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Conventional electron microscopy (EM) involves dehydration, fixation, and staining of biological samples, which distorts the native state of biological molecules and results in several artifacts. Also, the high-energy electron beam damages the sample and makes it difficult to obtain high-resolution images. These issues can be addressed using cryo-EM, which uses frozen samples and gentler electron beams. The technique was developed by Jacques Dubochet, Joachim Frank, and Richard Henderson, for...
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Optimization of Deep Eutectic Solvents Enables Green and Efficient Cryopreservation.

Wei He1,2,3, Taijie Zhan1,2,3, Hengxin Han1,2,3

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Summary

Deep eutectic solvents (DES) offer a low-toxicity alternative to dimethyl sulfoxide (DMSO) for cryopreservation. These biocompatible solvents effectively preserve human umbilical cord mesenchymal stem cells (HuMSC) with high viability, advancing cell therapy applications.

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Area of Science:

  • Biomedical Engineering
  • Cell Biology
  • Materials Science

Background:

  • Cryopreservation is vital for cell therapy, tissue engineering, and reproduction.
  • Dimethyl sulfoxide (DMSO) is a common cryoprotectant but has toxicity concerns.
  • Development of low-toxicity cryoprotectants is crucial for clinical applications.

Purpose of the Study:

  • To investigate deep eutectic solvents (DES) as biocompatible cryoprotectants.
  • To evaluate DES for efficient cryopreservation of human umbilical cord mesenchymal stem cells (HuMSC).
  • To compare DES performance against DMSO for cell viability and function.

Main Methods:

  • Explored six types of DES for thermal properties, toxicity, and cell permeability.
  • Utilized Raman spectroscopy and viscosity studies to analyze DES properties.
  • Optimized freezing protocols (cooling rate, incubation, loading) for DES cryopreservation.

Main Results:

  • DES demonstrated improved hydrogen-bonding at decreased temperatures.
  • Optimized DES protocols significantly improved mouse embryonic fibroblast (NIH-3T3) viability post-thaw.
  • HuMSC cryopreserved with DES showed high viability (94.65%), comparable to DMSO, retaining differentiation and enhancing proliferation.

Conclusions:

  • Deep eutectic solvents are effective, low-toxicity cryoprotective agents for stem cell preservation.
  • DES inhibit ice crystal growth by binding water, reducing mechanical cell damage.
  • DES represent a promising alternative to DMSO for clinical cell therapy applications.