In this project a flow assay has been developed and validated, incorporating the functions of blood platelets, coagulation and fibrinolysis factors and the contribution of red and white blood cells to the process of thrombus formation and fibrin clot formation. This test can be run in various modes, where coagulation is introduced at a defined extent, and has been applied to evaluate platelet and coagulation function defects, the effect of transfusion with blood components in patients with acquired haemodilution, and the effects of antithrombotic medication.

We have been able to demonstrate that the formation of a fibrin thrombus under flow is regulated in a manner markedly different from static conditions. Most striking are the prominent roles of platelets and local thrombin, reciprocally activating, and the sudden onset of massive fibrin formation, which is delayed by anticoagulants in a non-linear way
The project has resulted in an assay that can be run under a variety of optimized conditions to measure (and/or): (a) platelet deposition and formation of procoagulant platelets, (b) formation of fibrin fibres from the platelet surface, (c) triggering of the system with immobilized or solubilized tissue factor, (d) binding and incorporation of multiple (labelled) coagulation factors: strikingly, FVa, FXa, FIXa and plasminogen primarily bound to the surface of procoagulant platelets, but FIIa, FVIIIa/VWF, FXIIa and FXIIIa bound to domains in the fibrin fibres, (e) clot/thrombus contraction immediately before the formation of fibrin, (f) incorporation of erythrocytes and leukocytes (neutrophils) at the moment of flow reduction or stasis. All these processes have been functionally visualised in real time by multi-colour line-scanning confocal microscopy.
With the adjustments above, the efficacy and validity of the flow test(s) has been evaluated for relevant transfusion-related questions. Transfusion products (platelet concentrates, fresh-frozen plasma, factor concentrates, packed red cells) have been tested against the background of haemodilution both in vitro and in vivo (surgery patients with coagulopathy). Thresholds and endpoints have been determined.

Thesis 2016: Platelets and Coagulation; Partners in haemostasis, Frauke Swieringa