Freezing Chicken Primordial Germ Cells

- 2006 -

Cool cell suspension containing 20% ES-FBS 199 and approximately 2 x 10⁶ cells/ml to 4-5ºC for one hour.  Perform the remaining steps at to 4-5ºC. 

Centrifuge suspension for 5 minutes at 600 x g and discard supernatant.  Add 150µl of pre-cooled freeze media containing 10% ethylene glycol.  Load cell suspension in to a 0.5ml straw and heat seal.  The straws are frozen at a rate of 1ºC/minute until a final temperature of -85ºC is reached in a programmable freezer (Mini Digitcool UJ40, Cryo BioSystem, Paris, France).  The straws are then plunged in to liquid nitrogen for storage.  Straws are thawed in a 37ºC water bath for 30 seconds.

Recipe:
20% Embryonic Stem Cell-Qualified Fetal Bovine Serum
1% Penicillin/Streptomycin
10% Ethylene Glycol
69% media 199

Reference:
Moore, D.T., P.H. Purdy, and H.D. Blackburn. 2006. A Method for Preserving Chicken Primordial Germ Cells.  Poult. Sci. 85(10):1784-1790.

Obtain information from Harvey Blackburn at: hblackbu@lamar.colostate.edu