What is Cryopreservation? A Complete Guide
Cryopreservation is the process of cooling biological material cells, tissues, embryos, or organisms to sub-zero temperatures to halt all biological activity and preserve viability for future use. When stored correctly in liquid nitrogen at -196°C, biological samples can remain stable for decades without measurable degradation.
The process is used across reproductive medicine, fertility treatment, biobanking, scientific research, and veterinary science. It underpins IVF cycles worldwide, enables long-term sperm and embryo storage, and supports the preservation of cell lines in pharmaceutical and research laboratories.
Cryopreservation works by slowing or eliminating the formation of ice crystals, which would otherwise rupture cell membranes and destroy viability. Two primary methods are used: controlled-rate slow cooling and vitrification. Both require specialist cryogenic equipment designed to maintain precise temperatures during freezing, transport, and long-term storage.
How Does Cryopreservation Work?
At its core, cryopreservation relies on one principle: remove water activity from cells before ice crystals can form and cause damage. Two methods achieve this in different ways.
Controlled-Rate Slow Cooling
The sample is cooled gradually typically at 1°C to 2°C per minute using a programmable controlled-rate freezer. Cryoprotective agents (CPAs) such as DMSO or glycerol are introduced to the sample beforehand, drawing water out of the cells and replacing it with a solution that does not crystallise destructively at low temperatures. The sample is then transferred to liquid nitrogen storage at -196°C.
This method is widely used for sperm, red blood cells, and certain stem cell types.
Vitrification
Vitrification uses ultra-rapid cooling dropping the sample temperature so fast (up to -23,000°C per minute) that water molecules have no time to form crystals. Instead, the liquid transitions directly into a glass-like solid state. Higher concentrations of CPAs are used, and the sample is typically plunged directly into liquid nitrogen.
Vitrification has become the dominant method for oocyte and embryo cryopreservation in IVF, with significantly higher survival and pregnancy rates than slow cooling for these sample types.
Slow Cooling vs Vitrification at a Glance
| Factor | Controlled-Rate Slow Cooling | Vitrification |
|---|---|---|
| Cooling rate | 1 to 2°C per minute | Ultra-rapid, thousands of °C per minute |
| Ice crystal formation | Minimised through gradual dehydration | Eliminated; sample becomes glass-like solid |
| CPA concentration | Lower | Higher |
| Equipment | Programmable controlled-rate freezer | Vitrification device, direct liquid nitrogen plunge |
| Typical samples | Sperm, red blood cells, some stem cells | Oocytes, embryos, blastocysts |
| Long-term storage | Liquid nitrogen at -196°C | Liquid nitrogen at -196°C |
Cryopreservation Equipment for IVF and Research
Effective cryopreservation depends on the quality and reliability of the equipment used at every stage from initial freezing through to long-term storage and transport.
Liquid Nitrogen Storage Tanks
The primary storage vessel for cryopreserved samples. Cryolab supplies a full range of liquid nitrogen dewars and storage systems designed for IVF clinics, biobanks, sperm banks, and research laboratories built for long-term reliability at -196°C.
Dry Shippers
When samples need to be transported between sites, dry shippers maintain cryogenic temperatures without free liquid nitrogen, making them safe for air freight and courier use. Cryolab’s CryoStork dry shipper is purpose-built for the transport of reproductive samples including embryos, oocytes, and sperm straws. The CryoNest provides an additional option for secure sample transport.
Controlled-Rate Freezers
Programmable freezing units that automate the slow-cooling process, removing operator variability and ensuring reproducible results across every freeze cycle.
Cryogenic Accessories
Cryovials, straws, canes, goblets, and cryosleeves form the IVF consumables layer of any cryopreservation workflow. Cryolab supplies half canes, half cryosleeves, and a full range of storage accessories compatible with standard IVF and biobanking systems.
Applications of Cryopreservation
Reproductive Medicine and IVF
Embryo, oocyte, and sperm cryopreservation are central to modern IVF. Surplus embryos from stimulation cycles are frozen for future transfer, reducing the need for repeated stimulation and enabling cumulative pregnancy rates to build over time.
Biobanking
Biobanks store biological material blood, tissue, DNA, cell lines at cryogenic temperatures for research, diagnostics, and future therapeutic use. Cryopreservation infrastructure is the backbone of every biobank operation.
Sperm Banking
Men facing cancer treatment, surgery, or other fertility risks can have sperm cryopreserved prior to treatment. Donor sperm programmes also rely entirely on long-term cryogenic storage.
Stem Cell and Cell Line Preservation
Pharmaceutical and research laboratories preserve cell lines and stem cell populations in liquid nitrogen to maintain genetic stability and avoid culture drift over time.
Veterinary and Animal Breeding
Livestock genetics, endangered species conservation, and equine reproduction all use cryopreservation to store semen and embryos for artificial insemination programmes worldwide.
Expert Perspective: Paul Hague, Cryolab
Paul Hague has attended 42 consecutive ESHRE annual meetings more than any other delegate in the congress’s history. With over four decades in cryogenic science and reproductive medicine, he has witnessed the entire evolution of clinical cryopreservation, from early slow-cooling protocols to the widespread adoption of vitrification.
“The fundamentals have not changed protect the cell, control the temperature, trust the equipment. What has changed is our precision. The equipment available today gives clinics a level of control that simply was not possible 20 years ago.”
— Paul Hague, Director, Cryolab
Frequently Asked Questions About Cryopreservation
What is cryopreservation?
Cryopreservation is the storage of biological material including cells, tissues, sperm, eggs, and embryos at ultra-low temperatures, typically in liquid nitrogen at -196°C, to preserve viability indefinitely.
What temperature is used in cryopreservation?
Most cryopreservation protocols use liquid nitrogen at -196°C for long-term storage. During the cooling process, samples pass through intermediate temperatures depending on the method used slow cooling reduces temperature gradually, while vitrification drops it near-instantaneously.
What is the difference between vitrification and slow cooling?
Slow cooling reduces temperature at a controlled rate (typically 1-2°C per minute) to allow cellular dehydration before freezing. Vitrification uses ultra-rapid cooling to solidify the sample into a glass-like state with no ice crystal formation. For embryos and oocytes, vitrification now delivers superior outcomes.
How long can samples be cryopreserved?
In theory, samples stored correctly at -196°C can remain viable indefinitely. In clinical practice, human embryos have been stored for over 20 years and used successfully in IVF. Regulatory storage limits vary by country and sample type.
What cryoprotectants are used in cryopreservation?
Common cryoprotective agents (CPAs) include DMSO (dimethyl sulphoxide), glycerol, ethylene glycol, and propylene glycol. The choice of CPA depends on the sample type and the cooling method used.
Is cryopreservation used in IVF?
Yes. Cryopreservation is fundamental to modern IVF. Embryos, oocytes, and sperm are routinely cryopreserved to enable future frozen embryo transfer cycles, fertility preservation, and donor programmes.
What equipment is needed for cryopreservation?
A full cryopreservation workflow requires liquid nitrogen storage tanks, dry shippers for transport, controlled-rate freezers (for slow cooling protocols), and consumables including cryovials, straws, canes, and cryosleeves.
What is a dry shipper used for in cryopreservation?
A dry shipper is a specialised transport dewar that holds samples at cryogenic temperatures without free liquid nitrogen. This makes it safe for air transport and courier delivery between clinics, banks, and laboratories.
Can cryopreserved embryos be stored indefinitely?
Scientifically, yes at -196°C biological activity is effectively halted and degradation is negligible. Practically, storage duration is governed by clinic policies and national regulations. Many countries impose maximum storage periods, though extensions are often available in defined circumstances.
What is biobanking and how does cryopreservation support it?
Biobanking is the systematic collection and long-term storage of biological samples for research or clinical use. Cryopreservation is the primary method used to maintain sample integrity in biobanks, preserving DNA, tissue, blood, and cell lines for years or decades.
