Abstract: Various examples are provided related to nanofiber networks that can mimic in vivo physiology. In one example, a scaffold includes a nanofiber membrane extending across a scaffold opening. The nanofiber membrane can include a stack of nanofiber layers that include nanofibers disposed with a controlled orientation or direction and a controlled spacing. The nanofibers are cross-linked with nanofibers of an adjacent nanofiber layer. The scaffold can be incorporated into a transwell insert or plate. In another example, a microfluidic chip can include a first channel layer with a first fluid channel; a second channel layer comprising a second fluid channel; and a scaffold including a nanofiber membrane disposed between the first and second channel layers. The first and second channels cross each other.