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Freezing viable cells in 96-well format?


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#1 ramblingrat

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Posted 25 October 2013 - 03:34 PM

Hi guys,

 

I have generated (a lot!=hundreds) of different cell colonies and would like to freeze them to -80. Has anyone experience with freezing cells in 96-wells (so they survive at least over several weeks and could be thawed and re-cultured again)? Are 96 cell-cuture plates robust at -80 or would you use different plates? 

 

Many thanks!

 

R.



#2 bob1

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Posted 26 October 2013 - 12:43 AM

The usual way of freezing cells involves having them in suspension, so long as you can do this in the plate, I see no reason why this wouldn't work. The difficulty will be getting them to freeze down at an appropriate rate. The best way to do this will probably be wrapped in polystyrene somehow.

THe plates themselves should be fine at -80, but the cells will last only a few months (<6, cell line dependent) at most before there is significant die-off, perhaps shorter with the small volume for each well. For permanent storage you would need to have them in liquid nitrogen storage.

#3 ramblingrat

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Posted 27 October 2013 - 08:46 AM

Hi bob1,

 

Awesome, many thanks! I'll give it a try - my cells are suspension cells fortunately and I only need to keep them a few weeks :). Will put them in -20 first and then -80.

 

Best,

R.



#4 bob1

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Posted 27 October 2013 - 04:19 PM

I would strongly advise against freezing at -20 first, this causes ice crystal formation which is the major cause of death during the freezing process. If you are freezing following a standard protocol (10% DMSO in the final solution) then placing straight in the -80 is best, so long as they are wrapped in polystyrene (or floating on a pool of isopropanol) so that the freezing rate is about -1 deg C per minute.




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