What laboratory tests are required to diagnose antiphospholipid syndrome?

Laboratory Testing for Antiphospholipid Syndrome

To diagnose antiphospholipid syndrome, you must perform all three core antibody tests—lupus anticoagulant (LAC), anticardiolipin antibodies (aCL) IgG and IgM, and anti-β2-glycoprotein I antibodies (aβ2GPI) IgG and IgM—on the same blood sample, and if any test is positive, repeat the entire panel at least 12 weeks later to confirm persistent positivity. 1, 2

Required Laboratory Tests

The Three Mandatory Tests

You must order all three of the following tests simultaneously on the same sample 1, 3, 2:

• Lupus anticoagulant (LAC) detected by two parallel phospholipid-dependent clotting assays: dilute Russell's viper venom time (dRVVT) AND activated partial thromboplastin time (APTT) 4, 1, 2
• Anticardiolipin antibodies (aCL) IgG and IgM isotypes measured by ELISA or automated solid-phase assays, with positivity defined as levels greater than the 99th percentile of normal controls 1, 3
• Anti-β2-glycoprotein I antibodies (aβ2GPI) IgG and IgM isotypes measured by ELISA or automated solid-phase assays, with positivity defined as levels greater than the 99th percentile of normal controls 1, 3

Lupus Anticoagulant Testing Methodology

LAC testing follows a mandatory three-step process 1, 2:

1. Screening test using both dRVVT and APTT
2. Mixing study to differentiate inhibitor from factor deficiency
3. Confirmatory test with phospholipid correction to confirm phospholipid-dependent inhibition 4, 1

The result should be reported as definitively positive or negative with documentation of any interferences 1, 2

Critical Timing Requirements

If any test is positive on initial testing, you must repeat all three tests at least 12 weeks later to confirm persistent antibody positivity and exclude transient positivity. 1, 3, 2 This 12-week confirmation requirement applies only to positive results, not negative results 3, 2

The same antibody that was initially positive must remain positive on repeat testing for APS diagnosis 1

Absolute Contraindications to Testing

Never perform lupus anticoagulant testing while the patient is on anticoagulation therapy—this produces unreliable false-positive or false-negative results. 1, 2 Specifically:

• Vitamin K antagonists (warfarin) interfere with LAC testing 1
• Direct oral anticoagulants (DOACs) interfere with LAC testing 1
• Heparin interferes with LAC testing 1

If LAC testing is unavoidable in anticoagulated patients, DOAC-removal procedures may be attempted, but interpretation must be extremely cautious 1, 2

Additional Situations That Produce False Results

• During acute thrombosis: Antibody levels may decrease due to antibody deposition at the thrombotic site 3
• During pregnancy: Factor VIII increases can mask lupus anticoagulant by shortening APTT, producing false negatives 1, 3

If initial testing was performed under any of these conditions and was negative, repeat testing when the patient is off anticoagulation is warranted 3

Risk Stratification Based on Results

The antibody profile determines thrombotic risk and management intensity 1, 3, 2:

• Triple-positive patients (LAC + aCL + aβ2GPI of the same isotype): Highest risk of thrombosis and pregnancy complications, requiring the most aggressive management 1, 3, 2
• Double-positive patients (concordant aCL and aβ2GPI isotypes): Significantly increased diagnostic reliability and elevated risk 1
• Single LAC positivity alone without ELISA positivity: Actually carries low thrombotic risk 3
• IgG isotype: Clinically more relevant than IgM for both aCL and aβ2GPI; isolated IgM positivity is less clinically significant 1

Essential Interpretation Requirements

Laboratory results must be reviewed jointly by a clinical pathologist and a clinician experienced in APS interpretation, with full knowledge of the patient's anticoagulation status and clinical context. 1, 3, 2 Test results must always be correlated with clinical symptoms 1, 3

There is high inter-laboratory and inter-assay variability for aCL and aβ2GPI testing, so low-positive results near cutoffs should be interpreted cautiously 1, 3

Non-Criteria Tests (Not Recommended for First-Line Diagnosis)

• Anti-domain I β2GPI antibodies (aD1): Remain investigational for risk stratification; not recommended for first-line diagnosis 1, 3
• Antiphosphatidylserine/prothrombin antibodies (aPS/PT): May be useful when LAC testing is unreliable due to anticoagulation, but should not replace LAC 1, 3
• Other antiphospholipid antibody tests: Not recommended for routine diagnosis 1

Reporting Requirements

• LAC results: Report as positive/negative with warnings about potential interferences 1
• aCL and aβ2GPI results: Report with specific quantitative levels 1
• All results: Must include documentation of whether the patient was on anticoagulation at time of testing 1, 2