Initial Antibody Testing for Autoimmune Disease Evaluation
For initial evaluation of autoimmune conditions, anti-nuclear antibody (ANA) testing by indirect immunofluorescence assay (IIFA) on HEp-2 cells should be the first-line screening test, followed by specific antibody testing based on the ANA pattern, titer, and clinical presentation. 1
Primary Screening Test
- ANA testing is the first-level test for laboratory diagnosis of systemic autoimmune rheumatic diseases (SARD) and should be performed using IIFA on HEp-2 cells as the reference method 1
- A screening dilution of 1:160 on conventional HEp-2 substrates is generally suitable for detecting clinically significant ANA in adult populations 1, 2
- Both nuclear and cytoplasmic patterns should be reported and specified when possible, as they provide valuable information about potential autoantibody specificity 1, 2
Follow-up Testing Based on ANA Results
For Positive ANA Results:
- If ANA is positive, the pattern and highest dilution demonstrating reactivity should be reported to guide further testing 1
- Testing for anti-dsDNA antibodies is advised when there is clinical suspicion of SLE, particularly with a homogeneous nuclear pattern 1, 3
- Specific anti-ENA (extractable nuclear antigen) antibody testing should be performed based on the ANA pattern observed 3, 1
Pattern-Specific Follow-up Testing:
- Homogeneous/Diffuse pattern: Test for anti-dsDNA, anti-histone, and anti-nucleosome antibodies (associated with SLE) 3, 1
- Speckled pattern: Test for anti-SSA/Ro, anti-SSB/La (Sjögren's syndrome), anti-Sm, anti-RNP (SLE, MCTD), anti-Topo-1 (systemic sclerosis) 3, 1
- Nucleolar pattern: Test for anti-PM-Scl, anti-RNA polymerase I/III (systemic sclerosis) 2, 1
- Centromere pattern: Associated with limited cutaneous systemic sclerosis 1, 2
- Cytoplasmic patterns: Should be reported and may indicate specific conditions like myositis or primary biliary cholangitis 1, 2
Important Technical Considerations
- The method used for antibody detection should always be included in test results 1
- For anti-dsDNA antibody determination, the Farr assay and Crithidia luciliae immunofluorescence test (CLIFT) offer high clinical specificity 1, 3
- Results of anti-dsDNA antibody detection should be reported quantitatively (or semiquantitatively for CLIFT) 1, 3
- For monitoring SLE disease activity, the same anti-dsDNA antibody detection method should be used consistently 1, 3
Common Pitfalls and Caveats
- ANA testing is primarily intended for diagnostic purposes, not for monitoring disease progression 1, 3
- Up to 25% of sera from apparently healthy individuals can be ANA positive, depending on demographics, dilution, and other variables 1, 4
- A negative ANA result does not exclude autoimmune disease, as some specific autoantibodies may be present in ANA-negative patients 3, 5
- Alternative automated methods for ANA detection may have different sensitivity and specificity profiles; if clinical suspicion is strong and an alternative method is negative, IIFA should be performed 1, 3
- Positive ANA can occur in non-autoimmune conditions including infections, malignancies, and with certain medications 4, 5
Disease-Specific Antibody Panels
- Systemic Lupus Erythematosus: ANA, anti-dsDNA, anti-Sm, anti-RNP, anti-SSA/Ro, anti-SSB/La, anti-C1q (for lupus nephritis) 3, 6
- Sjögren's Syndrome: ANA, anti-SSA/Ro, anti-SSB/La 3, 7
- Systemic Sclerosis: ANA, anti-centromere, anti-Scl-70 (topoisomerase I), anti-RNA polymerase III 1, 6
- Inflammatory Myopathies: ANA, anti-Jo-1, anti-SRP, anti-Mi-2 3, 6
- Mixed Connective Tissue Disease: ANA, anti-U1-RNP 3, 7
- Rheumatoid Arthritis: Rheumatoid factor, anti-CCP antibodies 7, 6
By following this structured approach to autoantibody testing, clinicians can improve diagnostic accuracy and ensure appropriate management of patients with suspected autoimmune conditions.