What laboratory tests are used to diagnose Antiphospholipid Syndrome (APL) in a patient with Chronic Thromboembolic Pulmonary Hypertension (CTEPH)?

Laboratory Testing for Antiphospholipid Syndrome (APS)

The diagnostic laboratory workup for APS requires three mandatory tests performed in parallel: lupus anticoagulant (LA), anticardiolipin antibodies (aCL) IgG/IgM, and anti-β2 glycoprotein I antibodies (aβ2GPI) IgG/IgM, with all positive results requiring confirmation at least 12 weeks later to establish persistence. 1, 2

Core Mandatory Tests

Lupus Anticoagulant (LA)

• LA testing must use two parallel screening assays: dilute Russell's viper venom time (dRVVT) AND an LA-sensitive activated partial thromboplastin time (APTT) 1
• Omitting either dRVVT or APTT increases the risk of missing the diagnosis in up to 55% of triple-positive patients and 31% of APS patients 1
• Each screening test requires a 3-step methodology: screening test → mixing study → confirmatory test with phospholipid excess 1, 2
• Results are reported as positive or negative (not quantitative) 2, 3
• Critical pitfall: LA testing should NOT be performed during anticoagulation therapy as it causes false positives or false negatives 1

Anticardiolipin Antibodies (aCL)

• Measure both IgG and IgM isotypes by ELISA or validated solid-phase assays 1, 2
• Positive threshold is defined as values above the 99th percentile of normal controls 2, 3
• The 2023 ACR/EULAR criteria define moderate titers as >40 Units and high titers as >80 Units 1
• Report results with specific titer levels, not just positive/negative 3

Anti-β2 Glycoprotein I Antibodies (aβ2GPI)

• Measure both IgG and IgM isotypes by ELISA 1, 2
• Positive threshold is values above the 99th percentile of normal controls 2, 3
• Same titer thresholds as aCL: moderate >40 Units, high >80 Units 1
• Report with specific titer levels 3

Confirmation Testing Requirements

All positive results must be confirmed with repeat testing at least 12 weeks apart to rule out transient positivity 2, 3. This is mandatory for diagnosis and distinguishes persistent pathogenic antibodies from transient elevations.

Risk Stratification Based on Results

The antibody profile determines thrombotic risk and guides management intensity:

• Highest risk: Triple positivity (LA + aCL + aβ2GPI all positive) 2, 3
• High risk: Double positivity with concordant isotype (e.g., both aCL IgG and aβ2GPI IgG) 2, 3
• Moderate risk: Single LA positivity 1, 2
• Lower risk: Isolated IgM positivity at low-moderate titers 3

IgG isotypes carry greater clinical significance than IgM for thrombotic events, though IgM is more relevant in obstetric APS 3

Additional Considerations for CTEPH Patients

In the specific context of CTEPH, establishing the APS diagnosis is critical because:

• LA positivity is mandatory to identify high-risk triple-positive patients who require more aggressive anticoagulation 1
• The antibody profile informs prognosis and long-term anticoagulation decisions 1
• If the patient is already anticoagulated (which is typical in CTEPH), LA testing becomes extremely challenging and may need to be deferred or interpreted with extreme caution 1

Non-Criteria Tests (Not Routine)

• Antiphosphatidylserine-prothrombin (aPS/PT) antibodies may be considered only in patients with strong clinical suspicion but negative standard aPL panel 3, 4
• Anti-domain I β2GPI antibodies remain investigational 1
• Thrombin generation assays are not ready for routine clinical use 1, 3

Critical Pitfalls to Avoid

• Never test for LA during anticoagulation therapy - this is the most common source of false results 1
• Do not use classification criteria as diagnostic criteria - clinical diagnosis should be broader to optimize patient care 2, 4
• Low-positive results near threshold values require cautious interpretation due to 10% assay imprecision 2
• Single positive IgM without other antibodies has limited clinical significance and requires careful clinical correlation 3
• Results must always be interpreted in clinical context with knowledge of anticoagulation status 3

Ongoing Monitoring

For patients with confirmed APS, annual retesting of LA, aCL, and aβ2GPI is recommended to evaluate titer fluctuation and changes in antibody profile over time 1, 3, which may have therapeutic implications for anticoagulation intensity.