Example Metabolic Activity (nmoles/hr/106 cells)
Substrate | Enzyme | Concentration (µM) | Day 1 | Day 8 |
|---|---|---|---|---|
| 7-Ethoxycoumarin | Phase I (ECOD) | 100 | 0.234 | .239 |
| 7-Hydroxycoumarin | Phase II(UGT) | 100 | 88.8 | 104.4 |
| 7-Hydroxycoumarin | Phase II (SULT) | 100 | .980 | .836 |
Culture Condition and Morphology
Cryopreserved minipig 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 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 24-well at a 10x magnification
Day 7 – Morphology
Phase contrast image in a 24-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 24-well at 10x magnification