Cell Purification via Size-Exclusion Filtration and Crosslinking
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Solution Overview
Problem
Current methods for isolating cells in low concentrations, such as circulating tumor cells and fetal cells, face challenges in achieving high yield and purity, often altering the cells' biological properties and being unsuitable for large blood volumes or cells with unknown surface markers.
Innovation Solution
A filtration process using a high-precision membrane with specific pore sizes and coatings, combined with a crosslinking step to bind unwanted cells to the membrane, allowing for the isolation of cells based on physical properties while preserving their biological integrity and surface markers.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If magnetic bead methods are used to isolate target cells, then cell isolation can be achieved, but the biological properties and viability of the target cells are strongly altered
Solution Approach 1:
The patent replaces magnetic bead-based mechanical separation with a filtration system that uses physical pore size exclusion. Cells are separated based on their ability to pass through membranes with specific pore sizes, eliminating the need for magnetic beads that alter cell biology. The filtration process uses pressure-driven flow through layered membranes to achieve separation without cellular modification.
Solution Approach 2:
The patent employs layered membrane filtration systems with precisely controlled pore sizes (e.g., 3-10 µm for first membrane, 1-3 µm for second membrane) to separate cells based on physical dimensions. The porous structure allows smaller cells to pass through while retaining larger cells, achieving isolation without biochemical modification of the target cells.
2Reliability
If affinity membranes with immobilized antibodies are used, then target cells can be bound and isolated, but the cells lose viability or biological functionality
Solution Approach 1:
The patent replaces antibody-based affinity binding with purely physical filtration based on pore size exclusion. This mechanical separation method avoids the biochemical interactions that cause cell damage while maintaining isolation effectiveness through size-based discrimination of cells.
Solution Approach 2:
The patent uses membranes with defined pore sizes to physically exclude cells based on their dimensions. The porous structure provides size-selective separation without requiring cell surface marker recognition, thereby preserving cell viability and biological functionality while achieving specific isolation.
3Productivity
If physical separation approaches are used, then high throughput and biological integrity are maintained, but specificity is reduced resulting in low purity or low yield
Solution Approach 1:
The patent divides the separation process into multiple sequential filtration stages, each with membranes of different pore sizes. The first membrane (3-10 µm) performs initial separation, followed by a second membrane (1-3 µm) for refined isolation. This segmented approach increases both specificity and purity while maintaining high throughput capability.
Solution Approach 2:
The patent employs a hierarchy of porous membranes with progressively smaller pore sizes to achieve multi-level separation. The layered porous structure enables progressive filtering that enhances specificity at each stage, ultimately achieving high purity isolation without sacrificing throughput or biological integrity.
4Reliability
If filtration with small pore sizes is used to achieve high purity, then yield is reduced
Solution Approach 1:
The patent segments the filtration process into two stages: first membrane (3-10 µm) that captures most target cells with high yield, and second membrane (1-3 µm) that refines purity. This segmentation ensures that the majority of cells are recovered in the first stage while the second stage removes contaminants, achieving both high yield and high purity.
Solution Approach 2:
The patent performs preliminary separation with the first larger-pore membrane before applying the second smaller-pore membrane. This preliminary action captures the bulk of target cells that would be lost if only small-pore filtration were used, while subsequent refinement achieves high purity without sacrificing the yield already obtained.
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
This method enables the isolation of cells in high yield and purity, even at low concentrations, without modifying the cells' surface markers or biological functions, making it suitable for diagnostic purposes and large sample volumes.
Implementation Method 1
a) filtering the mixture M of cells over a membrane 10 having pores 11
Implementation Method 2
c) adding a crosslinking element 30 to the mixture M2 retained on the membrane 10, the crosslinking element 30 binding the polluting cells 20 to the membrane 10
Data Source
Figure 1~2b
Figure 3~4
Figure 5a~5c
AI summary
A process for isolating cells of interest from a sample comprising polluting cells. The process comprises a filtration step through a membrane provided with pores and with linkers, and a crosslinking step allowing to bind the polluting cells to the membrane, leaving the cells of interest unbound.