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Freezing HeK'n cells


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#1 bionick

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Posted 28 January 2005 - 04:40 AM

Hi Everybody!
How can I freezzing HeK'n cells? I don't have special medium and I can't find no additional informations :lol: . I'll be grateful for any ideas.
Thanks

#2 Simonsays

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Posted 31 January 2005 - 06:45 AM

I know how to freeze HEK cells... don't know what HeK'n are!?!? If HeK'n are adherent cells, I guess the protocol is just the same:

Trypsinize the cells
Count them
Resuspend them in 'freezing buffer' at about 1 million cell/ml.
Freeze them slowly down to -80°C (put the vials between two styrofoam racks taped tight... this way they don't freeze too quickly).
The day after, put them in liquid nitrogen.

Freezing buffer : DMEM (or any medium I guess) + 20% FBS + 10% sterile DMSO. NO ANTIBIOTICS.

Voilą!

Simon

#3 fred_33

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Posted 31 January 2005 - 08:36 AM

hi
for a 15cm diameter plate :
i trypsinise cells, centrifuge them and add 2ml of a [SERUM / 10%DMSO]solution
Then i but cells in cryotubes. Cryotubes are putted in polystyrene racks taped tight and put overnight at -80°C. The second day i freeze them in nitrogen.


i prepare 9ml SERUM + 1ml sterile DMSO and store it for max 2weeks at 4°C
2ml are used for a 15cm diameter plate where cells are 80% confluent.
No less efficiency if the plate is full.
experiments done on hela cells, hek 293 and hek 293pack cells




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