R9 - Dose-Dependent Analysis of Direct Anticoagulants by Acoustic Tweezing Thromboelastometry

Introduction: : Direct anticoagulants that target distinct points in the coagulation cascade, such as thrombin (e.g., heparin) and factor Xa (e.g., apixaban), are increasingly being used in clinical practice. Traditional coagulation assays (e.g., aPTT, PT/INR) and viscoelastic tests (e.g., TEG, ROTEM) often require large blood volumes and exhibit reduced sensitivity to direct anticoagulants. Integrated quasi-static acoustic tweezing thromboelastometry (i-QATT) is a non-contact technique that evaluates clot formation using a microliter-scale blood droplet, offering a rapid and low-volume alternative. This study investigates the ability of i-QATT to quantitatively assess the dose response of two mechanistically distinct direct anticoagulants: unfractionated heparin (UFH), a direct thrombin inhibitor, and apixaban, a direct factor Xa inhibitor.

Materials and

Methods: : Commercial pooled normal plasma was obtained from George King Bio-Medical (Overland Park, KS). Samples were spiked with either UFH (0.1–1.0 IU/mL) or apixaban (10–700 ng/mL), while untreated plasma served as the control. Clotting of the samples, following incubation at 36°C for 20 minutes, was initiated using a mixture of calcium chloride and either intrinsic (aPTT-XL reagent) or extrinsic (Thromboplastin-D reagent) pathway activator. A 6 μL drop of the resulting sample was deployed into the i-QATT device for coagulation analysis. Clot initiation time (CIT), a key coagulation parameter, was derived from i-QATT mechanical tweezographs (Figure 1A–D) to quantify the dose-dependent effects of each anticoagulant. The linear regression model was performed to assess this relationship, with the R² value indicating model’s goodness of fit.

Results, Conclusions, and Discussions:: CIT demonstrated a strong, linear, dose-dependent response with increasing concentrations of both anticoagulants. UFH-treated samples exhibited strong linearity, with R² values of 0.98 for intrinsic and 0.99 for extrinsic pathways (Figure 1E-F). Apixaban-spiked samples produced a similar response, with R² values of 0.81 for intrinsic and 0.98 for extrinsic pathways (Figure 1G-H). While UFH yielded high linearity in both pathways, apixaban demonstrated slightly reduced linearity in the intrinsic pathway, which is consistent given this pathway’s lower sensitivity to apixaban.

This study establishes CIT as a sensitive parameter for monitoring the effects of direct anticoagulants. The ability of i-QATT to quantify the dose response of both thrombin and factor Xa inhibition using microliter-scale plasma samples suggests its clinical potential for rapid, low-volume direct anticoagulation monitoring. Future studies will focus on validating these findings in patient samples to further define the clinical utility of i-QATT.

Acknowledgements and/or References (Optional):: This study was funded by NIH grant No. R21EB031401.