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The basic principles of cryopreservation and the importance of polyampholytes as a cryoprotectant

Year 2025, Volume: 9 Issue: 1, 47 - 56, 30.04.2025

Haydar Uğur , Saadet Kevser Pabuccuoğlu , Serhat Pabuccuoğlu

Abstract

Cryopreservation is an important process used to store the materials such as biological samples and food in liquid nitrogen (-196 °C) or in ultralow temperature freezer (-86 °C) for various biomedical, clinical and food-related applications. The main goal in the cryopreservation is to protect the materials against to damages during the freeze-thaw steps. Polyampholytes, which are the polymers containing both the cationic and anionic groups, have emerged as promising cryoprotective agents due to their unique properties. This review comprehensively discusses the history of cryopreservation, its fundamental principles, and the synthesis and cryoprotective properties of polyampholytes, with a focus on their mechanisms and applications.
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Keywords

Polyampholytes cell cryopreservation cryoprotectant non-permeable cryoprotectant

References

  • Agca, Y. (1994). Post-thaw survival and pregnancy rates of intact and biopsied and sexed in vitro produced bovine embryos after vitrification. Master's thesis, University of Wisconsin-Madison, US.
  • Ahmed, S., Fujita, S., & Matsumura, K. (2016). Enhanced protein internalization and efficient endosomal escape using polyampholyte-modified liposomes and freeze concentration. Nanoscale, 8(35), 15888-1590.
  • Ahmed, S., Miyawaki, O., & Matsumura, K. (2018). Enhanced adsorption of a protein-nanocarrier complex onto cell membranes through a high freeze concentration by a polyampholyte cryoprotectant. Langmuir, 34(6), 2352-2362.
  • Ahmed, S., Matsumura, K., & Hamada, T. (2019). Hydrophobic polyampholytes and nonfreezing cold temperature stimulate internalization of Au nanoparticles to zwitterionic liposomes. Langmuir, 35(5), 1740-1748.
  • Alfrey, T., Morawetz, H., Fitzgerald, E. B., & Fuoss, R. M. (1950). Synthetic electrical analog of proteins. Journal of the American Chemical Society, 72(4), 1864.
  • Alfrey, T., & Morawetz, H. (1952). Amphoteric polyelectrolytes. I. 2-Vinylpyridine-methacrylic acid copolymers. Journal of the American Chemical Society, 74(2), 436-438.
  • Al-Hasani, S., Diedrich, K., Wan der Ven, H., Reinecke, A., Hartje, M., & Krebs, D. (1987). Cryopreservation of human oocytes. Human Reproduction, 2(8), 695-700.
  • Armitage, W. J., & Juss, B. K. (1996). The influence of cooling rate on survival of frozen cells differs in monolayers and in suspensions. Cryo-Letters, 17, 213-218.
  • Benson, E. E. (2004). Cryoconserving algal and plant diversity: Historical perspectives and future challenges. In B. Fuller, N. Lane, & E. E. Benson (Eds.), Life in the frozen state (pp. 299-328). Baca Raton, US: CRC Press.
  • Bissoyi, A., Tomas, R. M. F., Gao, Y., Guo, Q., & Gibson, M. I. (2023). Cryopreservation of liver-cell spheroids with macromolecular cryoprotectants. ACS Applied Materials & Interfaces,15(2), 2630-2638.
  • Boquet, M., Selva, & J., Auroux, M. (1995). Effects of cooling and equilibration in DMSO, and cryopreservation of mouse oocytes on the rates of in vitro fertilization, development, and chromosomal abnormalities. Molecular Reproduction and Development, 40(1), 110-115.
  • Burkey, A. A., Hillsley, A., Harris, D. T., Baltzegar, J. R., Zhang, D. Y., Sprague, W. W., Rosales, A. M., & Lynd, A.A, (2020). Mechanism of polymer-mediated cryopreservation using poly(methyl glycidyl sulfoxide). Biomacromolecules, 21(11), 4668-4678.
  • Chen, Y., Sui, X., Zhang, T., Yang, J., Zhang, L., & Han, Y. (2023). Ice recrystallization inhibition mechanism of zwitterionic poly(carboxybetaine methacrylate). Physical Chemistry Chemical Physics, 25, 2752.
  • Chesné, C., & Guillouzo, A. (1988). Cryopreservation of isolated rat hepatocytes: A critical evaluation of freezing and thawing conditions. Cryobiology, 25(4), 323-330.
  • Ciferri, A., & Kudaibergenov, S. (2007). Natural and synthetic polyampholytes. Macromolecular Rapid Communications, 28, 1953-1968.
  • Colby, R. H., Ford, W. T., Kaur, B., Slater, L. A., Liang, S., Mourey, T. H., & D’Souza, L. (2011). Model random polyampholytes from nonpolar methacrylic esters. Macromolecules, 44(10), 3810-3816.
  • Dai, X., Zhao, D., Matsumura, K., & Rajan, R. (2023). Polyampholytes and Their Hydrophobic Derivatives as Excipients for Suppressing Protein Aggregation. ACS Applied Bio Materials,6(7), 2738-2746.
  • Dallman, R., Congdon, T. R., & Gibson, M. I. (2023). Cryopreservation of assay-ready hepatocyte monolayers by chemically-induced ice nucleation: preservation of hepatic function and hepatotoxicity screening capabilities. Biomaterials Science, 11, 7639-7654.
  • Du Prez, F., Kaya, N. U., Badi, N., Frank, D., & Resetco, C. (2017). Precisely alternating functionalized polyampholytes prepared in a single pot from sustainable thiolactone building blocks. ACS Macro Letters, 6(3), 277-280.
  • Ehrlich, G., & Doty, P. (1954). Macro-ions. III. The solution behavior of a polymeric ampholyte. Journal of the American Chemical Society, 76(14), 3764-3777.
  • Fahy, G. M. (2014). Principles of cryopreservation by vitrification. Methods in Molecular Biology , 1257, 21-82.
  • FAO. (2012). Cryoconservation of animal genetic resources (FAO Animal Production and Health Guidelines No. 12). Rome, Italy: FAO.
  • Friedler, S., Giudice, L. C., & Lamb, E. J. (1988). Cryopreservation of embryos and ova. Fertility and Sterility, 49(5), 743-764.
  • Gabaston, L., Furlong, S., Jackson, R., & Armes, S. (1999). Direct synthesis of novel acidic and zwitterionic block copolymers via TEMPO-mediated living free-radical polymerization. Polymer (Guildf.), 40(16), 4505-4514.
  • Hubel, A. (2009). Principles of cryopreservation. In A. Borini, G. Coticchio (Ed). Preservation of Human Oocytes 1th ed (pp 34-46). London, UK: CRC Press.
  • Ishibe, T., Gonzalez-Martinez, N., Georgiou, P. G., Murray, K. A., & Gibson, M. I. (2022). Synthesis of Poly(2-(methylsulfinyl) ethyl methacrylate) via Oxidation of Poly(2-(methylthio) ethyl methacrylate): Evaluation of the Sulfoxide Side Chain on Cryopreservation, ACS Polymers Au, 2, 449
  • Jain, M., Rajan, R., Hyon, S.-H., & Matsumura, K. (2014). Hydrogelation of dextran-based polyampholytes with cryoprotective properties via click chemistry. Biomaterials Science, 2(3), 308-317.
  • Jang, T. H. (2017). Cryopreservation and its clinical applications. Integrative Medicine Research, 6, 12-18.
  • Kamachi, M., Kurihara, M., & Stille, J. K. (1972). Synthesis of block polymers for desalination membranes. Preparation of block copolymers of 2-vinylpyridine and methacrylic acid or acrylic acid. Macromolecules, 5(2), 161-167.
  • Kamoshita, M., Kato, T., Fujiwara, K., Namiki, T., Matsumura, K., Hyon, S. H., Ito, J., & Kashiwazaki, N. (2017). Successful vitrification of pronuclear-stage pig embryos with a novel cryoprotective agent, carboxylated ϵ-poly-L-lysine. PLoS One, 12(4), 1-12.
  • Küçük, N., Raza, S., Matsumura, K., Uçan, U., Serin İ., Ceylan A., Aksoy M. (2021). Effect of different carboxylated poly l-lysine and dimethyl sulfoxide combinations on post thaw rabbit sperm functionality and fertility. Cryobiology,102, 127-132,
  • Li, G., Xue, H., Gao, C., Zhang, F., & Jiang, S. (2010). Nonfouling polyampholytes from an ion-pair comonomer with biomimetic adhesive groups. Macromolecules, 43(1), 14-16.
  • Liu, M., Zhang, X., Guo, H., Zhu, Y., Wen, C., Sui, X., Yang, J., & Zhang, L. (2019). Dimethyl sulfoxide-free cryopreservation of chondrocytes based on zwitterionic molecule and polymers. Biomacromolecules, 20 (10), 3980-3988.
  • Lovelock, J. E. (1953). The haemolysis of human red blood-cells by freezing and thawing. Biochimica et Biophysica Acta, 10(3), 414–426.
  • Maehara, M., Sato, M., Watanabe, M., Matsunari, H., Kokubo, M., Kanai, T., Sato, M., Matsumura, K., Hyon, S. H., Yokoyama, M., Mochida, J., & Nagashima, H. (2013). Development of a novel vitrification method for chondrocyte sheets. BMC Biotechnology, 13(1), 1.
  • Mandumpal, J. B., Kreck, C. A., & Mancera, R. L. (2010). A molecular mechanism of solvent cryoprotection in aqueous DMSO solutions. Physical Chemistry Chemical Physics, 13(9), 3839–3842.
  • Marton, H. L., Bhatt, A., Sagona, A. P., Kilbride, P., & Gibson, M. I. (2023a). Screening of Hydrophilic Polymers Reveals Broad Activity in Protecting Phages during Cryopreservation. Biomacromolecules, 25 (1), 413–424.
  • Marton, H. L., Kilbride, P., Ahmad, A., Sagona, A. P., & Gibson, M. I. (2023b). Anionic synthetic polymers prevent bacteriophage infection. Journal of the American Chemical Society, 45 (16), 8794–8799.
  • Matsumura, K., & Hyon, S. H. (2009). Polyampholytes as low toxic efficient cryoprotective agents with antifreeze protein properties. Biomaterials, 30, 4842–4849.
  • Matsumura, K., Hayashi, F., Nagashima, T., & Hyon, S. H. (2013a). Cryoprotective properties of polyampholytes. Cryobiology and Cryotechnology, 59(1), 23-28.
  • Matsumura, K., Hayashi, F., Nagashima, T., & Hyon, S. H. (2013b). Long-term cryopreservation of human mesenchymal stem cells using carboxylated poly-l-lysine without the addition of proteins or dimethyl sulfoxide. Journal of Biomaterials Science, Polymer Edition, 24(12), 1484–1497.
  • Matsumura, K., Kawamoto, K., Takeuchi, M., Yoshimura, S., Tanaka, D., & Hyon, S.H. (2016). Cryopreservation of a two-dimensional monolayer using a slow vitrification method with polyampholyte to inhibit ice crystal formation. ACS Biomaterials Science & Engineering, 2(6), 1023–1029.
  • Matsumura, K., Hayashi, F., Nagashima, T., Rajan, R., & Hyon, S. H. (2021). Molecular mechanisms of cell cryopreservation with polyampholytes studied by solid-state NMR. Communications Materials, 2, 15.
  • Mazur, P. (1963). Kinetics of water loss from cells at subzero temperatures and the likelihood of intracellular freezing. Journal of General Physiology, 47(2), 347–369.
  • Mazur, P. (1970). Cryobiology: The Freezing of Biological Systems, Science, 168(3934), 939-949.
  • Mazur, P., Leibo, S. P., & Chu, E. H. Y. (1972). A two-factor hypothesis of freezing injury: Evidence from chinese hamster tissue-culture cells. Experimental Cell Research, 71(2), 345–355.
  • Mazur, P. (1984). Freezing of living cells: mechanisms and implications. Physiological Reviews, 64(1), C125-C142.
  • Mazur, P., & Koshimoto, C. (2002). Is intracellular ice formation the cause of death of mouse sperm frozen at high cooling rates? Biology of Reproduction, 66, 1485–1490.
  • Mitchell, D. E., Lilliman, M., Spain, S. G., & Gibson, M. I. (2014). Quantitative study on the antifreeze protein mimetic ice growth inhibition properties of poly(ampholytes) derived from vinyl-based polymers. Biomaterials Science, 2(12), 1787–1795.
  • Mitchell, D. E., Cameron, N. R., & Gibson, M. I. (2015). Rational, yet simple, design and synthesis of an antifreeze-protein inspired polymer for cellular cryopreservation. 1 Chemical Communications, 51(65), 12977–12980.
  • Mumick, P. S., Welch, P. M., Salazar, L. C., & McCormick, C. L. (1994). Water-soluble copolymers. 56. Structure and solvation effects of polyampholytes in drag reduction. Macromolecules, 27(2), 323–331.
  • Murray, A., Congdon, T. R., Tomás, R. M. F., Kilbride, P., & Gibson, M. I. (2021). Red blood cell cryopreservation with minimal post-thaw lysis enabled by a synergistic combination of a cryoprotecting polyampholyte with DMSO/trehalose. Biomacromolecules, 23(2), 467–477.
  • Ock, S. A., & Rho, G. J. (2011). Effect of dimethyl sulfoxide (DMSO) on cryopreservation of porcine mesenchymal stem cells (pMSCs). Cell Transplantation, 20(8), 1231–1239.
  • Özkavukçu, S., & Erdemli, E. (2002). Cryopreservation: Basic knowledge and biophysical effects. Journal of Ankara Medical School, 24(4), 187-195.
  • Palasz, A. T., & Mapletoft, R. J. (1996). Cryopreservation of mammalian embryos and oocytes: Recent advances. Biotechnology Advances, 14, 127-149.
  • Pasch, J., Schiefer, A., Heschel, I., & Rau, G. (1999). Cryopreservation of keratinocytes in a monolayer. Cryobiology, 39(2), 158–168.
  • Patrickios, C. S., Hertler, W. R., Abbott, N. L., & Hatton, T. A. (1994). Diblock, ABC triblock, and random methacrylic polyampholytes: Synthesis by group transfer polymerization and solution behavior. Macromolecules, 27(4), 930–937.
  • Pegg, D. E., & Karow, A. M. (1987). The biophysics of organ preservation. New York, US: Plenum Press. Pesenti, T., Zhu, C., Gonzalez-Martinez, N., Tomás, R. M. F., Gibson, M. I., & Nicolas, J. (2022). Degradable polyampholytes from radical ring-opening copolymerization enhance cellular cryopreservation. ACS Macro Letters, 11 (7), 889–894.
  • Polge, C., Smith, A. & Parkes, A. (1949). Revival of Spermatozoa after Vitrification and Dehydration at Low Temperatures. Nature 164, 666. https://doi.org/10.1038/164666a0
  • Rajan, R., Jain, M., & Matsumura, K. (2013). Cryoprotective properties of completely synthetic polyampholytes via reversible addition-fragmentation chain transfer (RAFT) polymerization and the effects of hydrophobicity. Journal of Biomaterials Science, Polymer Edition, 24, 1767–1780.
  • Rajan, R., & Matsumura, K. (2014). Preparation of novel synthetic cryoprotectants. Cryobiology, 60(2), 99–103.
  • Rajan, R., Hayashi, F., Nagashima, T., & Matsumura, K. (2016). Toward a molecular understanding of the mechanism of cryopreservation by polyampholytes: Cell membrane interactions and hydrophobicity. Biomacromolecules, 17(5), 1882–1893.
  • Rajan, R., Kumar, N., & Matsumura, K. (2021). Design of an ice recrystallization-inhibiting polyampholyte-containing graft polymer for inhibition of protein aggregation. Biomacromolecules, 23 (2), 487–496.
  • Rajan, R., Kumar, N., Zhao, D., Dai, X., Kawamoto, K., & Matsumura, K. (2023). Polyampholyte‐Based Polymer Hydrogels for the Long‐Term Storage, Protection and Delivery of Therapeutic Proteins. Advanced Healthcare Materials, 12, 2203253.
  • Sağırkaya, H., & Bağış, H. (2003). Memeli embriyolarının kriyoprezervasyonu. Uludag Univ. J. Fac. Vet. Med., 22(1-2-3), 127-135.
  • Sambu, S. (2015). A bayesian approach to optimizing cryopreservation protocols. PeerJ, 3: e1039. https://doi.org/10.7717/peerj.1039
  • Shibao, Y., Fujiwara, K., Kawasaki, Y., Matsumura, K., Hyon, S.H., Ito, J., & Kashiwazaki, N. (2014). The effect of a novel cryoprotective agent, carboxylated ε-poly-l-lysine, on the developmental ability of re-vitrified mouse embryos at the pronuclear stage. Cryobiology, 68(2), 200–204.
  • Siebzehnruebl, E., Todorow, S., Van Uem, J., Koch, R., Wildt, L., & Lang, N. (1989). Cryopreservation of human and rabbit oocytes and one-cell embryos: A comparison of DMSO and propanediol. Human Reproduction, 4(3), 312–317.
  • Stubbs, C., Lipecki, J., & Gibson, M. I. (2017). Regioregular alternating polyampholytes have enhanced biomimetic ice recrystallization activity compared to random copolymers and the role of side chain versus main chain hydrophobicity. Biomacromolecules, 18(1), 295–302.
  • Stubbs, C., Bailey, T. L., Murray, K., & Gibson, M. I. (2020a). Polyampholytes as emerging macromolecular cryoprotectants. Biomacromolecules, 21(1), 7–17.
  • Stubbs, C., Murray, K. A., Ishibe, T., Mathers, R. T., & Gibson, M. I. (2020b). Combinatorial biomaterials discovery strategy to identify new macromolecular cryoprotectants. ACS Macro Letters, 9 (2), 290–294.
  • Tomás, R. M. F., Bissoyi, A., Congdon, T. R., & Gibson, M. I. (2022). Assay-ready cryopreserved cell monolayers enabled by macromolecular cryoprotectants. Biomacromolecules, 23 (9), 3948–3959.
  • Walters, E. M., Benson, J. D., Woods, E. J., Crister, J. K. (2009). The history of sperm cryopreservation. In A. A. Pacey, M. J. Tomlinson (Eds), Sperm banking theory and practice (pp. 1–17). London, UK: Cambridge University Press.
  • Watanabe, H., Kohaya, N., Kamoshita, M., Fujiwara, K., Matsumura, K., Hyon, S. H., Ito, J., & Kashiwazaki, N. (2013). Efficient production of live offspring from mouse oocytes vitrified with a novel cryoprotective agent, carboxylated ε-poly-L-lysine. PLoS One, 8(12), 10–14.
  • Yuan, L., Chen,B., Zhu, K., Ren, L., Yuan, X. (2024). Development of Macromolecular Cryoprotectants for Cryopreservation of Cells. Macromolecular Rapid Communication 45 (2400309), 1-15

The basic principles of cryopreservation and the importance of polyampholytes as a cryoprotectant

Year 2025, Volume: 9 Issue: 1, 47 - 56, 30.04.2025

Haydar Uğur , Saadet Kevser Pabuccuoğlu , Serhat Pabuccuoğlu

Abstract

Cryopreservation is an important process used to store the materials such as biological samples and food in liquid nitrogen (-196 °C) or in ultralow temperature freezer (-86 °C) for various biomedical, clinical and food-related applications. The main goal in the cryopreservation is to protect the materials against to damages during the freeze-thaw steps. Polyampholytes, which are the polymers containing both the cationic and anionic groups, have emerged as promising cryoprotective agents due to their unique properties. This review comprehensively discusses the history of cryopreservation, its fundamental principles, and the synthesis and cryoprotective properties of polyampholytes, with a focus on their mechanisms and applications.

Keywords

Polyampholytes cell cryopreservation cryoprotectant non-permeable cryoprotectant

References

  • Agca, Y. (1994). Post-thaw survival and pregnancy rates of intact and biopsied and sexed in vitro produced bovine embryos after vitrification. Master's thesis, University of Wisconsin-Madison, US.
  • Ahmed, S., Fujita, S., & Matsumura, K. (2016). Enhanced protein internalization and efficient endosomal escape using polyampholyte-modified liposomes and freeze concentration. Nanoscale, 8(35), 15888-1590.
  • Ahmed, S., Miyawaki, O., & Matsumura, K. (2018). Enhanced adsorption of a protein-nanocarrier complex onto cell membranes through a high freeze concentration by a polyampholyte cryoprotectant. Langmuir, 34(6), 2352-2362.
  • Ahmed, S., Matsumura, K., & Hamada, T. (2019). Hydrophobic polyampholytes and nonfreezing cold temperature stimulate internalization of Au nanoparticles to zwitterionic liposomes. Langmuir, 35(5), 1740-1748.
  • Alfrey, T., Morawetz, H., Fitzgerald, E. B., & Fuoss, R. M. (1950). Synthetic electrical analog of proteins. Journal of the American Chemical Society, 72(4), 1864.
  • Alfrey, T., & Morawetz, H. (1952). Amphoteric polyelectrolytes. I. 2-Vinylpyridine-methacrylic acid copolymers. Journal of the American Chemical Society, 74(2), 436-438.
  • Al-Hasani, S., Diedrich, K., Wan der Ven, H., Reinecke, A., Hartje, M., & Krebs, D. (1987). Cryopreservation of human oocytes. Human Reproduction, 2(8), 695-700.
  • Armitage, W. J., & Juss, B. K. (1996). The influence of cooling rate on survival of frozen cells differs in monolayers and in suspensions. Cryo-Letters, 17, 213-218.
  • Benson, E. E. (2004). Cryoconserving algal and plant diversity: Historical perspectives and future challenges. In B. Fuller, N. Lane, & E. E. Benson (Eds.), Life in the frozen state (pp. 299-328). Baca Raton, US: CRC Press.
  • Bissoyi, A., Tomas, R. M. F., Gao, Y., Guo, Q., & Gibson, M. I. (2023). Cryopreservation of liver-cell spheroids with macromolecular cryoprotectants. ACS Applied Materials & Interfaces,15(2), 2630-2638.
  • Boquet, M., Selva, & J., Auroux, M. (1995). Effects of cooling and equilibration in DMSO, and cryopreservation of mouse oocytes on the rates of in vitro fertilization, development, and chromosomal abnormalities. Molecular Reproduction and Development, 40(1), 110-115.
  • Burkey, A. A., Hillsley, A., Harris, D. T., Baltzegar, J. R., Zhang, D. Y., Sprague, W. W., Rosales, A. M., & Lynd, A.A, (2020). Mechanism of polymer-mediated cryopreservation using poly(methyl glycidyl sulfoxide). Biomacromolecules, 21(11), 4668-4678.
  • Chen, Y., Sui, X., Zhang, T., Yang, J., Zhang, L., & Han, Y. (2023). Ice recrystallization inhibition mechanism of zwitterionic poly(carboxybetaine methacrylate). Physical Chemistry Chemical Physics, 25, 2752.
  • Chesné, C., & Guillouzo, A. (1988). Cryopreservation of isolated rat hepatocytes: A critical evaluation of freezing and thawing conditions. Cryobiology, 25(4), 323-330.
  • Ciferri, A., & Kudaibergenov, S. (2007). Natural and synthetic polyampholytes. Macromolecular Rapid Communications, 28, 1953-1968.
  • Colby, R. H., Ford, W. T., Kaur, B., Slater, L. A., Liang, S., Mourey, T. H., & D’Souza, L. (2011). Model random polyampholytes from nonpolar methacrylic esters. Macromolecules, 44(10), 3810-3816.
  • Dai, X., Zhao, D., Matsumura, K., & Rajan, R. (2023). Polyampholytes and Their Hydrophobic Derivatives as Excipients for Suppressing Protein Aggregation. ACS Applied Bio Materials,6(7), 2738-2746.
  • Dallman, R., Congdon, T. R., & Gibson, M. I. (2023). Cryopreservation of assay-ready hepatocyte monolayers by chemically-induced ice nucleation: preservation of hepatic function and hepatotoxicity screening capabilities. Biomaterials Science, 11, 7639-7654.
  • Du Prez, F., Kaya, N. U., Badi, N., Frank, D., & Resetco, C. (2017). Precisely alternating functionalized polyampholytes prepared in a single pot from sustainable thiolactone building blocks. ACS Macro Letters, 6(3), 277-280.
  • Ehrlich, G., & Doty, P. (1954). Macro-ions. III. The solution behavior of a polymeric ampholyte. Journal of the American Chemical Society, 76(14), 3764-3777.
  • Fahy, G. M. (2014). Principles of cryopreservation by vitrification. Methods in Molecular Biology , 1257, 21-82.
  • FAO. (2012). Cryoconservation of animal genetic resources (FAO Animal Production and Health Guidelines No. 12). Rome, Italy: FAO.
  • Friedler, S., Giudice, L. C., & Lamb, E. J. (1988). Cryopreservation of embryos and ova. Fertility and Sterility, 49(5), 743-764.
  • Gabaston, L., Furlong, S., Jackson, R., & Armes, S. (1999). Direct synthesis of novel acidic and zwitterionic block copolymers via TEMPO-mediated living free-radical polymerization. Polymer (Guildf.), 40(16), 4505-4514.
  • Hubel, A. (2009). Principles of cryopreservation. In A. Borini, G. Coticchio (Ed). Preservation of Human Oocytes 1th ed (pp 34-46). London, UK: CRC Press.
  • Ishibe, T., Gonzalez-Martinez, N., Georgiou, P. G., Murray, K. A., & Gibson, M. I. (2022). Synthesis of Poly(2-(methylsulfinyl) ethyl methacrylate) via Oxidation of Poly(2-(methylthio) ethyl methacrylate): Evaluation of the Sulfoxide Side Chain on Cryopreservation, ACS Polymers Au, 2, 449
  • Jain, M., Rajan, R., Hyon, S.-H., & Matsumura, K. (2014). Hydrogelation of dextran-based polyampholytes with cryoprotective properties via click chemistry. Biomaterials Science, 2(3), 308-317.
  • Jang, T. H. (2017). Cryopreservation and its clinical applications. Integrative Medicine Research, 6, 12-18.
  • Kamachi, M., Kurihara, M., & Stille, J. K. (1972). Synthesis of block polymers for desalination membranes. Preparation of block copolymers of 2-vinylpyridine and methacrylic acid or acrylic acid. Macromolecules, 5(2), 161-167.
  • Kamoshita, M., Kato, T., Fujiwara, K., Namiki, T., Matsumura, K., Hyon, S. H., Ito, J., & Kashiwazaki, N. (2017). Successful vitrification of pronuclear-stage pig embryos with a novel cryoprotective agent, carboxylated ϵ-poly-L-lysine. PLoS One, 12(4), 1-12.
  • Küçük, N., Raza, S., Matsumura, K., Uçan, U., Serin İ., Ceylan A., Aksoy M. (2021). Effect of different carboxylated poly l-lysine and dimethyl sulfoxide combinations on post thaw rabbit sperm functionality and fertility. Cryobiology,102, 127-132,
  • Li, G., Xue, H., Gao, C., Zhang, F., & Jiang, S. (2010). Nonfouling polyampholytes from an ion-pair comonomer with biomimetic adhesive groups. Macromolecules, 43(1), 14-16.
  • Liu, M., Zhang, X., Guo, H., Zhu, Y., Wen, C., Sui, X., Yang, J., & Zhang, L. (2019). Dimethyl sulfoxide-free cryopreservation of chondrocytes based on zwitterionic molecule and polymers. Biomacromolecules, 20 (10), 3980-3988.
  • Lovelock, J. E. (1953). The haemolysis of human red blood-cells by freezing and thawing. Biochimica et Biophysica Acta, 10(3), 414–426.
  • Maehara, M., Sato, M., Watanabe, M., Matsunari, H., Kokubo, M., Kanai, T., Sato, M., Matsumura, K., Hyon, S. H., Yokoyama, M., Mochida, J., & Nagashima, H. (2013). Development of a novel vitrification method for chondrocyte sheets. BMC Biotechnology, 13(1), 1.
  • Mandumpal, J. B., Kreck, C. A., & Mancera, R. L. (2010). A molecular mechanism of solvent cryoprotection in aqueous DMSO solutions. Physical Chemistry Chemical Physics, 13(9), 3839–3842.
  • Marton, H. L., Bhatt, A., Sagona, A. P., Kilbride, P., & Gibson, M. I. (2023a). Screening of Hydrophilic Polymers Reveals Broad Activity in Protecting Phages during Cryopreservation. Biomacromolecules, 25 (1), 413–424.
  • Marton, H. L., Kilbride, P., Ahmad, A., Sagona, A. P., & Gibson, M. I. (2023b). Anionic synthetic polymers prevent bacteriophage infection. Journal of the American Chemical Society, 45 (16), 8794–8799.
  • Matsumura, K., & Hyon, S. H. (2009). Polyampholytes as low toxic efficient cryoprotective agents with antifreeze protein properties. Biomaterials, 30, 4842–4849.
  • Matsumura, K., Hayashi, F., Nagashima, T., & Hyon, S. H. (2013a). Cryoprotective properties of polyampholytes. Cryobiology and Cryotechnology, 59(1), 23-28.
  • Matsumura, K., Hayashi, F., Nagashima, T., & Hyon, S. H. (2013b). Long-term cryopreservation of human mesenchymal stem cells using carboxylated poly-l-lysine without the addition of proteins or dimethyl sulfoxide. Journal of Biomaterials Science, Polymer Edition, 24(12), 1484–1497.
  • Matsumura, K., Kawamoto, K., Takeuchi, M., Yoshimura, S., Tanaka, D., & Hyon, S.H. (2016). Cryopreservation of a two-dimensional monolayer using a slow vitrification method with polyampholyte to inhibit ice crystal formation. ACS Biomaterials Science & Engineering, 2(6), 1023–1029.
  • Matsumura, K., Hayashi, F., Nagashima, T., Rajan, R., & Hyon, S. H. (2021). Molecular mechanisms of cell cryopreservation with polyampholytes studied by solid-state NMR. Communications Materials, 2, 15.
  • Mazur, P. (1963). Kinetics of water loss from cells at subzero temperatures and the likelihood of intracellular freezing. Journal of General Physiology, 47(2), 347–369.
  • Mazur, P. (1970). Cryobiology: The Freezing of Biological Systems, Science, 168(3934), 939-949.
  • Mazur, P., Leibo, S. P., & Chu, E. H. Y. (1972). A two-factor hypothesis of freezing injury: Evidence from chinese hamster tissue-culture cells. Experimental Cell Research, 71(2), 345–355.
  • Mazur, P. (1984). Freezing of living cells: mechanisms and implications. Physiological Reviews, 64(1), C125-C142.
  • Mazur, P., & Koshimoto, C. (2002). Is intracellular ice formation the cause of death of mouse sperm frozen at high cooling rates? Biology of Reproduction, 66, 1485–1490.
  • Mitchell, D. E., Lilliman, M., Spain, S. G., & Gibson, M. I. (2014). Quantitative study on the antifreeze protein mimetic ice growth inhibition properties of poly(ampholytes) derived from vinyl-based polymers. Biomaterials Science, 2(12), 1787–1795.
  • Mitchell, D. E., Cameron, N. R., & Gibson, M. I. (2015). Rational, yet simple, design and synthesis of an antifreeze-protein inspired polymer for cellular cryopreservation. 1 Chemical Communications, 51(65), 12977–12980.
  • Mumick, P. S., Welch, P. M., Salazar, L. C., & McCormick, C. L. (1994). Water-soluble copolymers. 56. Structure and solvation effects of polyampholytes in drag reduction. Macromolecules, 27(2), 323–331.
  • Murray, A., Congdon, T. R., Tomás, R. M. F., Kilbride, P., & Gibson, M. I. (2021). Red blood cell cryopreservation with minimal post-thaw lysis enabled by a synergistic combination of a cryoprotecting polyampholyte with DMSO/trehalose. Biomacromolecules, 23(2), 467–477.
  • Ock, S. A., & Rho, G. J. (2011). Effect of dimethyl sulfoxide (DMSO) on cryopreservation of porcine mesenchymal stem cells (pMSCs). Cell Transplantation, 20(8), 1231–1239.
  • Özkavukçu, S., & Erdemli, E. (2002). Cryopreservation: Basic knowledge and biophysical effects. Journal of Ankara Medical School, 24(4), 187-195.
  • Palasz, A. T., & Mapletoft, R. J. (1996). Cryopreservation of mammalian embryos and oocytes: Recent advances. Biotechnology Advances, 14, 127-149.
  • Pasch, J., Schiefer, A., Heschel, I., & Rau, G. (1999). Cryopreservation of keratinocytes in a monolayer. Cryobiology, 39(2), 158–168.
  • Patrickios, C. S., Hertler, W. R., Abbott, N. L., & Hatton, T. A. (1994). Diblock, ABC triblock, and random methacrylic polyampholytes: Synthesis by group transfer polymerization and solution behavior. Macromolecules, 27(4), 930–937.
  • Pegg, D. E., & Karow, A. M. (1987). The biophysics of organ preservation. New York, US: Plenum Press. Pesenti, T., Zhu, C., Gonzalez-Martinez, N., Tomás, R. M. F., Gibson, M. I., & Nicolas, J. (2022). Degradable polyampholytes from radical ring-opening copolymerization enhance cellular cryopreservation. ACS Macro Letters, 11 (7), 889–894.
  • Polge, C., Smith, A. & Parkes, A. (1949). Revival of Spermatozoa after Vitrification and Dehydration at Low Temperatures. Nature 164, 666. https://doi.org/10.1038/164666a0
  • Rajan, R., Jain, M., & Matsumura, K. (2013). Cryoprotective properties of completely synthetic polyampholytes via reversible addition-fragmentation chain transfer (RAFT) polymerization and the effects of hydrophobicity. Journal of Biomaterials Science, Polymer Edition, 24, 1767–1780.
  • Rajan, R., & Matsumura, K. (2014). Preparation of novel synthetic cryoprotectants. Cryobiology, 60(2), 99–103.
  • Rajan, R., Hayashi, F., Nagashima, T., & Matsumura, K. (2016). Toward a molecular understanding of the mechanism of cryopreservation by polyampholytes: Cell membrane interactions and hydrophobicity. Biomacromolecules, 17(5), 1882–1893.
  • Rajan, R., Kumar, N., & Matsumura, K. (2021). Design of an ice recrystallization-inhibiting polyampholyte-containing graft polymer for inhibition of protein aggregation. Biomacromolecules, 23 (2), 487–496.
  • Rajan, R., Kumar, N., Zhao, D., Dai, X., Kawamoto, K., & Matsumura, K. (2023). Polyampholyte‐Based Polymer Hydrogels for the Long‐Term Storage, Protection and Delivery of Therapeutic Proteins. Advanced Healthcare Materials, 12, 2203253.
  • Sağırkaya, H., & Bağış, H. (2003). Memeli embriyolarının kriyoprezervasyonu. Uludag Univ. J. Fac. Vet. Med., 22(1-2-3), 127-135.
  • Sambu, S. (2015). A bayesian approach to optimizing cryopreservation protocols. PeerJ, 3: e1039. https://doi.org/10.7717/peerj.1039
  • Shibao, Y., Fujiwara, K., Kawasaki, Y., Matsumura, K., Hyon, S.H., Ito, J., & Kashiwazaki, N. (2014). The effect of a novel cryoprotective agent, carboxylated ε-poly-l-lysine, on the developmental ability of re-vitrified mouse embryos at the pronuclear stage. Cryobiology, 68(2), 200–204.
  • Siebzehnruebl, E., Todorow, S., Van Uem, J., Koch, R., Wildt, L., & Lang, N. (1989). Cryopreservation of human and rabbit oocytes and one-cell embryos: A comparison of DMSO and propanediol. Human Reproduction, 4(3), 312–317.
  • Stubbs, C., Lipecki, J., & Gibson, M. I. (2017). Regioregular alternating polyampholytes have enhanced biomimetic ice recrystallization activity compared to random copolymers and the role of side chain versus main chain hydrophobicity. Biomacromolecules, 18(1), 295–302.
  • Stubbs, C., Bailey, T. L., Murray, K., & Gibson, M. I. (2020a). Polyampholytes as emerging macromolecular cryoprotectants. Biomacromolecules, 21(1), 7–17.
  • Stubbs, C., Murray, K. A., Ishibe, T., Mathers, R. T., & Gibson, M. I. (2020b). Combinatorial biomaterials discovery strategy to identify new macromolecular cryoprotectants. ACS Macro Letters, 9 (2), 290–294.
  • Tomás, R. M. F., Bissoyi, A., Congdon, T. R., & Gibson, M. I. (2022). Assay-ready cryopreserved cell monolayers enabled by macromolecular cryoprotectants. Biomacromolecules, 23 (9), 3948–3959.
  • Walters, E. M., Benson, J. D., Woods, E. J., Crister, J. K. (2009). The history of sperm cryopreservation. In A. A. Pacey, M. J. Tomlinson (Eds), Sperm banking theory and practice (pp. 1–17). London, UK: Cambridge University Press.
  • Watanabe, H., Kohaya, N., Kamoshita, M., Fujiwara, K., Matsumura, K., Hyon, S. H., Ito, J., & Kashiwazaki, N. (2013). Efficient production of live offspring from mouse oocytes vitrified with a novel cryoprotective agent, carboxylated ε-poly-L-lysine. PLoS One, 8(12), 10–14.
  • Yuan, L., Chen,B., Zhu, K., Ren, L., Yuan, X. (2024). Development of Macromolecular Cryoprotectants for Cryopreservation of Cells. Macromolecular Rapid Communication 45 (2400309), 1-15
There are 75 citations in total.

Details

Primary Language English
Subjects Veterinary Sciences (Other)
Journal Section Review Articles
Authors

Haydar Uğur 0000-0003-1479-3104

Saadet Kevser Pabuccuoğlu 0000-0002-1793-0859

Serhat Pabuccuoğlu 0000-0002-6200-3018

Publication Date April 30, 2025
Submission Date April 12, 2025
Acceptance Date April 26, 2025
Published in Issue Year 2025 Volume: 9 Issue: 1

Cite

APA Uğur, H., Pabuccuoğlu, S. K., & Pabuccuoğlu, S. (2025). The basic principles of cryopreservation and the importance of polyampholytes as a cryoprotectant. Journal of Istanbul Veterinary Sciences, 9(1), 47-56.

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