Stem cells are remarkable biological assets defined by two fundamental properties: the capacity to self-renew and the potential to differentiate into many specialized cell types, such as blood, bone, or nerve cells. They function as a natural repair system for the body, making them highly valuable for medical treatments and research. Cryopreservation, the process of freezing biological material at extremely low temperatures, allows these living cells to be stored long-term while maintaining viability. This process suspends all metabolic activity, creating readily available cell banks for future therapeutic use.
Sources of Stem Cells for Preservation
The cells used for cryopreservation come from several primary sources, each offering different types of stem cells. Hematopoietic Stem Cells (HSCs) are the most clinically established type, responsible for generating all blood and immune system cells. HSCs are harvested primarily from bone marrow, collected surgically, or from peripheral blood after mobilization into the bloodstream via apheresis.
A non-invasive source is umbilical cord blood and tissue, collected after childbirth. Cord blood is rich in HSCs that are more primitive and adaptable than adult HSCs, often requiring less strict matching for transplantation. Cord tissue also contains Mesenchymal Stem Cells (MSCs), found in Wharton’s jelly, which are multipotent cells capable of differentiating into tissues like bone, cartilage, and fat.
A third category includes Induced Pluripotent Stem Cells (iPSCs), which are adult cells genetically reprogrammed to exhibit pluripotent qualities. These iPSCs, along with tissue-specific MSCs from sources like adipose tissue, are banked for future use in personalized medicine and drug development.
Therapeutic Applications of Cryopreservation
The ability to freeze and store stem cells provides a time-independent supply for established and emerging therapies. The most common and long-standing use is in hematopoietic stem cell transplantation, often referred to as a bone marrow transplant, for treating blood cancers like leukemia and lymphoma, and certain immune and metabolic disorders.
Cryopreservation allows a patient’s own cells (autologous) to be banked before high-dose chemotherapy or radiation, then reinfused afterward to restore the immune system. Cryopreserved cells from a donor (allogeneic) can also be stored and used when an autologous transplant is unsuitable, offering a potential cure for diseases like sickle cell anemia. Storing these cells provides time for extensive quality testing and ensures the cells are immediately available when needed.
Banked cord blood units have been particularly helpful for patients who require a transplant quickly or cannot wait for a fully matched adult donor. Beyond these established uses, cryopreservation is foundational for regenerative medicine.
Mesenchymal Stem Cells are being explored for their ability to modulate the immune system and promote tissue repair in conditions such as multiple sclerosis and heart failure. Induced pluripotent stem cells are banked to create patient-specific models for drug testing and to study how diseases like Parkinson’s or Alzheimer’s develop at the cellular level.
The Science of Freezing Stem Cells
Freezing living cells poses a substantial challenge because ice crystal formation causes mechanical damage to delicate cell membranes and organelles. To prevent this cellular destruction, a precisely controlled process begins with adding a Cryoprotective Agent (CPA).
The most widely used CPA is Dimethyl Sulfoxide (DMSO), which penetrates the cell membrane due to its low molecular weight. DMSO lowers the freezing point of the solution and reduces the amount of water that turns into ice inside the cells, minimizing damaging intracellular ice crystals. Because DMSO can be toxic at high concentrations, a carefully regulated protocol is necessary.
Following CPA addition, the cell product undergoes Controlled Rate Freezing (CRF) using specialized equipment. This process ensures the temperature decreases slowly, at a rate of 1 to 2 degrees Celsius per minute, down to about -40 degrees Celsius.
This slow cooling allows water to move out of the cell, dehydrating it and concentrating the protective CPA inside before the cooling rate is increased to rapidly bring the cells to the final storage temperature.
Storage and Revival of Cryopreserved Cells
Once the controlled freezing process is complete, the vials or cryo-bags of stem cells are transferred to a long-term storage facility. For indefinite preservation, the cells must be maintained at temperatures below -130 degrees Celsius, the point where all biological processes cease. This temperature is achieved using specialized liquid nitrogen tanks.
The most common method for clinical storage is in the vapor phase of liquid nitrogen, where temperatures are kept at or below -140 degrees Celsius. Storing cells in the vapor phase reduces the risk of cross-contamination between samples and prevents cryotube explosion upon retrieval, unlike direct immersion. Long-term stability depends on the constant monitoring of the liquid nitrogen level and temperature.
When the cells are needed for therapeutic use, the revival process must be executed with precision. The frozen sample is thawed rapidly, typically by immersion in a 37-degree Celsius water bath.
This rapid warming prevents the formation of ice crystals, a phenomenon known as recrystallization, and limits the time the cells are exposed to the potentially toxic CPA. Following thawing, the CPA is removed by washing the cells, and a viability assessment is performed to ensure a sufficient number of functional cells are available for treatment.