Cryopreserved Hepatocytes in Cell Culture

Cryopreserved PHH were purchased from BioreclamationIVT (Brussels, Belgium) and thawed using InVitroGRO CP medium or Cryopreserved Hepatocyte Recovery Medium (Thermo Fisher Scientific, USA). Cells were seeded in 96-well ultra-low attachment plates (Corning, USA) in culture medium (Williams’ medium E, supplemented with 2 mM L-glutamine, 100 U/ml penicillin, 100 mg/ml streptomycin, 10 mg/ml insulin, 5.5 mg/ml transferrin, 6.7 ng/ml sodium selenite, and 100 nM dexamethasone) with 10% fetal bovine serum (FBS) as previously described (Bell et al., 2016 (link)). Cryopreserved primary animal hepatocytes were obtained from BioreclamationIVT (Gottingen minipig, Wistar rat, rhesus monkey, CD1 mouse) or Lonza (USA; C57BL/6 mouse; hereafter called BL6) and were thawed according to the supplier’s instructions. Animal cells were seeded at 2000 (rhesus monkey) or 1000 cells/well (minipig, rat and mouse) and exposed as described earlier. Cell culture medium and all medium supplements were purchased from Sigma-Aldrich (Sweden) or Life Technologies (Sweden).