Tissue culture polystyrene flasks

The procedures were detailed in a previous study^{54 (link)}. The mouse embryonic fibroblast cell line NIH/3T3 was purchased from the Bioresource Collection and Research Center (BCRC), Taiwan. A complete cell culture medium composed of Dulbecco’s modified Eagle medium (DMEM, Gibco) and 10% calf serum (CS, Invitrogen) was used for cell culture. The cells were incubated in tissue culture polystyrene flasks (Corning) in 5% CO₂ at 37 °C until 90% confluence before seeding into the microfluidic chip.

The mouse embryonic fibroblast cell line NIH 3T3 was purchased from the Bioresource Collection and Research Center (BCRC), Hsinchu, Taiwan. A complete medium consisting of Dulbecco’s modified eagle medium (DMEM, Gibco, Waltham, MA, USA) and 10% calf serum (CS, Invitrogen, Carlsbad, CA, USA) was used for cell culture. The cells were incubated in tissue culture polystyrene flasks (Corning, Corning, NY, USA) in 5% CO₂ at 37 °C until a density of 5~6 × 10⁴ cells/cm² was reached. For surface coating, gelatin (Sigma, St. Louis, MO, USA) and poly-l-lysine (Sigma, St. Louis, MO, USA) were diluted in 1× phosphate-buffered saline (PBS) into concentrations of 0.1% (w/w) and 0.1 μg/mL, respectively, as recommended by the manufacturer. Coomassie Brilliant Blue (Bionovas, Toronto, ON, Canada) was diluted into a concentration of 2.5 g/L (in 0.1 L acetic acid, 0.3 L methanol, and 0.6 L ultrapure water) for verifying the presence of gelatin and poly-l-lysine.

Both the human lung carcinoma cells A549 and the fibroblasts NIH/3T3 were purchased from the Bioresource Collection and Research Center (BCRC) (Hsinchu City, Taiwan). Both types of cells were cultured in the complete medium consisting of Dulbecco’s modified Eagle medium (DMEM, Gibco, Waltham, MA, USA) and 10% calf serum (CS, Invitrogen, Carlsbad, CA, USA). Before seeding into the microfluidic chip, they were incubated in tissue culture polystyrene flasks (Corning, Corning, NY, USA) in 5% CO₂ at 37 °C until 90% confluence.