The performance of immunoassays for the detection of autoantibodies is of critical importance in the diagnosis and assessment of patients with autoimmune connective tissue diseases (ACTD). Our objective was to compare the features of two multiplexed assays—INNO-LIA ANA and Gennova-PictArray ENA ELISA—for measurement of multiple autoantibodies and their utility as a clinical tool in ACTD diagnosis. The antigens included SS-A/Ro (60 and 52), SSB/La, Sm, Sm/RNP, CENP-B, Jo-1, and Scl-70. Stored sera from 85 ACTD patients and 80 controls consisting of patients with vasculitis, rheumatoid arthritis and infectious diseases, as well as healthy subjects were analyzed jointly with clinical and laboratory data. Agreement between the two methods varied between 58 and 99% (Cohen’s kappa: 0.21–0.71) mostly for SSA and SSB. The frequency of specific autoantibodies measured using the two methods was more variable for SSA, SSB, and RNP/Sm. There were a higher number of ambiguous results when using INNO-LIA. The optimized cut-off values of the Gennova-PictArray resulted in over 99% specificities in samples obtained from the control group. Sensitivity patterns were more accurate in Gennova-PictArray than in INNO-LIA, as suggested in previously reported studies. A third method could be applied to determine which of the two methods is more accurate. 1. Introduction The detection of antinuclear antibodies (ANA) has long been an important tool in the early diagnosis of autoimmune connective tissue diseases [1]. The antigens used in their detection are purified by the saline extraction of animal origin nuclei, being termed as extractable nuclear antigens (ENA). The correct use and interpretation of serologic testing for diagnosing autoimmune diseases present a challenge to clinicians for several reasons: (a) the sensitivity and specificity of most laboratory tests for autoimmune disease are significantly less than 100% and (b) detection of autoantibodies using different techniques such as indirect immunofluorescence or multiplex bead assays give different results [2]. Multiplex microarray-based ELISA assays have been reported to provide concordant results when compared with ELISA-based tests [3] with the added advantage offered by multiplexing reduced labor and provision of the complete autoantibody profile with a single test. Autoantibodies play a role as biomarkers assisting in diagnosis and monitoring of disease activity, predicting disease onset, classifying disease subsets, and defining prognosis [4]. Their detection with high sensitivity and specificity is therefore of
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