"Visit the laboratory": Thoughts on navigating the clinical complexity of APS diagnosis

• APS diagnosis is extremely complex and critical to determining effective anticoagulation management
• Lupus anticoagulant testing is non-standardized, susceptible to false negatives and false positives
• Closing the knowledge gap between clinicians and laboratory professionals is critical for successful APS diagnosis

Part one of a two-part series with Professor Vittorio Pengo

Antiphospholipid Syndrome (APS) occupies a difficult corner of clinical medicine, sitting at the crossroads of autoimmunity, thrombosis, and obstetric disease. Accurate APS diagnosis matters profoundly, because it shapes whether a patient is committed to lifelong anticoagulation, how recurrent pregnancy loss is understood, and which class of anticoagulant is chosen after a thrombotic event. At the heart of the diagnostic workup sits the lupus anticoagulant (LA) assay, a test whose name itself is something of a misnomer. Despite the word "anticoagulant" in its title, a positive result most often signals a heightened risk of thrombosis rather than bleeding, and misinterpretation can send treatment in exactly the wrong direction.

LA testing is also notoriously difficult to perform and to read. The assay is not standardized across laboratories, it can generate false positives and false negatives depending on the clinical context, and reports of "weak" or "borderline" positivity routinely leave clinicians uncertain how to proceed. For physicians managing unexplained thrombosis, autoimmune disease, or obstetric complications, a working understanding of the clinical nuances of LA testing is not a luxury but a necessity.

In this two-part series, Professor Vittorio Pengo, Senior Scientist at the University of Padova, Italy, and a leading authority on APS diagnosis and LA testing, shares his perspective on the clinical and laboratory dimensions of the assay. The first installment focuses on the critical need for clinicians to understand what happens to their samples once they leave the ward and enter the laboratory, a process that is an essential component of effective LA testing.

LabLeaders: Why is lupus anticoagulant testing still considered so challenging for clinicians today?

Prof. Pengo: It is, at its core, a non-standardized and tricky test, and that point has to be clear for clinicians. To illustrate the scale of the problem: when our reference laboratory reviewed more than 300 patient samples that had been reported as positive for lupus anticoagulant by thrombosis centers across Italy, 71 of those turned out to be false positives, which is 23% of all submitted samples.¹

The test is susceptible to false negatives as well. The problem is particularly acute when results are reported as "weak" or "borderline." These classifications leave clinicians in a very difficult position, since they simply do not know what to do next for their patients. Reporting a weak or borderline result is itself a dangerous practice, because it offers no actionable guidance.

LabLeaders: In which clinical situations is lupus anticoagulant testing typically requested, and why does the context matter?

Prof. Pengo: There are three main scenarios. The first is unexplained thrombosis: arteriovenous thrombosis at uncertain sites in a young patient. The second is obstetric complications, such as recurrent pregnancy loss. The third is a more incidental finding: the patient is being assessed for surgery, or a routine test reveals an unexplained prolonged aPTT.

What is important to recognize is that all three of these scenarios may be associated with some degree of inflammation. And inflammation, specifically high C-reactive protein, is one of the causes of false positive results. This means the very situations that prompt lupus anticoagulant testing are the ones most likely to produce misleading results. That tension is at the heart of why this test is so difficult to interpret in practice.

LabLeaders: What are the main causes of false positive and false negative results?

Prof. Pengo: The primary cause of false positives is elevated C-reactive protein (CRP). When CRP is above 50 mg/L, it binds to phospholipids and can cause the test to read as falsely positive. Low-level CRP, below about 10 mg/L as you routinely see in stable patients, does not interfere.² The effect is specific to significantly elevated levels, which is precisely what you find in fresh thrombosis, the perioperative period, and active obstetric complications.

On the other side, the primary cause of false negatives is a very high Factor VIII level, above 150%.³ A second, less well-known cause involves the lupus cofactor, a phenomenon related to prothrombin. Clinicians do not need to master the analytical detail, but they should know that this mechanism exists, that it can suppress a lupus anticoagulant result that should be positive, and that it is explained fully in the laboratory literature. Both interferences should be kept in mind when a negative result does not fit the clinical picture.

LabLeaders: The term "lupus anticoagulant" seems counterintuitive — it prolongs clotting times in the lab but is associated with thrombosis. Can you explain?

Prof. Pengo: The term is genuinely a misnomer, and it causes real confusion among clinicians. "Anticoagulant" implies a tendency toward bleeding, but lupus anticoagulant is mainly associated with thrombosis, not bleeding. It prolongs the aPTT in the laboratory, which is how it got its name. But in the body, its predominant effect is pro-coagulant.

There are at least three distinct clinical entities that fall under this umbrella, and understanding them helps make sense of the name.

1. The first and most clinically dangerous is lupus anticoagulant occurring together with anti-cardiolipin and anti-β2GP1 antibodies: triple positivity. This combination is highly pro-coagulant and carries the greatest thrombotic risk.
2. The second is isolated lupus anticoagulant, where the LA is positive but anti-cardiolipin and anti-β2GP1 are negative. The pro-coagulant risk is real but considerably lower. It is also worth noting that single positive results are more common, partly because isolated positivity is more prone to false positives and transient reactions.
3. The third is the one true anticoagulant form: the lupus anticoagulant-hypoprothrombinemia syndrome. In these patients, anti-prothrombin antibodies cause a severe depletion of prothrombin, and the patient bleeds, sometimes massively. This syndrome mainly affects children, though it can occur in adults. It typically presents with unexpected bleeding such as ecchymosis or uterine bleeding in girls, with prothrombin levels falling below 10%.⁴ It is fortunately very rare, and primarily a concern for pediatricians and gynecologists.

LabLeaders: Should lupus anticoagulant testing be performed if a patient is already on long-term anticoagulation?

Prof. Pengo: Generally, no. If a patient is already on long-term oral anticoagulants, the test rarely changes the management plan — the medication stays the same regardless of the result, so there is little clinical reason to perform it.

However, there are specific situations where the test is critical. The most important is a young patient who arrives in the emergency room with proximal deep vein thrombosis (DVT) or pulmonary embolism. In that scenario, we face an urgent decision: should this patient be treated with Direct Oral Anticoagulants (DOACs) or Vitamin K Antagonists (VKA)? That decision is directly informed by lupus anticoagulant status and the overall antiphospholipid antibody profile.

The challenge is that many of these patients arrive already on heparin or DOACs, which interfere with the lupus anticoagulant test. This is why the single most important rule in this setting is: draw the blood before giving any anticoagulant. This must be absolutely clear to every clinician. If you miss that window, you lose your diagnostic opportunity.

LabLeaders: What should a clinician do if the patient is already anticoagulated when they arrive?

Prof. Pengo: If the lupus anticoagulant test cannot be performed because the patient is already anticoagulated, the first step is to order the immunological tests, specifically anti-cardiolipin and anti-beta 2 glycoprotein I (anti-β2GP1) antibodies. These are not coagulation-based assays; they are immunological tests and are unaffected by anticoagulants. They can be sent to the laboratory in standard serology tubes, without any special handling.

If both anti-cardiolipin and anti-β2GP1 are positive, you can suspect that the patient is triple positive, meaning all three antiphospholipid criteria are likely present. In that case, you should choose VKAs over DOACs. If the patient begins on a DOAC and antiphospholipid antibodies are subsequently confirmed, switching to a VKA for long-term treatment is the appropriate step, particularly in unprovoked DVT or pulmonary embolism where long-term anticoagulation is required.

There is also a useful surrogate test worth knowing about: anti-prothrombin antibodies. These antibodies mimic lupus anticoagulants in their clinical behavior, and a positive anti-prothrombin result provides a useful indirect signal that lupus anticoagulants may be present. Like the other immunological tests, it is not affected by ongoing anticoagulation and can be sent to the laboratory with routine serology.

LabLeaders: How significant is the risk of interference if a patient has only been on heparin or a DOAC for one or two days?

Prof. Pengo: It depends on the dose. At the start of treatment, patients typically receive therapeutic amounts of heparin, which is an initial bolus of 5,000 to 10,000 units. Some collection tubes contain a neutralizing agent that can quench heparin, but only up to a concentration of approximately one unit per milliliter. If the patient is on full therapeutic heparin, that quenching capacity is easily exceeded and the result cannot be trusted. When a patient is on full therapeutic anticoagulation, you should always maintain a degree of doubt about what is in the plasma.

LabLeaders: How important is retesting, and how should confirmed APS patients be monitored over time?

Prof. Pengo: Retesting is essential and is required by guidelines. A single positive lupus anticoagulant result is insufficient for APS diagnosis. According to the current ACR/EULAR classification criteria, a single positive test without confirmation earns only one point — not enough to classify a patient as having APS.⁵

Retesting should be performed at 12 weeks, because transient lupus anticoagulants do occur. They can appear with infections, medications, or inflammatory states, and then resolve. When we retested patients in our laboratory who had only a single positive lupus anticoagulant result, fewer than 50% remained positive at 12 weeks.⁶ This is clinically crucial: acting on a single positive result risks over-diagnosing APS and committing patients to long-term anticoagulation unnecessarily. The same 12-week confirmation requirement applies to women with recurrent pregnancy loss.⁵ The obstetric implications of an APS diagnosis are significant, and a single positive test is not sufficient.

Once APS is confirmed — laboratory criteria positive on two occasions at least 12 weeks apart with positive clinical domains — we then test annually. We monitor lupus anticoagulant alongside anti-cardiolipin and anti-β2GP1 because some patients do experience negativization of their antibodies over time.⁶

LabLeaders: If a patient's lupus anticoagulant becomes negative over time, can anticoagulation safely be stopped?

Prof. Pengo: This is an unresolved question, and I want to be honest about that. We do not yet have a randomized clinical trial to guide this decision, so no firm recommendation exists. Some treatments, for example hydroxychloroquine, can lead to negativization of lupus anticoagulant, but whether that negativization actually reduces the patient's underlying thrombotic risk is unknown.

Patients who become negative sometimes ask to stop anticoagulation. In a small number of carefully selected cases, particularly those where the original thrombotic event was provoked rather than idiopathic, or where arterial events could be attributed to competing cardiovascular risk factors such as hypertension or hypercholesterolemia, I have made this decision together with the patient, documented it in writing, and maintained very strict follow-up. But this cannot be taken as a general rule.

The negativization of lupus anticoagulant is also not that frequent, which itself limits how often this situation arises. Until a randomized trial answers the question, continuing anticoagulation remains the safer default, and any decision to stop must be made through careful shared decision-making.

LabLeaders: What pre-analytical information should clinicians always communicate to the laboratory when requesting lupus anticoagulant testing?

Prof. Pengo: Cross-talk between the clinician and the laboratory is fundamental, and it is where many diagnostic errors originate. The laboratory cannot properly interpret a lupus anticoagulant result without knowing what is circulating in the patient's blood at the time of the draw.

At minimum, the test request should clearly indicate whether the patient is currently receiving heparin, a DOAC, or a VKA. Ideally, it should also flag recent thrombotic events, surgery, or active inflammatory conditions. Many laboratory request forms do not currently accommodate this level of clinical detail, and that is something institutions should work to improve. A simple tick-box approach, noting which anticoagulant is ongoing, would be a meaningful step forward.

LabLeaders: What is your single most important message for clinicians caring for patients suspected of having APS?

Prof. Pengo: Visit the laboratory. A good internist, and more broadly any clinician managing these patients, should understand what happens to a sample after it leaves the ward. They should know about the interferences, the pre-analytical pitfalls, and what the request for a lupus anticoagulant test actually demands of the laboratory.

At a practical minimum, clinicians should ensure that test request forms clearly communicate whether the patient is currently receiving heparin, a DOAC, or a VKA. That information is fundamental to the laboratory's ability to interpret results correctly. Without it, the laboratory is working blind.

The gap between what a clinician orders and what the laboratory actually does is responsible for many of the problems we see, and closing that gap is a shared responsibility.

This is part one of a two-part series on lupus anticoagulant testing with Professor Vittorio Pengo. Part two will shift to the laboratory, examining why current guidelines insist on a three-step procedure with two parallel assays, the central and underappreciated role of phospholipids in assay performance, what Prof. Pengo believes the future of the coagulation laboratory should look like, and more.