Cryopreservation of mammalian cells

The cryopreservation of mammalian cells can avoid losses due to contamination, minimize genetic changes in continuous cell lines, and avoid aging and transformation of limited cell lines. in

Prior to preservation at cryogenic temperatures, the cells must be identified and checked for contamination. Listed below are several common media that can be used to freeze cells. For serum-containing media, commonly used media include:

Complete medium containing 10% glycerol.

Complete medium containing 10% dimethyl sulfoxide (DMSO).

Mixed medium of 50% cell conditioned medium and 50% fresh medium containing 10% glycerol.

Mixed medium of 50% cell conditioned medium and 50% fresh medium containing 10% DMSO.

For serum-free media, commonly used media include:

Mixed medium of 50% serum-free cell conditioned medium and 50% fresh serum-free medium containing 7.5% DMSO.

Fresh serum-free medium containing 7.5% DMSO and 10% cell culture grade BSA.

Suspension cultured cells

1. Calculate the number of living cells to be preserved at deep temperature. The cells should be in log phase. Centrifuge the cells at 200-400Xg for 5 minutes to pellet the cells. Aspirate the supernatant with a pipette and try to aspirate it without disturbing the cells.

2. For serum-containing medium, resuspend the cells in the freezing medium at a concentration of 1X107-5X107cells / ml; for serum-free culture; the resuspended concentration should be 0.5 × 107-1 × 107cells / ml.

3. Separately aliquot them into cryovials. Place the cryotube in an ice-water mixture, or store it in a refrigerator at 4 ° C, and start the following cryopreservation steps within 5 minutes.

4. Slowly freeze the cells at a rate of 1 ° C / min. You can use the adjusted freezer for cryopreservation, or put the cryopreservation tube in an insulated box, and then put it in -70 ℃ --90 ℃

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