Studies on the mechanisms of mammalian cell killing by a freeze-thaw cycle: conditions that prevent cell killing using nucleated freezing

Journal Article · Fri Apr 01 04:00:00 EST 1988 · Cryobiology; (United States)

DOI:https://doi.org/10.1016/0011-2240(88)90004-1· OSTI ID:6856102

Shier, W T

Normally a freeze-thaw cycle is a very efficient method of killing mammalian cells. However, this report describes conditions that prevent killing of cultured mammalian cells by nucleated freezing at -24 degrees C. Optimal protection from cell killing at -24 degrees C was obtained in isotonic solutions containing an organic cryoprotectant such as dimethyl sulfoxide (DMSO; 10%, v/v), a saccharide such as sucrose over a broad concentration range from 50 to 150 mM, and glucose. Glycerol was also an effective cryoprotectant but other organic solvents were ineffective, although in some cases they appeared to protect cell membranes, while not protecting other vital components. A wide variety of saccharide structures were effective at protecting cells from freeze-thaw killing, with trehalose being particularly effective. The degree of resistance to killing by a freeze-thaw cycle under these conditions varied widely among different cell lines. If toxicity of DMSO was responsible for this variability of cryoprotection, it must have been due to short-term, not longer term, toxicity of DMSO. Studies on the mechanism by which cells are protected from killing under these conditions indicated that neither vitrification of the medium nor the concentrating of components during freezing were involved. One model not eliminated by the mechanistic studies proposes that the organic solvent cryoprotectant component acts by fluidizing membranes under the thawing conditions, so that any holes produced by ice crystals propagating through membranes can reseal during the thawing process. In this model one of the mechanisms by which the saccharide component could act is by entering the cells and stabilizing vital intracellular components. Consistent with this, a freeze-thaw cycle promoted the uptake of labeled sucrose into cultured cells.

• Research Organization: Univ. of Minnesota, Minneapolis (USA)
• OSTI ID: 6856102
• Journal Information: Cryobiology; (United States), Journal Name: Cryobiology; (United States) Vol. 25:2; ISSN CRYBA
• Country of Publication: United States
• Language: English

Related Subjects

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59 BASIC BIOLOGICAL SCIENCES
63 RADIATION, THERMAL, AND OTHER ENVIRON. POLLUTANT EFFECTS ON LIVING ORGS. AND BIOL. MAT.
ALCOHOLS
ALDEHYDES
ANIMAL CELLS
ANIMALS
BIOCHEMICAL REACTION KINETICS
BIOLOGICAL EFFECTS
BIOLOGY
CARBOHYDRATES
CELL CONSTITUENTS
CELL CULTURES
CELL KILLING
CELL MEMBRANES
CRYOBIOLOGY
DMSO
FREEZING
GLUCOSE
GLYCEROL
HEXOSES
HYDROXY COMPOUNDS
ISOTOPE APPLICATIONS
ISOTOPES
KINETICS
MAMMALS
MEMBRANES
MONOSACCHARIDES
ORGANIC COMPOUNDS
ORGANIC SOLVENTS
ORGANIC SULFUR COMPOUNDS
RADIOISOTOPES
REACTION KINETICS
SACCHARIDES
SOLVENTS
SULFOXIDES
THAWING
TOXICITY
TRACER TECHNIQUES
VERTEBRATES