Auto-nucleating Device for Controlled Cryopreservation
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Solution Overview
Problem
Current cryopreservation methods face challenges with rapid cooling rates causing intracellular ice formation and variable sample survival rates due to thermal disequilibrium and random ice crystallization, necessitating cumbersome seeding procedures that can damage cells and tissues.
Innovation Solution
An auto-nucleating device with an elongated hollow tube coated with an ice-nucleating composition like cholesterol is introduced into the cryopreservation vessel, allowing spontaneous ice nucleation within the device and controlled extracellular ice crystal induction, reducing supercooling and minimizing cell damage.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Speed
If rapid cooling is used to achieve efficient cryopreservation, then freezing speed increases, but intracellular ice formation occurs causing cell death
Solution Approach 1:
The hollow fiber is pre-cooled to a temperature below the freezing point of the aqueous solution before the sample is introduced. This preliminary cooling action ensures that when the sample is added, ice nucleation occurs immediately at the fiber surface, preventing supercooling and controlling the freezing process to avoid intracellular ice formation while maintaining high freezing speed
Solution Approach 2:
The hollow fiber acts as an intermediary ice-nucleating structure between the cooling system and the sample. It provides a controlled interface for heat transfer and ice crystal formation, mediating the freezing process to achieve both rapid cooling and controlled extracellular ice formation that prevents cell damage
2Reliability
If manual seeding procedures are used to induce controlled ice formation, then ice crystal formation becomes controlled, but the procedure becomes cumbersome and can damage cells
Solution Approach 1:
The hollow fiber automatically performs the ice-nucleating function when pre-cooled and placed in the sample. The system self-regulates ice crystal formation through the pre-cooled fiber surface without requiring external manual seeding operations, eliminating the cumbersome steps and potential cell damage associated with manual procedures
Solution Approach 2:
The hollow fiber is pre-cooled before sample introduction, performing the ice-nucleation preparation in advance. This eliminates the need for manual seeding during the freezing process, simplifying the procedure while maintaining controlled ice formation
3Reliability
If pre-cooling of the hollow fiber is performed, then supercooling is reduced and controlled ice formation occurs, but additional cooling steps are required
Solution Approach 1:
The hollow fiber combines multiple functions: it serves as the cooling element, the ice-nucleating surface, and the structural support. By integrating these functions into a single pre-cooled component, the system achieves controlled ice formation without requiring separate complex cooling procedures or multiple devices
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
The auto-nucleating device enhances cryopreservation by reducing cell damage, improving sample survival rates, and enabling the preservation of low-motility samples, such as sperm, with higher post-thaw viability and reproducibility compared to traditional seeding methods.
Implementation Method 1
When the aqueous solutions are cooled further below their freezing point, the extent of supercooling increases. In the absence of intervention, the water molecules in the solution will, at a point usually no more than 15°C below the freezing point, spontaneously crystallize, and pure water will precipitate as ice.
Implementation Method 2
As these solutions are cooled to temperatures slightly below their freezing point, the solution remains in the liquid state. This condition in which the solution remains liquid below its phase transition temperature is termed supercooling.
Implementation Method 3
During this transition from the liquid to the solid state, the solution moves from a higher to a lower free energy state, resulting in an exothermic reaction. The heat produced during this phase transition causes a transient warming of the sample during which the sample temperature increases.
Implementation Method 4
Subsequently, as the heat in the sample dissipates, the thermal dis-equilibrium between the sample and cooling device created during this event causes the sample to undergo a rapid cooling rate to reestablish thermal equilibrium.
Data Source
Figure 1~3b
Figure 4~6
AI summary
A cryopreservation device (60) includes an adaptor (65) mounted to a port of a container (62) for receiving and storing a liquid sample. The adaptor includes a first tubular branch (67) closed by a needle septum (72), and a second tubular branch(69) including a flexible tube (74) terminating in a needle septum (75). The container is initially at below-atmospheric pressure. In use, the sample is injected through the septum in the second branch. The flexible tube is then heat sealed and cut. The sealed container may then be subject to a cryopreservation protocol. After thawing, the sample liquid may be withdrawn by a needle puncturing the septum in the first branch. An auto-nucleating device including a hollow tube containing a crystalline cholesterol matrix therein, wherein the ends of the tube are closed by a membrane impermeable to the cholesterol but permeable to liquids, may be introduced into the container and provides a site for ice nucleation during freezing of the liquid sample within the container.