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Cryobiology is the branch of biology that studies the effects of low temperatures on living things. The word cryobiology is derived from the Greek words "cryo" = cold, "bios" = life, and "logos" = science. It means the science of life at low temperature. In practice, cryobiology is the study of biological material or systems at temperatures below normal. Materials or systems studied may include proteins, cells, tissues, organs, or whole organisms. Temperatures may range from moderately hypothermic conditions to cryogenic temperatures.





The study of life at low temperatures.


The branch of physics and engineering that studies the production and use of very low temperatures. Cryogenics is not cryonics, although people often confuse them.


The low temperature preservation of humans and animals in a damaged state with the intention of future revival. Cryonics is not part of mainstream cryobiology because cryonics still depends heavily on speculative future technology.


Major areas of study in cryobiology


At least 6 major areas of study in cryobiology can be identified:

1.    Study of cold-adaptation of microorganisms, plants (= cold hardiness), invertebrates, and animals (= hibernation).

2.    Cryopreservation of cells, tissues, gametes, and embryos of animal and human origin for (medical) purposes of long-term storage. This usually requires the addition of substances which protect the cells during freezing and thawing (cryoprotectants).

3.    Preservation of organs under hypothermic conditions for transplantation.

4.    Lyophilization (freeze-drying) of pharmaceuticals.

5.    Cryosurgery, a (minimally) invasive approach for the destruction of unhealthy tissue using cryogenic gases/fluids.

6.    Physics of super cooling, ice nucleation/growth and mechanical engineering aspects of heat transfer during cooling and warming.


Historical background


Cryobiology history can be traced back to antiquity. As early as in 2500 BC low temperatures were used in Egypt in medicine. The use of cold was recommended by Hippocrates to stop bleeding and swelling. With the emergence of modern science, Robert Boyle studied the effects of low temperatures on animals.




In 1949 sperm was cryopreserved for the first time by a team of scientists led by Christopher Polge (1926-2006). This led to a much wider use of cryopreservation today, with many organs, tissues and cells routinely stored in low temperatures. Large organs such as hearts are usually stored and transported, for short times only, at cool but not freezing temperatures for transplantation. Cell suspensions (like blood and semen) and thin tissue sections can sometimes be stored almost indefinitely at liquid nitrogen temperature (cryopreservation). Human sperm, eggs and embryos are routinely stored in fertility research and treatments. In the early 2000s a baby was born from a cryopreserved egg fertilized by a cryopreserved sperm.


Cryosurgery (intended and controlled tissue destruction by ice formation) was carried out by James Arnott in 1845 in an operation on a patient with cancer. Although not very widespread, cryosurgery has its benefits. Hypothermia, e.g. during heart surgery on a "cold" heart (generated by cold perfusion without any ice formation) allows for much longer operations and improves recovery rates for patients.


Cryobiology in Humans


Cryobiology in humans with regards to infertility is also called vitrification which is a process in which a woman’s eggs are flash frozen in a matter of seconds. Freezing the eggs at such a rapid rate greatly improves their survival rate and prevents ice crystals from forming in the egg that could damage the DNA. Once the couple is ready to conceive the eggs are thawed and through the process of IVF sperm is placed with the eggs and together they are placed back into the uterus. Vitrification has its glitches and is not as reliable as freezing fertilized eggs or embryos because eggs alone are extremely sensitive to temperature. Many researchers are also freezing ovarian tissue in conjunction with the eggs in hopes that the ovarian tissue can be transplanted back into the uterus, stimulating normal ovulation cycles.


Applied cryobiology


Cryobiology as an applied science is primarily concerned with low temperature preservation. Hypothermic storage is typically above 0°C but below normothermic (32°C to 37°C) mammalian temperatures. Storage by cryopreservation, on the other hand, will be in the −80°C to −196°C temperature range. Organs and tissues are more frequently the objects of hypothermic storage, whereas single cells have been the most common objects cryopreserved.


A rule of thumb in hypothermic storage is that every 10°C reduction in temperature is accompanied by a 50% decrease in oxygen consumption.  Although hibernating animals have adapted mechanisms to avoid metabolic imbalances associated with hypothermia, hypothermic organs and tissues being maintained for transplantation require special preservation solutions to counter acidosis, depressed sodium pump activity and increased intracellular calcium. Special organ preservation solutions such as Viaspan (University of Wisconsin solution), HTK, and Celsior have been designed for this purpose.  These solutions also contain ingredients to minimize damage by free radicals, prevent edema, compensate for ATP loss, etc.


Cryopreservation of cells is guided by the "Two-Factor Hypothesis" of American cryobiologist Peter Mazur, which states that excessively rapid cooling kills cells by intracellular ice formation and excessively slow cooling kills cells by either electrolyte toxicity or mechanical crushing.  During slow cooling ice forms extracellularly, causing water to osmotically leave cells, thereby dehydrates them. Intracellular ice can be much more damaging than extracellular ice.


For red blood cells the optimum cooling rate is very rapid (nearly 100°C per second), whereas for stem cells the optimum cooling rate is very slow (1°C per minute). Cryoprotectants, such as DMSO (dimethyl sulfoxide) and glycerol, are used to protect cells from freezing. A variety of cell types are protected by 10% DMSO. Cryobiologists attempt to optimize cryoprotectant concentration (minimizing both ice formation and toxicity) as well as cooling rate. Cells may be cooled at an optimum cooling rate to a temperature between −30°C and −40°C before being plunged into liquid nitrogen.


Slow cooling methods rely on the fact that cells contain few nucleating agents, but contain naturally-occurring vitrifying substances that can prevent ice formation in cells that have been moderately dehydrated. Cryobiologists are increasingly using mixtures of cryoprotectants for full vitrification (zero ice formation) in preservation of cells, tissues and organs. Vitrification methods pose a challenge in the requirement to search for cryoprotectant mixtures that can minimize toxicity.


Scientific societies


The Society for Cryobiology was founded in 1964 to bring together those from the biological, medical and physical sciences who have a common interest in the effect of low temperatures on biological systems. As of 2007 the Society for Cryobiology had approximately 280 members from around the world, and one half of them are US based. The purpose of the Society is to promote scientific research in low temperature biology, to improve scientific understanding in this field, and to disseminate and apply this knowledge to the benefit of mankind. The Society organizes an annual scientific meeting dedicated to all aspects of low-temperature biology. This international meeting offers opportunities for presentation and discussion of the most up-to-date research in cryobiology as well as reviewing specific aspects through symposia and workshops. Members are also kept informed of news and forthcoming meetings through the Society newsletter, News Notes. The 2008-2009 President of the Society for Cryobiology is John K. Critser.


The Society for Low Temperature Biology was founded in 1964 and became a Registered Charity in 2003 (Charity Commission for England & Wales No. 1099747) with the purpose of promoting research into the effects of low temperatures on all types of organisms and their constituent cells, tissues and organs. As of 2006 the Society for Low Temperature Biology had approximately 130 (mostly British and European) members and holds at least one Annual General Meeting. The program usually includes both a symposium on a topical subject and a session of free communications on any aspect of low temperature biology. Recent symposia have included long-term stability, preservation of aquatic organisms, cryopreservation of embryos and gametes, preservation of plants, low temperature microscopy, vitrification (glass formation of aqueous systems during cooling), freeze drying and tissue banking. Members are informed through the Society Newsletter, which is presently published 3 times a year. Since 2005 the Chair of the Society for Low Temperature Biology has been Tiantian Zhang.


A list of additional scientific societies (mostly using "applied" cryobiology) can be found here.




CRYOBIOLOGY, (publisher: Elsevier) is the foremost scientific publication in this area, with approximately 60 refereed contributions published each year. Articles concern any aspect of low temperature biology and medicine (e.g. freezing, freeze-drying, hibernation, cold tolerance and adaptation, cryoprotective compounds, medical applications of reduced temperature, cryosurgery, hypothermia, and perfusion of organs).


CRYO LETTERS is an independent UK based rapid communication journal which publishes papers on the effects produced by low temperatures on a wide variety of biophysical and biological processes, or studies involving low temperature techniques in the investigation of biological and ecological topics.


CELL PRESERVATION TECHNOLOGY is a peer-reviewed quarterly scientific journal published by Mary Ann Liebert, Inc. dedicated to the diverse spectrum of preservation technologies including cryopreservation, dry-state (anhydrobiosis), glassy-state and hypothermic maintenance.


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