Cryopreservation

From Genome@Yale

Please follow directions at ATCC.

HeLa

1. Wash cells once with 1X PBS
2. 1-3mL 0.25X trypsin/EDTA for 5 minutes at RT
3. Detach cells by gentle rocking and tapping
4. Quench trypsin with DMEM + 10% FBS and transfer the cell suspension to 15mL falcon tube
5. Pellet the cells by centrifugation 3000 rpm 5 minutes at RT
6. Prepare freezing media: DMEM +10% FBS +5% DMSO (5mL media + 250uL DMSO)
7. Aspirate liquid from the cell pellet
8. Resuspend pellet in 4mL freezing media
9. Aliquot 1mL into each cryovial
10. Incubate cryovials on ice for 30 minutes
11. Transfer the cryovials to -80C for 12-24hours
12. Transfer the cryovials to liquid nitrogen tank