF-ANCA protocol led to a significant improvement in its positivity rate and titers. Key words: Antineutrophil cytoplasmic antibodies; ANCA-associated vasculitis; Autoimmune hepatitis; Ulcerative colitis; PR3ANCA; MPO-ANCA
ISSN:0100-879X
DOI:10.1590/1414-431X2023e12636