Cryopreservation

[Graham Dellaire]

In my lab we freeze cells as follows

trypsinize and spin down 1000 rpm 5 min
take cells up in 1 ml of DMEM with 20% FBS (cold)
add drop wise 1ml of Freezing medium (DMEM 20% DMSO)
resuspend gently and aliquote into two cryotubes (ie one ml each)
*pre chill cryotubes to atleast -4 but -20 or -80 is better

then there are many freezing/cooling chambers available
that utulize isopropyl alcohol etc...

the easiest is to put your cells in a styrofoam tube box (like for PCR products)
and put this at -80 c overnight 
in the morning place in liquid nitrogen

Ideally your cells should drop temperature by 1 degree a minute 
also you may want to change FBS content or resuspend cells in original 
supernatent as it may contain extracellular factors that may help your cells survive the
DEEP FREEZE <grin>

hope this helps


G>D>



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Graham Dellaire			    Snail Mail:
                                    Red Cross, Research		
McGill University                   Montreal Blood Services	  	
Faculty of Medicine                 3131 Sherbrooke St. East         
Div. of Experimental Medicine       Montreal, QC, Canada           
E-mail: popa0206 at po-box.mcgill.ca   H1W 1B2			   
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