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.

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