What is the purpose and interpretation of the Anti-Neutrophil Cytoplasmic Antibodies (ANCA) lab test?

ANCA Laboratory Testing: Purpose and Interpretation

ANCA testing is the mainstay diagnostic tool for detecting ANCA-associated vasculitides (AAV), particularly granulomatosis with polyangiitis (GPA), microscopic polyangiitis (MPA), and eosinophilic granulomatosis with polyangiitis (EGPA), using a combination of antigen-specific immunoassays (ELISA) for proteinase-3 (PR3) and myeloperoxidase (MPO), which have largely replaced indirect immunofluorescence as the preferred screening method. 1

What ANCA Tests Detect

Anti-neutrophil cytoplasmic antibodies (ANCAs) are autoantibodies, mainly of the IgG type, directed against antigens in the cytoplasm of neutrophil granulocytes and monocytes. 1

Two Main Testing Methods:

• Antigen-specific immunoassays (ELISA): High-quality ELISA tests for MPO-ANCA and PR3-ANCA are now the preferred screening method according to the 2017 revised international consensus. 1

• Indirect immunofluorescence (IIF): Historically used first, but increasingly being replaced by reliable antigen-specific immunoassays. 1

ANCA Patterns and Their Clinical Significance

c-ANCA (Cytoplasmic Pattern) with PR3 Specificity:

• Present in 80-90% of GPA patients with active systemic disease, confirming the diagnosis in up to 95% of cases. 1
• Specificity of 99% with sensitivity of 73% when using combined IIF and ELISA techniques. 1
• Sensitivity drops to 50% in limited forms of GPA or following corticosteroid therapy—a negative ANCA does not exclude GPA. 1
• Found in 20-40% of MPA and pauci-immune crescentic glomerulonephritis cases. 1

p-ANCA (Perinuclear Pattern) with MPO Specificity:

• More common in MPA and EGPA, detected in 30-40% of EGPA patients. 1, 2
• Approximately 5% of GPA patients can have positive p-ANCA/MPO, requiring careful differential diagnosis. 1, 2
• Clinical phenotype typically includes glomerulonephritis, peripheral neuropathy, and purpura. 2

Atypical ANCA Patterns:

• c-ANCA (atypical): Present in 80% of cystic fibrosis patients (targeting BPI), inflammatory bowel disease, primary sclerosing cholangitis, and rheumatoid arthritis. 1
• p-ANCA with non-MPO specificity: Associated with inflammatory bowel disease, autoimmune liver disease, drug-induced syndromes, and parasitic infections. 1, 2

When to Order ANCA Testing

Consider ANCA testing in any patient with suspicious clinical manifestations, particularly nasal crusting and bleeding, especially if they feel disproportionately unwell. 1

Key Clinical Scenarios:

• Pulmonary-renal syndrome: Simultaneous lung and kidney injury with rapidly declining GFR, microscopic hematuria with dysmorphic red cells, red cell casts, and moderate proteinuria (1-3 g/day). 1
• Upper respiratory tract symptoms: Chronic rhinosinusitis with nasal crusting, bleeding, and systemic symptoms (fatigue, weight loss, night sweats). 1
• Rapidly progressive glomerulonephritis: Declining kidney function over days to weeks. 1
• Multi-system involvement: Skin, eyes, nervous system, or other organ manifestations. 1

Diagnostic Algorithm

For Suspected Vasculitis:

1. Screen with antigen-specific ELISA for both MPO-ANCA and PR3-ANCA (preferred current approach). 1
2. Confirmatory IIF can be performed if ELISA is positive to assess pattern. 1
3. Do not delay treatment in rapidly deteriorating patients with compatible clinical presentation and positive MPO- or PR3-ANCA serology while waiting for biopsy results. 1

Complementary Testing:

• Elevated ESR and CRP. 1
• Renal function tests (proteinuria, microscopic hematuria, red cell casts). 1
• Chest imaging for pulmonary involvement. 1

Important Caveats and Pitfalls

False Positives:

• Cocaine/levamisole-induced vasculitis: Can cause positive c-ANCA and PR3, but ANCA reacts with human neutrophil elastase (HNE) rather than PR3, with massive apoptosis and abundant caspase-3/9 expression distinguishing it from autoimmune vasculitis. 1
• Non-vasculitic conditions: ANCA can be positive in inflammatory bowel disease, autoimmune hepatitis, primary sclerosing cholangitis, and drug-induced syndromes. 1, 2

Monitoring and Prognosis:

• Sequential c-ANCA monitoring can be useful in predicting relapse, though not all patients show classical fluctuation with disease activity. 1, 2
• ANCA-positive patients tend to have more frequent relapses than ANCA-negative patients, requiring vigilant monitoring. 2
• In EGPA, overall survival appears worse in ANCA-negative patients, while relapses are more frequent in ANCA-positive patients. 2

Sensitivity Limitations:

• About 10% of patients with small-vessel vasculitis or necrotizing crescentic glomerulonephritis are ANCA-negative—ANCA negativity does not exclude the diagnosis. 1
• Sensitivity is significantly reduced in limited disease forms and after corticosteroid treatment. 1

Not Recommended For:

ANCA testing is not included among recommended biomarkers for pleural effusion evaluation, where adenosine deaminase (ADA), interferon gamma, antinuclear antibodies (ANA), and NT-proBNP are the validated biomarkers. 3 ANCA should only be considered in pleural disease when there is clinical suspicion of systemic vasculitis secondarily affecting the pleura. 3