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Measurement of Intracellular Ice Formation Kinetics by High-Speed Video Cryomicroscopy

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Cryopreservation and Freeze-Drying Protocols

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1257))

Abstract

Quantitative information about the kinetics and cumulative probability of intracellular ice formation is necessary to develop minimally damaging freezing procedures for the cryopreservation of cells and tissue. Conventional cryomicroscopic assays, which rely on indirect evidence of intracellular freezing (e.g., opacity changes in the cell cytoplasm), can yield significant errors in the estimated kinetics. In contrast, the formation and growth of intracellular ice crystals can be accurately detected using temporally resolved imaging methods (i.e., video recording at sub-millisecond resolution). Here, detailed methods for the setup and operation of a high-speed video cryomicroscope system are described, including protocols for imaging of intracellular ice crystallization events, and stochastic analysis of the ice formation kinetics in a cell population. Recommendations are provided for temperature profile design, sample preparation, and configuration of the video acquisition parameters. Throughout this chapter, the protocols incorporate best practices that have been drawn from over a decade of experience with high-speed video cryomicroscopy in our laboratory.

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References

  1. Mazur P, Leibo SP, Chu EHY (1972) A two-factor hypothesis of freezing injury: evidence from Chinese hamster tissue-culture cells. Exp Cell Res 71:345–355

    Article  CAS  Google Scholar 

  2. Toner M (1993) Nucleation of ice crystals inside biological cells. In: Steponkus P (ed) Advances in low-temperature biology, vol 2. JAI Press, London, pp 1–51

    Google Scholar 

  3. Karlsson JOM, Cravalho EG, Toner M (1993) Intracellular ice formation: causes and consequences. Cryo-Letters 14:323–336

    Google Scholar 

  4. Göppert HR (1830) Ueber die Wärme-Entwickelung in den Pflanzen, deren Gefrieren und die Schutzmittel gegen dasselbe. Joseph Max, Breslau

    Google Scholar 

  5. Molisch H (1897) Untersuchungen über das Erfrieren der Pflanzen. Gustav Fischer, Jena

    Google Scholar 

  6. Diller KR, Cravalho EG (1971) A cryomicroscope for the study of freezing and thawing processes in biological cells. Cryobiology 7:191–199

    Article  Google Scholar 

  7. Brown MS, Reuter FW (1974) Freezing of nonwoody plant tissues. III. Videotape micrography and the correlation between individual cellular freezing events and temperature changes in the surrounding tissue. Cryobiology 11:185–191

    Article  CAS  Google Scholar 

  8. Pitt RE, Chandrasekaran M, Parks JE (1992) Performance of a kinetic model for intracellular ice formation based on the extent of supercooling. Cryobiology 29:359–373

    Article  CAS  Google Scholar 

  9. Toner M, Tompkins RG, Cravalho EG et al (1992) Transport phenomena during freezing of isolated hepatocytes. AIChE J 38:1512–1522

    Article  CAS  Google Scholar 

  10. Irimia D, Karlsson JOM (2005) Kinetics of intracellular ice formation in one-dimensional arrays of interacting biological cells. Biophys J 88:647–660

    Article  CAS  Google Scholar 

  11. Karlsson JOM (2010) Effects of solution composition on the theoretical prediction of ice nucleation kinetics and thermodynamics. Cryobiology 60:43–51

    Article  CAS  Google Scholar 

  12. Higgins AZ, Karlsson JOM (2013) Effects of intercellular junction protein expression on intracellular ice formation in mouse insulinoma cells. Biophys J 105:2006–2015

    Article  CAS  Google Scholar 

  13. Toner M, Cravalho EG, Stachecki J et al (1993) Nonequilibrium freezing of one-cell mouse embryos. Membrane integrity and developmental potential. Biophys J 64:1908–1921

    Article  CAS  Google Scholar 

  14. Karlsson JOM, Eroglu A, Toth TL et al (1996) Fertilization and development of mouse oocytes cryopreserved using a theoretically optimized protocol. Hum Reprod 11:1296–1305

    Article  CAS  Google Scholar 

  15. Karlsson JOM (2001) A theoretical model of intracellular devitrification. Cryobiology 42:154–169

    Article  CAS  Google Scholar 

  16. Toner M, Cravalho EG, Karel M (1990) Thermodynamics and kinetics of intracellular ice formation during freezing of biological cells. J Appl Phys 67:1582–1592

    Article  Google Scholar 

  17. Chambers R, Hale HP (1932) The formation of ice in protoplasm. Proc R Soc Lond B 110:336–352

    Article  CAS  Google Scholar 

  18. Luyet BJ, Gibbs MC (1937) On the mechanism of congelation and of death in the rapid freezing of epidermal plant cells. Biodynamica 1:1–18

    Google Scholar 

  19. Asahina E (1953) Analysis of the freezing process of living organisms. X. Freezing process of egg cell of sea-urchin. Low Temp Sci 10:81–92

    Google Scholar 

  20. Smith AU, Smiles J (1953) Microscopic studies of mammalian tissues during cooling to and rewarming from -79 °C. J R Microsc Soc 73:134–139

    Article  CAS  Google Scholar 

  21. Toner M, Cravalho EG, Karel M et al (1991) Cryomicroscopic analysis of intracellular ice formation during freezing of mouse oocytes without cryoadditives. Cryobiology 28:55–71

    Article  CAS  Google Scholar 

  22. Guenther JF, Seki S, Kleinhans FW et al (2006) Corrigendum to “Extra- and intra-cellular ice formation in Stage I and II Xenopus laevis oocytes”. Cryobiology 53:294

    Article  CAS  Google Scholar 

  23. Stott SL, Karlsson JOM (2009) Visualization of intracellular ice formation using high-speed video cryomicroscopy. Cryobiology 58:84–95

    Article  CAS  Google Scholar 

  24. Grocutt P (2014) Linkam Scientific Instruments, Ltd., Surrey, UK. Personal communication

    Google Scholar 

  25. Seki S, Mazur P (2010) The temperature and type of intracellular ice formation in preimplantation mouse embryos as a function of the developmental stage. Biol Reprod 82:1198–1205

    Article  CAS  Google Scholar 

  26. Aalen OO, Borgan Ø, Gjessing HK (2008) Survival and event history analysis: a process point of view. Springer, New York

    Book  Google Scholar 

  27. Scheiwe MW, Körber C (1987) Quantitative cryomicroscopic analysis of intracellular freezing of granulocytes without cryoadditive. Cryobiology 24:473–483

    Article  CAS  Google Scholar 

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Acknowledgments

The author gratefully acknowledges Peter Grocutt of Linkam Scientific Instruments for valuable discussions and technical support during the development of the high-speed video cryomicroscopy system and during the writing of this chapter. This work was supported by National Science Foundation grant CBET-1066619.

Author information

Authors and Affiliations

  1. Department of Mechanical Engineering, Villanova University, 800 E. Lancaster Ave, Villanova, PA, 19085-1681, USA

    Jens O. M. Karlsson Ph.D.

Authors
  1. Jens O. M. Karlsson Ph.D.
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Correspondence to Jens O. M. Karlsson Ph.D. .

Editor information

Editors and Affiliations

  1. Institute of Multiphase Processes, Leibniz Universität Hannover, Hannover, Germany

    Willem F. Wolkers

  2. Unit for Reproductive Medicine, Clinic for Horses, University of Veterinary Medicine Hannover, Hannover, Germany

    Harriëtte Oldenhof

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Karlsson, J.O.M. (2015). Measurement of Intracellular Ice Formation Kinetics by High-Speed Video Cryomicroscopy. In: Wolkers, W., Oldenhof, H. (eds) Cryopreservation and Freeze-Drying Protocols. Methods in Molecular Biology, vol 1257. Springer, New York, NY. https://doi.org/10.1007/978-1-4939-2193-5_7

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  • DOI: https://doi.org/10.1007/978-1-4939-2193-5_7

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  • Publisher Name: Springer, New York, NY

  • Print ISBN: 978-1-4939-2192-8

  • Online ISBN: 978-1-4939-2193-5

  • eBook Packages: Springer Protocols

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