Isolation and culture of primary hepatocytes

All the animal procedures were performed in accordance with the recommendations and guidelines from IACUC at University of Nebraska-Lincoln. Primary rat hepatocytes were isolated from male Sprague-Dawley rats (160–200 g weight) following a two-step collagenase perfusion protocol adapted from P.O Seglen.⁴³ Around 150–200 million cells were obtained at a viability greater than 90% as confirmed through Trypan blue dye exclusion test. Cells were seeded at a density of 100 000 cm⁻² on the collagen coated PDMS substrates and TCPS. Cells were maintained in a humidified 5% CO₂ incubator at 37 °C and cell culture media was replaced every 24 hours.