Trehalose-HEPES Cryopreservation Agent for Mitochondrial Integrity
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Solution Overview
Problem
Conventional cryoprotectants for cells are not suitable for cryopreservation of mitochondria, leading to swelling or breakage, which compromises their functionality post-thawing.
Innovation Solution
A cryopreservation agent comprising trehalose, HEPES, and serum albumin, without potassium chloride, ethylene glycol, ethylene glycol tetraacetic acid, or ethylenediaminetetraacetic acid, is used to protect mitochondrial integrity and maintain pH and osmotic pressure during cryopreservation.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If conventional cryoprotectants for cells are used for mitochondria cryopreservation, then cell preservation is achieved, but mitochondria become swollen or broken and lose functionality
Solution Approach 1:
The patent modifies the chemical composition parameters of the cryoprotectant by replacing conventional cell-based agents with a mitochondrial-specific formulation containing trehalose (10-30%), HEPES (5-20%), and serum albumin (0.1-5%), while excluding salts like KCl and NaCl that are harmful to mitochondria. This parameter change resolves the contradiction by creating a cryoprotectant that maintains mitochondrial integrity while preserving functionality.
Solution Approach 2:
The patent introduces trehalose as an intermediary substance that mediates protection between the cryopreservation process and mitochondrial structures. Trehalose forms a glassy matrix that physically supports mitochondria during freezing, preventing ice crystal formation and mechanical damage, thereby maintaining both integrity and functionality.
2Object-affected harmful factors
If conventional cryoprotectants are used, then freezing protection is provided, but mitochondrial swelling and breakage occur
Solution Approach 1:
The patent converts the potentially harmful freezing process into a beneficial protective mechanism by using trehalose to form a vitrified glassy state that prevents ice crystal formation. This glassy matrix transforms the damaging effects of freezing into a protective environment that maintains mitochondrial morphology while providing freezing protection.
Solution Approach 2:
The patent creates a composite cryoprotectant system combining trehalose (disaccharide), HEPES (buffer), and serum albumin (protein) in specific proportions. This composite formulation works synergistically: trehalose provides structural support and vitrification, HEPES maintains pH stability, and serum albumin offers additional protective binding, collectively preventing mitochondrial swelling and breakage.
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
The agent preserves mitochondrial integrity and ATP synthesis ability, ensuring better functionality of cryopreserved mitochondria post-thawing, suitable for mitochondrial transplantation.
Implementation Method 1
an agent for cryopreservation includes trehalose, HEPES (4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid) and serum albumin (SA)... preserves mitochondrial integrity... after thawing
Implementation Method 2
HEPES (4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid)... maintain pH... during cryopreservation
Implementation Method 3
cooling down the agent for cryopreservation and the mitochondria in the container... preserving the agent for cryopreservation and the mitochondria in the container which have been cooled down
Data Source
AI summary
An agent for cryopreservation includes trehalose, HEPES and serum albumin. The agent for cryopreservation does not include potassium chloride, sodium chloride, ethylene glycol, ethylene glycol tetraacetic acid and ethylenediaminetetraacetic acid.