Silk microtube fabrication and analysis of their ability to support platelet perfusion. (A) Silk microtubes are prepared by gel spinning aqueous silk solutions containing PEO porogen around a wire and functionalized via entrapment of ECM components. Resulting microtubes are freeze dried, removed from the wire, and soaked in water to leach out the PEO porogen. The resulting porous silk microtubes are fitted into the bioreactor chamber. (Ai) SEM cross sections of a silk microtube: microtube wall thickness was 50 ± 20 µm, with microtube wall pores diameter of 22 ± 4 μm to allow proplatelet elongation (scale bar = 20 µm). Arrows indicate silk microtubes borders. (Aii-Aiii) SEM images show pores on both the inner and outer surfaces of the silk microtubes, respectively. The inner and outer microtube wall pores diameter was 6 ± 2 μm (scale bars = 20 µm). (B) Whole blood (red) or peripheral blood platelets suspended in culture medium (pink) were perfused into functionalized silk microtubes. (C) Representative analysis of whole blood cells of 1 sample before (inlet) or after (outlet) perfusion. WBC, white blood cells; RBC, red blood cells; HCT, hematocrit; MCV, mean corpuscular volume; MCH, mean corpuscular hemoglobin; MCHC, mean corpuscular hemoglobin concentration; RDW, red blood cell distribution width; PLT, platelet. (D) Representative flow cytometry analysis of peripheral blood platelet basal activation before and after perfusion into silk microtube. Activation with ADP and thrombin demonstrated increased PAC-1 binding, indicating normal CD42b⁺ platelet functionality after the passage through the silk microtube lumen. (E) Confocal microscopy analysis of CD61⁺ platelet distribution within microtube lumen after passage of whole blood (green = CD61; blue = nuclei; scale bar = 50 μm). Silk fibroin microtubes were stained with Hoechst 33258 and visualized in blue.