Example Metabolic Activity (nmoles/hr/106 cells)
| Substrate | Enzyme | Concentration (µM) | Day 1 | Day 4 | Day 8 |
|---|---|---|---|---|---|
| 7-Ethoxycoumarin | Phase I (ECOD) | 100 | 0.853 | 0.289 | 0.115 |
| 7-Hydoxycoumarin | Phase II (UGT) | 100 | 78.33 | 71.5 | 84.5 |
| Phase II (SULT) | – | 8.68 | 8.57 | 16.13 |
Culture Condition and Morphology
Cryopreserved cynomologus primate hepatocytes were thawed and plated with HUREL PlatinumHeps™ Media supplemented with 10% serum and subsequently changed 24 hours post-seeding to HUREL PlatinumHeps™ maintenance Media. CYP substrate concentrations are given in the table above along with metabolite formation recorded as nmoles/hr/million cells. All incubations were carried out in triplicate on days 1, 4, and 8 (customer day) upon cell delivery and incubated for 60 minutes. Reactions took place in a humidified incubator at 37°C, in 5% CO2. Collected supernatants were stored at -20°C until further LC/MS/MS analysis.
Day 1 – Morphology
Phase contrast image in a 96-well at a 10x magnification
Day 7 – Morphology
Phase contrast image in a 96-well at a 10x magnification
Day 7 – Bile Canaliculi
Bile canaliculi assayed via 5-(and-6)-carboxy-2’, 7’-dichlorofluorescein diacetate (CDFDA) stain at a concentration of 5 µM and imaged in the GFP channel in a 96-well at 10x magnification