Hello! I'm a little bit confused - all of my life I was taught, that cells must be preserved in a mixture of 90% FBS + 10% DMSO (maybe 5% EG + 5% DMSO, for stem cells). But right now I'll have to cryopreserve a lot of cell lines in a big quantity at once. I made a little calculation, and I simply can not spend 0,6 L of serum for this task. For your interest - all of these lines are well characterized and well established cancer cultures.
Some of my colleagues insists that it is completely fine to cryoreserve cancer cells lines (not primary cultures) in a mixture of full growth media + 10% DMSO.
Is that true? Or I'll have a cryostorage full of dead cells at the end?
Hello Dmitry Lifanov
Yes, it is possible to cryopreserve cells in full growth media + DMSO.
The most common freezing medium is 90% FBS + 10% DMSO. FBS, with its high protein content, protects cells against shear forces and gives the medium a desirable osmotic environment with a physiological viscosity. As a result, cells frozen in 90% FBS and 10% DMSO have a better cell viability on thawing as compared to the cells cryopreserved in full growth media containing 10% FBS + 10% DMSO. Nevertheless, the use of a mixture of growth media containing 10% FBS + 10% DMSO to freeze cells is still a good option depending on the type of cell line.
You may cryopreserve the cancer cell lines in a mixture of full growth media + 10% DMSO. It is not true that you will finally end up with dead cells using full growth media + 10% DMSO. It is only that cells will revive better if you use a freezing medium containing 90% FBS +10% DMSO than using a freezing medium that contain 10% FBS + 80% growth medium +10% DMSO.
Best.
Malcom is correct, I used to freeze down run of the mill cancer cell lines with complete medium + 5-10% dmso. As an aside, I used to often heavily concentrate my vials to ensure I had a ton of cells post thaw, e.g. 3-4 vials per T175.
Hello Dmitry!
There is an alternative freezing medium that was published in Experimental Hematology back in the 1990s. It is called pentastarch freezing medium. It used serum albumin in place of FBS and has a reduced final DMSO concentration of 5% instead of 10%, and the frozen samples are stored at -80oC, not liquid nitrogen. IIRC, the lead author was Pat Stiff. The cancer institute I worked at used to use it routinely for freezing patient peripheral blood stem cell leukapheresis products. The hook was to never exceed a final cell concentration of 10^8 cells per milliliter. I've retired so I do not have the reference at hand.
Yes, we can freeze cells in the complete growth media along with 10% FBS + 10%DMSO. Cells stay healthy and survive well too on thawing afterwards.
Regular media (DMEM) with 10%FBS and 5%DMSO, absolutely will work, but use cryocan stepwise transfer from refrigerator to -80 degree celcius. Cells will survive minimum 4 -6 months.
Hello everybody! I just used full growth medium already with 10% FBS, then I took 1,5 grams of BSA, dissolved this in 45 ml of medium, filtered through 0,2um , added 5 ml of DMSO and used this mixture as a cryopreservation medium.
Yesterday I thawed some of the vials which were frozen almost 3 weeks ago in this mixture in liquid nitrogen. Viability is the same, as if I'd used 90% FBS + 10% DMSO.
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