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This protocol may be used to thaw and plate mouse embryonic stem cells.

Record History
Added on June 2, 2010 at 3:18 PM by Lindner, Jill
Modified on July 2, 2010 at 11:29 AM by Lindner, Jill
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TMESCSR: Protocols

Reagents, supplies, and equipment:

  • Previously prepared MEF feeders (usually a T25 or 60 mm culture dish)
  • cES cell media
  • 15 ml conical tube


  1. Quickly (~2 min) thaw one 1 ml vial ES cells in 37°C water bath.
  2. Remove thawed ES cells to a 15 ml conical tube.  Add 9 ml warm cES cell media.
  3. Centrifuge at 1000 rpm for 5 min.
  4. Aspirate supernatant from centrifuged conical tube and media from MEF plate.
  5. Resuspend cells in 5 ml cES cell media.
  6. Add suspension to MEF plate.  Gently rock plate to evenly disperse cells.
  7. Incubate at 37°C.

In approximately 2 days, plate(s) should be ready to expand to various concentrations.

Source: TMESCSR 4/10