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Comparison of Antinuclear Antibody Testing Methods: Immunofluorescence Assay versus Enzyme Immunoassay

Comparison of Antinuclear Antibody Testing Methods: Immunofluorescence Assay versus Enzyme Immunoassay,RICHARD A. GNIEWEK,DANIEL P. STITES,THOMAS M. M

Comparison of Antinuclear Antibody Testing Methods: Immunofluorescence Assay versus Enzyme Immunoassay   (Citations: 12)
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Performances of anti-nuclear antibody testing by immunofluorescence assay (ANA-IFA) and enzyme immu- noassay (ANA-EIA) were compared in relation to patient diagnosis. A total of 467 patient serum samples were testedbyANA-IFA(Kallestad;Sanofi)andANA-EIA(RADIAS;Bio-Rad),andtheirage,sex,diagnosis,disease status, and medications were obtained through chart review. Reference ranges were established by testing 98 healthy blood donor samples. Eighty-six samples came from patients with diffuse connective tissue diseases, including systemic lupus erythematosus, discoid lupus erythematosus, or drug-induced lupus (n 5 71); systemic sclerosis, CREST syndrome (calcinosis, Raynaud's phenomenon, esophageal motility abnormalities, sclerodactyly, and telangiectasia), or Raynaud's syndrome (n 5 8); Sjogren's syndrome (n 5 5); mixed connective tissue disease (n 5 5); and polymyositis or dermatomyositis (n 5 3). The sensitivity, specificity, positive predictive value, and negative predictive value for ANA-IFA were 87.2, 48.0, 29.1, and 93.9%, respec- tively, for the reference range of <1:160. For ANA-EIA, they were 90.7, 60.2, 35.8, and 96.4%, respectively, for thereferencerangeof<0.9.ANA-EIAoffersequivalentsensitivityandhigherspecificitycomparedtoANA-IFA. Anti-nuclear antibody (ANA) testing is widely used as a screening test in connective tissue diseases (CTD) such as systemic lupus erythematosus (SLE), scleroderma, CREST syndrome (calcinosis, Raynaud's phenomenon, esophageal motility abnormalities, sclerodactyly, and telangiectasia), Sjogren's syndrome, mixed connective tissue disease (MCTD), polymyositis, and dermatomyositis. However, positive ANA results are seen in a significant proportion of the elderly pop- ulation (6, 17, 18, 20) and sensitivity of ANA testing varies widelyfromoneclinicaldiseasetoanother.Forexample,ANA testing has been reported to be positive in .95% of patients with SLE but in only 10 to 50% of patients with dermatomy- ositis and polymyositis (20). The first description of ANA was made by Hargraves and colleagues in 1948 when they observed LE (lupus erythemato- sus) cells in the bone marrow of patients with SLE (4). Cur- rently, the most commonly used method for ANA testing is ANA-immunofluorescence assay (ANA-IFA) in which slides prepared from human epithelioid cells (HEp-2 cells) as a sub- strate are incubated with diluted serum. The presence of au- toantibodies is detected by fluorescent antiimmunoglobulin antibody, and characteristic morphologic patterns of fluores- cent staining are observed. Certain ANA-IFA patterns are associated with the presence of autoantibodies to certain nu- clear antigens which in turn are associated with certain clinical states (7, 13, 17, 20). For example, a diffuse or homogenous pattern is associated with such clinical states as SLE, rheuma- toid arthritis, scleroderma, Sjogren's syndrome, and drug-in- duced lupus. The ANA-IFA is a subjective assay requiring skilled personnel and is a manual assay with a significant amount of hands-on time. Therefore, an ANA-enzyme immu- noassay (ANA-EIA) is an attractive alternative to ANA-IFA, especially when the operation is automated. ANA-EIA should be able to reduce training time and hands-on time as well as eliminate the subjectivity in interpreting results. Studies on concordance of ANA-IFA and ANA-EIA results in which se- rum samples were tested by both methods have been reported previously (8, 11). However, the correlations between ANA results and the presence of CTD were not described in these studies. In this study, we compared the performance of ANA- IFA and ANA-EIA based on patient diagnosis. We conclude thatANA-EIAoffersequivalentsensitivityandincreasedspec- ificity compared to ANA-IFA. from the institution's Committee on Human Research was obtained for the use of these patient and blood donor serum samples for the purpose of this study. For the blood donors, age and sex were obtained. Assays.ANA-IFA (Kallestad QUANTAFLUOR; Sanofi Diagnostics Pasteur, Inc., Chaska, Minn.) was performed according to the manufacturer's instruc- tions. The serum samples were kept frozen (2208C) until they were ready to be tested by ANA-EIA by using the automated analyzer RADIAS (Bio-Rad Lab- oratories, Hercules, Calif.). The ANA-EIA plates for the RADIAS are coated with a HEp-2 cell extract containing ANA antigens which include double- strandedDNA(dsDNA),Sjogren'ssyndromeantigensAandB(SS-AandSS-B)
Published in 1997.
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