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Updated: Aug 9, 2026
Measurement of Factor V Activity in Human Plasma Using a Microplate Coagulation Assay
Published on: September 9, 2012
Una prueba de generación de trombina quimioluminiscente microfluídica para el monitoreo en tiempo real en plasma de
Theodora J Steeghs1, Lisa van Engelshoven2, Nerissa M M Coolen2
1Enzyre B.V, Novio Tech Campus, Nijmegen, the Netherlands; Department of Haematology, Radboud university medical center, Nijmegen, the Netherlands.
Background:
The thrombin generation assay (TGA) is a global coagulation test able to determine the amount of thrombin formed in plasma samples. TGA has shown to be useful to monitor the effect of procoagulant and anticoagulation therapies in patients with bleeding diatheses or an increased thrombosis risk.
Objective:
This study describes a new miniaturized platform to measure thrombin generation using a microfluidic chemiluminescent-based approach (TGAlum).
Methods:
In the TGAlum, generated thrombin cleaves a β-Ala-Gly-Arg tripeptide from a caged-aminoluciferin moiety which almost instantaneously results in the generation of a photon by luciferase. The amount of photons is proportional to the activity of thrombin in the microfluidic TGAlum system. By printing the TGA, luciferase components and CaCl2, a microfluidic cartridge was functionalized for TGAlum. By only applying 20 μL citrated plasma to the microfluidic cartridges, thrombin generation was monitored successfully with the characteristic peak profile in 110 nL detection chambers using in a prototype readout device.
Results:
The TGAlum was shown to be sensitive to therapeutic levels of anticoagulants (DOACs) to prevent thrombosis, as well as to the reversal effects of the antidotes thereof. Furthermore, the TGAlum was able to detect the defect in plasma of patients with hemophilia A, as well as the hemostatic replacement effect of FVIII infusion in patients with hemophilia A.
Conclusion:
This study provides proof-of-concept that the present microfluidic TGAlum is applicable for monitoring therapy of hemophilia A patients at a global level as well as monitoring anticoagulant therapy with DOACs using a portable readout device.
