Two-Step Magnetic Cell Sorting with Dual Iron Content Particles
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Solution Overview
Problem
Current cell separation methods, such as magnetic cell sorting and gradient density centrifugation, are time-consuming and laborious, especially when isolating target cells from whole blood, and often result in low purity due to the need for multiple steps and intermediate processing, which complicates the efficient removal of non-target cells.
Innovation Solution
A method involving the use of magnetic particles with different iron contents and antigen recognizing moieties for two-step magnetic cell sorting, where the first step aggregates non-target cells and the second step enriches target cells using a magnetic field gradient, allowing for a one-step incubation and subsequent separation without intermediate processing.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Manufacturing precision
If multiple process steps including centrifugation are used to remove non-target cells and purify target cells, then the purity of target cells is improved, but the processing time and operational complexity increase
Solution Approach 1:
The patent combines erythrocyte aggregation and magnetic cell sorting into a single integrated process. The aggregation reagent and magnetic particles are mixed together to form a dual-functional reagent that simultaneously aggregates non-target cells and enables magnetic separation, eliminating the need for separate centrifugation steps while maintaining high purity.
Solution Approach 2:
The magnetic particles serve multiple functions: they enable magnetic separation of target cells, and when combined with the aggregation reagent, they also facilitate removal of aggregated non-target cells. This multi-functional approach reduces the number of process steps while achieving both purification and concentration.
2Manufacturing precision
If multiple process steps including centrifugation are used to remove non-target cells and purify target cells, then the purity of target cells is improved, but the operational complexity increases
Solution Approach 1:
The patent merges the aggregation reagent and magnetic particles into a single combined reagent system. This integration eliminates the need for separate aggregation and magnetic sorting steps, reducing operational complexity from multiple manual interventions to a single incubation followed by magnetic separation.
Solution Approach 2:
The aggregation reagent is designed to pre-aggregate non-target cells during the incubation period before magnetic separation is applied. This preliminary aggregation action simplifies the subsequent magnetic sorting step by pre-concentrating non-target cells into easily removable aggregates, reducing the complexity of the separation process.
3Reliability
If only negative selection is performed to deplete non-target cells, then the cells remain untouched and viable, but the purity of specific target cell subpopulations is low
Solution Approach 1:
The patent segments the cell population into three distinct groups using different mechanisms: aggregated non-target cells removed by centrifugation, target cells enriched by magnetic sorting, and untouched cells remaining in the supernatant. This segmentation allows simultaneous achievement of high purity for specific subpopulations while maintaining overall cell viability.
Solution Approach 2:
Different separation mechanisms are applied to different cell populations within the same sample: aggregation for non-target cells, magnetic sorting for target cells, and no treatment for untouched cells. This localized approach to different cell subsets enables high purity isolation of specific target populations while preserving the viability of all cells through gentle, non-destructive methods.
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 approach significantly reduces processing time and increases purity by effectively removing non-target cells and enriching target cells, such as regulatory T-cells and hematopoietic stem cells, from whole blood with high efficiency, achieving purities of up to 91% and 67% respectively within a short timeframe.
Implementation Method 1
The magnetic force Fm acting on a magnetic particle in a magnetic field gradient can be described as follows: Fm=Vp*(xm-xm,medium)*grad(H2)
Implementation Method 2
contacting the sample with a cell aggregation agent and first magnetic particles
Data Source
Figure 1(a)~1(d)
Figure 2
Figure 3
AI summary
The invention is directed to a method for enriching target cells from a sample of cells characterized by: a) contacting the sample with a cell aggregation agent and first magnetic particles having an iron content of 0.1 pg to 5000 pg, coupled to a first antigen recognizing moiety; and second magnetic particles having an iron content of 0.05 fg to 100 fg and coupled to a second antigen recognizing moiety to obtain mixture a) b) applying a first magnetic field gradient to the mixture a) thereby removing the cells bound to the first antigen recognizing moiety coupled to the first magnetic particles, to obtain a mixture b) and obtaining an agglomerate comprising the cells of mixture a) bound to the cell aggregation agent c) applying a second magnetic field gradient to the mixture b) thereby immobilizing the cells bound to the second antigen recognizing moiety coupled to the second magnetic particles d) recovering the immobilized cells from the second magnetic field gradient as target cells.