Comparison of the procedures for drug metabolism studies with CCHHs and MMHHs
The time requirement for CCHHs will increase significantly if multiple individual lots (e.g., comparison of different hepatocyte donors) are used since each lot will require efforts in cell recovery, viability and cell concentration determination, and preparation of cell suspensions at the desired cell density. These additional steps are not required for MMHHs.
| Procedure | CCHH | MMHH |
|---|---|---|
| Vial retrieval of from storage | From liquid nitrogen storage (3–5 minutes) | From −80°C storage (3–5 minutes) |
| Thawing of the cryopreserved hepatocytes in a 37°C water bath | 1.5–2 minutes | 1.5–2 minutes |
| Addition of the thawed hepatocytes to recovery medium followed by centrifugation for 10 minutes for cell recovery | 10–15 minutes | Not required |
| Microscopic examination of thawed hepatocytes for the quantification of viability and cell concentration | 10–15 minutes | Not required |
| Adjustment of cell suspension to contain 2× the final desired cell density | 5 minutes | Not required |
| Addition to vessels containing 2× test compoundsa | ||
| Total time from freezer to incubation | 30.5–45 minutes | 7–10 minutes |
↵a For example, addition of 50 μl of hepatocytes to a 96-well plate with 50 μl of test articles at 2× the final concentration, followed by incubation for the desired time duration at 37°C: 1–3 minutes.