Thrombophilia Screening: Not So Straightforward.

Although inherited thrombophilias are lifelong risk factors for a first thrombotic episode, progression to thrombosis is multifactorial and not all individuals with inherited thrombophilia develop thrombosis in their lifetimes. Consequently, indiscriminate screening in patients with idiopathic thrombosis is not recommended, since presence of a thrombophilia does not necessarily predict recurrence or influence management, and testing should be selective. It follows that a decision to undertake laboratory detection of thrombophilia should be aligned with a concerted effort to identify any significant abnormalities, because it will inform patient management. Deficiencies of antithrombin and protein C are rare and usually determined using phenotypic assays assessing biological activities, whereas protein S deficiency (also rare) is commonly detected with antigenic assays for the free form of protein S since available activity assays are considered to lack specificity. In each case, no single phenotypic assay is capable of detecting every deficiency, because the various mutations express different molecular characteristics, rendering thrombophilia screening repertoires employing one assay per potential deficiency, of limited effectiveness. Activated protein C resistance (APCR) is more common than discrete deficiencies of antithrombin, protein C, and protein S and also often detected initially with phenotypic assays; however, some centres perform only genetic analysis for factor V Leiden, as this is responsible for most cases of hereditary APCR, accepting that acquired APCR and rare F5 mutations conferring APCR will go undetected if only factor V Leiden is evaluated. All phenotypic assays have interferences and limitations, which must be factored into decisions about if, and when, to test, and be given consideration in the laboratory during assay performance and interpretation. This review looks in detail at performance and limitations of routine phenotypic thrombophilia assays.

Towards harmonized interpretation of anticardiolipin and anti-ß2-glycoprotein I antibody detection for diagnosis of antiphospholipid syndrome using defined level intervals and likelihood ratios: Communication from the ISTH SSC Subcommittee on Lupus Anticoagulant/Antiphospholipid Antibodies.

BACKGROUND: Improving harmonization of the clinical interpretation of anticardiolipin (aCL) and anti-ß2-glycoprotein I antibodies (aß2GPI) IgG/IgM in the diagnosis of antiphospholipid syndrome (APS) is desirable. Likelihood ratios (LR) with corresponding test-result intervals can identify the power of a test to discriminate between a diseased and non-diseased patient and may be useful for the semiquantitative interpretation of aCL/aß2GPI results. OBJECTIVES: To determine moderate and high thresholds for aCL and aß2GPI IgG/IgM determined with chemiluminescent immunoassay, enzyme-linked immunosorbent assay, fluorescence enzyme immunoassay, and multiplex flow immunoassay. PATIENTS/METHODS: aCL and aß2GPI IgG/IgM were determined with four solid-phase systems in a case-control study population including 381 APS patients and 727 controls. Interval-specific LR (IS-LR) were determined for ranges determined by prespecified specificity and sensitivity levels. Three methods were used for determining thresholds that separated low, moderate, and high positive antibody levels. Inter-assay agreement was checked with Cohen's kappa statistics. RESULTS: Assay- and antibody-specific thresholds demonstrated increasing IS-LR, reflecting different clinical significance for low, moderate, and high levels, especially for IgG aCL and aß2GPI and in thrombotic APS. IS-LR per antibody and unit range were comparable across solid-phase platforms resulting in enhanced harmonization of result interpretation. Agreement between assays for identifying high levels improved by semiquantitative interpretation compared to quantitative reporting. CONCLUSIONS: aCL and aß2GPI IgG/IgM moderate and high thresholds were determined for four analytical platforms. Thresholds improve harmonized interpretation of aCL/aß2GPI levels across platforms. The proposed thresholds should be verified in an independent case-control study to check interlaboratory transferability.

Taipan snake venom time has high sensitivity for lupus anticoagulants in non-anticoagulated, triple positive antiphospholipid syndrome patients.

INTRODUCTION: Dilute Russell's viper venom time (dRVVT) and activated partial thromboplastin time (APTT) are the mainstay assays in lupus anticoagulant (LA) detection yet they have limitations, particularly in relation to interferences and specificity. The recently validated Taipan snake venom time (TSVT) screening with ecarin time (ET) confirmatory assays overcome many of those limitations due to the innate specificity engendered from direct prothrombin activation, and insensitivity to the effects of vitamin K antagonists (VKA). The present study aimed to further evidence diagnostic utility of TSVT/ET by performing them in samples from 116 nonanticoagulated patients with established triple-positive antiphospholipid syndrome (APS). METHODS: Samples were identified in three expert centres who performed dRVVT, APTT and solid phase antiphospholipid antibody assays with reagents from a variety of manufacturers. All samples additionally received TSVT/ET analysis using standardised reagents. RESULTS: Ninety seven of 116 (83.6%) were dRVVT- and APTT-positive, 85/97 (87.6%) of which were TSVT/ET-positive, 9/116 (7.8%) were dRVVT-positive only, 6 of which were TSVT/ET-positive, and 10/116 (8.6%) were APTT-positive only, 5 of which were TSVT/ET-positive. 96/116 TSVT/ET-positivity returned a high sensitivity for LA of 82.8%. Low coefficients of determination revealed weak relationships between LA potency and anticardiolipin and anti-ß2-glycoprotein I antibody titres for all three LA assays. CONCLUSIONS: TSVT/ET has high sensitivity for the clinically significant LA found in triple positive APS patients. TSVT/ET can establish multiple LA assay positivity in nonanticoagulated patients negative for one of dRVVT or APTT, and is the only assay pairing insensitive to VKAs, the recommended anticoagulation for APS.

International Council for Standardization in Haematology Field Study Evaluating Optimal Interpretation Methods for Activated Partial Thromboplastin Time and Prothrombin Time Mixing Studies.

CONTEXT.­: The prothrombin time (PT) and activated partial thromboplastin time (APTT) are screening tests used to detect congenital or acquired bleeding disorders. An unexpected PT and/or APTT prolongation is often evaluated using a mixing test with normal plasma. Failure to correct ("noncorrection") prolongation upon mixing is attributed to an inhibitor, whereas "correction" points to factor deficiency(ies). OBJECTIVE.­: To define an optimal method for determining correction or noncorrection of plasma mixing tests through an international, multisite study that used multiple PT and APTT reagents and well-characterized plasma samples. DESIGN.­: Each testing site was provided 22 abnormal and 25 normal donor plasma samples, and mixing studies were performed using local PT and APTT reagents. Mixing study results were evaluated using 11 different calculation methods to assess the optimal method based on the expected interpretation for factor deficiencies (correction) and noncorrection (inhibitor effect). Misprediction, which represents the failure of a mixing study interpretation method, was assessed. RESULTS.­: Percentage correction was the most suitable calculation method for interpreting PT mixing test results for nearly all reagents evaluated. Incubated PT mixing tests should not be performed. For APTT mixing tests, percentage correction should be performed, and if the result indicates a factor deficiency, this should be confirmed with the subtraction III calculation where the normal pooled plasma result (run concurrently) is subtracted from the mixing test result with correction indicated by a result of 0 or less. In general, other calculation methods evaluated that performed well in the identification of factor deficiency tended to have high misprediction rates for inhibitors and vice versa. CONCLUSIONS.­: No single method of mixing test result calculation was consistently successful in accurately distinguishing factor deficiencies from inhibitors, with between-reagent and between-site variability also identified.

Triple-positive antiphospholipid syndrome does not guarantee positivity in each lupus anticoagulant assay.

BACKGROUND: Triple positivity for all 3 criteria antiphospholipid antibodies confers high risk of symptom development in carriers, and recurrence in antiphospholipid syndrome (APS). Most triple-positivity studies report lupus anticoagulant (LA) testing as positive without distinguishing between positivity with dilute Russell's viper venom time (dRVVT) and activated partial thromboplastin time (APTT) and single-assay positivity or only perform dRVVT. Single LA assay repertoires remain in use in some centers, which risks missing some triple positives. Positivity with both assays may identify higher risk. OBJECTIVES: The aim of this study is to investigate the frequency of single LA assay positivity in triple-positive patients. METHODS: Three hundred forty-two triple-positive profiles from nonanticoagulated patients (237 APS, 45 systemic lupus erythematosus without APS symptoms, and 60 nonclinical criteria) were identified from laboratory databases and assessed for LA positivity by dRVVT and/or APTT. RESULTS: Seventy-three of 237 (30.8%) APS samples were LA-positive with 1 assay, 40/237 (16.9%) by dRVVT only, and 33/237 (13.9%) with APTT only. Nineteen of 45 (42.2%) were LA-positive with 1 assay in the systemic lupus erythematosus cohort; 12/45 (26.7%) with dRVVT only and 7/45 (15.5%) with APTT only. Thirty-three of 60 (55.0%) were LA-positive with 1 assay in the nonclinical criteria cohort; 24/60 (40.0%) with dRVVT only and 9/60 (15.0%) with APTT only. The most common solid-phase assay profile was elevated immunoglobulin G aCL and aß2GPI. CONCLUSION: Up to 55.0% of triple-positive samples were positive in 1 LA assay, representing significant potential for misdiagnosis and inappropriate management via single LA assay repertoires.

Lupus Anticoagulant Testing: Taipan Snake Venom Time with Ecarin Time as Confirmatory Test.

Testing for lupus anticoagulants (LA) in the presence of therapeutic anticoagulation is largely discouraged because of the risk of false-positive and false-negative results, although the ability to detect LA in this setting can be clinically valuable. Strategies such as mixing tests and anticoagulant neutralization can be effective, but have their own limitations. The prothrombin activators in venoms from Coastal Taipan and Indian saw-scaled viper snakes provide an additional analytical avenue in that they are insensitive to the effects of vitamin K antagonists and inevitably bypass the effects of direct factor Xa inhibitors. Oscutarin C in Coastal Taipan venom is phospholipid- and Ca2+-dependent, so the venom is used in a dilute phospholipid design as an LA screening test called the Taipan snake venom time (TSVT). The ecarin fraction of Indian saw-scaled viper venom is cofactor-independent and operates as a prothrombin-activated confirmatory test, the ecarin time, because the absent phospholipid precludes inhibition by LAs. Bypassing all coagulation factors except prothrombin and fibrinogen renders the assays innately more specific than other LA assays, while TSVT as a screening test has good sensitivity to LAs detected in other assays, as well as occasional antibodies unreactive in other assays.

Lupus Anticoagulant Testing: Dilute Prothrombin Time (dPT).

Lupus anticoagulants (LA) rarely affect routine prothrombin time assays because the high phospholipid (PL) content in thromboplastin reagents tends to overwhelm the antibodies. Dilution of thromboplastin to create a dilute prothrombin time (dPT) screening test renders the assay sensitive to the presence of LA. Technical and diagnostic performances are enhanced if recombinant thromboplastins are employed in place of tissue-derived reagents. Presence of an LA cannot be deduced from an elevated screening test alone since other coagulation disturbances can prolong clotting times. Confirmatory testing with less dilute or undiluted thromboplastin reveals the PL-dependent nature of LA by reducing the clotting time relative to that of the screening test. Where appropriate, such as a known or suspected coagulation factor deficiency, mixing tests are valuable in correcting factor deficiencies and evidencing inhibitory properties of LA, to increase diagnostic specificity. Although LA testing is commonly restricted to dilute Russell's viper venom time and activated partial thromboplastin time, dPT is sensitive to LA unreactive in those assays, and inclusion in routine testing increases detection rates of clinically significant antibodies.

ADAMTS13 Activity: Screening Test Protocol.

Accurate estimation of ADAMTS13 (a disintegrin-like and metalloprotease with thrombospondin type 1 motif, member 13) activity level is necessary for diagnosis and management of thrombotic microangiopathies (TMA). In particular, it permits distinction between thrombotic thrombocytopenic purpura (TTP) and other TMAs, prompting disorder appropriate treatment. Manual and automated quantitative assays of ADAMTS13 activity are commercially available, some providing results within less than an hour, but they require specialist equipment and personnel and tend to only be available in specialized diagnostic facilities. Technoscreen ADAMTS13 Activity is a rapid, commercially available, semiquantitative screening test employing flow-through technology and an ELISA activity assay principle. It is a simple to perform screening tool, not requiring specialist equipment or personnel. The colored end point is compared to a reference color chart containing four color intensity indicators corresponding to ADAMTS13 activity levels of 0, 0.1, 0.4, or 0.8 IU/mL. Reduced levels detected in the screening test should be confirmed by quantitative assay. The assay lends itself to use in nonspecialized laboratories, remote, and point-of-care settings.

ADAMTS13 Activity Measurement by ELISA and Fluorescence Resonance Energy Transfer Assay.

Accurate estimation of ADAMTS13 (a disintegrin-like and metalloprotease with thrombospondin type 1 motif, member 13) activity level is crucial in the diagnostic setting of differentiation between thrombotic thrombocytopenic purpura (TTP) and other thrombotic microangiopathies. The original assays were too cumbersome and time-consuming for use in the acute situation, and treatment was often based on clinical findings alone, with confirmatory laboratory assays following days or weeks later. Rapid assays are now available that can generate results fast enough to impact on immediate diagnosis and management. Assays based on fluorescence resonance energy transfer (FRET) or chemiluminescence principles can generate results in less than an hour, although they require specific analytical platforms. Enzyme-linked immunosorbent assays (ELISA) can generate results in about 4 h, but do not require specialized equipment beyond ELISA plate readers that are in regular use in many laboratories. The present chapter describes principles, performance, and practical aspects of an ELISA and a FRET assay, for quantitative measurement of ADAMTS13 activity in plasma.

ADAMTS13 Antibody and Inhibitor Assays.

A finding of an ADAMTS13 (a disintegrin-like and metalloprotease with thrombospondin type 1 motif, member 13) activity level of <10% of normal is usually sufficient to distinguish thrombotic thrombocytopenic purpura (TTP) from other thrombotic microangiopathies. TTP can be congenital or acquired, the most common form being acquired immune-mediated TTP caused by autoantibodies than inhibit ADAMTS13 function and/or increase its clearance. Basic 1 + 1 mixing tests can detect the presence of inhibitory antibodies, and quantification can be achieved with Bethesda-type assays that measure loss of function in a series of mixtures of test plasma and normal plasma. Not all patients present with inhibitory antibodies, and here the ADAMTS13 deficiency may be caused by clearing antibodies alone, which are not detectable in functional assays. ELISA assays are commonly used to detect clearing antibodies via capture with recombinant ADAMTS13. Since they also detect inhibitory antibodies, they are the preferred assay, although they cannot distinguish between inhibitory and clearing antibodies. The present chapter describes principles, performance, and practical aspects of a commercial ADAMTS13 antibody ELISA and a generic approach to Bethesda-type assays for detecting inhibitory ADAMTS13 antibodies.

Added value of antiphosphatidylserine/prothrombin antibodies in the workup of obstetric antiphospholipid syndrome: communication from the ISTH SSC Subcommittee on Lupus Anticoagulant/Antiphospholipid Antibodies.

BACKGROUND: The added value of antiphosphatidylserine/prothrombin antibodies (aPS/PT) in the diagnostic workup of antiphospholipid syndrome (APS) is unclear. Currently, diagnosis of thrombotic APS (TAPS) and obstetric APS (OAPS) requires persistent presence of lupus anticoagulant (LAC), anticardiolipin (aCL) immunoglobulin (Ig) G/IgM, or anti-ß2-glycoprotein I (aß2GPI) IgG/IgM antibodies. OBJECTIVES: To evaluate the role of aPS/PT IgG and IgM in OAPS. METHODS: aPS/PT IgG/IgM, aCL IgG/IgM, aß2GPI IgG/IgM, and LAC were determined in 653 patients (OAPS, TAPS, and controls). In-house aPS/PT cut-off values were calculated, titers and prevalence were compared between OAPS, TAPS, and controls and type of pregnancy morbidity. Sensitivity, specificity, likelihood ratios, and odds ratios (OR) with 95% CI were calculated. RESULTS: In OAPS, aPS/PT IgG and IgM showed an OR of 4.32 (95% CI, 2.54-7.36) and 3.37 (95% CI, 1.93-5.89), respectively, but the association was not independent of LAC. Prevalence and titers of aPS/PT IgG and IgM were lower in OAPS than in patients with TAPS. aPS/PT were more prevalent and showed higher titers in patients with late pregnancy loss than in patients with early pregnancy loss with a positivity of 86.4% and 39.3%, respectively. Higher aPS/PT titers did not increase the likelihood of having OAPS. CONCLUSION: The added value of aPS/PT testing in the current diagnostic workup of OAPS seems limited compared with LAC, aCL, and aß2GPI. aPS/PT might be useful in specific subsets of patients with OAPS. However, future multicentric studies are needed to elucidate the risk of less frequent and most severe obstetrical manifestations.

Factor V Leiden-independent activated protein C resistance: Communication from the plasma coagulation inhibitors subcommittee of the International Society on Thrombosis and Haemostasis Scientific and Standardisation Committee.

Activated protein C resistance (APC-R) due to the single-nucleotide polymorphism factor V Leiden (FVL) is the most common cause of hereditary thrombophilia. It is found predominantly in Caucasians and is uncommon or absent in other populations. Although FVL is responsible for >90% of cases of hereditary APC-R, a number of other F5 variants that also confer various degrees of APC-R and thrombotic risk have been described. Acquired APC-R due to increased levels of coagulation factors, reduced levels of inhibitors, or the presence of autoantibodies occurs in a variety of conditions and is an independent risk factor for thrombosis. It is common for thrombophilia screening protocols to restrict assessment for APC-R to demonstrating the presence or absence of FVL. The aim of this Scientific and Standardisation Committee communication is to detail the causes of FVL-independent APC-R to widen the diagnostic net, particularly in situations in which in vitro APC-R is encountered in the absence of FVL. Predilution clotting assays are not FVL specific and are used to detect clinically significant F5 variants conferring APC-R, whereas different forms of acquired APC-R are preferentially detected using the classical activated partial thromboplastin time-based APC-R assay without predilution and/or endogenous thrombin potential APC-R assays. Resource-specific recommendations are given to guide the detection of FVL-independent APC-R.

Activated Partial Thromboplastin Time and Prothrombin Time Mixing Studies: Current State of the Art.

Mixing studies have long been in the clinical laboratory armamentarium for investigating unexpected, prolonged activated partial thromboplastin time (aPTT) or prothrombin time (PT). The purpose of the mixing study is to identify whether the aPTT/PT prolongation is secondary to a factor deficiency versus an inhibitor, which would present as a "corrected" and "noncorrected" mixing study, respectively. The differentiation between a factor deficiency and inhibitor may likely further direct clinical decisions, including additional diagnostic testing or factor replacement therapy. While aPTT/PT mixing studies are simple tests to perform, there is a lack of standardization for both the testing protocol and the interpretation of what is considered to be a corrected or noncorrected mixing study result. This review will describe the common indications for the mixing test, preanalytic variables that may affect mixing study performance, and describe several methods for interpreting the results of aPTT and PT mixing tests.

Added value of antiphosphatidylserine/prothrombin antibodies in the workup of thrombotic antiphospholipid syndrome: Communication from the ISTH SSC Subcommittee on Lupus Anticoagulant/Antiphospholipid Antibodies.

BACKGROUND: Diagnosis of antiphospholipid syndrome (APS) requires persistent presence of lupus anticoagulant (LAC), anticardiolipin (aCL) IgG/IgM, or anti-ß2 glycoprotein I (aß2GPI) IgG/IgM antibodies. Other antiphospholipid antibodies (aPL) such as antiphosphatidylserine/prothrombin antibodies (aPS/PT) are promising in assessment of thrombotic APS (TAPS). AIM: To evaluate the added value of aPS/PT IgG and IgM in TAPS. MATERIAL AND METHODS: aPS/PT IgG/IgM, aCL IgG/IgM, aß2GPI IgG/IgM, and LAC were determined in 757 patients (TAPS and controls). aPS/PT cut-off values were calculated, and aPS/PT titers and positivity were compared between TAPS and controls, type of thrombosis, and antibody profiles. Likelihood ratios (LR), odds ratios (OR), and aPL score were determined. RESULTS: aPS/PT IgG and IgM were associated with TAPS and triple positivity. In-house calculated cut-offs were higher for IgM (43 units), compared to manufacturer's cut-off (30 units). Thresholds of 90 (IgG) and 200 (IgM) units were determined as high-titer cut-off. Higher aPS/PT titers were observed in triple positive patients and showed higher LR and OR for TAPS. aPS/PT was independently associated with TAPS when adjusted for aCL/aß2GPI, but not when adjusted for LAC. In isolated LAC positive patients, aPS/PT was positive in 27.1% TAPS patients and in 77.3% patients with autoimmune disease. Diagnostic value of aPL score did not differ with and without including aPS/PT. CONCLUSION: aPS/PT positivity, especially with high antibody titer, is associated with TAPS diagnosis. Analysis on top of current laboratory criteria is not essential in TAPS diagnosis, but aPS/PT could be useful in patients with thrombosis and a double positive aPL profile (aCL+/aß2GPI+).

Testing for Lupus Anticoagulants.

Lupus anticoagulant (LA) is one of the three criteria antiphospholipid antibodies (aPLs) employed in classification, and by default diagnosis, of antiphospholipid syndrome (APS). Detection of LA is not via calibrated assays but is based on functional behavior of the antibodies in a medley of coagulation assays. A prolonged clotting time in a screening test is followed by demonstration of phospholipid dependence and inhibitory properties in confirmatory and mixing tests, respectively, which are modifications of the parent screening test. Complications arise because no single screening test is sensitive to every LA, and no test is specific for LA, because they are prone to interference by other causes of elevated clotting times. Several screening tests are available but the pairing of dilute Russell's viper venom time (dRVVT) with LA-sensitive activated partial thromboplastin time (aPTT) is widely used and recommended because it is proven to have good detection rates. Nonetheless, judicious use of other assays can improve diagnostic performance, such as dilute prothrombin time to find LA unreactive with dRVVT and aPTT, and the recently validated Taipan snake venom time with ecarin time confirmatory test that are unaffected by vitamin K antagonist and direct factor Xa inhibitor anticoagulation. Expert body guidelines and their updates have improved harmonization of laboratory practices, although some issues continue to attract debate, such as the place of mixing tests in the medley hierarchy, and areas of data manipulation such as assay cut-offs and ratio generation. This article reviews current practices and challenges in the laboratory detection of LA.

Statewide trends and factors associated with genetic testing for hereditary cancer risk in Arkansas 2013-2018.

BACKGROUND: Early identification of hereditary cancer risk would save lives, but genetic testing (GT) has been inadequate. We assessed i) trends for hereditary breast and ovarian cancer (HBOC), Lynch syndrome, and other GT and ii) factors associated with receipt of GT. METHODS: We used data from the Arkansas All-Payer Claims Database from January 2013 through June 2018 (commercial, Medicaid), December 2017 (state employee), or December 2016 (Medicare) and identified enrollees with ≥1 month of enrollment. Using Current Procedural Terminology (CPT-4) codes, rates for GT were calculated per 100,000 person-quarters and time series regressions estimated. Second, GT and covariate information for enrollees with 24 months of continuous enrollment were used to estimate separate logistic regression models for each GT category. RESULTS: Among 2,520,575 unique enrollees, HBOC testing rates were 2.2 (Medicaid), 22.0 (commercial), 40.4 (state employee), and 13.1(Medicare) per 100,000 person-quarters and increased linearly across all plans. Older age (OR=1.24; 95%CI 1.20 - 1.28), female sex (OR=18.91; 95%CI 13.01 - 28.86), higher comorbidity burden (OR=1.08; 95%CI 1.05 - 1.12), mental disorders (OR=1.53; 95%CI 1.15 - 2.00), and state employee coverage (OR=1.65; 95%CI 1.37 - 1.97) were positively associated with HBOC testing. Less than 1 of 10,000 enrollees received Lynch syndrome testing, while < 5 of 10,000 received HBOC testing. CONCLUSION: GT rates for hereditary cancer syndromes have increased in Arkansas but remain low. Receipt of GT was explained with high discrimination by sex and plan type. IMPACT: Expansion of GT for hereditary cancer risk in Arkansas is needed to identify high-risk individuals who could benefit from risk-reduction strategies.

Effects of regional citrate anticoagulation on thrombin generation, fibrinolysis and platelet function in critically ill patients receiving continuous renal replacement therapy for acute kidney injury: a prospective study.

BACKGROUND: Regional citrate anticoagulation (RCA) is recommended for continuous renal replacement therapy (CRRT). However, filter life varies and premature filter clotting can occur. The aims of this explorative prospective study were to investigate the effects of RCA on thrombin generation, fibrinolysis and platelet function in critically ill patients receiving CRRT, to compare clotting parameters between systemic and intra-circuit blood samples, and to screen participants for coagulation disorders. We recruited critically ill adult patients admitted to a 30-bedded Intensive care unit in a tertiary care hospital who required CRRT with RCA for acute kidney injury (AKI). Patients with pre-existing thrombotic, bleeding tendencies or a CRRT duration less than 48 h were excluded. We measured coagulation and thrombophilia parameters at baseline. Thrombin generation, D-dimer and platelet function were measured pre-CRRT and at 12, 24, 36, 48 and 72 h after commencing CRRT using blood samples taken from the arterial line and the circuit. RESULTS: At baseline, all eleven patients (mean age 62.4 years, 82% male) had Factor VIII and von Willebrand Factor concentrations above reference range and significantly increased peak thrombin generation. During CRRT, there were no significant changes in systemic maximum peak thrombin generation, time to peak thrombin generation, fibrinogen, D-dimer and platelet function analysis. We observed no significant difference between paired samples taken from the patient's arterial line and the circuit. CONCLUSIONS: Critically ill patients with AKI requiring CRRT are hypercoagulable. Citrate used for anticoagulation during CRRT does not affect thrombin generation, D-dimer or platelet function. Systemic clotting parameters reflect intra-circuit results. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT02486614. Registered 01 July 2015-Registered after recruitment of first patient. https://clinicaltrials.gov/ct2/show/NCT02486614.

Direct oral anticoagulants-Remove versus Taipan snake venom time for detection of a lupus anticoagulant in patients taking oral direct factor Xa inhibitors.

BACKGROUND: The optimal method of detecting a lupus anticoagulant (LA) for patients taking direct factor Xa inhibitor (DFXaI) direct oral anticoagulants (DOACs) remains controversial. Methods include charcoal adsorption of the DOACs to allow testing with the activated partial thromboplastin time (APTT) and dilute Russell viper venom time (dRVVT), or use of the DFXaI-insensitive Taipan snake venom time (TSVT) and Ecarin time (ET) assays on neat plasma. OBJECTIVES: The objective was to compare the utility of APTT and dRVVT analysis following DOAC Remove against TSVT/ET on untreated plasma for LA detection in spiked plasmas and routine clinical samples for patients on DFXaIs. PATIENTS/METHODS: Various LA-negative and LA-positive samples were assayed by APTT, dRVVT, and TSVT/ET, and then separately spiked with rivaroxaban, apixaban, and edoxaban calibrators to a concentration of ~190 ng/ml and the assays repeated on spiked plasma before and after DOAC Remove treatment. Testing of 284 consecutive samples from DFXaI-anticoagulated patients by APTT/dRVVT and TSVT/ET before and after DOAC Remove treatment was undertaken. RESULTS: In the spiking model, we found that both TSVT/ET and DOAC Remove strategies generally distinguished LA-negative and LA-positive samples, but some false-positive LA results occurred. In the investigation of 284 consecutive patient samples on DFXaIs, the percentage agreement for LA detection in neat samples tested by TSVT/ET versus APTT and dRVVT after DOAC Remove treatment was 90% (Cohen kappa 0.12). CONCLUSION: Our data highlight uncertainty and disagreement for testing LA in patients on DFXaI. Further studies are required.

Semiquantitative interpretation of anticardiolipin and antiß2glycoprotein I antibodies measured with various analytical platforms: Communication from the ISTH SSC Subcommittee on Lupus Anticoagulant/Antiphospholipid Antibodies.

BACKGROUND: Antiß2glycoprotein I (aß2GPI) and anticardiolipin (aCL) IgG/IgM show differences in positive/negative agreement and titers between solid phase platforms. Method-specific semiquantitative categorization of titers could improve and harmonize the interpretation across platforms. AIM: To evaluate the traditional 40/80-unit thresholds used for aCL and aß2GPI for categorization into moderate/high positivity with different analytical systems, and to compare with alternative thresholds. MATERIAL AND METHODS: aCL and aß2GPI thresholds were calculated for two automated systems (chemiluminescent immunoassay [CLIA] and multiplex flow immunoassay [MFI]) by receiver operating characteristic curve analysis on 1108 patient samples, including patients with and without antiphospholipid syndrome (APS), and confirmed on a second population (n = 279). Alternatively, regression analysis on diluted standard material was applied to identify thresholds. Thresholds were compared to 40/80 threshold measured by an enzyme-linked immunosorbent assay (ELISA). Additionally, likelihood ratios (LR) were calculated. RESULTS: Threshold levels of 40/80 units show poor agreement between ELISA and automated platforms for classification into low/moderate/high positivity, especially for aCL/aß2GPI IgG. Agreement for semiquantitative interpretation of antiphospholipid antibodies (aPL) IgG between ELISA and CLIA/MFI improves with alternative thresholds. LR for aPL IgG increase for thrombotic and obstetric APS based on 40/80 thresholds for ELISA and adapted thresholds for the other systems, but not for IgM. CONCLUSION: Use of 40/80 units as medium/high thresholds is acceptable for aCL/aß2GPI IgG ELISA, but not for CLIA and MFI. Alternative semiquantitative thresholds for non-ELISA platforms can be determined by a clinical approach or by using monoclonal antibodies. Semiquantitative reporting of aPL IgM has less impact on increasing probability for APS.