The Role of Fibrinogen Spacing and Patch Size on Platelet Adhesion Under Flow

Platelet adhesion to the vessel wall during vascular injury is mediated by platelet glycoproteins binding to their respective ligands on the vascular wall. In this study we investigated the roles that ligand patch spacing and size play in regulating platelet interactions with fibrinogen under hemodynamic flow conditions. To regulate the size and distance between patches of fibrinogen we developed a photolithography based technique to fabricate patterns of proteins surrounded by a protein repellant layer of poly(ethylene glycol). We demonstrate that when mepacrine labeled whole blood is perfused at a shear rate of 100 s^-1 over substrates patterned with micron-sized wide lines of fibrinogen, platelets selectively adhere to the areas of patterned fibrinogen. Using fluorescent and scanning electron microscopy we demonstrate that the degree of platelet coverage (3% - 35%) and the ability of platelet aggregates to grow laterally was dependent upon the distance (6 - 30 μm) between parallel lines of fibrinogen. We also report on the effects of fibrinogen patch size on platelet adhesion by varying the size of the protein patch (2 – 20 μm) available for adhesion, demonstrating that the downstream length of the ligand patch is a critical parameter in platelet adhesion under flow. We expect that these results and protein patterning surfaces to be useful in understanding the spatial and temporal dynamics of platelet adhesion under physiologic flow, and in the development of novel platelet adhesion assays.