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cryopreservation and nitrogen - cells in vapors or in liquid (Jun/03/2005 )

hi all.
i have quite a basical question. for storing cells in a long term with liquid nitrogen, is better that cells are in nitrogen or is it ok if they are in vapors of nitrogen? no one in the lab can tell me a good answer....

may you share your experinece? or maybe give one or two references?

thanks.
Fred

-fred_33-

Hi Fred,

I think our lab store our samples in boxes, which are then stacked in towers, and then submerge into tank half-filled with liquid nitrogen. I hope this helps.

-Spate-

QUOTE (fred_33 @ Jun 3 2005, 10:12 AM)
hi all.
i have quite a basical question. for storing cells in a long term with liquid nitrogen, is better that cells are in nitrogen or is it ok if they are in vapors of nitrogen? no one in the lab can tell me a good answer....

may you share your experinece? or maybe give one or two references?

thanks.
Fred



I think they both have advantages and disadvantages. When you store the cells in the liquid then there is a chance that it will get into the bottles and give explosion. The temparature in liquid is colder than in vapor so the cells are better stored there. When you store in the vapor then you don't need to be afraid that the liquid will get into the bottle. The problem is you have to remember to fill the tank more often to ensure enough vapor for storage. So here is the danger, if there is not enough vapor then the cells may get warmer.

-indoubt-

well thanks for sharing ideas.
I store cells in cryotubes that are good capped and never encounter an introduction of nitrogen in it. i assume that vapors are very cold (don't have right value but at least -150°C...) that is enough to ensure good conservation. but as people i asked for it tell me contrary opinions (and the fact they are very sure of them) made me in trouble...

-fred_33-

Hi

All tube and cryogenic storage companies say to store your cells in the vapour phase. Basically this is impractical unless you have a vast storage capacity, so almost all labs in the world store the cells in dewars with some of the tubes in the liquid and some in the vapour phase above.

The lab i am in at the moment has been having a huge problem with a particular batch of cryotubes that were allowing the LN2 in and causing the tubes to explode, it took us a while (2-3 months) to work out it was the tubes in particular and replace them with a new brand.

Bob

-bob1-

A few extra thoughts ...

For those of you who have cells stored in tubes that do explode, remember to loosen the lid a little after removing the cells from the tank. This allows the Liq. N2 to escape and not expand inside the vial, leading to an explosion.

Another point not mentioned here ... because the liquid phase does move freely through vials (even if you think you screwed the lid on tight) , there is a risk of spreading cell and mycoplasma contamination. This is an important reason why some labs store only in vapour phase AND why most labs (in Australia) only allow mycoplasma-free cells in the liquid phase.

-AussieUSA-

QUOTE (AussieUSA @ Aug 9 2005, 10:47 AM)
Another point not mentioned here ... because the liquid phase does move freely through vials (even if you think you screwed the lid on tight) , there is a risk of spreading cell and mycoplasma contamination. This is an important reason why some labs store only in vapour phase AND why most labs (in Australia) only allow mycoplasma-free cells in the liquid phase.


Can't mycoplasm (or other) contamination also come from the vapour spreading it around? Both liquid N2 and vapour move around, right?

-vairus-

QUOTE (AussieUSA @ Aug 9 2005, 07:47 PM)
A few extra thoughts ...

For those of you who have cells stored in tubes that do explode, remember to loosen the lid a little after removing the cells from the tank. This allows the Liq. N2 to escape and not expand inside the vial, leading to an explosion.

Another point not mentioned here ... because the liquid phase does move freely through vials (even if you think you screwed the lid on tight) , there is a risk of spreading cell and mycoplasma contamination. This is an important reason why some labs store only in vapour phase AND why most labs (in Australia) only allow mycoplasma-free cells in the liquid phase.

What do you mean with move freely? Do you mean N2 diffuse through the cryovials?
If so how would if affect contamination (as mycoplasma are way way bigger then N2 molecules)?
Or do you mean vials are immersed in liquid nitrogen and material from broken vials float freely in the liquid (as it would in water)?

Just some thoughts,.....

-IronLunatic-

Both types of tanks will keep the cells from degrading for years. My tank only needs to be filled every 7-9 months with liquid nitrogen because it is a high duration liquid type dewar tank. But the vials will be pressurized until the temperature outside is great enough for the gas to be released. A gas type tank will stop any cross contamination of particulates into the vials if the vials are not filled to the top. It is better to store the cells in a gas type tank if you have the money to spend each month to fill it.

-dave2-