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Using the HEMOCLOT direct thrombin inhibitor assay to determine plasma concentrations of dabigatran

Stangier, Joachima; Feuring, Martinb

Blood Coagulation & Fibrinolysis: March 2012 - Volume 23 - Issue 2 - p 138–143
doi: 10.1097/MBC.0b013e32834f1b0c
ORIGINAL ARTICLES

The objective of the present study was to assess the suitability of an accurate, sensitive, standardized, chronometric blood coagulation test to determine the anticoagulation activity of dabigatran and to quantify concentrations of dabigatran in plasma. Dabigatran was spiked at increasing concentrations in pooled citrated normal human plasma to measure diluted thrombin time with the HEMOCLOT THROMBIN INHIBITOR assay. Calibration curve linearity, inter-assay and intra-assay precision, and assay accuracy were investigated. Dabigatran stability in plasma and the feasibility of lyophilized dabigatran standards for assay calibration were assessed. Data are presented as back-calculated plasma concentrations of dabigatran using regression analysis. Dabigatran's calibration curve for thrombin clotting time was linear over the concentration range 0–4000 nmol/l (0–1886 ng/ml). The R 2 was 0.99. Total assay imprecision for dabigatran was 4.7–12.0% coefficient of variation, with 1.2–3.1% for intra-run imprecision, 4.0–10.0% for inter-run precision and 0.3–8.3% for between-day imprecision. Assay accuracy was determined at three dabigatran concentrations; deviation from sample target concentrations ranged from −20.7% (100 nmol/l; 47.15 ng/ml) to 5.6% (1500 nmol/l; 707.3 ng/ml). Assay robustness was determined by analysing identical dabigatran samples in two independent laboratories. The mean bias of dabigatran coagulation times between laboratories was 6.6%. The HEMOCLOT Thrombin Inhibitors assay is suitable for determining the anticoagulant activity and calculating plasma concentrations of dabigatran using simple and widely available chronometric coagulation devices. The use of this rapid, established, standardized and calibrated assay should provide accurate and consistent results when assessing the anticoagulant activity of dabigatran.

aDrug Metabolism and Pharmacokinetics, Boehringer Ingelheim Pharma GmbH & Co. KG, Biberach an der Riss

bDepartment of Medical Affairs, Boehringer Ingelheim GmbH, Ingelheim am Rhein, Germany

Correspondence to Joachim Stangier, PhD, Boehringer Ingelheim GmbH & Co. KG, Birkendorferstr. 69, 88397 Biberach an der Riss, Germany Tel: +49 7351 548 443; fax: +49 7351 838 443; e-mail: joachim.stangier@boehringer-ingelheim.com

Received 8 June, 2011

Revised 28 September, 2011

Accepted 31 October, 2011

© 2012 Lippincott Williams & Wilkins, Inc.