How is a screening test for ANA usually performed in a clinical laboratory?
An indirect immunofluorescence test is the most widely accepted and studied assay used to detect ANA. The patient’s serum is diluted and then layered onto a slide with human epithelial type 2 (HEp-2) cells. A fluorescein-tagged antibody reagent directed to human immunoglobulin (Ig) is added. Any antibodies (from the patient) bound to the nucleus will be stained, and the nucleus will fluoresce when viewed under a fluorescence microscope ( Fig. 16.1 ). The results are registered as positive or negative and the strength of a positive reaction at a particular serum dilution. A dilution of at least 1:160 is required to consider a test significantly positive since >10% of the population can have a low-titer ANA. Other methods (enzyme-linked immunosorbent assay, multiplex immunoassays) to screen for ANAs are used in some laboratories. It is important to know which method is used.
The laboratory also reports a pattern of nuclear staining (rim, diffuse, speckled, or nucleolar). The peripheral or rim pattern (corresponding to autoantibodies to deoxynucleoproteins) is the most specific pattern for SLE, whereas a speckled pattern, which is the most common pattern in both SLE and other diseases, is the least specific. A nucleolar pattern should raise suspicion for scleroderma. Currently patterns have less significance because a positive test is usually followed by an ANA profile, which tests for specific types of autoantibodies including those that are highly specific for SLE (e.g., anti-dsDNA, anti-Sm).
