Ula round bottomed plates

HepG2 cells were seeded using a Multiflo FX multi-mode automated reagent dispenser (Biotek, UK) at 2.0 × 10 5 cells per well (2D) in Ibidi microplate 96 well black plates (Ibidi, USA) and incubated for 24 h at 37 °C in a Forma Steri-Cycle CO 2 incubator (Thermo Scientific, UK) or 400 cells per well (3D) in 96-well ULA round-bottomed plates (Corning, UK) in 180μL of complete DMEM. Cells were incubated for 4, 11 and 21 days 37°C in a Forma Steri-Cycle CO 2 incubator (Thermo Scientific, UK). Post-seeding CYP1A2 and CYP2E1 mRNA expression in HepG2 2D and 3D cultures were measured relying on RT-PCR. The results are expressed as percent mean mRNA content normalised to GAPDH and are the mean ± SD of n = 3 experiments.

HepG2 cells were seeded using a Multiflo FX multi-mode automated reagent dispenser (Biotek, UK) at 2.0 × 10 5 cells per well (2D) in Ibidi 96 well black microplates (Ibidi, USA) and 200 cells per well (3D) in 96well ULA round-bottomed plates (Corning, UK). Cells were incubated for 24 h (2D) and 7 days (3D) at 37 °C, 5% CO 2 . 2D and 3D cell cultures were treated with 30 mM of APAP for 24 h. Cells were subsequently stained with Calcein AM (1.5μM) and Ethidium homodimer-1 (10μM) and cell death and viability data were measured using Operetta CLS High Content Analysis System (PerkinElmer, UK).
For cells assessed on the Mera system, HepG2 cells were cultured in ULA microplates for 7 days and transferred to Mera for treatment and staining. 3D microtissues were stained with Calcein AM (3μM) and Ethidium homodimer-1 (10μM) on Mera and were then transferred to a ULA microplate for imaging consistency. Pixel intensity was measured using an IX70 Olympus inverted microscope (Olympus, Tokyo, Japan).