When should I order an anti‑streptolysin O (ASO) titer versus an antinuclear antibody (ANA) test?

When to Order ASO vs ANA Testing

Order ASO titer when evaluating for post-streptococcal sequelae (acute rheumatic fever or post-streptococcal glomerulonephritis), and order ANA when screening for systemic autoimmune rheumatic diseases like SLE, Sjögren's syndrome, or systemic sclerosis.

ASO Testing: Indications and Interpretation

Primary Clinical Scenarios for ASO

• Order ASO titer when you suspect acute rheumatic fever or post-streptococcal glomerulonephritis following a latent period after Group A streptococcal (GAS) infection 1
• ASO testing provides evidence of preceding GAS infection, which is required to diagnose non-suppurative post-streptococcal diseases 1

Critical Testing Strategy for ASO

• Always order at least two sequential ASO measurements rather than relying on a single value, as timing relative to infection is often unknown and single measurements are liable to misdiagnosis 1
• Simultaneously order anti-DNase B antibody along with ASO to optimize diagnosis of preceding GAS infection, as this dual approach significantly improves diagnostic accuracy 1
• Age-specific upper limits of normal for children ages 2-12 in the United States are: ASO ≤240 and anti-DNase B ≤640, with both increasing with age 2

Important Caveats for ASO Interpretation

• Elevated ASO titers (≥1:160) can be found in various clinical conditions beyond post-streptococcal diseases and do not necessarily correlate with positive streptococcal cultures, ESR, CRP, or rheumatoid factor 3
• In healthy adults, virtually no subjects have undetectable ASO titers between ages 15-60 years, and fewer than 5% have titers <10 TU after age 5, making ASO a poor screening test for general inflammatory conditions 4
• Do not use ASO testing as a general inflammatory marker or to evaluate non-specific rheumatic complaints without specific clinical suspicion for post-streptococcal sequelae 3

ANA Testing: Indications and Interpretation

Primary Clinical Scenarios for ANA

• Order ANA as the first-level screening test when evaluating for systemic autoimmune rheumatic diseases (SARD), including SLE, Sjögren's syndrome, systemic sclerosis, inflammatory myopathies, and mixed connective tissue disease 5, 6
• ANA testing is indicated when patients present with compatible clinical symptoms such as persistent joint pain/swelling, photosensitive rash, oral ulcers, pleuritic chest pain, unexplained fever, Raynaud's phenomenon, dry eyes/mouth, or muscle weakness 6
• ANA is primarily intended for diagnostic purposes, not for monitoring disease progression once a diagnosis is established 5, 6

Critical Testing Strategy for ANA

• Indirect immunofluorescence assay (IIFA) on HEp-2 cells is the reference method for ANA screening, with sensitivity of 95.8% and specificity of 86.2% at 1:160 dilution 5, 6
• A screening dilution of 1:160 provides optimal balance for detecting clinically significant ANA in adults, as lower titers have poor specificity: 31.7% of healthy individuals are positive at 1:40,13.3% at 1:80, and only 5.0% at 1:160 6
• Always report both the titer and the immunofluorescence pattern, as different patterns suggest different autoantibodies and disease associations 5, 6

Follow-up Testing Algorithm After Positive ANA

• If ANA titer is ≥1:160 with any pattern, order specific extractable nuclear antigen (ENA) panel including anti-Sm, anti-RNP, anti-SSA/Ro, anti-SSB/La, anti-Scl-70, and anti-Jo-1 6
• If clinical suspicion for SLE exists, add anti-dsDNA antibody testing using both Crithidia luciliae immunofluorescence test (CLIFT) for specificity and solid phase assays for sensitivity 6
• For homogeneous patterns specifically, anti-dsDNA and anti-histone antibodies are mandatory follow-up tests 6
• For speckled patterns, the specific ENA panel guides diagnosis toward particular autoimmune diseases 6

Important Caveats for ANA Interpretation

• In cases of high clinical suspicion, order specific antibody testing regardless of ANA result, as some autoantibodies (anti-Jo-1, anti-ribosomal P, anti-SSA/Ro) may be present in ANA-negative patients 6, 7
• ANA titers of 1:80 or lower have limited diagnostic value with only 74.7% specificity and should be interpreted with great caution 6
• Do not repeat ANA testing for disease monitoring—once positive, use quantitative anti-dsDNA and complement levels instead 6
• Different laboratory methods (IIFA vs. solid phase assays) have fundamentally different test characteristics and can yield discrepant results, so always ensure the laboratory specifies the method used 5, 6

Direct Comparison: Key Decision Points

Use ASO when:

• Evaluating for acute rheumatic fever (Jones criteria suspected)
• Investigating post-streptococcal glomerulonephritis (hematuria, proteinuria, edema following pharyngitis)
• Confirming recent streptococcal infection in specific post-infectious syndromes

Use ANA when:

• Screening for any systemic autoimmune rheumatic disease
• Evaluating multi-system inflammatory symptoms without clear infectious etiology
• Assessing patients with unexplained cytopenias, proteinuria, or inflammatory arthritis
• Following up on clinical suspicion of lupus, scleroderma, Sjögren's, or myositis

Never use ASO for:

• General screening for autoimmune diseases 3
• Monitoring disease activity in established rheumatic conditions 3
• Evaluating non-specific arthralgias or myalgias without post-streptococcal context 3

Never use ANA for:

• Diagnosing or monitoring post-streptococcal sequelae 5
• Disease activity monitoring once diagnosis is established 5, 6
• Screening asymptomatic patients without clinical suspicion 8