title: Detection of specific antinuclear reactivities in patients with negative anti-nuclear antibody immunofluorescence screening tests.
abstract: BACKGROUND
  For detection of anti-nuclear antibodies (ANAs) and antibodies to extractable nuclear antigens (ENAs), samples frequently are screened with indirect immunofluorescence (IIF); further determination of anti-ENA antibodies is performed only when the result is positive. However, because anti-ENA reactivities are found in samples with low fluorescence intensities, we determined anti-ENA antibodies in samples with negative IIF and thus calculated the sensitivity of IIF for specific ANAs.
  
  
  METHODS
  We collected 494 samples consecutively referred by rheumatologists for routine ANA testing. IIF on HEp-2 and HEp-2000 (HEp-2 cells transfected with Ro60 cDNA) and line immunoassay (LIA) for the detection of specific ANAs were performed on all samples.
  
  
  RESULTS
  Fluorescence intensities and patterns on HEp-2 were strongly correlated with those on HEp-2000 [Spearman rho = 0.852 (P <0.001) and 0.838 (P <0.001), respectively]. Sixty-eight of 494 samples were positive on LIA, of which only 72% (confidence interval, 68-76%) were detected with HEp-2 and 75% (confidence interval, 70-78%) with HEp-2000. Of 291 samples negative on both substrates, 12 were positive on LIA. Connective tissue diseases were diagnosed in four of these patients and suspected in at least three others.
  
  
  CONCLUSIONS
  The HEp-2 and HEp-2000 substrates perform comparably for fluorescence intensities and patterns and for detecting specific ANAs, but some patients with negative IIF show reactivity on LIA. We recommend testing for fine reactivities, regardless of the IIF result, when the clinical suspicion for rheumatic connective tissue disease is high.
authors.affiliation: Department of Rheumatology, Ghent University Hospital, De Pintelaan 185, Ghent 9000, Belgium. Ilse.Hoffman@rug.ac.be
authors.forename: Ilse E A
authors.initials: IE
authors.lastname: Hoffman
authors.forename: Isabelle
authors.initials: I
authors.lastname: Peene
authors.forename: Eric M
authors.initials: EM
authors.lastname: Veys
authors.forename: Filip
authors.initials: F
authors.lastname: De Keyser
mesh_terms: D000974:Antibodies, Antinuclear / Q000097:blood*; D003240:Connective Tissue Diseases / Q000175:diagnosis; D004797:Enzyme-Linked Immunosorbent Assay; D019084:Fluorescent Antibody Technique, Indirect; D006801:Humans; D011446:Prospective Studies; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
chemical_list: D000974:Antibodies, Antinuclear