Updated 12/10/07 MDR

CELL THAWING1)

MATERIALS

1. Cell-appropriate culture medium
2. 15mL tube
3. Tissue culture flask
4. Cryogloves
5. Frozen cells (from Dewar #5, in room 227)

PROTOCOL

• *ALWAYS FOLLOW SUPPLIER’S RECOMMENDATIONS**

1. Prewarm culture medium to 37C in water bath. Place 10mL warm medium in 15mL tube
2. Retrieve frozen cells. Be sure to use cryogloves to prevent personal injury.
3. Thaw cells rapidly, within 1-3 minutes of removal from tank, by swirling vigorously in 37C water bath
4. Transfer cells to 15mL tube of warm medium. Mix thoroughly by gently pipetting solution up and down.
5. Pellet the cells by centrifugation at 700 rpm for 5 minutes
6. Aspirate off medium to remove DMSO-containing freezing medium.
7. Resuspend cells in 5mL of warm culture medium by gently pipetting up and down.
8. Transfer to T25. Write cell line, Po date, and initials on flask.
9. Examine cells under microscope. Place in 37C incubator.
10. Update frozen cell log online if taken from frozen stock.