Figure 3.

Air-liquid interface Bridge and Immunostaining of A549 cells. (A) Two sets of 2 microscope slides (1” × 3”) were space 6 mm apart and adhered together using silicone sealant and 1 silicone strip (87 mm long × 5–6 mm wide × 1.6 mm thick). This was placed in a 150 mm petri dish and autoclaved. Two lung gasket membranes were placed on top of the bridge on Day 3. 20 mL of device medium was pipetted below the bridge, every other day 50% of the medium was exchanged until Day 10 (device assembly). A red arrow indicates where the lung cells (A549) were plated and how the air-liquid interface sits on top of the Bridge. (B) Fluorescent micrographs (400× magnification) of A549 cells on Day 10 were observed after immunostaining under a fluorescent microscope for tight junctions (protein ZO-1, green). The nuclei were counter-stained (DAPI, blue). Scale bars (10 μ) are present on the micrographs.