On Tue, 14 Apr 1998 16:56:14 -0400, K_Vora at lac.jci.tju.edu (Kalpit A.
Vora) wrote:
>In article <Pine.GSO.3.96.980413132953.29697B-100000 at dilbert.ucdavis.edu>,
>"L.A. Pike-Nobile" <ez063669 at mailbox.ucdavis.edu> wrote:
>>> I am getting extremely variable rates of viability from my B cell lines
>> when I thaw them after storage in liquid N2. I freeze them in 50% media/
>> 30% FCS/20% DMSO. I freeze them gradually (I use on of those little
>> ethanol-jacked freezing things that claims to give 1oC/hour) and I thaw
>> very rapidly (37 water bath) and rinse them thoroughly.
>>>> AND YET--I still get some vials with less than 1% viability while others
>> are fine. Some lines are better than others, but I see this problem even
>> within the same line, frozen at the same time. I'm worried about
>> completely losing some of my more fragile lines.
>>>> Are there any tricks I should know about?
>>>> Thanks,
>> Larry
>Dear Larry,
>We routinely freeze our hybridoma in fetal calf serum containing 5%DMSO
>and nothing else. Also We pellet the cells resuspend in the above freezing
>medium and leave it overnigt at -70 before transfering to liquid nitrogen.
>It can remain at -70 for 3-4days without effecting the recovery. Also try
>and freeze the cells when they are young and growing in exponential phase.
>I donot see any problem in the way you thaw them out.
>Hope this helps.
>Kalpit
Hi Larry
This mathod of Kalpit's is quite OK
I would add: wrap your cells in a nice packet of newspaper when you
put them at -70 °C, that makes them cool down slowly as if you use a
sophisticated machine.
Wolfgang
