Visualization of intracellular ice crystals formed in very rapidly frozen cells at −27 °C

Measurement of the size of intracellular ice crystals in mouse oocytes using a melting point depression method and the influence of intracellular solute concentrations

“…As shown in Table 3, the values of r range from ~ 4 to 28 nm with different initial solute concentrations, and these are on the same order of magnitude as the results from the previous theoretical investigations or electron microscopic observations [12,24,32]. During our experiments, it was observed that there exists a probability that some cells partially freeze and recrystallize during warming.…”

“…However, for the detection of the size of intracellular ice crystals, there exist serious technical and theoretical difficulties. For example, electron microscopy has been used to visualize the ice crystals in frozen cells in a few studies [24]. The complicated sample preparation processes for electron microscopy, esp.…”

“…The Gibbs-Thomson relation has been widely used to calculate or estimate the size of micro-particles by measuring the depression of their melting points [4,15,27]. Since the size of intracellular ice crystals generally ranges from several nanometers to less than 0.1μm [12,13,24,32], the melting point depression of intracellular ice would be predicted to be approximately 1 to 10°C. Although these facts indicate that the melting depression method may be a promising approach, the current thermal analytical instruments cannot be used for such measurement due to the small volume of intracellular solutions.…”

Cryobank of plant genetic resources in Russian Academy of Sciences

“…Most volume of plant cell is occupied by central vacuole that contains freezable water. Cell damages during freezing and subsequent thawing can be caused, on the one hand, by the formation of intracellular ice crystals with acute facets disintegrating cell membranes [3,4], and, on the other hand, by dehydration. Therefore the most dangerous process, which may occur during freezing is intracellular ice formation.…”

A rapid technique for the histological examination of large ovarian follicles

“…Figure 5 shows the effect of freezing an ovary that was fixed but not glycerolized ; the basal membrane is disorganized and the nuclei of the granulosa cells have been markedly altered. Glycerol may protect the tissues by lowering the freezing temperature of intracellular water, by modifying the shape of ice crystals (Shimada and Asahina, 1975 ;Sherman and Liu, 1976) and by stabilizing the membranes (Maurer, 1978 …”