Why can antiphospholipid antibodies be falsely positive during an acute illness?
Many readers have likely heard that one should avoid sending antiphospholipid antibody tests during active inflammation. Is the reason related to thrombosis? Is it related to the inflammatory state? Or is it instead related to issues related to the lab tests themselves. For this episode, we aimed to identify an answer.
To begin, antiphospholipid antibody syndrome (APS) is a syndrome of recurrent thrombosis (classically both venous and arterial) and miscarriages associated with autoantibodies against specific phospholipids or phospholipid-binding proteins (e.g., β2 glycoprotein I, cardiolipin, and prothrombin) or one of a heterogeneous group of coagulation factor inhibitors called lupus anticoagulants.
Why are the coagulation factor inhibitors called lupus anticoagulants? First, though can be associated with systemic lupus erythematosus they can also arise after a variety of events that trigger the immune system (e.g., infections, other autoimmune conditions, malignancy, and drugs. Lupus anticoagulant is one subtype of the syndrome, but together they are called antiphospholipid antibodies. As to why they are referred to as anticoagulants, the key is that they present with a prolongation of the activated partial thromboplastin time (aPTT) even though what they actually represent is a procoagulable state.
To understand why they lead to prolongation of the aPTT while being associated with elevated risk of thrombosis one must recognize that lab tests and our circulation are two different environments and produce different results. The key principle: in vivo veritas. In vivo, antibodies against phospholipids create a hypercoagulable state via a variety of mechanisms (e.g., endothelial injury, activation of platelets and complement, or interacting with the coagulation factors themselves). In vivo, however, none things factors are present. Instead, we utilize the aPTT. In doing so we aim to assess the intrinsic clotting cascade. The tube in which we draw the lab has citrate to bind the calcium and prevent the clotting cascade from initiation. Then, we reintroduce calcium and artificial phospholipids into the sample and determine how fast a clot forms. When an antibody is present and interfering with the phospholipids in the assay you may see a slowed activation of clotting and falsely prolonged test.
To distinguish a prolongation of the aPTT due to APS from other causes one must use both clinical criteria (i.e., presence of a clot or multiple spontaneous abortions, or pre-eclampsia/placental insufficiency) and at least 2 positive tests for an antiphospholipid antibody at least 12 weeks apart. There are two big categories of lab tests that can show APS:
- Direct tests. These measure one of the common antibodies (e.g., β2 glycoprotein I, cardiolipin
- Indirect tests. These aim to prove that there is a phospholipid-dependent prolongation of the aPTT.
The of the more interesting examples of an indirect test is the Daboia russelii viper venom test (dRVVT). This test uses a viper venom known to cause clot formation and therefore speed up the clotting assays. The tests are then assessed to see if they correct with phospholipid or the addition of normal plasma.
Returning to the original question, why might acute inflammation lead to a false elevation in the APS testing? One study looked at 51 critically ill patients who did not have an underlying coagulopathy, liver disease, or were receiving therapeutic anticoagulation. Twenty-seven of 51 patients developed positive testing for APS, with a prolonged aPTT and confirmation with dRVVT. Interestingly, though anti-cardiolipin and anti-β2 glycoprotein I were sent in all patients, none of them returned positive. None of the patients developed thromboembolic complications and the prolongation in aPTT resolved spontaneously during the ICU stay in 63% of the patients. For almost all patients, the lupus anticoagulant testing returned to normal within 4 weeks.
When compared to patients without positive testing, those with a positive lupus anticoagulant:
- Had a higher mean c-reactive protein (14.7 vs 6.0)
- Were more likely to have had sepsis
- Were more likely to receive “catecholamine treatment” (i.e., norepinephrine, epinephrine, or dobutamine)
Could the difference in c-reactive protein (CRP) level provide an explanation for this difference? Possibly. CRP is of course an acute phase protein that has an affinity for phospholipids, and particularly phosphatidylcholine. Another study published in 2010 studied 39 patients who had had a transiently positive lupus anticoagulant. The investigators compared the CRP from the positive to subsequent negative samples. As expected, the CRP was higher in the initial (positive) samples. They then added CRP to donor serum at a variety of concentrations and repeated the tests. Amazingly the aPTT became elevated above 2.4 mg/dL and increased proportionally with the CRP concentration.
It appears that CPR interferes with the aPPT assay. This helps explain why 12-week confirmatory testing is so important, and why we shouldn’t send these tests during an acute illness unless there is high suspicion for catastrophic APS.
Take Home Points
- In vivo veritas! Not all lab tests reflect their in vivo counterparts.
- APS testing can be falsely elevated during acute illness because of interference of CRP with the PTT and confirmatory assays.
- Don’t send APS testing during an acute event unless you have a high suspicion for catastrophic APS or APS causing a new stroke and need to make an urgent decision on anticoagulation.
CME/MOC
Click here to obtain AMA PRA Category 1 Credits™ (1.00 hours), Non-Physician Attendance (1.00 hours), or ABIM MOC Part 2 (1.00 hours).
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Credits & Citation
◾️Episode written by Hannah Abrams
◾️Show notes written by Tony Breu and Hannah Abrams
◾️Audio edited by Clair Morgan of nodderly.com
Abrams HR, Cooper AZ, Breu AC. In Vivo Veritas. The Curious Clinicians Podcast. February 16, 2022
Image credit: https://bangkokherps.wordpress.com/2011/05/05/russells-viper/
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