Based on Katenz et al 2006

Cryopreservation media for preserving primary hepatocytes

10% DMSO, 10% FBS, 0.2M Trehalose dihydrate

• 1mL DMSO
• 1mL Fetal Bovine Serum
• 760mg trehalose dihydrate
• Cells in Supplemented William’s E media (or appropriate growth media) to 10mL

1. Aliquot in cryotubes.
2. Note estimated number of cells on the tube.
3. Freeze 24 hours at -80 in Nalgene Cryo 1C freezing container (to control rate of temperature change and minimize formation of ice crystals).
4. Transfer to liquid nitrogen for long-term storage.

To use cells:

1. Thaw in 37C water bath.
2. Centrifuge at 50G for 1 minute.
3. Remove cryopreservation media and replace with appropriate growth media.
4. Optionally, repeat spin/media change to wash.