How should I test for lupus anticoagulant (LA) in a patient with an unexplained prolonged activated partial thromboplastin time (aPTT)?

Testing for Lupus Anticoagulant in Patients with Prolonged aPTT

Perform two parallel screening tests—dilute Russell's viper venom time (dRVVT) and an LA-sensitive activated partial thromboplastin time (aPTT)—followed by mixing studies and phospholipid confirmatory tests when either screening test is prolonged. 1

Required Testing Algorithm

Step 1: Dual Screening Tests (Mandatory)

• Run both dRVVT and LA-sensitive aPTT simultaneously as your initial screening tests 1, 2
• The dRVVT is the most robust first-line test for detecting LA in high-risk thrombotic patients 1
• An aPTT performed with silica as activator and low phospholipid content is the second test of choice due to its sensitivity 1
• Omitting either test misses up to 55% of triple-positive antiphospholipid antibody samples and 31% of APS patients 1, 2
• Do NOT use kaolin-activated aPTT (problematic in automated coagulometers) or ellagic acid-activated aPTT (insensitive for LA) 1

Step 2: Mixing Study (If Screening Prolonged)

• Mix patient plasma 1:1 with pooled normal plasma without pre-incubation 2
• Failure of the clotting time to correct indicates an inhibitor (LA) rather than a factor deficiency 3, 2
• Critical distinction: A prolonged mixing test could indicate LA, heparin contamination, or specific clotting factor inhibitors 1
• Perform a thrombin time (TT) on test plasma to identify heparin contamination 1
• Check clinical history for bleeding, which suggests specific factor inhibitors (e.g., factor VIII inhibitor) rather than LA 1, 4

Step 3: Confirmatory Test (Phospholipid-Dependent)

• Add high concentrations of phospholipids to the prolonged screening test 2
• LA-positive samples will show shortening of clotting time with excess phospholipids, confirming phospholipid-dependent inhibition 1, 2
• The clotting time may not always shorten to within normal range; calculate percentage of shortening against mean of normal controls to avoid false negatives 1
• Use pooled normal plasma prepared by double centrifugation with minimal residual platelets (<10^7/mL) and 100% clotting factor activity 1

Critical Pre-Analytical Requirements

Sample Collection Timing

• Collect blood BEFORE initiating any anticoagulant therapy 3, 2, 5
• If patient is already anticoagulated, testing is strongly discouraged due to false-positive and false-negative results 1, 3, 2
• Testing close to an acute thrombotic event requires caution due to elevated acute-phase reactants (e.g., factor VIII) 2

Sample Handling

• Use 0.109 M sodium citrate at 9:1 blood-to-anticoagulant ratio 2
• Perform double centrifugation to obtain platelet-poor plasma 3, 2
• If testing is delayed, freeze plasma promptly and thaw at 37°C before analysis 2
• Store pooled normal plasma frozen at -70°C in small aliquots 1

Testing During Anticoagulation (When Unavoidable)

Warfarin/VKA

• Draw blood when INR <1.5, ideally 1-2 weeks after discontinuation with LMWH bridging (last LMWH dose >12 hours before draw) 2
• Report LA as absent with comment recommending repeat testing when obtained during VKA therapy 1, 2

Direct Oral Anticoagulants (DOACs)

• Employ commercial DOAC-removal agents before testing, though availability is limited and removal may be incomplete 1, 2
• Activated charcoal is an inexpensive alternative but has more heterogeneous effects 1
• Compare clotting test results before and after DOAC adsorption to prevent misinterpretation 1
• If clotting times remain prolonged after DOAC removal, residual interference cannot be ruled out 1

Low Molecular Weight Heparin (LMWH)

• Screening is possible with reagents containing heparin neutralizers up to 0.8 U/mL 1, 2
• Effect on LA assays varies depending on the FXa to FIIa activity ratio of each LMWH preparation 1

Unfractionated Heparin

• LA screening is not feasible if heparin content exceeds reagent neutralization capacity 1, 2
• Commercial dRVVTs and aPTTs with neutralizers can quench heparin up to 0.8 U/mL 1

Result Interpretation and Follow-Up

Positive Result Actions

• Do NOT start long-term anticoagulation based on a single positive LA result 2
• Schedule repeat LA testing after >12 weeks to confirm persistence before making any treatment decision 1, 3, 2, 5
• Transient positivity is common; only persistent positivity (>12 weeks apart) supports APS diagnosis 2, 5
• Obtain a full antiphospholipid antibody profile (anticardiolipin IgG/IgM and anti-β₂-glycoprotein I IgG/IgM) for proper risk stratification 2

Risk Stratification

• LA positivity is mandatory for identifying high-risk triple-positive aPL patients (LA + anticardiolipin + anti-β₂-glycoprotein I) 1, 2
• Persistent isolated LA carries lower thrombotic risk than triple positivity 1, 2
• Triple positivity with medium-high titers of the same isotype (usually IgG) indicates highest thrombotic risk 2

Reporting Standards

• Report quantitative results with interpretative comment indicating presence/absence of LA 1
• Avoid comments such as "borderline" or "dubious LA"; instead state "to be tested again in 1 week" 1

Common Pitfalls to Avoid

• Never diagnose antiphospholipid syndrome on a single positive LA test; persistence >12 weeks is mandatory 2, 5
• Do not rely on a single screening assay (dRVVT or aPTT alone); both are required to avoid missing true LA cases 1, 2
• Avoid testing during any anticoagulant therapy whenever possible; all anticoagulants interfere with LA assays 1, 3, 2
• Do not use freeze/thawed platelets as phospholipid source for confirmatory tests due to poor batch-to-batch consistency 1
• Recognize that hydroxychloroquine may weakly interfere with LA testing by affecting IgG-β₂GPI complex formation 1
• Aspirin and clopidogrel do not interfere with LA testing 1