Submitted by Lauren French (download pdf)



  • Wear glasses and gloves before opening the liquid nitrogen tank.
  • Take the cell vial out from the liquid nitrogen tank, put the vial into an ice bucket immediately.
  • Put 4 mls complete media into a new tissue culture T25 plate or P10 if densely frozen (collagen-coated).
  • Place cells in a 37°C water bath, thaw as quickly as possible, less than 60 seconds.
  • Transfer to waiting T25 flask.
  • The next day, replace all media with new complete media.  Watch closely and expand to P10 or T25 when ready.


Note: Growth rate will be slow after a freeze, and cell death always occurs in some fraction.  Growing on a smaller surface will more readily allow cells to become healthy.

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