Sack: Freezing CHO cells
Revision as of 18:42, 8 October 2012 by Jon Sack (New page: Freezing CHO Cells<br> Christina McGee 4-5-2012<br> # Grow 10 cm dish to 50-90% confluency in T-75 flask # Digest cells with 3 ml Trypsin/EDTA, wait for cells to detach. # Add 7 ml med...)
Freezing CHO Cells
Christina McGee 4-5-2012
- Grow 10 cm dish to 50-90% confluency in T-75 flask
- Digest cells with 3 ml Trypsin/EDTA, wait for cells to detach.
- Add 7 ml media, transfer to 15 ml sterile tube.
- Centrifuge 250 G/5 min.
- Remove supernatant. Gently resuspend cell pellet in 2 ml freezing medium*.
- Transfer 0.5 ml to 4 freezing vials labeled with name of cell line, date and your initials.
- Place vial in freezing box in -80°C freezer.
- Transfer to liquid nitrogen 1-4 days later.
- Record location of cells in Liquid N2 excel spreadsheet
- Freezing medium: Cell medium (F-12 or DMEM/F-12) without selection agents or antibiotics plus 20% FBS and 10% DMSO (dimetylsulfoxide, sterile).
- Return to Sack:Protocols