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For immunofluorescence analysis, cells were seeded into 4-well chamber slides and allowed to reach confluence. They were then washed with PBS and fixed in 4% PFA in PBS for 20 min at room temperature (RT). Cells were then permeabilized and blocked for nonspecific antigen binding with PBS + 10% fetal bovine serum (FBS) + 0.1% Triton X-100 for 1 hour at RT. The cells were then incubated with primary antibodies against platelet and endothelial cell adhesion molecule 1 (CD31; Abcam, ab28364, Cambridge, UK), vascular endothelial (VE)-cadherin (CD144; Santa Cruz Biotechnology, sc-6458, Dallas, TX, USA), endothelial nitric oxide synthase (eNOS; Abcam, ab5589), von Willebrand factor (vWF; Abcam, ab6994) overnight at 4 °C. The next day, cells were washed 3 times with 1× PBS and incubated with corresponding secondary antibody (1:500) for 2 h at RT. After washing 3 times with 1× PBS, mounting medium–containing 4,6-diamidino-2-phenylindole (DAPI) (Vector Laboratories, Burlingame, CA, USA) was applied to slides to visualize nuclei, and coverslips were placed to seal slides. The slides were visualized using a Leica camera (Wetzlar, Germany) DMI6000 B fluorescence microscope. Confocal microscopy was used to visualize luminal structures in microvessel PDMS devices by taking z-stacks of the entire width of the device (200 μm) with slice widths of approximately 5 μm.