Tissue culture media freezing medium

There has been an increase in the use of cadaveric heart valves for patients with valvular defects. The valves are best stored by freezing but some success has been achieved by simple cold storage in an antibiotic medium made up of ingredients common to most tissue culture solutions. At a storage temperature of 4 °C there is a continual loss of viability of fibroblasts so that by three weeks there are practically no viable cells and the valves cannot be used. 

To freeze uninfected and infected HL60 cells, transfer media and cells from tissue-culture flask into 50 mL conical tubes. Centrifuge at 450 x g for 5 min to pellet cells. Decant or aspirate supernatant and discard. Resuspend 1 mL of cells at a final concentration of 5 x 10 cells/mL in freezing medium. Transfer contents ( 0.5 mL per tube) into NUNC cryovials. Freeze at -80 °C overnight, then transfer vials to Uquid nitrogen the next day. Freeze infected HL60 cells at 90% infection level. 

The nucleotide patterns of cultured animal cells and ascites tumor cells are generally less vulnerable than those of tissues however, if washing is required, a medium capable of supporting energy metabolism should be employed. If cells are to be packed by centrifugation prior to extraction, cultures or ascitic fluids may first have to be cooled. Cells are extracted in the cold some procedures employ alternate freezing and thawing in the presence of acidic extractants. Extraction of cultured mouse embryo cells with 60% methanol for 16 hours at — 20 C has been employed in the analysis of deoxyribonucleoside triphosphates 9). 

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