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Cited by 5Systemic Lupus Erythematosus ResearchBlood Coagulation and Thrombosis MechanismsClinical Laboratory Practices and Quality Control

Analytical dilemmas in lupus anticoagulant detection

Open access
Moore, Gary W.·2023·Exploration of Immunology
DOI 10.37349/ei.2023.00104
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Abstract

Accurate lupus anticoagulant (LA) detection is crucial to antiphospholipid syndrome (APS) diagnosis. Detection is based on LA functional behavior in coagulation assays irrespective of epitope specificity. LA screening tests employ dilute phospholipids to accentuate in vitro inhibition by LAs, although they are not LA-specific and can be elevated by other coagulation abnormalities. Elevated screening tests are reflexed to mixing tests to distinguish between factor deficiency and inhibition. Confirmatory tests with high phospholipid concentration swamp LA to generate shorter clotting times than screening tests, whilst prolongation persists with non-phospholipid-dependent inhibitors. LA heterogeneity means that no single screening test detects every LA and the screen/mix/confirm medley must be applied to at least two assay types, usually dilute Russell’s viper venom time (dRVVT) and an LA-sensitive activated partial thromboplastin time (aPTT). Most laboratories restrict LA testing to these two assays, yet others, such as dilute prothrombin time (dPT), can perform with equal diagnostic efficacy, and additionally detect LA unreactive with dRVVT and aPTT. Converting clotting times to normalized ratios improves assay performance, and practitioners must choose between normal pooled plasma (NPP) clotting time denominators to reflect on-the-day assay performance, or reference interval (RI) mean clotting times to negate the effects of NPP variation. Cut-offs can be generated parametrically from normally distributed data, or different percentiles applied depending on the preferred balance between sensitivity and specificity. Sourcing sufficient donors for accurate cut-off estimations is problematic and transference exercises can be undertaken on low donor numbers. Analytical limitations of mixing tests have led to the adoption of alternative algorithms to the screen/mix/confirm test order, whilst some continue to rigidly apply the latter despite those limitations. Strategies to reduce or eliminate the effects of therapeutic anticoagulation have limitations, whilst the Taipan snake venom time (TSVT) screening test with an ecarin time (ET) confirmatory test is insensitive to vitamin K antagonist (VKA) and direct activated factor X anticoagulation.

AI snapshot

Written from the paper’s full text.

TL;DR

This narrative review examines the analytical dilemmas in laboratory detection of lupus anticoagulants (LA), arguing that reliance on the dRVVT/aPTT pairing, rigid screen/mix/confirm algorithms, and inconsistent normalization and cut-off practices leave clinically significant antibodies undetected. It surveys alternative assays (dPT, ASLA, VLVT, textarin time, TSVT/ET) and testing strategies for anticoagulated patients, concluding that no single algorithmic solution fits all cases.

Key findings

  • No LA assay is LA-specific; elevated screening tests only indicate prolonged clotting times, and antibody heterogeneity means no single assay detects every LA, warranting at least two assay types (typically dRVVT plus an LA-sensitive aPTT).
  • Other assays detect LA sub-populations unreactive with dRVVT and aPTT: dPT (with recombinant thromboplastin) has been associated with a higher thrombosis association than dRVVT/aPTT in some studies, and TSVT/ET is insensitive to VKA and direct FXa inhibitors because direct FII activation bypasses FXa inhibition.
  • Cut-off derivation is contested: ISTH recommends blanket non-parametric 99th percentile cut-offs, while BSH and CLSI permit parametric RIs; the paper notes mean +2SD equals the 97.5th percentile for normally distributed data and the 99th percentile equals mean +2.3SD, so differences are minimal unless distributions are markedly skewed.
  • Mixing tests have unacceptably high false-negative rates due to the dilution effect; ISTH now recommends mixing test-specific cut-offs over the index of circulating anticoagulant (Rosner index), and BSH/CLSI permit deviation from the classical algorithm. Percentage correction cut-offs are typically 10–20%.

Methods

  • Narrative review of the analytical literature on LA detection, covering the three-step screen/mix/confirm medley, alternative assays, normalization approaches, cut-off statistics, interpretive algorithms, and testing in anticoagulated patients.
  • Discusses pre-analytical requirements (platelet count < 10 × 10⁹/L, double centrifugation), normalization denominators (NPP clotting time vs. reference interval mean clotting time), and statistical requirements for cut-offs (minimum 120 normal donor plasmas for statistically valid RIs; as few as 39 for a parametric estimate; 20–40 donors for transference validation, with ≤10% of results outside 95% reference limits permitting adoption of the manufacturer's RI).
  • Summarizes guideline positions from ISTH, BSH, and CLSI, and presents suggested testing hierarchies for VKA, DFXaI, DTI, and heparin anticoagulation (Figure 3).

Limitations

  • Stated in the paper: no reference plasmas or gold-standard LA-specific assays exist; reagent and analyser variability mean common RIs cannot be applied; mixing tests are not 100% effective and have analytical limitations including the unavoidable dilution effect; sourcing sufficient donors for accurate cut-off estimation is problematic; strategies to reduce or eliminate anticoagulant interference have limitations, and a negative result at each step of the assay hierarchy may not fully exclude LA.
  • Evident from the text: this is a narrative review without primary data or systematic search methodology, so its conclusions rest on the author's synthesis of cited literature; the author reports consultancy fees from Technoclone, a reagent manufacturer, which is a potential competing interest.

Model-written summary. Check it against the paper before relying on it.

Integrity signals

Registry

Crossref · 10.37349/ei.2023.00104

Cites

177 of 177 references verified0 citation errors · 0 not checkable

Statistics

Nothing to recompute.

Claims the paper makes

0 of 213 cited sources checked · 0 supported · 0 flagged

  • Premisereasoning

    LA screening tests employ dilute phospholipids to accentuate in vitro inhibition by LAs.

    “LA screening tests employ dilute phospholipids to accentuate in vitro inhibition by LAs, although they are not LA-specific and can be elevated by other coagulation abnormalities.”
    • Also stated in: § The three-step medley · plain
      “the most effective screening tests for LA employ dilute PL to increase competition for limited binding sites, which accentuates the inhibitory effect of any LA present”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites [22]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [23]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • Premisereasoning

    Converting clotting times to normalized ratios improves assay performance.

    “Converting clotting times to normalized ratios improves assay performance”
    • Also stated in: § From data to diagnosis · plain
      “The solution, which is a recommended practice in all guidelines, is the conversion of screen, mix, and confirm clotting times to normalized ratios [19][20][21].”
    Quote in the paperSubstantial claim0 of 3 sources checked
    • cites [19]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [20]

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    • cites [21]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • Premisecitation

    Sourcing sufficient donors for accurate cut-off estimations is problematic.

    “Sourcing sufficient donors for accurate cut-off estimations is problematic”
    • Also stated in: § Which statistical model for cut-off determination? · plain
      “Sourcing such large donor numbers is difficult for most diagnostic facilities”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [88]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • Premisereasoning

    No single screening test detects every LA.

    “LA heterogeneity means that no single screening test detects every LA”
    • Also stated in: § Introduction · plain
      “no single assay, or assay type, will detect every LA”
    Quote in the paperSubstantial claim0 of 3 sources checked
    • cites [7]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [14]

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    • cites [16]

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  • Interpretationreasoning

    The screen/mix/confirm medley must be applied to at least two assay types, usually dRVVT and an LA-sensitive aPTT.

    “the screen/mix/confirm medley must be applied to at least two assay types, usually dilute Russell's viper venom time (dRVVT) and an LA-sensitive activated partial thromboplastin time (aPTT)”
    Quote in the paperSubstantial claim0 of 3 sources checked
    • cites [7]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [14]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [16]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • May combine two claims. 33
  • Classified as probably not a substantial claim. 17

  • May combine two claims.

    Premisecitation

    dPT can perform with equal diagnostic efficacy to dRVVT/aPTT and can detect LA unreactive with dRVVT and aPTT.

    “others, such as dilute prothrombin time (dPT), can perform with equal diagnostic efficacy, and additionally detect LA unreactive with dRVVT and aPTT”
    • Also stated in: § The other LA assays · hedged
      “some studies have evidenced that LAs detected with dPTs employing recombinant thromboplastin exhibit a higher association with thrombosis than those detected by dRVVT and aPTT [58,62]”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites [58]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [62]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation

    Cut-offs can be generated parametrically from normally distributed data, or different percentiles applied depending on the preferred balance between sensitivity and specificity.

    “Cut-offs can be generated parametrically from normally distributed data, or different percentiles applied depending on the preferred balance between sensitivity and specificity.”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [19]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [88]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • Classified as probably not a substantial claim.

    Premisecitation

    LA detection is based on LA functional behavior in coagulation assays irrespective of epitope specificity.

    “Detection is based on LA functional behavior in coagulation assays irrespective of epitope specificity.”
    • Also stated in: § Introduction · plain
      “LAs are defined through functional properties irrespective of epitope specificities [4,14].”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites [4]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [14]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • May combine two claims.

    Premisereasoning

    LA screening tests are not LA-specific and can be elevated by other coagulation abnormalities.

    “although they are not LA-specific and can be elevated by other coagulation abnormalities”
    • Also stated in: § The three-step medley · plain
      “none are LA-specific”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premisecitation

    NPP clotting time denominators reflect on-the-day assay performance, whereas RI mean clotting time denominators negate the effects of NPP variation.

    “practitioners must choose between normal pooled plasma (NPP) clotting time denominators to reflect on-the-day assay performance, or reference interval (RI) mean clotting times to negate the effects of NPP variation”
    Quote in the paperSubstantial claim0 of 9 sources checked
    • cites [14]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [20]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [28]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [70]

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    • cites [75]

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    • cites [77]

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    • cites [80]

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    • cites [81]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [82]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • Probably not a substantial claim: may not pass S4 (based).

    Premisenone given

    Analytical limitations of mixing tests have led to adoption of alternative algorithms to the screen/mix/confirm test order.

    “Analytical limitations of mixing tests have led to the adoption of alternative algorithms to the screen/mix/confirm test order”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Premisecitation

    LA functional activity is mainly attributed to autoantibodies against β2GPI and prothrombin, each separately contributing to overall LA activity.

    “LA functional activity is mainly attributed to autoantibodies against β2GPI and prothrombin, each separately contributing to the overall LA activity [14,15].”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [14]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [15]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • Classified as probably not a substantial claim. May combine two claims.

    Premisecitation

    Plasma filtration through 0.22 μm filters is not recommended as it can generate microparticles and may reduce or remove some coagulation factors.

    “plasma filtration through 0.22 µm filters will remove all platelets, but is not recommended as it can also generate microparticles, and additionally, may reduce or remove some coagulation factors [19,20]”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites [19]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [20]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • May combine two claims.

    Premisecitation

    Platelet counts in plasma being tested for LA must be < 10 × 10^9/L, otherwise residual available PL may shorten the clotting time.

    “Platelet counts in plasma being tested for LA must be < 10 × 10 9 /L, otherwise the residual available PL may provide sufficient additional binding sites for coagulation factors to shorten the clotting time.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [27]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • Classified as probably not a substantial claim.

    Premisecitation

    Ultracentrifugation is not recommended because the severity of centrifugation conditions can generate PL-containing microparticles.

    “ultracentrifugation might appear a tempting alternative, it is not recommended because the severity of centrifugation conditions can generate PL-containing microparticles [19,29]”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites [19]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [29]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • Classified as probably not a substantial claim.

    Premisecitation

    LAs are detected by inference based on their behavior in a medley of coagulation assays.

    “LAs are detected by inference based on their behavior in a medley of coagulation assays [14,16]”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites [14]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [16]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • May combine two claims.

    Premisecitation

    The combination of elevated screening and mixing tests narrows the interpretation to that of an inhibitor, and the confirmatory test secures the distinction between factor-specific inhibition and factor-non-specific/PL-dependent inhibition of LAs.

    “The combination of elevated screening and mixing tests narrows the interpretation to that of an inhibitor, and it is the confirmatory test that then secures the distinction between factor-specific inhibition and the factor-non-specific/PL-dependent inhibition of LAs [14].”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [14]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • Probably not a substantial claim: may not pass S4 (based).

    Premisenone given

    Strategies to reduce or eliminate the effects of therapeutic anticoagulation have limitations.

    “Strategies to reduce or eliminate the effects of therapeutic anticoagulation have limitations”
    Quote in the paperNot a substantial claim

  • Classified as probably not a substantial claim.

    Premisecitation

    Double centrifugation is recommended practice to achieve the requisite platelet count reduction.

    “Recommended practice to achieve the requisite platelet count reduction is double centrifugation”
    Quote in the paperSubstantial claim0 of 3 sources checked
    • cites [19]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [20]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [21]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    Persistent isolated LA is reported as a high-risk profile in some recent studies.

    “persistent isolated LA is reported as a high-risk profile in some recent studies [11][12][13]”
    Quote in the paperNot a substantial claim0 of 3 sources checked
    • cites [11]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [12]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [13]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    APS is a systemic autoimmune disorder characterized by arterial or venous thrombosis and/or pregnancy morbidity in patients with persistent aPL.

    “Antiphospholipid syndrome (APS) is a systemic autoimmune disorder characterized by the non-specific clinical manifestations of arterial or venous thrombosis and/or pregnancy morbidity in patients with persistent antiphospholipid antibodies (aPL) [1].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [1]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • May combine two claims.

    Premisereasoning

    The primary roles of aPTT reagents designed for routine coagulation screening are detection of factor deficiencies and heparin monitoring, and the PL component is not necessarily optimized for LA detection.

    “The primary roles of aPTT reagents designed for routine coagulation screening are detection of factor deficiencies and heparin monitoring, and the PL component is not necessarily optimized for LA detection.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premisecitation

    Other factors contribute to variable reliability of mixing tests: reagent sensitivity and specificity, variability in NPP quality and suitability, and interpretive inconsistencies.

    “Principal amongst them is the unavoidable dilution effect that can lead to false-negatives, but other factors also contribute to variable reliability of mixing tests, such as reagent sensitivity and specificity, variability in NPP quality and suitability, (i.e. an NPP with a short clotting time requires more potent LA to prolong the result beyond the cut-off), and interpretive inconsistencies [20,”
    Quote in the paperSubstantial claim0 of 3 sources checked
    • cites [20]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [76]

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    • cites [96]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • May combine two claims.

    Interpretationcitation

    Because LA testing does not stop at the screening test, any false-positive screening test will be subjected to the rest of the three-step process, where a false-positive composite interpretation is less likely.

    “LA testing does not stop at the screening test, so any false-positive screening test will be subjected to the rest of the three-step process, where a falsepositive interpretation of the composite is less likely [14,20,35]”
    Quote in the paperSubstantial claim0 of 3 sources checked
    • cites [14]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [20]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [35]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • May combine two claims.

    Premisecitation

    Other assays have been shown to detect small LA sub-populations that do not manifest in dRVVT and aPTT, and can increase detection rates.

    “other assays have been shown to detect small LA sub-populations that do not manifest in dRVVT and aPTT, and can increase detection rates [43][44][45][46][47][48][49][50]”
    Quote in the paperSubstantial claim0 of 8 sources checked
    • cites [43]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [44]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [45]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [46]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [47]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [48]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [49]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [50]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • Probably not a substantial claim: may not pass S4 (based).

    Premisenone given

    Every intrinsic pathway-based LA assay is no more than a modified aPTT.

    “Every intrinsic pathway-based LA assay is no more than a modified aPTT”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Interpretationreasoning

    The low and high PL screen and confirm assay design remains effective for prothrombin-directed and LA-β2GPI antibodies because their inhibition occurs only at limiting PL concentrations, and not at high PL concentrations.

    “the low and high PL screen and confirm assay design remains effective for these antibodies because their inhibition occurs only at limiting PL concentrations, and not at high PL concentrations”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S2 (checkable).

    Interpretationreasoning

    LA testing is best done with an aPTT reagent specifically formulated for LA detection, even if it means having a separate reagent for routine coagulation screening.

    “LA testing is best done with an aPTT reagent specifically formulated for LA detection, even if it means having a separate reagent for routine coagulation screening.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Interpretationreasoning

    A factor deficiency or natural statistical outlier would generate an elevated screening test, yet the mixing test would be normal and the confirmatory test similarly prolonged, so a false-positive composite interpretation would not ensue.

    “a factor deficiency or natural statistical outlier would generate an elevated screening test result, yet the mixing test would be normal and the confirmatory test similarly prolonged, so a false-positive composite interpretation would not ensue”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premisecitation

    In situations with elevated confirmatory test, a mixing test can correct factor deficiencies and effects of some anticoagulants, and can make PL-dependence more obvious by diluting antibody titer.

    “In such situations, a mixing test can be invaluable as it will correct factor deficiencies and the effects of some anticoagulants, and can make PL-dependence more obvious by diluting the antibody titer [103,109].”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites [103]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [109]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • May combine two claims.

    Interpretationreasoning

    The iterative algorithm permits conclusion of LA if confirm ratio significantly corrects elevated screen ratio and confirm ratio is normal, with normal coagulation screening and no anticoagulation.

    “Given the analytical inconsistencies of mixing tests, the iterative algorithm permits the conclusion of the presence of an LA if the confirm ratio significantly corrects the elevated screen ratio and the confirm ratio is normal, the latter excluding other causes of elevated clotting times in conjunction with normal coagulation screening and confirmation that the patient is not anticoagulated.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationreasoning

    Performing a mixing test increases specificity, especially when other or co-existent abnormalities are suspected; it also increases diagnostic certainty in straightforward cases.

    “Performing a mixing test increases specificity, especially in situations where other or co-existent abnormalities are suspected, but they also increase diagnostic certainty and confidence in otherwise straightforward cases if desired.”
    Quote in the paperSubstantial claim

  • Classified as probably not a substantial claim.

    Premisecitation

    The ISTH guideline recommends performance of both mixing test and confirmatory test in response to elevated screening test instead of strict sequential testing.

    “In acknowledgment of the occurrence of false-negative mixing tests, the ISTH guideline recommends performance of both mixing test and confirmatory test in response to the finding of an elevated screening test in place of strict adherence to sequential testing [21].”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [21]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • May combine two claims.

    Interpretationcitation

    Integrated testing oversimplifies LA detection in some patients, particularly where there are co-existing abnormalities; there is no one-size-fits-all algorithmic solution to LA testing.

    “Whilst this is certainly true for some patients, integrated testing oversimplifies LA detection in others, particularly where there are co-existing abnormalities, emphasizing that there is not a one-size-fits-all algorithmic solution to LA testing [14,103].”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites [14]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [103]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • May combine two claims.

    Premisecitation

    In VKA anticoagulated patients, an elevated screen mixing test accompanied by significant discordance with a normal confirm mixing test is diagnostic for the presence of an LA, but negative mixing tests do not exclude an LA due to the dilution effect.

    “In VKA anticoagulated patients, an elevated screen mixing test accompanied by significant discordance with a normal confirm mixing test is diagnostic for the presence of an LA, but crucially, negative mixing tests do not exclude an LA due to the dilution effect [14,19,20,73].”
    Quote in the paperSubstantial claim0 of 3 sources checked
    • cites [14]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [19]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [20]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • Probably not a substantial claim: may not pass S2 (checkable).

    Premisecitation

    Attempting to detect LA in anticoagulated patients can be valuable for decisions regarding therapy duration and choice of type of anticoagulant in view of the aPL profile.

    “However, attempting to detect LA in anticoagulated patients can be valuable for decisions regarding therapy duration and choice of type of anticoagulant in view of the aPL profile [17,123,124].”
    Quote in the paperNot a substantial claim0 of 3 sources checked
    • cites [17]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [123]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [124]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • May combine two claims.

    Premisecitation

    Some potent LAs give concordant screen and confirm results on undiluted plasma; dilution in mixing test permits PL-dependence to manifest, and integrated testing returns a false-negative result.

    “Some LAs are so potent that testing on undiluted plasma gives concordant screen and confirm results, whereupon dilution of antibody potency in the mixing test permits PL-dependence to manifest, and integrated testing returns a false-negative result [109].”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [109]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • May combine two claims.

    Interpretationcitation

    Integrated testing improves detection of weaker antibodies but oversimplifies complex cases unless a mixing step is included.

    “Integrated testing immediately assesses for phospholipid dependence irrespective of whether the screening test is elevated or not, which improves the detection of weaker antibodies, but oversimplifies more complex cases unless a mixing step is included”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premisecitation

    An elevated confirmatory test in the scenario of elevated screening test can be due to a co-existing abnormality, such as a mild factor deficiency, undisclosed anticoagulation, or an LA sufficiently potent to resist high PL concentration confirm reagent.

    “An elevated confirmatory test in this scenario can be due to a co-existing abnormality, such as a mild factor deficiency missed in routine coagulation screening, undisclosed anticoagulation, or an LA sufficiently potent to partially or completely resist the overwhelming effect of a high PL concentration confirm reagent [20,31,103,108,109].”
    Quote in the paperSubstantial claim0 of 3 sources checked
    • cites [20]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [31]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [103]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • Classified as probably not a substantial claim.

    Premisecitation

    Testing for LA in anticoagulated patients is largely discouraged.

    “This complicates LA testing and increases the possibility of false positive and negative interpretations, and testing for LA in anticoagulated patients is largely discouraged [17,[19][20][21].”
    Quote in the paperSubstantial claim0 of 3 sources checked
    • cites [17]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [19]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [20]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • Classified as probably not a substantial claim.

    Premisecitation

    The most recently published expert guideline, from the ISTH, recommends mixing test-specific cut-off over ICA because it produces less false-negative mixing test results.

    “The most recently published expert guideline, from the ISTH, recommends mixing test-specific cut-off over ICA because it produces less false-negative mixing test results [21].”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [21]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • May combine two claims.

    Premisecitation

    Applying integrated testing with a higher normalized LA ratio cut-off than for non-anticoagulated patients improves specificity but reduces sensitivity.

    “Applying integrated testing with a higher normalized LA ratio cut-off than for non-anticoagulated patients has been shown to improve specificity, but with a reduction in sensitivity [130,131].”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites [130]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [131]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • May combine two claims.

    Interpretationreasoning

    Mixing tests can be invaluable where other abnormalities are present, but a hindrance when design limitations of the mixing test principle give misleading information.

    “There are variations on how to apply the composite testing model, particularly regarding the place of mixing tests, which can be invaluable where other abnormalities are present, but a hindrance when design limitations of the mixing test principle give misleading information.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Premisecitation

    Increased C-reactive protein can interfere with aPTT-based assays, giving rise to false-positive results, whilst elevated FVIII and fibrinogen can shorten aPTT, giving rise to the potential for false-negatives.

    “The increased C-reactive protein can interfere with aPTT-based assays, giving rise to false-positive results [162], whilst the elevated FVIII and fibrinogen [20,21,163,164] can shorten aPTT, giving rise to the potential for false-negatives.”
    Quote in the paperSubstantial claim0 of 3 sources checked
    • cites [162]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [20]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [21]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • May combine two claims.

    Interpretationreasoning

    Applying composite testing to each assay type improves specificity but interpretation can be challenging when trying to unravel the possible presence of an LA where there are other abnormalities that may mask, mimic, or co-exist.

    “Applying composite testing to each assay type improves specificity but interpretation can be challenging when trying to unravel the possible presence of an LA where there are other abnormalities that may mask, mimic, or co-exist.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premisecitation

    If heparin is the only cause of an elevated clotting time, screen and confirm results are normally concordant and the mixing test may or may not be elevated depending on dose and reagent heparin sensitivity, and interpretation of the composite would not suggest the presence of an LA.

    “If heparin is the only cause of an elevated clotting time, screen and confirm results are normally concordant and the mixing test may or may not be elevated depending on dose and reagent heparin sensitivity, and interpretation of the composite would not suggest the presence of an LA [134].”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [134]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • May combine two claims.

    Premisecitation

    If the screen is elevated but the confirm is normal, and discordance significant, the normal confirm result evidences quenching of the heparin by the neutraliser, and of the LA by the confirm reagent.

    “If the screen is elevated but the confirm is normal, and discordance significant, the normal confirm result evidences quenching of the heparin by the neutraliser, and of the LA by the confirm reagent [14,16,20].”
    Quote in the paperSubstantial claim0 of 3 sources checked
    • cites [14]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [16]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [20]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • May combine two claims.

    Interpretationreasoning

    Antibody heterogeneity and reagent variability affect all assays, including the favored dRVVT and aPTT pairing, yet overreliance on two familiar and well-performing assays solves some problems but ignores others.

    “Antibody heterogeneity and reagent variability affect all assays, including the favored dRVVT and aPTT pairing, yet overreliance on two familiar and well-performing assays solves some problems but ignores others.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premisecitation

    Activated charcoal adsorption products are reported to be effective in facilitating reliable dRVVT and aPTT analysis, but less beneficial for dPT.

    “They are reported to be effective in facilitating reliable dRVVT and aPTT analysis [149-153], but less beneficial for dPT [154].”
    Quote in the paperSubstantial claim0 of 6 sources checked
    • cites [149]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [150]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [151]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [152]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [153]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [154]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • May combine two claims.

    Premisecitation

    Transient LAs in response to infections are well described, whilst infections are reported as a common precipitating factor of catastrophic APS in patients with existing APS.

    “Transient LAs in response to infections are well described [169-171], whilst infections are reported as a common precipitating factor of catastrophic APS in patients with existing APS [172].”
    Quote in the paperNot a substantial claim0 of 4 sources checked
    • cites [169]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [170]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [171]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [172]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • Classified as probably not a substantial claim. May combine two claims.

    Premisecitation

    Recent recommendations state that LA testing on patients receiving DOAC should not be done unless an adsorption procedure has been performed, and that the adsorption should only be done on samples from patients known to be DOAC anticoagulated.

    “Consequently, the recent recommendations state that LA testing on patients receiving DOAC should not be done unless an adsorption procedure has been performed, and that the adsorption should only be done on samples from patients known to be DOAC anticoagulated [17,155].”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites [17]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [155]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • Rests on an assumption: DOACs and parenteral DTIs are unaffected by heparin neutralizers.

  • May combine two claims.

    Premisecitation

    False-positive interpretations due to greater elevation of screen results compared to confirm results have been reported for dRVVT and aPTT-based assays with DTI and DFXaI, and for TSVT/ET with DTI.

    “False-positive interpretations due to greater elevation of screen results compared to confirm results have been reported for dRVVT and aPTT-based assays with DTI and DFXaI [116, 137, 138, 141, 142], and for TSVT/ET with DTI [70].”
    Quote in the paperSubstantial claim0 of 3 sources checked
    • cites [116]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [137]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites [138]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

47 of 47

  • The authors state this as an assumption.

    When interpreting coagulation assays for LA, no other causes of abnormal clotting times are present.

    “interpretation is necessarily based on the assumption that no other causes of abnormal clotting times are present, which is not always the case”

  • The paper relies on this without support.

    Normal pooled plasma replenishes deficient clotting factors and corrects factor deficiencies in mixing tests.

    “The theory behind such mixing tests is that the NPP will replenish any deficient clotting factors such that the mixing test result corrects back to normality”

  • The paper relies on this without support.

    DOACs and parenteral DTIs are unaffected by heparin neutralizers.

    “The DOACs (dabigatran and the three DFXaIs) and parenteral DTI are unaffected by heparin neutralizers.”

  • The paper relies on this without support.

    Accurate LA detection is crucial to effective diagnosis and management of APS.

    “Accurate LA detection is crucial to effective diagnosis and management of APS but it remains hampered by the lack of specific assays.”
4 of 4

Cites

Flagged first, then by citations

Research works printed but not linked: 4

  1. Moore GW. Clinical utility of the less commonly employed assays for lupus anticoagulant detection: the evidence. J Coag Dis. 2010;2:69–79.
  2. Goldford MD. Lupus anticoagulant screen and confirm reagents utilizing the FX activating venom from Vipera lebetina. J Thromb Haemost. 2013;11:61.
  3. Jouhikainen T. Detection of lupus anticoagulant by means of dilute Russell’s viper venom time is affected by oral anticoagulant therapy. Blood Coagul Fibrinolysis. 1990;1:627–32.
  4. Dunois C, Peyrafitte M, De Muynck S, Villedieu V, Amiral J. New dRVVT assays for lupus anticoagulant (LA) detection with improved specificity and sensitivity: HEMOCLOT LA-S and LA-C. Res Pract Thromb Haemost. 2018;2:207.

Cited by

Sorted by citations

Related

Authors
Moore, G. W.
Affiliations
Cambridge University Hospitals NHS Foundation Trust (GB)
Corresponding author
Moore, Gary W.
ORCID
Moore, Gary W.
Venue
Exploration of Immunology
Publisher
Open Exploration Publishing
Published
2023-08-31
Pages
300-324
ISSN
2768-6655 (ISSN-L)
DOI
10.37349/ei.2023.00104
OpenAlex
W4386331100
Cited by
5
References
169
Citations by year
2026: 5
FWCI
0.86
Type
journal article
Language
en
Open access
Licence
Locations
  • Exploration of Immunology (publishedVersion, cc-by)
  • DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion, cc-by-sa)
Topics
  • Health Sciences › Medicine › Rheumatology › Systemic Lupus Erythematosus Research
  • Health Sciences › Medicine › Hematology › Blood Coagulation and Thrombosis Mechanisms
  • Health Sciences › Medicine › Physiology › Clinical Laboratory Practices and Quality Control
Keywords
lupus anticoagulant, antiphospholipid syndrome, dilute Russell's viper venom time, activated partial thromboplastin time, mixing test, factor deficiency, vitamin K antagonists

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