Frozen Dreams: How Cryopreservation Technologies Work

Cryonauts who sleep for decades on their journey to distant star systems are a recurring theme in science fiction. Today, cryopreservation technologies are advancing rapidly—could humanity be on its way to the stars? Let’s explore cryonics, cryobiology, and other “cryo” sciences in this article.

The concept of freezing people has long fascinated science fiction writers. Special chambers for long-term storage of human bodies frequently appear in literature and film. Sometimes, freezing occurs accidentally as a result of extreme frostbite. This trope can be found in serious works like Escape of Mr. McKinley, James Cameron’s Avatar, and the newer films in Ridley Scott’s Alien franchise and satirical productions such as Idiocracy, South Park, and Futurama.  

Neil R. Jones’s short story The Jameson Satellite is a particularly influential work about cryopreservation. Its protagonist, Professor Jameson, sends his body into Earth’s orbit to preserve it in the vacuum of space and survive into the distant future. He succeeds, and millions of years later, a race of mechanized beings discovers Jameson and brings him back to life.  

At the age of 12, this story captured the imagination of Robert Ettinger, who would later become the leading advocate for cryonics.

Cryonics: Hope or Hoax?

Cryonics is the belief that the bodies of deceased individuals can be frozen and revived in the distant future when technology has advanced enough to repair all physical defects. However, the scientific community broadly considers cryonics a pseudoscience, and providing such services is often deemed fraudulent—organizations offering cryonics charge exorbitant fees for their services.  

Robert Ettinger, a key figure in the cryonics movement, was also a science fiction writer. In 1964, he published The Prospect of Immortality, a book outlining his vision of how cryopreservation technology, combined with other future inventions, could fundamentally transform human society. While the book has some artistic merit, its scientific basis is tenuous at best. The horizons Ettinger described remain far beyond the current reach of cryobiology.

Is It Possible?

Cryopreservation is a well-established method in cryobiology for storing biological materials at liquid nitrogen’s temperature of -196°C. It is considered successful if the specimen retains its full viability after thawing.  

Experiments in lowering the storage temperatures of biological samples began in the 20th century but never aimed to freeze humans or other animals in their entirety. One of the pioneers in this field was the Russian scientist Ilya Ivanov, a specialist in artificial insemination. In the 1910s and 1920s, he frequently used cooling techniques to preserve horse semen at around -15°C.

The development of effective cryopreservation became possible with the discovery of cryoprotectants—substances that shield cells from the damaging effects of freezing. These compounds lower the crystallization temperature and protect cell membranes. The first known cryoprotectant, glycerol, was identified by British scientists Christopher Polge, Audrey Smith, and Alan Parks in 1949. They successfully froze sperm from various animal species at liquid nitrogen temperatures and later thawed it for use in artificial insemination.  

This groundbreaking discovery led to the widespread adoption of freezing agricultural animal semen in many countries beginning in the mid-20th century.

Scientific Foundations

Cryopreservation involves two primary approaches for preserving biological specimens: controlled-rate freezing and vitrification. Controlled-rate freezing is based on the work of Peter Mazur and his team, who discovered that by carefully regulating the cooling rate of a specimen saturated with cryoprotectants, intracellular ice crystals can be minimized in size, preventing them from rupturing cell membranes. Conversely, Vitrification was first explored in the 1930s by a group of scientists led by Basil Luyet. Their research showed that at very high concentrations of cryoprotectants and rapid cooling rates, biological specimens can bypass the crystallization phase altogether, transitioning directly into a glass-like state.

Before freezing, the biological specimen must be saturated with cryoprotectants. In vitrification, this saturation requires a higher concentration of these chemicals. The specimen is then placed in a container and gradually infused with cryoprotectants over several stages. In controlled-rate freezing, the specimen is cooled using a programmable freezer that follows a precise schedule. The process begins with cooling at a rate of 1–2 degrees Celsius per minute, slowing to fractions of a degree per minute after seeding—a process that initiates controlled crystallization. Once the specimen reaches a target temperature, typically between -30 and -45 degrees Celsius, it is submerged in liquid nitrogen for long-term storage. In vitrification, the specimen, already saturated with cryoprotectants, is directly plunged into liquid nitrogen. This immediate cooling prevents ice formation, preserving the specimen in a glass-like state. Despite its apparent simplicity, vitrification requires considerable expertise, as the specimen must be immersed in liquid nitrogen quickly and carefully to avoid damage.

Thawing a cryopreserved specimen is similar for both methods, though the speed of warming differs. Vitrified specimens must be thawed rapidly to prevent ice crystal formation, which can destroy cells. Specimens preserved through controlled-rate freezing are gradually thawed, allowing for better control of temperature changes. Thawing methods include immersing the specimen in warm water, exposing it to room-temperature air, or using warm airflow, depending on the freezing and storage conditions. Research by Peter Mazur highlighted that the rate of warming is as critical to successful cryopreservation as the cooling rate. Rapid warming is essential for vitrified specimens to prevent ice crystals’ formation during the transition.

In 2017, researchers from John Bishof’s lab achieved a breakthrough in cryopreservation by successfully preserving zebrafish embryos. Previously, this was impossible due to the unique structure of their membranes. The embryos were cooled to liquid nitrogen temperatures at an extraordinary rate of 90,000 degrees Celsius per minute. Thawing was achieved with a laser pulse immediately upon removal from liquid nitrogen, aided by gold nanoparticles in the cryoprotectant. This method allowed for volumetric heating at an unprecedented rate of 14 million degrees Celsius per minute, ensuring the survival of the embryos.

After thawing, cryoprotectants must be carefully removed, as they are often toxic at room temperature and can harm the biological specimen. For example, while glycerol is relatively safe in low concentrations used in cosmetics and food, the high concentrations required for cryopreservation can cause significant damage. Other cryoprotectants, such as ethylene glycol, propylene glycol, and dimethyl sulfoxide, pose similar risks. Additionally, cryopreserved specimens are often severely dehydrated due to the cryoprotectants, necessitating careful washing to remove the chemicals while rehydrating the cells.

Experiments

The primary focus of modern cryobiology is freezing reproductive cells and embryos of living organisms. These materials are relatively simple structures capable of withstanding sudden and extreme temperature changes without significant damage—however, even these present challenges. For example, oocytes (female reproductive cells) are far more difficult to preserve than sperm cells. Embryos, being multicellular organisms, pose additional complications, as selecting suitable preservation methods for certain vertebrates can be particularly challenging.

Cryopreservation technology is in high demand among professionals working with animals, as it is critical in preserving biodiversity. Many wild mammals are on the brink of extinction, and their survival in natural habitats is no longer viable in some cases. Cryobanks have been established worldwide to safeguard these species’ genetic resources. These facilities, resembling large storage tanks filled with liquid nitrogen, contain racks holding containers of biological material. Cryobanks store reproductive cells, embryos, and certain other cell types, ensuring their preservation for future use.

Cryopreservation is also instrumental in optimizing vivariums that house laboratory animals. Thousands of mouse and rat strains exist, with more expected in the future, making cryo-archiving indispensable. Large genetic research centers preserve many mouse lines in the form of sperm or embryos, maintaining only the most commonly used strains as live colonies.

Cryopreservation has also proven helpful in animal breeding and selection. Freezing genetic material enables the storage and use of reproductive cells from individuals with desirable traits, such as higher fertility, greater body weight, faster fur growth, and other advantageous characteristics. Additionally, cryobanks providing services for the storage of human reproductive cells have gained popularity, though their purpose is strictly reproductive.

Despite its achievements, cryobiology still faces numerous unresolved challenges. For instance, effective methods for freezing amphibian and fish embryos and larvae remain elusive. This is why the 2017 breakthrough by American scientists demonstrating the vitrification of zebrafish (Danio rerio) embryos was so significant. Challenges also arise when working with mammals, even though reliable techniques for preserving gametes and embryos exist for many species. For example, the embryos and oocytes of wild cats and domestic pigs are particularly problematic due to their high lipid content. Lipid granules play a key role in cellular nutrition, and low temperatures can alter their physical state, disrupting metabolism and leading to cell death.

A considerable amount of research is focused on cooling and, ultimately, freezing larger biological structures such as organs and tissues. One milestone involved cooling a rabbit kidney to -45°C, after which it retained functionality and was successfully transplanted.

Another notable experiment was published in 2006, demonstrating that rat hippocampal brain cells could survive vitrification. However, the results were inconclusive. First, the brain tissue was reduced to thin slices, only 500 microns thick. Second, the criterion for cell viability was maintaining high intracellular potassium levels, which does not necessarily indicate normal neuronal function or the maintenance of synaptic connections.

These achievements are remarkable but raise whether Robert Ettinger’s dream of human cryopreservation—or even the preservation of the human brain—could become a reality in the foreseeable future.

What Discoveries Are Needed to Make This a Reality?

Freezing a human is currently impossible, and cryopreservation is not yet feasible for most multicellular organisms. Successful cryopreservation has been limited to the roundworm Caenorhabditis elegans (along with a few other small parasitic worms) and free-floating larvae of certain small coral species, which are small and have relatively simple body structures.

The main obstacle to freezing large and complex organisms lies in the diversity of cell and tissue types within their bodies. Intercellular connections are susceptible to external disturbances, making preserving the structural integrity of cells, tissues, and organs during cryopreservation is an immense challenge. Each cell type requires its specific protocol, and sometimes, the optimal conditions for cryopreservation vary significantly across different cell types. Without addressing these complexities, successfully freezing whole organs or organisms will remain unattainable.

It is difficult to predict when this challenge might be overcome, but unlocking the secrets of anabiosis could hold the key. Many organisms—including insect larvae, molluscs, amphibians, and reptiles—can survive cold periods in a frozen state, reviving when conditions improve. While none of these organisms naturally reach the extreme temperatures of -196°C, their abilities are remarkable. For example, the wood frog can survive winter hibernation by converting 35% of the water in its body into ice, halting breathing, feeding, and even its heartbeat.

Similarly, it is known that lowering the human body’s temperature can significantly slow metabolism, increasing the chances of survival in extreme conditions. One of the most striking examples occurred in 2006 with Mitsutaka Uchikoshi, a 35-year-old Japanese man. During a hiking trip with friends, he got lost, suffered a fractured pelvis, and lost consciousness. He remained in the mountains for 24 days. The low ambient temperature caused Uchikoshi’s body temperature to drop to 22°C. Remarkably, after receiving medical treatment, he made a full recovery. Doctors believe he entered a state resembling hibernation, which allowed him to survive so long without food or water.

These cases provide hope that understanding the mechanisms of anabiosis could eventually bring us closer—though still far from imminent—to the human cryopreservation described in science fiction.

Something to Consider

Even if we fully restore an organism’s biological functions after thawing, this does not guarantee the revival of a person as they were before. Currently, we lack sufficient understanding of how brain activity shapes human personality. Cryonics advocates argue that the cessation of the brain’s electrical activity does not equate to its formal death. They claim that if brain activity can be restored after cryopreservation, the individual would return to life as though nothing had happened.

However, no conclusive evidence supports this claim, and conducting experiments to confirm or refute it is currently impossible. Beyond the technological breakthroughs required for cryonics, progress in this field will also depend heavily on the work of neuroscientists and philosophers of consciousness. These experts will need to develop criteria for personal identity to ensure that an individual’s personality and sense of self can be fully restored. 

Without such criteria, we risk two undesirable outcomes: either reviving a living body devoid of key cognitive functions or bringing forth an entirely new personality, potentially retaining only fragments of the original individual’s memories.