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Related Experiment Video

Updated: Apr 14, 2026

Tandem High-pressure Freezing and Quick Freeze Substitution of Plant Tissues for Transmission Electron Microscopy

Tandem High-pressure Freezing and Quick Freeze Substitution of Plant Tissues for Transmission Electron Microscopy

Published on: October 13, 2014

A proof-of-concept study on high-pressure freezing for cryopreservation.

Fang Song¹, Mayuko Sato², Yuya Toyama¹

¹Department of Chemical System Engineering, Graduate School of Engineering, The University of Tokyo, 113-8656 Tokyo, Japan.

|April 13, 2026

Summary

High-pressure freezing (HPF) reduces cytotoxic cryoprotective agent (CPA) concentrations needed for cell vitrification. This method improves cell viability and tissue structure preservation, advancing cryopreservation techniques.

Keywords:

cryopreservation cryoprotective agents fracture high-pressure freezing vitrification

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Last Updated: Apr 14, 2026

Tandem High-pressure Freezing and Quick Freeze Substitution of Plant Tissues for Transmission Electron Microscopy

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Published on: October 13, 2014

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Published on: October 25, 2019

Plunge Freezing: A Tool for the Ultrastructural and Immunolocalization Studies of Suspension Cells in Transmission Electron Microscopy

Plunge Freezing: A Tool for the Ultrastructural and Immunolocalization Studies of Suspension Cells in Transmission Electron Microscopy

Published on: May 5, 2017

Area of Science:

Biotechnology
Cryobiology
Cell Biology

Background:

Vitrification for cryopreservation typically requires high concentrations (30-50%) of cytotoxic cryoprotective agents (CPAs).
High CPA concentrations limit the broader application of vitrification due to toxicity and impact on cell viability.
Ice crystal formation during freezing and thawing is a major challenge in cryopreservation.

Purpose of the Study:

To investigate the potential of high-pressure freezing (HPF) to reduce CPA concentrations during vitrification.
To assess the impact of HPF on the viability and structural integrity of cryopreserved 2D cell monolayers and 3D cell spheroids.
To explore HPF as a novel approach for improved cryopreservation.

Main Methods:

Utilized a high-pressure freezing (HPF) device, typically used for cryofixation.
Cryopreserved 2D cell monolayers and 3D cell spheroids using reduced CPA concentrations (20-30 v/v%) with HPF.
Compared HPF with conventional plunge freezing for cell monolayers and spheroids.

Main Results:

HPF enabled successful cryopreservation with significantly lower CPA concentrations (20-30 v/v%) compared to standard methods.
HPF cell monolayers showed higher post-thaw viability and better substrate retention, supporting subsequent proliferation.
HPF cell spheroids demonstrated enhanced viability, metabolic activity, and preserved cell-cell adhesion.
HPF significantly improved cell viability and structural integrity compared to plunge freezing.

Conclusions:

High-pressure freezing (HPF) is a promising technique for reducing CPA concentrations in vitrification, thereby enhancing cell viability.
HPF offers a viable alternative to conventional cryopreservation methods, particularly for sensitive cell types and tissues.
Further development of HPF devices and integration with advanced warming techniques could lead to cryopreservation with minimal or no CPA.