Correlation of ANA Characteristics with pANCA IFA Interference

Abstract

Abstract Background Antineutrophil cytoplasmic antibody (ANCA) testing by the indirect immunofluorescence assay (IFA) is important for the diagnosis of autoimmune vasculitis. A common analytical interference for ANCA-IFA is the presence of an antinuclear antibody (ANA), which can cause an apparent perinuclear ANCA (pANCA) result on ethanol-fixed neutrophils. Here, the association of ANA patterns, titers, and concentrations with pANCA interference is investigated. Methods Samples positive for ANA by IFA with homogeneous, speckled, dense fine speckled (DFS), and centromere patterns were tested for ANA by enzyme immunoassay (EIA)] and for ANCA by IFA on ethanol-fixed neutrophils. Titers and concentrations were determined for the ANA-IFA and EIA, respectively, and correlated with the frequency of pANCA interpretations. Results For ANA-EIA positive samples (≥1.1U), 20.0% led to a pANCA interpretation compared to 5.1% for negative samples (≤1.0U). For samples positive by ANA-IFA, 12.9% resulted in a pANCA interpretation. Interference on pANCA correlated with ANA-IFA titer, with ANA titers ≥1:1280 identified as pANCA positive in 20.9% of samples compared to 9.7% for titers <1:1280. There was also a correlation with ANA pattern, as homogeneous samples were most likely to be called positive for pANCA (31.7%), followed by speckled (8.8%), DFS (6.8%), and centromere (3.6%). Conclusions Positivity for ANA by EIA is associated with increased prevalence of pANCA interpretation. Samples positive for ANA by IFA also demonstrated this association, particularly with higher-titer, homogeneous patterns. Laboratories can use this information to determine an optimal workflow for when investigating potential pANCA interferences.