6 These include antibodies to Golgi apparatus, mitochondria, Jo-1, ribonuclear protein, and others.
12 Using HEp-2 cells as the substrate, the IFA allows detection of more than 50 autoantibodies to 30 different nuclear and cytoplasmic antigens. In August 2009, the ACR issued a statement declaring HEp-2 IFA as the preferred method for ANA screening. In 2008, the American College of Rheumatology (ACR) initiated a task force to investigate and collect information from physicians to evaluate the extent of the problem. These new methods do not agree 100% with IFA, and the lack of standardization and inconsistencies among these assays have frustrated many clinicians and laboratory professionals. During the past decade, this increasing volume has led to the development of less labor-intensive methods, including enzyme linked immunoassay and multiplex microsphere immunofluorescent assays, for detecting ANAs or extractable nuclear antigen antibodies. At ARUP Laboratories, Salt Lake City, UT, more than 14,000 ANA tests are performed monthly and more than 160,000 per year. 1–6 The difficulty in diagnosing CTD dramatically increases the number of ANA tests ordered. A negative ANA test is thought to rule out systemic lupus erythematosus (SLE) however, a positive result is not specific for CTD. The ANA test should be ordered only when the patient’s history, symptoms, and physical examination findings are suggestive of a connective tissue disease (CTD). Patients with autoimmune disease often produce antinuclear antibodies (ANAs) that can be detected by indirect fluorescent antibody (IFA) techniques using a HEp-2 cell substrate. Variations in slide and substrate quality were also noted (ie, clarity, consistency of fluorescence, cell size, number and quality of mitotic cells).Īlong with subjectivity of interpretation, HEp-2 IFA assays are also vulnerable to standardization issues similar to other methods for ANA screening.
Within the specific groups of serum samples, agreement ranged from 44% in scleroderma serum samples to 93% in healthy control subjects, with 72% agreement in the SLE group. Standardization among the HEp-2 IFA assays occurred when they exhibited the same titer ± 1 doubling dilution.Īgreement of the 5 assays was 78%. In addition, 12 defined serum samples from the Centers for Disease Control and Prevention and 100 patient serum samples sent to ARUP Laboratories (Salt Lake City, UT) for ANA IFA testing were also examined (n = 372). We evaluated 5 commercially available HEp-2 antinuclear antibody (ANA) indirect fluorescent antibody (IFA) assays using patient serum samples from 45 patients with rheumatoid arthritis, 50 with systemic lupus erythematosus (SLE), 35 with scleroderma, 20 with Sjögren syndrome, 10 with polymyositis, and 100 healthy control subjects.
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