Freezing Cells: FGB

10/8/98

Reagents:

Freezing Medium (FM) is10 % DMSO / 90 % Media (with 10% FBS)

Aliquot and keep frozen at -20 °C

Briefly, thaw at 37’ C and keep it on ice until used.

1. Transfer cells from TC flask to 50 mL tube.

2. Wash TC flask with 10 mL media and transfer to the same conical tube.

3. Spin at 1000 rpm for 10 minutes at 4 °C.

4. Combine like cell pellets if low cell number.

5. Spin at 1000 rpm/4'C/10 minutes.

6. Aspirate supernatant then resuspend pellet in ~5 ml ice cold FM.

The goal is 10-20 million cells/mL of FM. Do not over dilute!

7. Transfer the FM + cells to pre-labeled freezing vials (1 mL per vial).

7. Freeze in blue alcohol freezer box (with fresh isopropanol in bottom) O/N at -80 °C. Never place in a blue box already in freezer.

8. Move to -140 °C the next day.