Cryopreservation Medium Composition for High Cell Survivability
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Solution Overview
Problem
Cryopreservation of living cells is challenging due to harsh freezing and thawing conditions, leading to low survivability rates and cell damage, particularly from osmotic dehydration and intracellular freezing.
Innovation Solution
A cryopreservation medium comprising dimethyl sulfoxide (DMSO), disaccharides, human serum, and cytokines like IL-7 or IL-15, with specific concentration ranges, is used to preserve mammalian cells, followed by controlled cooling and thawing.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Duration of action of stationary object
If conventional freezing methods are used to preserve cells, then cells can be stored for extended periods, but cell survivability is low due to osmotic dehydration and intracellular freezing damage
Solution Approach 1:
The patent modifies the chemical composition parameters of the freezing medium by incorporating specific concentrations of DMSO (1-10%), disaccharides (0.25-5%), human serum (10-90%), and human serum albumin (0.5-30%), along with cytokines IL-7 and IL-15. These parameter changes optimize the medium's ability to protect cells during freezing and thawing, resolving the contradiction between extended storage duration and cell survivability.
Solution Approach 2:
The invention creates a composite cryopreservation medium that combines multiple protective agents: DMSO as a cryoprotectant, disaccharides for osmotic balance, human serum and albumin for membrane protection, and cytokines for cellular recovery. This composite formulation synergistically addresses both the need for long-term storage stability and high cell survivability after thawing.
2Duration of action of stationary object
If freezing is used to preserve cells, then cells can be stored long-term, but cell membrane damage occurs due to osmotic dehydration
Solution Approach 1:
The patent introduces human serum and human serum albumin as intermediary substances that mediate between the freezing environment and cell membranes. These intermediaries form protective layers around cells, reducing direct exposure to harmful freezing conditions and preventing osmotic dehydration-induced membrane damage while enabling long-term storage.
Solution Approach 2:
The cryopreservation medium provides beforehand cushioning by incorporating DMSO and disaccharides that preemptively protect cells from freezing damage. These substances accumulate around cells before freezing occurs, cushioning against osmotic stress and intracellular ice formation that would otherwise cause membrane damage during storage.
3Temperature
If intracellular freezing occurs, then cells can be preserved at low temperatures, but cell injuries result from ice crystal formation
Solution Approach 1:
The patent converts the harmful effect of freezing into a beneficial preservation method by using DMSO and disaccharides to control ice crystal formation. These substances modify the freezing process so that extracellular ice forms first, creating a protective environment that prevents intracellular ice formation. The cytokines IL-7 and IL-15 further convert the stress of freezing into a controlled differentiation signal, transforming potential cell injury into beneficial cellular adaptation.
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 method achieves high survivability of thawed cells, suitable for applications such as adoptive cell transfer, with enhanced cell survival both in vitro and post-transplantation.
Implementation Method 1
dimethyl sulfoxide (DMSO)...allows for freezing and subsequent thawing of viable cells
Implementation Method 2
disaccharide...cells attempt to maintain osmotic equilibrium across the membrane
Implementation Method 3
human serum...human serum albumin...damage to cells plasma membrane which results from osmotic dehydration
Implementation Method 4
IL-7 or IL-15...thawed cells using the media and methods described herein retain high survivability
Data Source
AI summary
The present disclosure provides, among other things, a cryopreservation medium for cryopreserving mammalian cells, the medium comprising: dimethyl sulfoxide (DMSO), disaccharide, human serum, and IL-7 and/or IL-15. The present disclosure also provides, among other things, a cryopreservation medium for cryopreserving mammalian cells, the medium comprising: between about 1 w/v % and 10 w/v % dimethyl sulfoxide (DMSO), between about 0.25 w/v % and 5 w/v % disaccharide, and between about 10 w/v % and 90 w/v % human serum. The present disclosure also provides, among other things, a cryopreservation medium for cryopreserving mammalian cells, the medium comprising: between about 1 w/v % and 10 w/v % dimethyl sulfoxide (DMSO), between about 0.25 w/v % and 5 w/v % disaccharide, and between about 0.5 w/v % and 30 w/v % human serum albumin.