Background: Thrombin generation assay was developed several years ago to mimic physiological coagulation mechanisms but it had important limitations. Thrombogram-Thrombinoscope assay using a fluorogenic substrate, allows obtaining thrombin generation curves in non-defibrinated platelet rich plasma (PRP) in a fully automated manner.

To cite this article: Depasse F., Gerotziafas G. T., Busson J., Van Dreden P., Samama M.M. Assessment of three chromogenic and one clotting assays for the measurement of synthetic pentasaccharide fondaparinux (Arixtra Synthetic pentasaccharide fondaparinux (Arixtra 1) is the ®rst indirect (antithrombin-dependent) synthetic speci®c factor (F)Xa inhibitor approved in several countries for the prevention of deep vein thrombosis (DVT) in major orthopedic surgery. In addition, ongoing clinical trials aim to prove its ef®cacy for the treatment of DVT and in coronary heart disease (for review see [1,2]). Fondaparinux plasma concentrations obtained in patients treated at prophylactic and therapeutic doses range from 0.1 to 0.5 mg mL À1 and from 0.6 to 1.5 mg mL À1, respectively; in healthy volunteers, the subcutaneous injection of the established prophylactic dose (2.5 mg) is associated with plasma levels at 0.34 AE 0.04 mg mL À1 [3±5]. Although no coagulation monitoring is advocated, accurate assays dedicated to anti-Xa activity measurement should be available. This could be useful in practical use at least in some particular groups of patients, e.g. with renal impairment, during pregnancy, body weight < 50 kg as recommended for LMWHs or in the elderly. Nevertheless, no special assay has been developed for this purpose and the commercial available assays are designed for either unfractionated or low molecular weight heparins. The aim of this work was to evaluate the possibility of using commercially available assays. Three different chromogenic and one clotting assays for the measurement of fondaparinux anti-Xa activity in plasma were evaluated.Normal pool plasma (Normapool 1 , Hyphen BioMed, Neuville sur Oise, France) was spiked with increasing amounts of fondaparinux (Sano®, Paris, France) in order to obtain plasma concentrations ranging from 0 to 10 mg mL À1 . The antithrombin activity level was measured in this range of concentrations using the Berichrom 1 Heparin was used with addition of exogenous bovine antithrombin as provided with the assay and without addition of exogenous antithrombin. In addition to the results expressed in LMWH anti-Xa IU mL À1 , results were registered as raw data, i.e. optical density variation per minute (DOD min À1 ) and clotting time for the chromogenic and the clotting assays, respectively. FXa inhibition for the chromogenic assays was calculated by reporting the DDO min À1 of the considered sample to the DDO min À1 of the normal pool plasma free of fondaparinux considered as baseline. A statistical analysis was performed using an analysis of variance (ANOVA, F-test).All the antithrombin activity levels measured remained in the normal range and were not signi®cantly affected by the addition of fondaparinux.Results expressed in LMWH anti-Xa IU/mL differed signi®cantly (P < 0.01) from an assay to another for the same fondaparinux plasma concentration, e.g. (mean AE SD, n 10) 0.93 AE 0.03 IU mL A linear relationship was obtained for fondaparinux plasma concentrations ranging from 0 up ...