Cryostorage involves preserving biological materials at extremely low temperatures to maintain their integrity over long periods. This process suspends biological activity, allowing cells, tissues, and other biological constructs to remain viable for future use. The ability to halt degradation through deep freezing has broad implications across medicine, research, and conservation efforts.
What is Cryostorage?
Cryostorage is the practice of preserving biological samples by cooling them to temperatures around -196°C (-320°F) using liquid nitrogen. This extremely cold environment halts all metabolic processes within the cells or tissues. The aim is to store these materials without degradation, maintaining their structural and functional integrity for extended durations. The scientific term for this preservation method is cryopreservation.
The Science Behind Cryopreservation
The challenge in cryopreservation is preventing the formation of ice crystals within cells, which can cause severe damage and lead to cell death. To mitigate this, cryoprotective agents (CPAs) are introduced to the biological material. These “antifreeze” compounds, such as dimethyl sulfoxide (DMSO) and glycerol, work by increasing the solute concentration within and around the cells, reducing ice formation.
Two main cooling methods are employed in cryopreservation. Slow freezing involves a gradual reduction in temperature. As the temperature slowly decreases, water is drawn out of the cells by osmosis, and extracellular ice forms, concentrating the solutes inside the cells. This controlled dehydration helps prevent the formation of intracellular ice crystals.
Alternatively, vitrification involves rapid cooling, transforming the cell’s internal solution into a glassy, non-crystalline solid. This flash-freezing technique, often achieved by plunging samples directly into liquid nitrogen, prevents ice crystal formation entirely. Vitrification requires higher concentrations of CPAs compared to slow freezing. However, some CPAs, like DMSO, can be toxic at high concentrations, requiring careful management.
Applications in Medicine and Research
Cryostorage is a routine practice with widespread applications across various fields, enabling long-term preservation of biological materials. In medicine, it is used for human reproductive cells, including sperm, eggs, and embryos, supporting fertility treatments like in vitro fertilization (IVF). Stem cells, particularly those for regenerative medicine and bone marrow transplants, blood components, and certain organ tissues intended for transplantation are also cryopreserved. This allows for the banking of these materials until needed.
Research laboratories rely on cryostorage for the long-term preservation of cell lines, tissue samples, and microbial strains. This ensures biological models are available for future studies, allowing researchers to revisit samples from decades ago with new questions or technologies. By halting metabolic activity, cryostorage prevents genetic drift and changes that would occur if cells were continuously cultured. In conservation efforts, cryostorage is used to create “biobanks” that store genetic material from endangered species, providing a safeguard against extinction and a resource for future breeding or reintroduction programs.
Ensuring Viability Post-Thaw
The success of cryostorage depends on the viability of the biological material after it is thawed. The thawing process is a carefully controlled step, often involving rapid warming in a 37°C water bath. This swift rewarming is important to minimize the formation of large ice crystals, which can damage the cells.
Following thawing, cryoprotective agents must be carefully removed from the cells to prevent their toxicity, typically through a series of washes with solutions of decreasing CPA concentration to prevent osmotic shock. Assessing viability post-thaw is an important step to confirm success. Methods for assessment include cell culture to observe growth and function, and various functional assays to measure metabolic activity or cell membrane integrity. Immediate post-thaw viability measurements may not always predict long-term functionality, so some protocols include culturing cells for a period to observe sustained viability and functionality.