*2.4. Cell Culture and Flow Experiments*

Human liver hepatocellular carcinoma cells (HepG2) were obtained from Korean Cell Line Bank. HepG2 cells in suspension containing 1 × 104 cell/chamber were seeded on the nanofiber scaffold in the microfluidic chip under static conditions, and allowed to form a stable attachment for 24 h. After the cells were allowed to settle, the medium was continuously perfused to the chip at a flow rate of 0.1 mL/h using a commercial syringe pump (New Era Pump Systems, Farmingdale, NY, USA). The perfused culture medium was Dulbecco's modified Eagle's medium (Life Technologies, ThermoFisher Scientific, Waltham, MA, USA) supplemented with 10% fetal bovine serum (Life Technologies, ThermoFisher Scientific, Waltham, MA, USA) and 1% penicillin streptomycin. The cultures were stored for 14 days in a humidified atmosphere of 5% CO2 in air at 37 ◦C. We could not find any deterioration in the device operation nor change in the nanofiber morphology during the culture. Moreover, the nanofibers stuck tightly in the chamber and embedded well on the chamber bottom.
