Polyampholyte Cryopreservation Stabilizer for Cell Viability
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Solution Overview
Problem
Current cryopreservation solutions for animal cells, particularly cell sheets and three-dimensional structures, face challenges with damage due to ice crystal formation and dewatering shrinkage during freezing, leading to cytotoxicity and recrystallization issues upon thawing.
Innovation Solution
A novel animal cell cryopreservation solution utilizing a polyampholyte with amino and carboxyl groups, combined with an epichlorohydrin-crosslinked sucrose polymer macromolecule, which stabilizes the vitreous state and prevents crystallization during freezing and recrystallization during thawing.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If conventional cryoprotective agents (DMSO, glycerin) are used for cell sheets and three-dimensional structures, then cell dispersion cryopreservation is achieved, but damage occurs after thawing due to ice crystal formation and dewatering shrinkage
Solution Approach 1:
The patent changes the chemical composition parameters by introducing polyampholytes (with specific amino acid sequences and charges) and sucrose polymer macromolecules into the cryopreservation solution. These parameter changes enable the solution to achieve vitrification at higher temperatures and maintain the vitreous state more effectively, preventing ice crystal formation and dewatering shrinkage damage while improving cell viability after thawing.
Solution Approach 2:
The patent uses composite cryoprotective agents combining polyampholytes (which provide vitrification capability) with sucrose polymer macromolecules (which enhance structural stability). This composite approach creates a synergistic effect that improves vitrification ability and maintains the vitreous state more effectively than single agents, thereby reducing ice crystal formation and dewatering shrinkage damage.
2Stability of the object's composition
If high concentration vitrifying solutions (DAP 213 with 2 M DMSO, 1 M acetamide, 3 M propylene glycol) are used, then vitreous state is achieved, but cytotoxicity increases and damage occurs due to recrystallization during thawing
Solution Approach 1:
The patent changes the chemical composition by using polyampholytes with specific amino acid sequences and charges, combined with sucrose polymer macromolecules. This allows achieving vitrification at lower concentrations than conventional high-concentration solutions, thereby reducing cytotoxicity while maintaining vitreous state stability and preventing recrystallization damage during thawing.
Solution Approach 2:
The patent employs biocompatible polyampholytes and sucrose polymer macromolecules that are less toxic than conventional cryoprotective agents. These agents provide effective vitrification protection without the high cytotoxicity associated with high-concentration DMSO and propylene glycol solutions, enabling safer cryopreservation of cell sheets and three-dimensional structures.
3Stability of the object's composition
If rapid freezing rate and high solute concentration are used for vitrification, then aqueous solution vitrification is achieved, but the method becomes complex and toxic
Solution Approach 1:
The patent modifies the chemical parameters of the cryopreservation solution by incorporating polyampholytes with specific amino acid sequences and charges, along with sucrose polymer macromolecules. This enables the solution to achieve vitrification at more manageable freezing rates and lower solute concentrations, simplifying the freezing process and reducing toxicity while maintaining effective vitreous state formation.
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 solution enhances vitrification ability, allowing for the stable maintenance of the vitreous state at higher temperatures, thereby improving the viability of animal cells during freezing and thawing processes.
Implementation Method 1
a vitrifying method has been attempted, which controls ice crystallization and allows solidification in an amorphous state
Implementation Method 2
Based on consideration that the damage might be caused by ice crystal formation or dewatering shrinkage during freezing
Data Source
AI summary
Provided are a vitrification stabilizer for an animal cell cryopreservation fluid, and an animal cell cryopreservation fluid which exhibits superior vitrification capabilities due to the animal cell cryopreservation fluid containing the vitrification stabilizer for an animal cell cryopreservation fluid. The vitrification stabilizer for an animal cell cryopreservation fluid contains: an amphoteric polymer compound selected from the group consisting of (a) a carboxylated amphoteric polymer compound obtained by reacting ε-poly-L-lysine with butyl succinic anhydride, (b) a carboxylated amphoteric polymer compound obtained by reacting ε-poly-L-lysine with butyl succinic anhydride and succinic anhydride, or (c) a carboxylated amphoteric polymer compound obtained by reacting ε-poly-L-lysine with a compound represented by formula I; and (d) a sucrose polymer macromolecule to which epichlorohydrin has been crosslinked.