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细胞微液滴可控快速降温及复温的冻存系统研究
Research on the Cryopreservation System of Controlled Rapid Cooling and Rewarming of Cellular Micro-droplets
- 默认刊物名称 2025年
- 作者机构：
上海理工大学,上海市上海市200082,中国
- 作者简介：
- 基金信息：国家自然科学基金(52076140)
DOI：
中图分类号：收稿日期：2024-11-27，
修回日期：2024-12-22，
录用日期：2025-01-22，
- 稿件说明：
移动端阅览
胡健. 细胞微液滴可控快速降温及复温的冻存系统研究[J/OL]. 默认刊物名称, 2025.
Research on the Cryopreservation System of Controlled Rapid Cooling and Rewarming of Cellular Micro-droplets[J/OL]. Moren journal, 2025.
摘要
细胞微液滴的玻璃化保存方法在低温保存领域受到广泛关注，但微液滴玻璃化过程还存在降温/复温过程不可控等局限性。本文采用移动滑台升降，结合焦耳热复温方法搭建用于细胞微液滴玻璃化过程可控快速降温及复温的冻存系统， 该系统通过调节浸入液氮的速度实现快速、可控降温，并通过控制焦耳热的加热时间、电流大小实现高速率复温过程，实现微液滴的玻璃化过程并显著避免了液滴复温过程的反玻璃化对细胞的损伤。结果表明：本系统可控制降温速率可以达到1.8×10
4
℃/min，能够实现细胞微液滴较低浓度保护剂下的玻璃化保存；复温速率控制可以达到4.0×10
4
℃/min，能有效避免较低浓度保护剂复温过程反玻璃化及冰晶再生长的发生；通过微液滴A549细胞低温保存效果，证实了本系统冷冻复苏后的存活率显著高于传统麦管保存。本文研究工作有望为细胞微液滴自动玻璃化保存及复温方法提供新的解决方案。
Abstract
The vitrification preservation method of cell microdroplets has received extensive attention in the field of cryopreservation. However
the vitrification process of microdroplets still has limitations such as uncontrollable cooling/rewarming processes. In this paper
a cryopreservation system for controllable rapid cooling and rewarming during the vitrification process of cell microdroplets was constructed by combining the lifting of a moving slide table with the Joule heating rewarming method. This system achieves rapid and controllable cooling by adjusting the speed of immersion in liquid nitrogen
and realizes a high-rate rewarming process by controlling the heating time and current intensity of Joule heating
thus realizing the vitrification process of microdroplets and significantly avoiding the damage to cells caused by devitrification during the droplet rewarming process. The results show that the cooling rate controlled by this system can reach 1.8 × 10⁴ °C/min
enabling the vitrification preservation of cell microdroplets with a relatively low concentration of cryoprotectant. The rewarming rate control can reach 4.0 × 10⁴ °C/min
which can effectively avoid the occurrence of devitrification and ice crystal regrowth during the rewarming process of the relatively low concentration of cryoprotectant. Through the cryopreservation effect of A549 cells in microdroplets
it has been confirmed that the survival rate after cryopreservation and resuscitation of this system is significantly higher than that of the traditional straw preservation. The research work in this paper is expected to provide new solutions for the automatic vitrification preservation and rewarming methods of cell microdroplets.
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references
MENEGHEL J , KILBRIDE P , MORRIS G J . Cryopreservation as a Key Element in the Successful Delivery of Cell-Based Therapies-A Review [J ] . Front Med (Lausanne) , 2020 , 7 : 592242 .
TAYLOR M J , WEEGMAN B P , BAICU S C , et al . New Approaches to Cryopreservation of Cells, Tissues, and Organs [J ] . Transfus Med Hemother , 2019 , 46 ( 3 ): 197 - 215 .
WENG L , BEAUCHESNE P R . Dimethyl sulfoxide-free cryopreservation for cell therapy: A review [J ] . Cryobiology , 2020 , 94 : 9 - 17 .
AMINI M , BENSON J D . Technologies for Vitrification Based Cryopreservation [J ] . Bioengineering (Basel) , 2023 , 10 ( 5 ).
AONO A , NAGATOMO H , TAKUMA T , et al . Dynamics of intracellular phospholipid membrane organization during oocyte maturation and successful vitrification of immature oocytes retrieved by ovum pick-up in cattle [J ] . Theriogenology , 2013 , 79 ( 8 ): 1146 - 52 e1 .
XU Y , WANG T , DENG X , et al . Analysis on the Leidenfrost effect of cryoprotectant microdroplets on the liquid nitrogen surface [J ] . SCIENTIA SINICA Technologica , 2017 , 47 ( 2 ): 190 - 6 .
SHARMA Y , SHARMA M . Biophysics of Cryopreservation [J ] . International Journal of Thermodynamics , 2022 : 1 - 11 .
VAJTA G , HOLM P , KUWAYAMA M , et al . Open pulled straw (OPS) vitrification: a new way to reduce cryoinjuries of bovine ova and embryos [J ] . Molecular Reproduction and Development: Incorporating Gamete Research , 1998 , 51 ( 1 ): 53 - 8 .
KUWAYAMA M . Highly efficient vitrification for cryopreservation of human oocytes and embryos: the Cryotop method [J ] . Theriogenology , 2007 , 67 ( 1 ): 73 - 80 .
LANE M , SCHOOLCRAFT W B , GARDNER D K , et al . Vitrification of mouse and human blastocysts using a novel cryoloop container-less technique [J ] . Fertility and sterility , 1999 , 72 ( 6 ): 1073 - 8 .
LIMA G L , LUZ V B , LUNARDI F O , et al . Effect of cryoprotectant type and concentration on the vitrification of collared peccary (Pecari tajacu) ovarian tissue [J ] . Anim Reprod Sci , 2019 , 205 : 126 - 33 .
ZHU W , PAN P , HUANG Y , et al . [Droplet freeze-thawing system based on solid surface vitrification and laser rewarming] [J ] . Sheng Wu Yi Xue Gong Cheng Xue Za Zhi , 2023 , 40 ( 5 ): 973 - 81 .
LANDA V , TEPLA O . Cryopreservation of mouse 8-cell embryos in microdrops [J ] . Folia biologica , 1990 , 36 ( 3-4 ): 153 - 8 .
DEMIRCI U , MONTESANO G . Cell encapsulating droplet vitrification [J ] . Lab Chip , 2007 , 7 ( 11 ): 1428 - 33 .
ZHANG X , KHIMJI I , SHAO L , et al . Nanoliter droplet vitrification for oocyte cryopreservation [J ] . Nanomedicine (Lond) , 2012 , 7 ( 4 ): 553 - 64 .
CUI M , ZHAN T , YANG J , et al . Droplet Generation, Vitrification, and Warming for Cell Cryopreservation: A Review [J ] . ACS Biomater Sci Eng , 2023 , 9 ( 3 ): 1151 - 63 .
XU Y , GUO N , YANG G , et al . Cryobiology for biobanking [J ] . SCIENTIA SINICA Vitae , 2023 .
韩恒鑫 , 胥义 , 刘宝林 . 复杂组织器官低温保存研究进展 [J ] . 真空科学与技术学报 , 2022 , 42 ( 01 ): 1 - 13 .
Han Hengxin , Xu Yi , Liu Baolin . Research Progress in Cryopreservation of Complex Tissues and Organs [J ] . Chinese Journal of Vacuum Science and Technology , 2022 , 42 ( 01 ): 1 - 13 .
SEKI S , MAZUR P . The dominance of warming rate over cooling rate in the survival of mouse oocytes subjected to a vitrification procedure [J ] . Cryobiology , 2009 , 59 ( 1 ): 75 - 82 .
HAN Z , BISCHOF J C . Critical cooling and warming rates as a function of CPA concentration [J ] . CryoLetters , 2020 , 41 ( 4 ): 185 - 93 .
KLEINHANS F W , SEKI S , MAZUR P. Simple , inexpensive attainment and measurement of very high cooling and warming rates [J ] . Cryobiology , 2010 , 61 ( 2 ): 231 - 3 .
GUO Z , ZUCHOWICZ N , BOUWMEESTER J , et al . Conduction-Dominated Cryomesh for Organism Vitrification [J ] . Adv Sci (Weinh) , 2024 , 11 ( 3 ): e2303317 .
ZHAN L , HAN Z , SHAO Q , et al . Rapid joule heating improves vitrification based cryopreservation [J ] . Nat Commun , 2022 , 13 ( 1 ): 6017 .
HAN H , ZHAN T , CUI M , et al . Investigation of Rapid Rewarming Chips for Cryopreservation by Joule Heating [J ] . Langmuir , 2023 , 39 ( 31 ): 11048 - 62 .
XUAN X . Joule heating in electrokinetic flow [J ] . Electrophoresis , 2008 , 29 ( 1 ): 33 - 43 .
HAN Z H , BISCHOF J C . Critical Cooling and Warming Rates as a Function of Cpa Concentration [J ] . Cryoletters , 2020 , 41 ( 4 ): 185 - 93 .
MAZUR P , SEKI S . Survival of mouse oocytes after being cooled in a vitrification solution to -196 degrees C at 95 degrees to 70,000 degrees C/min and warmed at 610 degrees to 118,000 degrees C/min: A new paradigm for cryopreservation by vitrification [J ] . Cryobiology , 2011 , 62 ( 1 ): 1 - 7 .
SEKI S , MAZUR P . Effect of warming rate on the survival of vitrified mouse oocytes and on the recrystallization of intracellular ice [J ] . Biol Reprod , 2008 , 79 ( 4 ): 727 - 37 .
ARAV A , NATAN Y , KALO D , et al . A new, simple, automatic vitrification device: preliminary results with murine and bovine oocytes and embryos [J ] . J Assist Reprod Genet , 2018 , 35 ( 7 ): 1161 - 8 .
SEKI S , KAWABE T , YAMAZAKI W , et al . Cryopreservation of rat embryos at all developmental stages by small-volume vitrification procedure and rapid warming in cryotubes [J ] . Sci Rep , 2023 , 13 ( 1 ): 20903 .
ZHU Y , ZHANG Q J , FENG H L , et al . Automation in vitrification and thawing of mouse oocytes and embryos [J ] . Front Cell Dev Biol , 2023 , 11 : 1330684 .
STEIF P S , PALASTRO M C , RABIN Y . The effect of temperature gradients on stress development during cryopreservation via vitrification [J ] . Cell Preserv Technol , 2007 , 5 ( 2 ): 104 - 15 .
WARKENTIN M , STANISLAVSKAIA V , HAMMES K , et al . Cryocrystallography in capillaries: critical glycerol concentrations and cooling rates [J ] . J Appl Crystallogr , 2008 , 41 ( Pt 4 ): 791 - 7 .
HOPKINS J B , BADEAU R , WARKENTIN M , et al . Effect of common cryoprotectants on critical warming rates and ice formation in aqueous solutions [J ] . Cryobiology , 2012 , 65 ( 3 ): 169 - 78 .
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