To test that neurons remain in their distinct compartments, one excitatory neuronal population was seeded against another. Neurons in the left chamber were infected with lentivirus encoding tdTomato prior to seeding, to aid in visualization of their position within the two-compartment device. Cells were given five hours to attach before filling the well with fresh media. After approximately four weeks in culture, tdTomato-labeled axonal projections were seen traversing the microchannels. Importantly, tdTomato-labeled neuronal cell bodies were retained on the left side of the device (Fig. S1a and b†). Examination of an entire well revealed that the seeded neurons grow uniformly within the entire chamber, but do not migrate into the adjacent chamber (Fig. S1c†).
