Cryopreservation: Meaning, Process, Steps, Benefits & Application

The word cryo comes from the Greek word "kayos" meaning "snow". It means being saved in "frozen state". It is a process of cooling and keeping cells, tissues, or organs at very low temperatures to maintain their function. Cryopreservation is a method by which low temperatures are used to preserve living cells and tissues. In this process, the tissues can be stored for a very long time. The science of cryopreservation is known as "cryobiology". 

Key takeaways: Cryopreservation, oocytes, spermatozoa, tissues, ovarian tissue, pre-transplant embryos, organs.

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What is Cryopreservation?

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• Cryopreservation is a method of storing live cells, tissues and other biological samples in deep freezing temperatures for storage or storage. The sample is usually stored at − 196 ° C.
• It can be done over the temperature: Solid carbon dioxide (at -79^oC), Deep refrigerators with low temperatures (-80^oC), In the vapor phase nitrogen (at 150^oC), In liquid nitrogen (196^oC).
• At such low temperatures, all cellular biological functions cease and the cell dies. Cryopreservation helps cells survive cold and thaw.
• The formation of ice inside the cells can break down cell membranes. This can be prevented by controlling the freezing point and carefully selecting the freezing point.

Read Also: Extinction

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Cryopreservation Process

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In this process, living organisms including cells, oocytes, spermatozoa, tissues, ovarian tissue, pre-transplant embryos, organs, etc. they are kept at extremely cold temperatures without interfering with cell function. Dry ice and liquid nitrogen are commonly used in this method.

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Cryopreservation Steps

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The process followed by plant regeneration involves the following steps.

Priority Selection: For cryopreservation, selection of appropriate plants is essential. Two key factors depend on it as nature and density. Any tissue can be selected for this purpose, for example embryo, meristem, ovules sperm, etc. Congestion should be high.

Addition of Cryoprotectant: Chemical properties are important as they prevent the destruction of cryo. Some examples of cryoprotectants are alcohol, other amino acids such as proline, and dimethyl sulfoxide. In particular two cryoprotectants should be used together instead of one as they are considered to be more effective.

Cold: Different types of plants show different types of low temperature sensitivity. There are several types of methods:

• Slow Freezing - In this process, the tissue or plant material freezes slightly with a slow cooling rate. The great advantage is that the plant cells are slightly hydrated and function better.
• How to Freeze Fast - Containers are immersed in liquid nitrogen. In this process, the temperature drops from 300 to 1000 degrees immediately.
• Drying Method - This way hydrated cells and seeds are stored.

Storage for Liquid Nitrogen: It is also important to keep the sale or goods at a certain temperature. Generally, temperatures are maintained - 70 to - 196 ° C. Long-term storage is carried out at a temperature of 196 ° C in liquid nitrogen. Continuous nitrogen supply is required to prevent damage.

Melting: The melting process is usually done by throwing the dishes in a hot water bath with a strong rotation. It also causes the dishes to be moved or moved to another bath at 0 ° C.

Washing and refining: Stored items are washed to remove cryoprotectant. In addition, the material is recycled into a new environment.

Performance Rate: Due to the final pressure, the cells may die. The presence of functionality can be seen in most cases.

Calculated by formula: (No. of growing cells / No. of melted cells) × 100

Crop Renewal: Thereafter, live seeds are grown in indirect growing areas. Appropriate environmental conditions are maintained.

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Cryopreservation of Embryos

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During fertility treatment, hormones are used to promote egg development. The eggs are then removed and fertilized in a laboratory. Many embryos can be created and implanted in a woman's uterus. These embryos can be kept cryopreserved and can be used over time. With this, a woman can get more embryos in the future, without having to spend another IVF cycle.

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Oocyte Cryopreservation

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By means of vitrification, the eggs freeze quickly so that there is little time available for the formation of ice crystals. New cryoprotectants are used with many anti-freeze products.

The oocyte is first placed in a bathtub containing a low anti-freeze component like a cryoprotectant. Some sucrose is added to help drain the egg. The egg is then transferred to a cryoprotectant anti-freeze at high anti-frotectant concentration for a few seconds and then immediately transferred to liquid nitrogen. When the egg melts and is used for female implantation.

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Cryopreservation of Sperm

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The sperm sample is mixed with the solution, which provides protection during freezing and thawing. This is followed by transfer to plastic containers, which are then stored in liquid nitrogen to be frozen.

This procedure ensures the chances of future pregnancy. Sperm can be re-inserted, frozen and stored in cryonks for less than a year. These sperm may be used for specific infertility treatments.

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Applications of Cyropreservation

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In Medical Science

1. Cryopreservation gained prominence in traditional medicine after its use in the treatment of infertility. Since then, gamete cryopreservation has been developed to combat childlessness.
2. Sperm was the first cell that was frozen successfully and remained frozen easily due to its small cytoplasm as well as its low water content. Also, the nuclear material of the sperm is compressed and protected from damage. For these reasons, cryopreservation of sperm cells is often used in human medicine today.
3. Live births resulting from assisted reproductive cycles using frozen sperm or embryos have been observed in recent years. Human oocytes and ovarian tissue have also been preserved. Studies and research of lymphoid cells in the memory of immunology, allograft of the aortic roots, and osteoblasts for bone preservation are still ongoing.
4. Folk medicine also now usually performs cryopreservation of the cornea, umbilical cord, and hematopoietic cells, as well as sperm retention.
5. Cryopreservation of bull sperm has been used to produce rare and endangered species. Each year, more than 25 million calves are fertilized by frozen bull semen. Tissues, cell lines, DNA, and serum samples can also be stored in cryogenic banks.

In Biological Science

1. Cryopreservation is one of the most reliable ways to preserve plant genetics for a long time.
2. In agriculture, germplasm cryopreservation is used to improve the genetic makeup of domestic species and to adapt to natural conditions. Although the practice of preserving plant germplasm at cryogenic temperatures is relatively new, scientists have been doing processes to preserve plant cells and plant tissue for over 40 years now.
3. These techniques can now be used for plant genotype as well. New cryogenic methods using cryoplates (V and D) have recently been developed. This technology has advantages such as ease of use and excellent recovery rates after cryopreservation.
4. Aquatic biotechnologies rely on cryopreservation storage for gametes, embryos, and embryonic cells to spread important species, protect endangered species, and preserve genetic diversity.
5. Research shows that marine fish sperm cryopreservation is more effective than freshwater fish cryopreservation and that pregnancy rates are similar to those of mammals.

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Benefits of Cryopreservation

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• Once the material has been successfully stored at a certain temperature, it can be stored in an efficient way.
• No mutation or contamination of mold or bacteria occurs after the completion of the storage process and the stored materials.
• Limited space is required for cryopreservation purposes.
• A little work is needed for cryopreservation purposes.
• Cryopreservation enhances the effectiveness of assisted reproductive therapies by allowing all extracted and / or fertilized cells to be stored for future use.
• Cryopreservation allows people who are losing their fertility to maintain their reproductive cells and possibly become pregnant in assisted ways in the future. Women who want to delay childbirth or who have a family history of premature menopause can use it.
• Cryopreservation is a powerful tool for storing viruses that are endangered. It can also help maintain plant fertility.
• By freezing the embryos between cycles, ovarian stimulation is not necessary each time, and if the female ovaries are overly stimulated, implantation may be delayed without wasting regenerated oocytes.
• Cryopreservation allows pregnant women in their first course of treatment to donate their unused frozen embryos for research.
• Currently only one or two embryos are implanted, and any embryos that remain in the cryopreserve for future medical cycles.

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Cryopreservation of Animal cells

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The formation of animal cell lines is expensive, time-consuming, and labour-intensive. Continuous cell line has several advantages over cross-fertilizing cell lines such as:

• They live forever.
• They grow very fast.
• They can easily combine.

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Cryopreservation Plant Cell

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Due to the gradual extinction of species and rare species the need for genetic conservation is growing. The castle journal's method of storage fails to prevent the losses caused by:

• Attack of pathogen and insects
• Climate change
• Environmental disturbances
• Political and economic reasons

Items to be stored are kept at low temperatures because the cell growth rate slows down. As a result, biological activities are long overdue

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Things to Remember

• Cryopreservation is a method of storing live cells, tissues and other biological samples in deep freezing temperatures for storage or storage. The sample is usually stored at − 196 ° C.
• In this process, living organisms including cells, oocytes, spermatozoa, tissues, ovarian tissue, pre-transplant embryos, organs, etc. they are kept at extremely cold temperatures without interfering with cell function. 
• Cryopreservation gained prominence in traditional medicine after its use in the treatment of infertility. Since then, gamete cryopreservation has been developed to combat childlessness.
• Cryopreservation of bull sperm has been used to produce rare and endangered species. Each year, more than 25 million calves are fertilized by frozen bull semen. 
• Cryopreservation is a powerful tool for storing viruses that are endangered. It can also help maintain plant fertility.