A Simple Cryomicroscopic Method to Measure the Size of Intracellular Ice Crystals

2009 ◽  
Author(s):  
Xu Han ◽  
Hongbin Ma ◽  
John K. Critser
Keyword(s):  
Melting Point ◽  
Ethylene Glycol ◽  
Cooling Rate ◽  
Ice Crystals ◽  
Ice Formation ◽  
Melting Point Depression ◽  
Critical Importance ◽  
Factors Influencing ◽  
Intracellular Ice Formation ◽  
Intracellular Ice

Investigating the factors influencing the characteristics of intracellular ice formation (IIF) is of critical importance for cryopreservation and cryosurgery techniques. However, for the detection of the size of intracellular ice crystals, ∼10nm-0.1μm, there exist serious technical and theoretical difficulties. In this study, a cryomicroscopic method was established to measure the size of intracellular ice crystals in mouse oocytes during their warming processes by investigating the melting point depression of the intracellular ice crystals from extracellular ones. Using the Gibbs-Thomson relation, the size of intracellular ice crystals was calculated and the results range from 4–28 nm, when the molality of the extracellular ethylene glycol and NaCl ranges from 0 to 4m and 0.15 to 0.6m, respectively, and the cooling rate is 100K/min.

scholarly journals Intracellular ice formation in mouse zygotes and early morulae vs. cooling rate and temperature-experimental vs. theory

Cryobiology ◽  
2016 ◽  
Vol 73 (2) ◽  
pp. 181-186 ◽  
Author(s):  
Bo Jin ◽  
Shinsuke Seki ◽  
Estefania Paredes ◽  
Juan Qiu ◽  
Yanbin Shi ◽  
...  
Keyword(s):  
Cooling Rate ◽  
Ice Formation ◽  
Intracellular Ice Formation ◽  
Intracellular Ice ◽  
Mouse Zygotes

Quantifying the Growth Rate and Morphology of Ice Crystals Growing in Cryoprotectants via High-Speed Camera and Cryo-Microscope

2013 ◽  
Author(s):  
Xu Xue ◽  
Zhi Zhu He ◽  
Jing Liu
Keyword(s):  
Growth Rate ◽  
Cooling Rate ◽  
High Speed ◽  
Ice Crystals ◽  
Experimental Methodology ◽  
Impact Factors ◽  
High Speed Camera ◽  
Intracellular Ice Formation ◽  
Intracellular Ice ◽  
Near Future

Recently, several significant progresses have been made on the studies of extracellular and intracellular ice formation based on high-speed camera and cryomicroscope. This experimental methodology could accurately capture rapid formation process of ice crystals at micro-scale. In this paper, we are dedicated to quantify and comparatively investigate the growth rate and morphology of ice crystals growing in DMSO, sucrose and trehalose, respectively via high-speed camera and cryo-microscope. Several impact factors such as the concentration of cryoprotectants and the cooling rate have been investigated. The results indicate that the species and concentration of cryoprotectants and the cooling rate could significantly affect the growth rate and morphology of ice crystals. DMSO is better than trehalose and sucrose as cryoprotectant because of its molecular structure. This work may enhance current understanding of the factors for ice crystals formation and help optimize the cryopreservation process in the near future.

A Study of Intracellular Ice Formation in Jurkat Cells

2008 ◽  
Author(s):  
Tathagata Acharya ◽  
Ram V. Devireddy
Keyword(s):  
Cooling Rate ◽  
Jurkat Cells ◽  
Freezing Rate ◽  
Ice Formation ◽  
Substantial Evidence ◽  
Cooling Rates ◽  
Cryoprotective Agents ◽  
Intracellular Ice Formation ◽  
Intracellular Ice ◽  
Phosphate Buffered Saline

The objective of this study was to characterize the IIF behavior of Jurkat cells in isotonic conditions in the absence of any cryoprotective agents. The Jurkat cells were collected from culture and then washed and re-suspended in Dulbecco’s Phosphate Buffered Saline (PBS). The freezing experiments were carried out at defined freezing protocols and at various freezing rates of 5, 20, 30 and 50 °C/min. The results suggest there was no substantial evidence of intracellular ice formation at lower cooling rates of 5, 20 and 30° C/min. The first conspicuous indication of intracellular ice formation (IIF) was observed at a freezing rate of 50 °C/min. At this cooling rate, unlike the usual sudden blackening of cells, the cells suddenly grew and exploded suggesting the formation of intracellular ice, which was reminiscent of a prior observed phenomenon for IIF.

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