Microfluidic Point-of-Care Ecarin-Based Clotting and Chromogenic Assays for Monitoring Direct Thrombin Inhibitors

Abstract

Direct thrombin inhibitors (DTIs), such as bivalirudin and dabigatran, have maintained steady inpatient and outpatient use as substitutes for heparin and warfarin, respectively, because of their high bioavailability and relatively safe “on-therapy” range. Current clinical methods lack the capacity to directly quantify plasma DTI concentrations across wide ranges. At present, the gold standard is the ecarin clotting time (ECT), where ecarin maximizes thrombin activity and clotting time is evaluated to assess DTIs’ anticoagulation capability. This work focused on the development of a microfluidic paper analytic device (µPAD) that can quantify the extent of anticoagulation as well as DTI concentration within a patient’s whole blood sample. Capillary action propels a small blood sample to flow through the nitrocellulose paper channels. Digital images of whole blood migration are then captured by our self-coded Raspberry Pi and/or the Samsung Galaxy S8 smartphone camera. Both the flow length and the blue absorbance from the plasma front on the μPAD were measured, allowing simultaneous, dual assays: ecarin clotting test (ECT) and ecarin chromogenic assay (ECA). Statistically significant (p < .05) changes in flow and absorbance were observed within our translational research study. Currently, there are no quantitative, commercially available point-of-care tests for the ECT and ECA within the United States. Both the ECT and ECA assays could be instrumental to differentiate between supratherapeutic and subtherapeutic incidents during bridging anticoagulant therapy and limit the unwarranted use of reversal agents.

Novel Substituted Azoloazines with Anticoagulant Activity

“…Meizothrombin has up to 97% thrombin activity (compared with 10% without ecarin), and it converts fibrinogen to fibrin much more efficiently. Because direct thrombin inhibitors can inhibit not only thrombin, but also meizothrombin, ecarin-based assays are quite appropriate to estimate anticoagulation capability under optimal conditions, compared with conventional coagulation analyses [44]. Therefore, as the next step of our study, the effect of compound 5d on ecarin time was estimated.…”

JECTAuthors: a Diverse, Highly Prized Group

“…In this issue, we have Adam Blakey, a recent graduate from the University of Nebraska who described the losses in heater-cooler efficiency with the extended water lines that some have implemented in light of recent infectious concerns (2). In the previous issue, my own students, Vina Nguyen and Ben Alouidor were able to publish their hard work collaborating with a biomedical engineering laboratory to develop novel anticoagulation testing platforms for heparin and direct thrombin inhibitors (3). * Research projects like these require a process that follows well-established guidelines to produce new knowledge.…”