Full Length Article| Volume 164, P32-39, April 2018

Comprehensive characteristics of the anticoagulant activity of dabigatran in relation to its plasma concentration

Published:February 17, 2018DOI:


      • ROTEM® clotting times correlated well with dabigatran plasma concentrations.
      • Ecarin chromogenic assay and diluted thrombin time correlated with concentrations.
      • Coagulability similar in dabigatran-spiked samples and those from treated patients.
      • Storage for one year at −80 °C changed neither dabigatran levels nor coagulability.



      Issues with laboratory measurement of dabigatran include: 1. Do coagulation assays reflect dabigatran plasma concentrations? 2. Do samples from patients treated with dabigatran have the same coagulability as dabigatran-spiked samples from healthy volunteers? 3. What is the long-term stability of dabigatran after storage at −80 °C? This study aims to evaluate these questions.

      Materials and methods

      Ecarin chromogenic assay (ECA), a laboratory-developed diluted thrombin time (LD-dTT), prothrombin time (PT) and activated partial thromboplastin time (APTT) and ROTEM® were used to measure dabigatran anticoagulant activity and liquid chromatography-tandem mass spectrometry (LC-MS/MS) to measure dabigatran plasma concentrations. ROTEM® (EXTEM, INTEM, FIBTEM) was performed in whole blood and the other assays in platelet poor plasma (PPP), both in samples spiked with dabigatran (0, 25, 50, 100, 250, 500 and 1000 ng/mL) from healthy donors and in ex vivo samples from patients treated with dabigatran etexilate. Citrated PPP samples were frozen and stored at −80 °C, 1, 3, 6 and 12 months until analysis.


      EXTEM and FIBTEM clotting time (CT), ECA and LD-dTT correlate well with dabigatran plasma concentrations. With the exception of few ROTEM® parameters, there were no differences between spiked and patient samples. Samples were stable for at least 12 months at −80 °C.


      EXTEM and FIBTEM CT, ECA and LD-dTT are suitable for measuring the effect of dabigatran in treated patients. In general, results from spiked plasma samples are similar to those of patient samples. Storage of dabigatran plasma samples for up to 12 months does not influence measured levels.


      APTT (activated partial thromboplastin time), APTT-Cepha (APTT using Cephascreen® (Diagnostica Stago) reagent), APTT-PSL (APTT using Pathromtin®SL (Siemens Healthcare) reagent), ANOVA (analysis of variance), CAT (calibrated automated thrombinography), CT (clotting time), DMSO (dimethyl sulfoxide), DTI (direct thrombin inhibitor), ECA (ecarin chromogenic assay), ETP (endogenous thrombin potential), EXTEM (thromboelastometry testing the extrinsic haemostatic system), HCl (hydrochloric acid), INTEM (thromboelastometry testing the intrinsic haemostatic system), FIBTEM (thromboelastometry testing the fibrin part of clot formation), LC-MS/MS (liquid chromatography-tandem mass spectrometry), LD-dTT (a laboratory developed diluted thrombin time), MCF (maximum clot firmness), MRM (multiple reaction monitoring), NOAC (non-vitamin K antagonist oral anticoagulants), PT (prothrombin time), PT-INN (PT using Dade® Innovin® (Siemens Healthcare) reagent), PT-Owren (PT using Owrens (Medirox) reagent), ROTEM (rotational thromboelastometry), SE (standard error), SEE (standard error of the estimates), TF (tissue factor), TQD (tandem quadrupole), UPLC (ultra-performance liquid chromatography), VKA (vitamin K-antagonist)


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        • Ten Cate V.
        • Ten Cate H.
        • Verheugt F.W.
        The global anticoagulant registry in the FIELD-atrial fibrillation (GARFIELD-AF): exploring the changes in anticoagulant practice in patients with non-valvular atrial fibrillation in the Netherlands.
        Neth. Hear. J. Oct 2016; 24: 574-580
        • Boehringer Ingelheim Canada Ltd
        Pradaxa™ Product Monograph.
        Jan 27 2012
        • Kirchhof P.
        • Benussi S.
        • Kotecha D.
        • Ahlsson A.
        • Atar D.
        • Casadei B.
        • et al.
        2016 ESC guidelines for the management of atrial fibrillation developed in collaboration with EACTS.
        Eur. Heart J. Oct 7 2016; 37: 2893-2962
        • Sjogren V.
        • Bystrom B.
        • Renlund H.
        • Svensson P.J.
        • Oldgren J.
        • Norrving B.
        • et al.
        Non-vitamin K oral anticoagulants are non-inferior for stroke prevention but cause fewer major bleedings than well-managed warfarin: a retrospective register study.
        PLoS One. Jul 10 2017; 12e0181000
        • Reilly P.A.
        • Lehr T.
        • Haertter S.
        • Connolly S.J.
        • Yusuf S.
        • Eikelboom J.W.
        • et al.
        The effect of dabigatran plasma concentrations and patient characteristics on the frequency of ischemic stroke and major bleeding in atrial fibrillation patients: the RE-LY trial (randomized evaluation of long-term anticoagulation therapy).
        J. Am. Coll. Cardiol. Feb 4 2014; 63: 321-328
        • Antovic J.P.
        • Skeppholm M.
        • Eintrei J.
        • Boija E.E.
        • Soderblom L.
        • Norberg E.M.
        • et al.
        Evaluation of coagulation assays versus LC-MS/MS for determinations of dabigatran concentrations in plasma.
        Eur. J. Clin. Pharmacol. Nov 2013; 69: 1875-1881
        • Kuhn J.
        • Gripp T.
        • Flieder T.
        • Dittrich M.
        • Hendig D.
        • Busse J.
        • et al.
        UPLC-MRM mass spectrometry method for measurement of the coagulation inhibitors dabigatran and rivaroxaban in human plasma and its comparison with functional assays.
        PLoS One. Dec 23 2015; 10e0145478
        • Hawes E.M.
        • Deal A.M.
        • Funk-Adcock D.
        • Gosselin R.
        • Jeanneret C.
        • Cook A.M.
        • et al.
        Performance of coagulation tests in patients on therapeutic doses of dabigatran: a cross-sectional pharmacodynamic study based on peak and trough plasma levels.
        J. Thromb. Haemost. Aug 2013; 11: 1493-1502
        • Skeppholm M.
        • Al-Aieshy F.
        • Berndtsson M.
        • Al-Khalili F.
        • Ronquist-Nii Y.
        • Soderblom L.
        • et al.
        Clinical evaluation of laboratory methods to monitor apixaban treatment in patients with atrial fibrillation.
        Thromb. Res. 2015 Jul; 136: 148-153
        • Boonen K.
        • Schmitz E.
        • Rozestraten F.
        • van den Heuvel D.
        • Brunsveld L.
        • van der Voort P.
        • et al.
        Real life dabigatran and metabolite concentrations, focused on inter-patient variability and assay differences in patients with atrial fibrillation.
        Clin. Chem. Lab. Med. Oct 2017; 55 (2002-2009)
        • Sinigoj P.
        • Malmstrom R.E.
        • Vene N.
        • Ronquist-Nii Y.
        • Bozic-Mijovski M.
        • Pohanka A.
        • et al.
        Dabigatran concentration: variability and potential bleeding prediction in “real-life” patients with atrial fibrillation.
        Basic Clin. Pharmacol. Toxicol. Nov 2015; 117: 323-329
        • Bozic-Mijovski M.
        • Malmstrom R.E.
        • Malovrh P.
        • Antovic J.P.
        • Vene N.
        • Sinigoj P.
        • et al.
        Diluted thrombin time reliably measures low to intermediate plasma dabigatran concentrations.
        Ann. Clin. Biochem. Jul 2016; 53: 446-451
        • Schmitz E.M.
        • Boonen K.
        • van den Heuvel D.J.
        • van Dongen J.L.
        • Schellings M.W.
        • Emmen J.M.
        • et al.
        Determination of dabigatran, rivaroxaban and apixaban by ultra-performance liquid chromatography - tandem mass spectrometry (UPLC-MS/MS) and coagulation assays for therapy monitoring of novel direct oral anticoagulants.
        J. Thromb. Haemost. Oct 2014; 12: 1636-1646
        • Douxfils J.
        • Mullier F.
        • Loosen C.
        • Chatelain C.
        • Chatelain B.
        • Dogne J.M.
        Assessment of the impact of rivaroxaban on coagulation assays: laboratory recommendations for the monitoring of rivaroxaban and review of the literature.
        Thromb. Res. Dec 2012; 130: 956-966
        • Helin T.A.
        • Pakkanen A.
        • Lassila R.
        • Joutsi-Korhonen L.
        Laboratory assessment of novel oral anticoagulants: method suitability and variability between coagulation laboratories.
        Clin. Chem. May 2013; 59: 807-814
        • Stangier J.
        • Feuring M.
        Using the HEMOCLOT direct thrombin inhibitor assay to determine plasma concentrations of dabigatran.
        Blood Coagul. Fibrinolysis. Mar 2012; 23: 138-143
        • Loeffen R.
        • Kleinegris M.C.
        • Loubele S.T.
        • Pluijmen P.H.
        • Fens D.
        • van Oerle R.
        • et al.
        Preanalytic variables of thrombin generation: towards a standard procedure and validation of the method.
        J. Thromb. Haemost. Dec 2012; 10: 2544-2554
        • Douxfils J.
        • Mullier F.
        • Robert S.
        • Chatelain C.
        • Chatelain B.
        • Dogne J.M.
        Impact of dabigatran on a large panel of routine or specific coagulation assays. Laboratory recommendations for monitoring of dabigatran etexilate.
        Thromb. Haemost. May 2012; 107: 985-997
        • Wagenvoord R.J.
        • Deinum J.
        • Elg M.
        • Hemker H.C.
        The paradoxical stimulation by a reversible thrombin inhibitor of thrombin generation in plasma measured with thrombinography is caused by alpha-macroglobulin-thrombin.
        J. Thromb. Haemost. Jun 2010; 8: 1281-1289
        • Lassen J.F.
        • Brandslund I.
        • Antonsen S.
        International normalized ratio for prothrombin times in patients taking oral anticoagulants: critical difference and probability of significant change in consecutive measurements.
        Clin. Chem. Mar 1995; 41: 444-447
        • Comuth W.J.
        • Faaborg L.
        • Henriksen L.Ø.
        • Münster A.B.
        Measurement of dabigatran: previously demonstrated Hemoclot® Thrombin Inhibitor assay reagent instability on Sysmex CS-2100i is no longer an issue.
        Scand. J. Clin. Lab. Invest. Feb-Apr 2018; 78 (Epub 2017 Nov 16): 149-152
        • Solbeck S.
        • Ostrowski S.R.
        • Stensballe J.
        • Johansson P.I.
        Thrombelastography detects dabigatran at therapeutic concentrations in vitro to the same extent as gold-standard tests.
        Int. J. Cardiol. Apr 1 2016; 208: 14-18
        • Taune V.
        • Wallen H.
        • Agren A.
        • Gryfelt G.
        • Sjovik C.
        • Wintler A.M.
        • et al.
        Whole blood coagulation assays ROTEM and T-TAS to monitor dabigatran treatment.
        Thromb. Res. May 2017; 153: 76-82
        • Levy J.H.
        • Ageno W.
        • Chan N.C.
        • Crowther M.
        • Verhamme P.
        • Weitz J.I.
        • et al.
        When and how to use antidotes for the reversal of direct oral anticoagulants: guidance from the SSC of the ISTH.
        J. Thromb. Haemost. Mar 2016; 14: 623-627
        • Brinkman H.J.
        Global assays and the management of oral anticoagulation.
        Thromb J. Feb 2015; 10 (eCollection 2015)9-015-0037-1
        • Reilly P.A.
        • van Ryn J.
        • Grottke O.
        • Glund S.
        • Idarucizumab Stangier J.
        A specific reversal agent for dabigatran: mode of action, pharmacokinetics and pharmacodynamics, and safety and efficacy in phase 1 subjects.
        Am J Emerg Med. Sep 2016; 28
        • Furugohri T.
        • Morishima Y.
        Paradoxical enhancement of the intrinsic pathway-induced thrombin generation in human plasma by melagatran, a direct thrombin inhibitor, but not edoxaban, a direct factor Xa inhibitor, or heparin.
        Thromb. Res. Sep 2015; 136: 658-662
        • Gribkova I.V.
        • Lipets E.N.
        • Rekhtina I.G.
        • Bernakevich A.I.
        • Ayusheev D.B.
        • Ovsepyan R.A.
        • et al.
        The modification of the thrombin generation test for the clinical assessment of dabigatran etexilate efficiency.
        Sci. Rep. Jul 5 2016; 629242
        • Walker J.B.
        • Hughes B.
        • James I.
        • Haddock P.
        • Kluft C.
        • Bajzar L.
        Stabilization versus inhibition of TAFIa by competitive inhibitors in vitro.
        J. Biol. Chem. Mar 14 2003; 278: 8913-8921
        • Skeppholm M.
        • Hjemdahl P.
        • Antovic J.P.
        • Muhrbeck J.
        • Eintrei J.
        • Ronquist-Nii Y.
        • et al.
        On the monitoring of dabigatran treatment in “real life” patients with atrial fibrillation.
        Thromb. Res. Oct 2014; 134: 783-789
        • Rabinovitch R.
        • Barnes P.
        • Curcio K.
        • Dorman J.
        • Huisman A.
        • Nguyen L.
        • et al.
        Validation, Verfication, and Quality Assurance of Automated Hematology Analyzers; Approved Standard - Second edition.
        Clinical and Laboratory Standards Institute, 2010
        • Faaborg L.
        • Comuth W.J.
        • Bloch-Münster A.M.
        • Henriksen L.Ø.
        • Husted S.
        Handling of the Hemoclot Thrombin Inhibitor Assay Reagents on Sysmex CS-2100i when Monitoring Dabigatran in Acute Clinical Situations. Poster.
        XXV International Society on Thrombosis and Haemostasis Congress, Toronto, Canada2015 (Abstract(June))
        • Rasmuson A.
        • Berndtsson M.
        • Soderblom L.
        • Skeppholm M.
        • Malmstrom R.
        • Antovic J.
        Evaluation of Different Coagulation Laboratory Methods to Monitor Apixaban, Rivaroxaban and Dabigatran Treatment.
        The XXVI Congress of the International Society on Thrombosis and Haemostasis, 2017 (poster presentation)
        • Comuth W.J.
        • Faaborg L.
        • Henriksen L.O.
        • Munster A.B.
        Measurement of dabigatran: previously demonstrated Hemoclot ((R)) thrombin inhibitor assay reagent instability on Sysmex CS-2100i is no longer an issue.
        Scand. J. Clin. Lab. Invest. Nov 16 2017;
        • Eikelboom J.W.
        • Connolly S.J.
        • Brueckmann M.
        • Granger C.B.
        • Kappetein A.P.
        • Mack M.J.
        • et al.
        Dabigatran versus warfarin in patients with mechanical heart valves.
        N. Engl. J. Med. Sep 26 2013; 369: 1206-1214