Cryopreservation - AbsoluteAstronomy.com

Cryopreservation is a process where cells

Cell (biology)

The cell is the basic structural and functional unit of all known living organisms. It is the smallest unit of life that is classified as a living thing, and is often called the building block of life. The Alberts text discusses how the "cellular building blocks" move to shape developing embryos....

or whole tissues

Biological tissue

Tissue is a cellular organizational level intermediate between cells and a complete organism. A tissue is an ensemble of cells, not necessarily identical, but from the same origin, that together carry out a specific function. These are called tissues because of their identical functioning...

are preserved by cooling to low sub-zero temperature

Temperature

Temperature is a physical property of matter that quantitatively expresses the common notions of hot and cold. Objects of low temperature are cold, while various degrees of higher temperatures are referred to as warm or hot...

s, such as (typically) 77 K or −196 °C (the boiling point of liquid nitrogen

Liquid nitrogen

Liquid nitrogen is nitrogen in a liquid state at a very low temperature. It is produced industrially by fractional distillation of liquid air. Liquid nitrogen is a colourless clear liquid with density of 0.807 g/mL at its boiling point and a dielectric constant of 1.4...

). At these low temperatures, any biological activity, including the biochemical reactions that would lead to cell death, is effectively stopped. However, when cryoprotectant

Cryoprotectant

A cryoprotectant is a substance that is used to protect biological tissue from freezing damage . Arctic and Antarctic insects, fish, amphibians and reptiles create cryoprotectants in their bodies to minimize freezing damage during cold winter periods. Insects most often use sugars or polyols as...

solutions are not used, the cells

Cell (biology)

being preserved are often damaged due to freezing

Freezing

Freezing or solidification is a phase change in which a liquid turns into a solid when its temperature is lowered below its freezing point. The reverse process is melting....

during the approach to low temperatures or warming to room temperature.

Temperature

Cryogenic storage at very low temperatures is presumed to provide an indefinite, if not near infinite, longevity to cells although the actual “shelf life” is rather difficult to prove. In experiments with dried seeds, researchers found that there was noticeable variability in deterioration when samples were kept at different ‘frozen’ temperatures–even ultra cold ones. Temperatures below the glass transition point (Tg) of polyol's water

Water

Water is a chemical substance with the chemical formula H2O. A water molecule contains one oxygen and two hydrogen atoms connected by covalent bonds. Water is a liquid at ambient conditions, but it often co-exists on Earth with its solid state, ice, and gaseous state . Water also exists in a...

solutions (around minus 136°C) appear to be accepted as the range where biological activity

Metabolism

Metabolism is the set of chemical reactions that happen in the cells of living organisms to sustain life. These processes allow organisms to grow and reproduce, maintain their structures, and respond to their environments. Metabolism is usually divided into two categories...

very substantially slows down, and minus 196°C (liquid phase of liquid nitrogen

Liquid nitrogen

) is the preferred temperature for the storage of important specimens. While fridges, deep freezers and extra cold deep freezers, all similar to domestic ones, are used for many items, generally the ultra cold of liquid nitrogen at -196°C is required for successful preservation of the more complex biological structures to virtually stop all biological activity.

Risks

Phenomena which can cause damage to cells during cryopreservation mainly occur during the freezing stage, and include: solution effects, extracellular

Extracellular

In cell biology, molecular biology and related fields, the word extracellular means "outside the cell". This space is usually taken to be outside the plasma membranes, and occupied by fluid...

ice formation, dehydration and intracellular

Intracellular

Not to be confused with intercellular, meaning "between cells".In cell biology, molecular biology and related fields, the word intracellular means "inside the cell".It is used in contrast to extracellular...

ice formation. Many of these effects can be reduced by cryoprotectant

Cryoprotectant

s.

When having reached the frozen stage, the preserved material is relatively safe from further damage. However, estimates based on the accumulation of radiation-induced DNA damage during cryogenic storage have suggested a maximum storage period of 1000 years.

Solution effects

As ice crystals grow in freezing water, solutes are excluded, causing them to become concentrated in the remaining liquid water. High concentrations of some solutes can be very damaging.

Extracellular ice formation

When tissues are cooled slowly, water migrates out of cells

Cell (biology)

and ice

Ice

Ice is water frozen into the solid state. Usually ice is the phase known as ice Ih, which is the most abundant of the varying solid phases on the Earth's surface. It can appear transparent or opaque bluish-white color, depending on the presence of impurities or air inclusions...

forms in the extracellular space. Too much extracellular ice can cause mechanical damage to the cell membrane due to crushing.

Dehydration

The migration of water causing extracellular ice formation can also cause cellular dehydration. The associated stresses on the cell can cause damage directly.

Intracellular ice formation

While some organism

Organism

In biology, an organism is any contiguous living system . In at least some form, all organisms are capable of response to stimuli, reproduction, growth and development, and maintenance of homoeostasis as a stable whole.An organism may either be unicellular or, as in the case of humans, comprise...

s and tissues

Biological tissue

can tolerate some extracellular ice, any appreciable intracellular ice is almost always fatal to cells.

Main methods to prevent risks

The main techniques to prevent cryopreservation damages are a well established combination of controlled rate and slow freezing on one hand, and a newer flash-freezing process known as vitrification on the other.

Slow programmable freezing

Controlled-rate and slow freezing, also called slow programmable freezing (SPF), is a set of well established techniques pioneered in the early 1970s which enabled the first human embryo

Embryo

An embryo is a multicellular diploid eukaryote in its earliest stage of development, from the time of first cell division until birth, hatching, or germination...

frozen birth Zoe Leyland in 1984. Since then machines that freeze biological samples using programmable steps, or controlled rates, have been used all over the world for human, animal and cell biology – 'freezing down' a sample to better preserve it for eventual thawing, before it is deep frozen, or cryopreserved, in liquid nitrogen. Such machines are used for freezing oocyte, skin, blood products, embryo, sperm, stem cells and general tissue preservation in hospitals, veterinary practices and research labs around the world. As an example, estimates put the number of live births from frozen embryos 'slow frozen' at some 300,000 to 400,000 or 20% of the estimated 3 million IVF births.

Lethal intracellular freezing can be avoided if cooling is slow enough to permit sufficient water to leave the cell during progressive freezing of the extracellular fluid. That rate differs between cells of differing size and water permeability

Semipermeable membrane

A semipermeable membrane, also termed a selectively permeable membrane, a partially permeable membrane or a differentially permeable membrane, is a membrane that will allow certain molecules or ions to pass through it by diffusion and occasionally specialized "facilitated diffusion".The rate of...

: a typical cooling rate around 1°C/minute is appropriate for many mammalian cells after treatment with cryoprotectant

Cryoprotectant

s such as glycerol

Glycerol

Glycerol is a simple polyol compound. It is a colorless, odorless, viscous liquid that is widely used in pharmaceutical formulations. Glycerol has three hydroxyl groups that are responsible for its solubility in water and its hygroscopic nature. The glycerol backbone is central to all lipids...

or dimethyl sulphoxide, but the rate is not a universal optimum. The 1°C/minute rate can be easily achieved by using a rate-controlled freezer or a benchtop portable freezing container such as CoolCell.

Several independent studies have provided evidence that frozen embryos stored using slow-freezing techniques may in some ways be 'better' than fresh in IVF. The studies were presented at the American Society for Reproductive Medicine conference in San Francisco, US, 2008. The studies indicate that using frozen embryos rather than fresh embryos reduced the risk of stillbirth and premature delivery though the exact reasons are still being explored.

Vitrification

Researchers who have developed a new technique, vitrification, as of 2000 claim to provide the benefits of cryopreservation without damage due to ice crystal formation. In clinical cryopreservation, vitrification usually requires the addition of cryoprotectant

Cryoprotectant

s prior to cooling. The cryoprotectants act like antifreeze

Antifreeze

Antifreeze is a freeze preventive used in internal combustion engines and other heat transfer applications, such as HVAC chillers and solar water heaters....

: they lower the freezing temperature. They also increase the viscosity. Instead of crystal

Crystal

A crystal or crystalline solid is a solid material whose constituent atoms, molecules, or ions are arranged in an orderly repeating pattern extending in all three spatial dimensions. The scientific study of crystals and crystal formation is known as crystallography...

lizing, the syrupy solution turns into an amorphous ice

Amorphous ice

Amorphous ice is an amorphous solid form of water, meaning it consists of water molecules that are randomly arranged like the atoms of common glass. Everyday ice is a crystalline material where the atoms are regularly arranged in a lattice whereas amorphous ice is distinguished by a lack of...

—i.e., it vitrifies. Rather than a phase change from liquid to solid by crystallization, the amorphous state is like a "solid liquid", and the transformation is over a small temperature range described as the glass transition

Glass transition

The liquid-glass transition is the reversible transition in amorphous materials from a hard and relatively brittle state into a molten or rubber-like state. An amorphous solid that exhibits a glass transition is called a glass...

temperature.

Vitrification of water is promoted by rapid cooling, and can be achieved without cryoprotectants by an extremely rapid drop in temperature (megakelvins per second). The rate that is required to attain glassy state in pure water was considered to be impossible until 2005.

Two conditions usually required to allow vitrification are an increase in the viscosity and a depression of the freezing temperature. Many solutes do both, but larger molecules generally have larger effect, particularly on viscosity. Rapid cooling also promotes vitrification.

In established methods of cryopreservation, the solute must penetrate the cell membrane in order to achieve increased viscosity and depress freezing temperature inside the cell. Sugars do not readily permeate through the membrane. Those solutes that do, such as dimethyl sulfoxide

Dimethyl sulfoxide

Dimethyl sulfoxide is an organosulfur compound with the formula 2SO. This colorless liquid is an important polar aprotic solvent that dissolves both polar and nonpolar compounds and is miscible in a wide range of organic solvents as well as water...

, a common cryoprotectant, are often toxic in high concentration. One of the difficult compromises faced in vitrifying cryopreservation is limiting the damage produced by the cryoprotectant itself due to cryoprotectant toxicity. Mixtures of cryoprotectants and the use of ice blockers have enabled Twenty-First Century Medicine

Twenty-First Century Medicine

21st Century Medicine is a California cryobiological research company which has as its primary focus the development of perfusates and protocols for viable long-term cryopreservation of human organs, tissues and cells at cryogenic temperatures through the use of vitrification. Dr. Gregory M...

to vitrify a rabbit

Rabbit

Rabbits are small mammals in the family Leporidae of the order Lagomorpha, found in several parts of the world...

kidney

Kidney

The kidneys, organs with several functions, serve essential regulatory roles in most animals, including vertebrates and some invertebrates. They are essential in the urinary system and also serve homeostatic functions such as the regulation of electrolytes, maintenance of acid–base balance, and...

to -135°C with their proprietary vitrification cocktail. Upon rewarming, the kidney was successfully transplanted into a rabbit, with complete functionality and viability, able to sustain the rabbit indefinitely as the sole functioning kidney.

Freezable tissues

In general, cryopreservation is easier for thin samples and small clumps of individual cells, because these can be cooled more quickly and so require lower doses of toxic cryoprotectants. Therefore, the goal of cryopreserving human liver

Liver

The liver is a vital organ present in vertebrates and some other animals. It has a wide range of functions, including detoxification, protein synthesis, and production of biochemicals necessary for digestion...

s and heart

Heart

The heart is a myogenic muscular organ found in all animals with a circulatory system , that is responsible for pumping blood throughout the blood vessels by repeated, rhythmic contractions...

s for storage and transplant

Organ transplant

Organ transplantation is the moving of an organ from one body to another or from a donor site on the patient's own body, for the purpose of replacing the recipient's damaged or absent organ. The emerging field of regenerative medicine is allowing scientists and engineers to create organs to be...

is still some distance away.

Nevertheless, suitable combinations of cryoprotectants and regimes of cooling and rinsing during warming often allow the successful cryopreservation of biological materials, particularly cell suspensions or thin tissue samples. Examples include:

Semen
Semen
Semen is an organic fluid, also known as seminal fluid, that may contain spermatozoa. It is secreted by the gonads and other sexual organs of male or hermaphroditic animals and can fertilize female ova...
Blood
Blood
Blood is a specialized bodily fluid in animals that delivers necessary substances such as nutrients and oxygen to the cells and transports metabolic waste products away from those same cells....
- Special cells for transfusion
- Stem cell
  Stem cell
  This article is about the cell type. For the medical therapy, see Stem Cell TreatmentsStem cells are biological cells found in all multicellular organisms, that can divide and differentiate into diverse specialized cell types and can self-renew to produce more stem cells...
  
  s. It is optimal in high concentration of synthetic serum, stepwise equilibration and slow cooling.
- Umbilical cord blood Further information: Cord blood bank#Cryopreservation
Tissue samples like tumor
Tumor
A tumor or tumour is commonly used as a synonym for a neoplasm that appears enlarged in size. Tumor is not synonymous with cancer...

s and histological cross sections
Histology
Histology is the study of the microscopic anatomy of cells and tissues of plants and animals. It is performed by examining cells and tissues commonly by sectioning and staining; followed by examination under a light microscope or electron microscope...
Eggs (oocyte
Oocyte
An oocyte, ovocyte, or rarely ocyte, is a female gametocyte or germ cell involved in reproduction. In other words, it is an immature ovum, or egg cell. An oocyte is produced in the ovary during female gametogenesis. The female germ cells produce a primordial germ cell which undergoes a mitotic...

s) See oocyte cryopreservation
Oocyte cryopreservation
Human oocyte cryopreservation is a novel technology in which a woman’s eggs are extracted, frozen and stored. Later, when she is ready to become pregnant, the eggs can be thawed, fertilized, and transferred to the uterus as embryos.-History:Cryopreservation itself has always played a central role...
Embryos that are 2, 4 or 8 cells when frozen
Ovarian tissue
Plant
Plant
Plants are living organisms belonging to the kingdom Plantae. Precise definitions of the kingdom vary, but as the term is used here, plants include familiar organisms such as trees, flowers, herbs, bushes, grasses, vines, ferns, mosses, and green algae. The group is also called green plants or...

seeds or shoots may be cryopreserved for conservation
Conservation biology
Conservation biology is the scientific study of the nature and status of Earth's biodiversity with the aim of protecting species, their habitats, and ecosystems from excessive rates of extinction...

purposes.

In addition, efforts are underway to preserve humans cryogenically, known as cryonics

Cryonics

Cryonics is the low-temperature preservation of humans and animals who can no longer be sustained by contemporary medicine, with the hope that healing and resuscitation may be possible in the future. Cryopreservation of people or large animals is not reversible with current technology...

. In such efforts either the brain within the head or the entire body may undergo the above process. Cryonics is in a different category from the aforementioned examples, however: while countless cryopreserved cells, vaccines, tissue and other biologial samples have been thawed and successfully used, this has not yet been the case at all for cryopreserved brains or bodies. At issue are the criteria for defining "success". Proponents of cryonics claim that cryopreservation using present technology, particularly vitrification of the brain, may be sufficient to preserve people in an "information theoretic

Information theoretical death

Information-theoretic death is the destruction of the information within a human brain to such an extent that recovery of the original person is theoretically impossible by any physical means...

" sense so that they could be revived and made whole by hypothetical vastly advanced future technology.

Semen

Semen is an organic fluid, also known as seminal fluid, that may contain spermatozoa. It is secreted by the gonads and other sexual organs of male or hermaphroditic animals and can fertilize female ova...

can be used successfully almost indefinitely after cryopreservation. The longest reported successful storage is 22 years. It can be used for sperm donation

Sperm donation

Sperm donation is the provision by a man, , of his sperm, with the intention that it be used to impregnate a woman who is not usually the man's sexual partner, in order to produce a child....

where the recipient wants the treatment in a different time or place, or for men undergoing a vasectomy

Vasectomy

Vasectomy is a surgical procedure for male sterilization and/or permanent birth control. During the procedure, the vasa deferentia of a man are severed, and then tied/sealed in a manner such to prevent sperm from entering into the seminal stream...

to still have the option to have children.

Testicular tissue

Cryopreservation of immature testicular tissue is a developing method to avail reproduction to young boys who need to go through gonadotoxic therapies. Animal data look promising, since healthy offsprings have been obtained after transplantation of frozen testicular cell suspensions or tissue pieces. However, none of the fertility restoration options from frozen tissue, i.e. cell suspension transplantation, tissue grafting and in vitro maturation

In vitro maturation

In vitro maturation is the technique of letting ovarian follicles mature in vitro.-Techniques available:The ability of in IVM depends on how mature the follicle already is...

(IVM) has proved efficient and safe in humans as yet.

Oocytes

Human Oocyte cryopreservation is a new technology in which a woman’s eggs (oocytes) are extracted, frozen and stored. Later, when she is ready to become pregnant, the eggs can be thawed, fertilized, and transferred to the uterus as embryos.

Embryos

Cryopreservation for embryos are used for embryo storage, e.g. when in vitro fertilization has resulted in more embryos than is currently needed.

Pregnancies have been reported from embryos stored for 16 years. Many studies have evaluated the children born from frozen embryos, or “frosties”. The result has uniformly been positive with no increase in birth defects or development abnormalities. A study of more than 11,000 cryopreserved human embryos showed no significant effect of storage time on postthaw survival for IVF or oocyte donation cycles, or for embryos frozen at the pronuclear or cleavage stages. In addition, the duration of storage had no significant effect on clinical pregnancy, miscarriage, implantation, or live birth rate, whether from IVF or oocyte donation cycles. Rather, oocyte age, survival proportion, and number of transferred embryos are predictors of pregnancy outcome.

Ovarian tissue

Cryopreservation of ovarian tissue is of interest to women who want to preserve their reproductive function beyond the natural limit, or whose reproductive potential is threatened by cancer therapy, for example in hematologic malignancies or breast cancer. The procedure is to take a part of the ovary and carry out slow freezing before storing it in liquid nitrogen whilst therapy is undertaken. Tissue can then be thawed and implanted near the fallopian, either orthotopic (on the natural location) or heterotopic (on the abdominal wall), where it starts to produce new eggs, allowing normal conception to take place. The ovarian tissue may also be transplanted into mice that are immunocompromised (SCID mice) to avoid graft rejection, and tissue can be harvested later when mature follicles have developed.

Moss

Cryopreservation of whole moss

Moss

Mosses are small, soft plants that are typically 1–10 cm tall, though some species are much larger. They commonly grow close together in clumps or mats in damp or shady locations. They do not have flowers or seeds, and their simple leaves cover the thin wiry stems...

plant

Plant

Plants are living organisms belonging to the kingdom Plantae. Precise definitions of the kingdom vary, but as the term is used here, plants include familiar organisms such as trees, flowers, herbs, bushes, grasses, vines, ferns, mosses, and green algae. The group is also called green plants or...

s, especially Physcomitrella patens

Physcomitrella patens

Physcomitrella patens is a moss used as a model organism for studies on plant evolution, development and physiology.-Model organism:...

, has been developed by Ralf Reski

Ralf Reski

Ralf Reski is a German Professor of Plant Biotechnology and former Dean of the Faculty of Biology of the University of Freiburg...

and coworkers and is performed at the International Moss Stock Center

International Moss Stock Center

The International Moss Stock Center , located at the Albert-Ludwigs-University of Freiburg, is an international resource center which is specialized in collecting, preserving and distributing moss plants of a high value of scientific research.-Moss collection:The moss collection of the IMSC...

. This biobank

Biobank

A biobank is a cryogenic storage facility used to archive biological samples for use in research and experiments. Ranging in size from individual refrigerators to warehouses, biobanks are maintained by institutions such as hospitals, universities, nonprofit organizations, and pharmaceutical...

collects, preserves, and distributes moss mutant

Mutant

In biology and especially genetics, a mutant is an individual, organism, or new genetic character, arising or resulting from an instance of mutation, which is a base-pair sequence change within the DNA of a gene or chromosome of an organism resulting in the creation of a new character or trait not...

s and moss ecoptype

Ecotype

In evolutionary ecology, an ecotype,Greek: οίκος = home and τύπος = type, coined by Göte Turesson in 1922 sometimes called ecospecies, describes a genetically distinct geographic variety, population or race within species , which is adapted to specific environmental conditions.Typically, ecotypes...

Natural cryopreservation

Water bears (Tardigrada), microscopic multicellular organisms, can survive freezing

Freezing

Freezing or solidification is a phase change in which a liquid turns into a solid when its temperature is lowered below its freezing point. The reverse process is melting....

at low temperatures by replacing most of their internal water with the sugar

Sugar

Sugar is a class of edible crystalline carbohydrates, mainly sucrose, lactose, and fructose, characterized by a sweet flavor.Sucrose in its refined form primarily comes from sugar cane and sugar beet...

trehalose

Trehalose

Trehalose, also known as mycose or tremalose, is a natural alpha-linked disaccharide formed by an α,α-1,1-glucoside bond between two α-glucose units. In 1832, H.A.L. Wiggers discovered trehalose in an ergot of rye, and in 1859 Marcellin Berthelot isolated it from trehala manna, a substance made...

, preventing it from crystallization that otherwise damage cell membranes. Mixtures of solutes can achieve similar effects. Some solutes, including salts, have the disadvantage that they may be toxic at high concentrations. In addition to the Water bear, wood frogs can tolerate the freezing of their blood and other tissues. Urea is accumulated in tissues in preparation for overwintering, and liver glycogen is converted in large quantities to glucose in response to internal ice formation. Both urea and glucose act as "cryoprotectants" to limit the amount of ice that forms and to reduce osmotic shrinkage of cells. Frogs can survive many freeze/thaw events during winter if not more than about 65% of the total body water freezes. Research exploring the phenomenon of "Freezing frogs" has been primarily carried out by the Canadian researcher, Dr. Kenneth B. Storey

Kenneth B. Storey

-Boards:* Journal of Thermal Biology * Journal of Comparative Physiology B * Past member: American Journal of Physiology, Molecular Physiology, Copeia, J...

.

Freeze tolerance, in which organisms survive the winter by freezing solid and ceasing life functions, is known in a few vertebrates: five species of frogs (Rana sylvatica, Pseudacris triseriata, Hyla crucifer, Hyla versicolor, Hyla chrysoscelis
Hyla chrysoscelis
The Cope's Grey Tree Frog is a species of tree frog which is found in the United States. It is almost indistinguishable from the Grey Tree Frog, Hyla versicolor, and shares much of its geographic range. Both species are variable in color, mottled gray to gray-green, resembling the bark of trees...

), one salamander (Hynobius
Hynobius
Hynobius or Asian Salamanders is a genus of salamander in the Hynobiidae family, endemic to Japan, Korea, China and Far East Russia.It contains the following species:-Species:...

keyserlingi), one snake (Thamnophis sirtalis) and three turtles (Chrysemys picta, Terrapene carolina
Terrapene carolina
The common box turtle is a species of box turtle with six existing subspecies. It is found throughout the eastern United States and Mexico. The box turtle has a distinctive hinged lowered shell that allows it to completely enclose itself...

, Terrapene ornata
Terrapene ornata
Terrapene ornata is a species of North American box turtle sometimes referred to as the western box turtle or ornate box turtle.- Taxonomy :There are two subspecies of T. ornata:...

). Snapping turtle Chelydra serpentina and wall lizard Podarcis muralis also survive nominal freezing but it has not been established to be adaptive for overwintering. In the case of Rana sylvatica one cryopreservant is ordinary glucose, which rises in concentration by approximately 19 mmol/l when the frogs are slowly cooled;

History

One of the most important early workers on the theory of cryopreservation was James Lovelock

James Lovelock

James Lovelock, CH, CBE, FRS is an independent scientist, environmentalist and futurologist who lives in Devon, England. He is best known for proposing the Gaia hypothesis, which postulates that the biosphere is a self-regulating entity with the capacity to keep our planet healthy by controlling...

of Gaia theory fame. He suggested that damage to red blood

Blood

Blood is a specialized bodily fluid in animals that delivers necessary substances such as nutrients and oxygen to the cells and transports metabolic waste products away from those same cells....

cells during freezing was due to osmotic

Osmosis

Osmosis is the movement of solvent molecules through a selectively permeable membrane into a region of higher solute concentration, aiming to equalize the solute concentrations on the two sides...

stresses. Lovelock in early 1950s had also suggested that increasing salt concentrations in a cell as it dehydrates to lose water to the external ice might cause damages to the cell. Cryopreservation of tissue in recent times started with the freezing of fowl sperm, which in 1957 was cryopreserved by a team of scientists in the UK led by Christopher Polge. The process moved into the human world in the 1950s with pregnancies obtained after insemination of frozen sperm. However, the rapid immersion of the samples in liquid nitrogen did not, for certain of these samples–such as types of embryos, bone marrow and stem cells–produce the necessary viability to make them usable on thawing. Increased understanding of the mechanism of freezing injury to cells emphasised the importance of controlled or slow cooling to obtain maximum survival on thawing of the living cells. A controlled rate cooling process, allowing biological samples to equilibrate to optimal physical parameters osmotically in a cryoprotectant (a form of anti-freeze) before cooling in a predetermined, controlled way proved necessary. The ability of cryoprotectants, in the early cases glycerol, to protect cells from freezing injury was discovered accidentally. Freezing injury has two aspects–direct damage from the ice crystals and secondary damage caused by the increase in concentration of solutes as progressively more ice is formed. In 1963 Peter Mazur, at Oak Ridge National Laboratory in the USA, showed that lethal intracellular freezing could be avoided if cooling was slow enough to permit sufficient water to leave the cell during progressive freezing of the extracellular fluid. That rate differs between cells of differing size and water permeability: a typical cooling rate around 1°C/minute is appropriate for many mammalian cells after treatment with cryoprotectants such as glycerol or dimethyl sulphoxide, but the rate is not a universal optimum.

External links

The source of this article is wikipedia, the free encyclopedia. The text of this article is licensed under the GFDL.

Temperature

Risks

Solution effects

Extracellular ice formation

Dehydration

Intracellular ice formation

Main methods to prevent risks

Slow programmable freezing

Vitrification

Freezable tissues

Semen

Testicular tissue

Oocytes

Embryos

Ovarian tissue

Moss

Natural cryopreservation

History

See also

External links