freezing CHO flasks

[Seth Crosby]

Hullo all you CHOsters!
Just wanted to relate a little experiment I (and probably many others) =

did which might save you some time. I have frozen flasks of CHO cells, =

both sub and fully confluent. Freezing the flask is an alternative to =

carrying cells that aren=B9t immediately needed, while you=B9re on vacation=
, =

etc. =

To freeze a flask:
1) remove media
2) for a T75 add 6ml of freezing media (mine is F12 with 10%FCS and 5% =

DMSO)
3) lay flask flat and wiggle it a bit to work the mix into the cells
3) pop flask into a -70=9AC, laying it so that the cells are submerged

to thaw:
1) pour in 100ml of warmed PBS (w/ Ca&Mg)
2) tilt flask so that PBS covers the cell layer (do not lay flask flat =

or your cup will runneth over)
3) within a minute everything will be thawed...after thawing (ice will =

tear up the cells) wiggle the flask a bit to fully dissolve the DMSO =

layer
4) right flasks and aspirate (the residual DMSO hasn=B9t seemed to do =

any harm, but you might want to add an extra PBS rinse)
5) add media

The cells look fine and grow well. I have not tested anything else.

-- =

Seth Crosby, MD
Abbott Labs
Senior Research Scientist
Growth Factors Group