Exosome Isolation via Two-Phase Fluid System
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Solution Overview
Problem
Existing exosome isolation methods, such as ultracentrifugation, filtration, and solubility-precipitation, are costly, time-consuming, inefficient, and result in low purity due to contamination, especially when dealing with small or large sample volumes, and are not suitable for isolating exosomes from diverse biological materials like plant lysates.
Innovation Solution
A two-phase fluid system using PEG and dextran is employed to separate exosomes from contaminants, involving centrifugation and washing steps to achieve high purity, with optional dextran removal via alcohol precipitation, allowing isolation from various biological materials including plant lysates.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Manufacturing precision
If ultracentrifuge based methods are used for exosome isolation, then exosomes can be isolated, but the method is expensive, laborious, time consuming, and results in low efficiency with low purity due to contamination
Solution Approach 1:
The patent changes the isolation parameter from centrifugal force (ultracentrifugation) to solubility difference (precipitation). By using polymeric precipitation, exosomes are isolated based on their solubility characteristics rather than particle size, enabling faster isolation without requiring high-speed centrifugation while achieving high purity
Solution Approach 2:
The patent replaces the mechanical ultracentrifugation system with a chemical precipitation system. Instead of using mechanical force to separate exosomes, the method uses chemical principles (solubility differences between exosomes and contaminants in polymeric solutions) to achieve separation, thereby reducing time and cost while improving purity
2Manufacturing precision
If ultracentrifuge based methods are used for exosome isolation, then exosomes can be isolated, but the method is expensive and requires expensive equipment
Solution Approach 1:
The patent replaces expensive mechanical ultracentrifuge equipment with simple chemical precipitation techniques using polymeric solutions. This substitution eliminates the need for expensive equipment while maintaining high isolation purity, making the method cost-effective for routine use
Solution Approach 2:
The patent uses inexpensive polymeric materials (such as polyethylene glycol or other water-soluble polymers) as the isolation medium. These cheap polymers enable high-purity exosome isolation without requiring expensive equipment, significantly reducing the overall cost of the isolation process
3Productivity
If filtration based methods are used for exosome isolation, then exosomes can be isolated faster and with less costly devices, but the forces applied cause exosome structure to deteriorate and disintegrate
Solution Approach 1:
The patent replaces mechanical filtration (which applies physical force) with chemical precipitation (which uses solubility differences). This substitution eliminates mechanical stress on exosomes during isolation, preventing structural deterioration and disintegration while maintaining high isolation speed
Solution Approach 2:
The patent changes the isolation mechanism from physical force (filtration pressure) to chemical property (solubility). By using polymeric precipitation, exosomes are separated based on their solubility characteristics rather than being forced through membranes, thereby preserving their structural integrity while maintaining fast isolation speed
4Ease of operation
If polymeric precipitation is used for exosome isolation, then the method is easier and faster, but there is an excess amount of contaminant besides the exosomes
Solution Approach 1:
The patent segments the isolation process into multiple steps: initial precipitation to concentrate exosomes, followed by selective removal of contaminants based on their different solubility characteristics. This segmentation allows the method to first achieve easy concentration and then systematically remove contaminants to achieve high purity
Solution Approach 2:
The patent uses changes in solution parameters (such as polymer concentration, temperature, or solvent composition) to differentially precipitate exosomes from contaminants. By adjusting these parameters, the method achieves both ease of operation and high purity, as different components separate based on their specific solubility properties under controlled conditions
5Productivity
If ultracentrifuge based methods are used for exosome isolation, then exosomes can be isolated, but the method does not have any selectivity and isolates many undesired substances
Solution Approach 1:
The patent changes the isolation parameter from particle size (centrifugation) to solubility (precipitation). This parameter change provides inherent selectivity, as exosomes and contaminants have different solubility characteristics in polymeric solutions, enabling the method to efficiently isolate exosomes while excluding undesired substances based on their solubility differences
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-purity exosome isolation from small and large samples with ease and cost-effectiveness, removing 98% of protein contaminants and maintaining exosome integrity, suitable for diverse biological sources.
Implementation Method 1
Solubility-Precipitation based methods: Exosomes can be precipitated in solutions where their solubility is low without any need for ultracentrifugation. Polymers such as polyethylene glycol (PEG) used in these methods retains the water molecules and enable precipitation of the molecules with lower-solubility including the exosomes.
Implementation Method 2
Jongmin Kim et al. in their discloses a method to isolate extracellular vesicles (EVs) by using a polyethylene glycol/dextran aqueous two-phase system (ATPS).
Implementation Method 3
separating the mixture formed for isolation into two phases by means of the two phase fluid extraction system
Implementation Method 4
dextran removal method comprising the following sub-steps: a. adding purification solution containing monohydric alcohol or acetone in order to remove dextran from the collected lower phase
Data Source
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Figure 1(c)~2(a)
Figure 2(b)~2(c)
AI summary
The present invention is an exosome isolation method which enables to obtain high purity exosomes inexpensively from samples of large and small amounts and types of samples from which exosomes could not be obtained before.