freezing down T cells?

Tony Schountz schountzwa at bioax1.bio.ornl.gov
Fri May 8 09:10:12 EST 1998

janjan wrote:

> Does anyone know of a technique (if one exists) for effectively freezing
> and thawing T cells-- and getting them to proliferate after thawing?
> Thanks.
> Janet

I routinely froze mouse T cell clones 2-3 days after stimulating them with
irradiated APC and peptide.  Just use 5% FBS-Click's (Irvine Scientific) with
10% tissue culture grade DMSO (Sigma) as your freezing medium.  Important thing
is to LSM your T cells, then wash a couple of times with 5% Click's, resuspend
the pellet, put the tube on ice and drop-wise add ice-cold Click's/DMSO with
gentle shaking.  I just packed a beaker with ice, plunged the tube into the ice,
added my freezing medium while swirling the beaker (and consequently the
cells).  Once you've got them to 10^6/ml, aliquot into cryovials, put them in a
box with a lid and freeze them at -70 overnight.  Then transfer to liquid
nitrogen.  To be safe, pull out a tube from LN2 after overnight and check for
viability.  Should be >90% by trypan blue, and should be able to put them up
with APC and peptide.

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