title: A study on ice crystal formation behavior at intracellular freezing of plant cells using a high-speed camera.
abstract: Intracellular ice crystal formation (IIF) causes several problems to cryopreservation, and it is the key to developing improved cryopreservation techniques that can ensure the long-term preservation of living tissues. Therefore, the ability to capture clear intracellular freezing images is important for understanding both the occurrence and the IIF behavior. The authors developed a new cryomicroscopic system that was equipped with a high-speed camera for this study and successfully used this to capture clearer images of the IIF process in the epidermal tissues of strawberry geranium (Saxifraga stolonifera Curtis) leaves. This system was then used to examine patterns in the location and formation of intracellular ice crystals and to evaluate the degree of cell deformation because of ice crystals inside the cell and the growing rate and grain size of intracellular ice crystals at various cooling rates. The results showed that an increase in cooling rate influenced the formation pattern of intracellular ice crystals but had less of an effect on their location. Moreover, it reduced the degree of supercooling at the onset of intracellular freezing and the degree of cell deformation; the characteristic grain size of intracellular ice crystals was also reduced, but the growing rate of intracellular ice crystals was increased. Thus, the high-speed camera images could expose these changes in IIF behaviors with an increase in the cooling rate, and these are believed to have been caused by an increase in the degree of supercooling.
authors.affiliation: Graduate School of Mechanical Engineering, Kanagawa Institute of Technology, 1030 Shimo-ogino, Atsugi, Kanagawa, 243-0292, Japan. Electronic address: s1291001@cco.kanagawa-it.ac.jp.
authors.forename: Takako
authors.initials: T
authors.lastname: Ninagawa
authors.affiliation: Toyo Engineering Works, LTD., 1634, Shimotsuruma, Yamato, Kanagawa, 242-0001, Japan.
authors.forename: Akemi
authors.initials: A
authors.lastname: Eguchi
authors.affiliation: Cryobiofrontier Research Center, Faculty of Agriculture, Iwate University, Ueda 3-18-8, Morioka, Iwate, 020-8550, Japan. Electronic address: ykawa@iwate-u.ac.jp.
authors.forename: Yukio
authors.initials: Y
authors.lastname: Kawamura
authors.affiliation: Department of Mechanical Engineering, Oita National College of Technology, 1666 Maki, Oita, 870-0152, Japan. Electronic address: konishi@oita-ct.ac.jp.
authors.forename: Tadashi
authors.initials: T
authors.lastname: Konishi
authors.affiliation: Graduate School of Mechanical Engineering, Kanagawa Institute of Technology, 1030 Shimo-ogino, Atsugi, Kanagawa, 243-0292, Japan. Electronic address: narumi@me.kanagawa-it.ac.jp.
authors.forename: Akira
authors.initials: A
authors.lastname: Narumi
mesh_terms: D000818:Animals; D015925:Cryopreservation / Q000379:methods*; D005615:Freezing; D007053:Ice; D010784:Photomicrography / Q000379:methods*; D059828:Plant Cells
chemical_list: D007053:Ice
keywords: High-speed camera; Ice crystal; Intracellular freezing; Plant; Plasma membrane; Supercooling