I am growing mESCs in 2i medium. I have cryopreserved mESCs cultured in DMEM+FBS in 90%FBS+10% DMSO but would like to know how to cryopreserve cells grown in 2i medium which is free of serum. I prepared few freezedowns in synth-a-freeze medium but the viability on thawing is very poor. Can you suggest me what freezing medium can I use?
HI Neeraj,
We use a product from StemCell Technologies called CryoStor. It is zeno free and works great here is the link:
http://www.stemcell.com/en/Products/All-Products/CryoStorCS10.aspx
A friend of mine has recommended a product from Essential Pharma
( http://www.cryo-ess.com/) We have ordered some but not tried it yet.
I'm curious how the 2i is working for you perhaps we could talk offline about it? ([email protected]).
Kind regards,
Steve
Hi Neeraj
Best practice is to cryopreserve in commercially available freezing media.
If its not available. Try to use the used growth media from the flask + 10 percent DMSO, rather than using the fresh media . It has good effect on cell viability upon thawing. Always consider to count cell number when freezing.
Hope it helps.
Kind regards
Bilal
Hi Neeraj. You can either use Serum Replacement solution plus 10% DMSO or normal serum+Lif media plus 10% DMSO. In Both cases I found the cells recover.
Hello
To Cryopreservation medium is 80% culture medium (FBS or serum free ES cell medium, depending on the medium used for the mESC line), 10% (v/v) FBS and 10% (v/v) DMSO. Store medium at 4°C for up to one week.
http://online.liebertpub.com/doi/pdf/10.1089/scd.2011.0283
I am freezing the cells in 90% ES-FBS and 10% DMSO, and the cells are fine. The revival is also good. Upon thaw, I dilute them with DMEM, give them a spin to remove FBS and DMSO, and plate them in 2i medium.
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