Platelets obtained from MK exposure to high shear rates can be activated by thrombin. Cell effluents in the flow-through of MKs exposed to high shear rates were analyzed in a fibrinogen adhesion assay in static conditions, followed by confocal microscopy, with cell staining with phalloidin–Alexa 546 to visualize actin and Alexa 488 to visualize αIIbβ3. Washed blood platelets (A) and MK-derived platelets generated by shear exposure (B) adhered to fibrinogen in the absence (left panels) or in the presence (right panels) of thrombin. Nonactivated cells display diffuse actin staining and αIIbβ3 membrane localization. After thrombin activation, actin filaments are organized as stress fibers. They display a similar cytoskeletal organization to washed blood platelets. The bars represent 10 μm. Flow cytometry assay (C): Samples were labeled with anti–CD62P-FITC (FL1, P-selectin) and anti–CD41-PE (FL2, αIIb). Settings of FSC-SSC profiles of washed blood platelets were used to analyze flow-through cells. Histogram plot of activated platelets in the flow-through, in the absence or presence of thrombin, after labeling with nonimmune IgG (thin line, gray background), anti-CD62P, or anti-CD41a (thick line). Electron microscopy: (D) in the presence of thrombin, the platelet-sized fragment displayed morphologic changes characteristic of activated platelets, namely a spherical shape, surface pseudopods (p), dense material within dilated cisternae of SCCS, no granulation in the cytoplasm, and a central bundle of microfilaments, reminiscent of activated blood platelets. The bar represents 2 μm.