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Variation in antinuclear antibody detection by automated indirect immunofluorescence analysis

Pisetsky et al reported on assay variation in the detection of antinuclear antibodies (ANA) in sera of patients with established systemic lupus erythematosus (SLE).1 The authors determined ANA in sera from 103 patients with established SLE using three different validated and widely used immunofluorescence assays (IFA) (from ImmunoConcepts, Inova Diagnostics and Bio-Rad), an ELISA and a bead-based multiplex assay (both from Bio-Rad). With IFA, the frequency of ANA negativity varied from 4.9% to 22.3%. Part of the samples (11.7% and 13.6%) were negative with ELISA and multiplex assay, respectively. This study showed differences among IFA ANA kits which might have implications for eligibility for clinical trials.1

In a correspondence to the paper of Pisetsky et al .1 Dr Mahler discussed some relevant points related to ANA detection,2 including the substantial interobserver variability inherent to IFA. Accordingly, Dr Mahler recommended to use automated interpretation systems to reduce variability and subjectivity.2

We evaluated variation in ANA detection by automated IFA systems. Three samples (sample 1: homogeneous 1:320; sample 2: fine speckled 1:80; sample 3: negative) were distributed to 31 Belgian laboratories that use automated IFA systems and 27 laboratories participated (NOVA View n=12 (Inova Diagnostics, San Diego, USA); EUROPattern n=6 (Euroimmun, Lubeck, Germany); G-Sight n=7 (Menarini, Firenze, Italy); Image Navigator n=2 (ImmunoConcepts, Sacramento, California, USA)). Each sample was determined at least four times in different runs (the majority was determined at least eight times) according to the instructions of the manufacturer. The fluorescence intensity units (reported as light intensity units (LIU) or probability index) are shown in figure 1, except for EUROPattern that does not report fluorescence intensity. The results not only show variation in the way fluorescence intensity is reported by different companies …