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Study on freezing process of killifish egg: utilizing the undercooled state for cryopreservation
M Ujihira1, N Aizawa, K Tanishita
1Department of Mechanical Engineering, Keio University, Kanagawa, Japan.
Bio-Medical Materials and Engineering
|January 1, 1994
Summary
Preserving large cells and tissues requires maintaining an undercooled state during freezing. This method, tested on killifish eggs, avoids intracellular ice crystal damage, improving cryopreservation success.
Area of Science:
- Cryobiology
- Cellular Biology
- Biophysics
Background:
- Intracellular ice crystal formation during freezing destroys cells and tissues.
- Cryopreservation aims to preserve biological materials by preventing such damage.
Purpose of the Study:
- To investigate the feasibility of preserving large cells and tissues by maintaining an undercooled state.
- To determine optimal conditions for cryopreservation by avoiding intracellular ice crystal growth.
Main Methods:
- Utilized fertilized killifish eggs as biological test samples.
- Employed a cooling system with Peltier devices, capable of reaching -50°C.
- Observed egg morphology and evaluated damage rates (hatching rate) at cooling rates from 0.1 to 10°C/min between 0°C and -40°C.
Main Results:
- Intracellular undercooled states were observed under both extracellular undercooling and freezing conditions.
- Extracellular undercooling demonstrated a protective effect on the eggs.
- Extracellular freezing frequently led to egg damage.
Conclusions:
- Cryopreservation is achieved by maintaining an undercooled state until -40°C, followed by rapid freezing with liquid nitrogen.
- Maintaining a stable extracellular undercooled state is crucial for achieving intracellular undercooling and successful cryopreservation.
- Dehydration or stabilizing the extracellular undercooled state is desirable for effective cryopreservation.