Cryopreservation

Cryopreservation is a method to preserve cells and tissues for longer periods at the freezing points typically at -196 degrees Celsius. However, this cooling is carried out carefully and involves the use of cryoprotectants that prevent the crystallization of cellular materials. In this article, we will learn about Cryopreservation, its procedure, methods, and applications.

What is Cryopreservation?

Cryopreservation is a process of preservation of organelles, cells, tissues, and other biological materials in extremely freezing conditions. The word “cryo” is derived from the Greek word kayos which means ‘frost.’ The science that deals with Cryopreservation is termed “Cryobiology“. Cryopreservation is carried out when we need to preserve any biological specimen like eggs, sperm, stem cells, or other biological components for an extended period without killing them. Cryopreservation involves the preservation of specimens at the freezing points typically at -196 degrees Celsius. However, this cooling is carried out carefully and involves the use of cryoprotectants that prevent the crystallization of cellular materials.

Cryopreservation Procedure

Cryopreservation is done carefully and involves steps that prevent cell death while arresting its metabolic activities. This is typically done by the use of dry ice or liquid nitrogen. The steps involved in Cryopreservation are discussed below.

• Selection of material: Select the material based on factors such as size, shape, mass, and acidity to ensure that the biological materials are not damaged.
• Addition of cryoprotectants: Glycerol, FBS, salts, sugars, and glycols are added to cryo-protected substances. This helps to lower the freezing point and slow down the cooling process and prevents the risk of formation of ice crystals.
• Freezing: Various freezing methods protect cells from damage and prevent death by exposing them to warm solutions of Cryogenic Protective Agents.
• Storage in Cold Conditions: Keep the cryopreserved samples in extremely cold places, like -80°C in a freezer, for at least 5 to 24 hours before moving them to storage containers.
• Thawing: Warm the biological samples when necessary to control the cooling rate and avoid cell damage from crystal formation.

Diagram of Cryopreservation

The proper labelled diagram with the potocol of cryopreservation is given below:

Different Methods of Cryopreservation

Different specimen need different methods of preservation. Hence the Cryopreservation method has been modified in different ways to suit the needs of different biological material. Some of these methods are:

• Slow freezing: The temperature is reduced gradually. It is done gradually to minimize the ice crystal formation.
• Vitrification: Rapid cooling is applied to prevent ice crystal formation. It is used for embryos, oocytes and small tissues.
• Direct plunging into liquid nitrogen: The samples are directly immersed in the liquid nitrogen. There is no controlled cooling.
• Freeze drying: Biological materials are preserved in a dry state. Water is removed from the sample with the help of sublimation.
• Dry ice storage: It is done for short-term storage with the help of solid carbon dioxide. It is a cost-effective and simple method.

Usage of Cryopreservation

The usage of Cryopreservation are as follows:

Egg Freezing

Eggs are frozen very rapidly in the vitrification procedure, which makes it difficult for ice crystals to form. A large amount of new cryoprotectants with antifreeze are used. In this process, the egg is placed in a liquid that contains some antifreeze. To remove water from the egg, a small amount of sugar is mixed in. For a few seconds, it was moved up in the high antifreeze liquids and placed immediately into liquid nitrogen. The egg is going to be placed in a woman when it thaws.

Embryo Freezing

Hormones help the eggs to grow in treatment for infertility. These eggs are fertilized in the laboratory. Embryos can be made and implanted into the female’s uterus. It is possible to freeze these embryos and use them in the future. So, without going through a whole new treatment, the woman could have another embryo implanted.

Sperm Freezing

When frozen and thawed, the sperm mixture is added to a solution for cryo-protection. Then it’s inserted into plastic containers, which freeze in liquid nitrogen. This increases the chances of having a baby in the future. Sperm may be stored in a cryobank for less than one year and used to treat it.

Benefits of Cryopreservation

Cryopreservation has several benefits as follows:

• Fertility Treatments
• Keeps Genetics Safe
• Protects Important Genes
• Saves Endangered Species
• Requires Little Space and Work
• Keeps Samples Healthy
• Preserves Samples Longer
• Stops Genetic Changes

Applications of Cryopreservation

Cryopreservation is the process of storing things for long periods at low temperatures. This method is used in a lot of different areas such as medicine, biology, genetics, etc.

• Gene Bank
• Blood Transfusion
• Artificial Insemination
• In Vitro Fertilization
• Freezing Cell Cultures
• Organ Transplantation
• Storage of Rare Germplasm
• Seed Bank
• Conservation of Endangered Plants
• Biodiversity Conservation

Advantages and Disadvantages of Cryopreservation

The advantages and disadvantages of Cryopreservation are as follows:

Cryopreservation has following advantages:

• It can be used for fertility treatments
• It saves endangered species
• Cryopreservation keeps the samples healthy
• It helps to preserve the samples for a longer duration
• It also helps to stop genetic changes
• It helps in organ transplantation by preserving donor organs

The disadvantages of Cryopreservation are as follows:

• Ice crystals can form inside the cell which can cause damage to the cells
• The cell viability can be affected if unsuitable cryoprotectants are used
• Fluctuations in temperature during storage can impact the preserved samples
• It is expensive

Conclusion – Cryopreservation

• Cryopreservation is a method to preserve cells and tissues for longer periods.
• The cells, tissues, and organs are preserved at extremely low temperatures up to −196°C.
• This method is used in the treatment of infertility, gene banks, seed banks etc.
• The method has several advantages such as the preservation of endangered species, freezing eggs and sperm etc.

FAQs- Cryopreservation

What is Cryopreservation?

Cryopreservation is the freezing and storage of biological samples at extremely low temperatures. Usually, the samples are stored at −196°C, which is the liquid nitrogen temperature.

What temperature is used in Cryopreservation?

Cryopreservation is done below 0°C up to −196°C. The usual range of temperature is -80 to −140°C. Cryogenic temperatures fall within a well-defined range of -238 °F to -460 °F.

Which chemical is used in Cryopreservation?

Cryoprotective substances that are frequently employed in cryopreservation include propylene glycol, glycerol, and dimethyl sulfoxide (DMSO). Additional substances employed in cryopreservation encompass liquid nitrogen and dry ice.

What are the steps of Cryopreservation?

Cryopreservation involves 5 steps: Selection of material, Addition of cryo-protectants, Freezing, Storage in Cold Conditions, and Thawing.

What are the methods of Cryopreservation?

The two basic methods of Cryopreservation include the conventional slow freezing (SF) and the rapid freezing method or vitrification (Vit).

What is Plant Cryopreservation?

Plant Cryopreservation is the process of storing plant material in liquid nitrogen indefinitely. The substance may consist of pollen, seeds, latent buds, or shoot tips.

What is Cryopreservation and its Advantages?

The process of quickly freezing biological materials in order to prolong their shelf life is known as Cryopreservation. Benefits of Cryopreservation are Fertility treatment and Genetic contamination protection.

Why nitrogen is used in Cryopreservation?

Liquid nitrogen is utilised in Cryopreservation due to its low temperature (-196°C).

Why is DMSO used in Cryopreservation?

Cryoprotectant dimethyl sulfoxide (DMSO) is frequently employed in Cryopreservation in order to stop ice from forming and cells from dying during the freezing process.

What is the principle of Cryoprotectant?

Cryoprotectants are substances that are used in order to avoid the production of ice when freezing organs.