An apparatus for magnetic sorting of cells

Abstract

This application provides a device for magnetic cell sorting, belonging to the field of magnetic cell sorting technology. The device includes an outer shell, inside which are a liquid inlet, a receiving cavity, and a liquid outlet connected sequentially from top to bottom along the fluid flow direction. An upper filter layer, a magnetic field amplification layer, and a lower filter layer are sequentially arranged from top to bottom within the receiving cavity. The magnetic field amplification layer consists of several magnetic matrix particles and flow channels arranged in a predetermined pattern. The magnetic matrix particles are composed of balls and a layer of thermoplastic polymer material disposed on the outer surface of the balls. Through a special structural design, especially the synergistic design of the internal structure of the magnetic field amplification layer, this device can significantly improve cell sorting efficiency and cell viability protection.

Description

Technical Field

[0001] This application relates to the field of cell magnetic sorting technology, specifically to a device for cell magnetic sorting. Background Technology

[0002] In the field of cell immunotherapy, T cells are the core effector cells of CAR-T therapy, and their efficient separation technology is crucial. Among the current mainstream T cell sorting technologies, density gradient centrifugation suffers from low sorting accuracy and impaired cell viability, while flow cytometry is difficult to meet the needs of large-scale clinical use due to its expensive equipment and high technical requirements. Immunomagnetic sorting, on the other hand, has become the preferred technology for separating target T cell subsets in CAR-T therapy due to its advantages such as ease of operation and good cell viability. The main principle of immunomagnetic sorting is that cells are attached to superparamagnetic magnetic particles through bioactive molecules. Under the influence of a magnetic field, cells labeled by the magnetic particles are retained in the magnetic field, while unlabeled cells are collected by flowing out or being drawn out. When the magnetic field is removed, the labeled cells can be eluted and collected, thereby separating target cells from non-target cells. Based on the size of the magnetic particles used, magnetic sorting can be divided into two systems: micrometer-scale (1-5 μm) and nanometer-scale (50-200 nm). While micrometer-scale magnetic sorting has high magnetic field responsiveness and sorting efficiency, its larger size may lead to problems such as mechanical damage to cells, increased non-specific adsorption, and decreased cell activity, especially when sorting rare cells. On the other hand, nanometer-scale magnetic particles have a higher specific surface area and better biocompatibility, which can significantly reduce cell damage and improve sorting specificity. In addition, due to their smaller size, they are easier to penetrate biological membranes and be effectively processed by cells and organisms, which can simplify the sorting process and reduce cell damage.

[0003] Although nanoscale magnetic sorting has significant advantages, it still faces many technical bottlenecks in practical applications. The key device for sorting, the sorting column, suffers from problems such as uneven magnetic field distribution leading to unstable sorting efficiency, magnetic matrix particles agglomerating and falling off, uneven pore size affecting cell sorting purity, and unreasonable flow channel design causing shear force damage to fragile cells and easy blockage during the sorting process. Therefore, there is an urgent need to provide a new type of cell magnetic sorting device to solve the above problems. Utility Model Content

[0004] The purpose of this application is to provide an apparatus for magnetic sorting of cells.

[0005] The embodiments of this application can be implemented through the following technical solutions:

[0006] A device for magnetic cell sorting includes an outer shell. Inside the outer shell, there are an inlet, a receiving cavity, and an outlet that are sequentially connected from top to bottom along the fluid flow direction. An upper filter layer, a magnetic field amplification layer, and a lower filter layer are sequentially arranged from top to bottom within the receiving cavity. The magnetic field amplification layer consists of a plurality of magnetic matrix particles and channels arranged in a predetermined pattern. The magnetic matrix particles consist of balls and a layer of thermoplastic polymer material disposed on the outer surface of the balls.

[0007] Furthermore, the diameter of the ball is 0.2 to 1.0 mm, and / or the ball may be made from iron, cobalt, nickel and their alloys.

[0008] Furthermore, the thickness L of the thermoplastic polymer material layer is 5–30 μm.

[0009] Furthermore, the arrangement of the magnetic matrix particles is selected from close-packed arrangement or simple cubic arrangement.

[0010] Furthermore, the diameter d2 of the magnetic matrix particles is 0.3 to 0.7 mm.

[0011] Furthermore, the filling rate of the magnetic matrix particles is 40% to 60%.

[0012] Furthermore, the gap width D of the flow channel is 0.15 to 0.75 mm.

[0013] Furthermore, the upper and lower filter layers are filter structures with a pore channel range of 0.25 to 0.5 mm.

[0014] Furthermore, the receiving cavity is cylindrical, and the diameters of the liquid inlet and the liquid outlet are smaller than the receiving cavity; and / or, the outer shell includes an upper cover and a bottom shell connected to the bottom end of the upper cover, the liquid inlet is located at the top of the upper cover, and the receiving cavity and the liquid outlet are located inside the bottom shell.

[0015] Furthermore, the upper cover and the bottom shell are connected by ultrasonic welding.

[0016] The device for magnetic cell sorting provided in the embodiments of this application has at least the following beneficial effects:

[0017] The device for magnetic cell sorting provided in this application significantly improves cell sorting efficiency and cell viability protection through a special structural design, especially the synergistic design of the internal structure of the magnetic field amplification layer. A uniform magnetic field is formed through a specific arrangement of magnetic matrix particles. The design of the polymer material layer can effectively prevent metal corrosion and mechanical damage to cells, while reducing non-specific adsorption. In addition, the optimized design of the diameter, filling rate and flow channel gap width of the magnetic matrix particles ensures sufficient magnetic field strength while allowing cells to pass through in a mild fluid environment, reducing the damage to cells caused by fluid shear force. Attached Figure Description

[0018] Figure 1 A schematic diagram of a device for magnetic cell sorting provided in an embodiment of this application;

[0019] Figure 2 This is a schematic diagram of the internal filling matrix layer arrangement according to one embodiment of this application;

[0020] Figure 3 This is a schematic diagram of the internal filling matrix layer arrangement in another embodiment of this application;

[0021] Figure 4 for Figure 2 A magnified view of a portion of the image;

[0022] Figure 5 A schematic diagram illustrating the application principle of the device for magnetic cell sorting provided in the embodiments of this application;

[0023] Numbers in the diagram

[0024] 1-Upper cap, 2-Bottom shell, 3-Upper filter layer, 4-Magnetic field amplification layer, 41-Magnetic matrix particles, 411-Ball bearings, 412-Thermoplastic polymer material layer; 42-Flow channel, 5-Lower filter layer, 6-Liquid inlet, 7-Liquid outlet; Detailed Implementation

[0025] The present application will now be further described based on preferred embodiments and with reference to the accompanying drawings.

[0026] Furthermore, for ease of understanding, various components on the drawings have been enlarged or reduced, but this is not intended to limit the scope of protection of this application.

[0027] Singular forms of words also include plural meanings, and vice versa.

[0028] In the description of the embodiments of this application, it should be noted that if terms such as "upper," "lower," "inner," or "outer" are used to indicate the orientation or positional relationship based on the orientation or positional relationship shown in the accompanying drawings, or the orientation or positional relationship commonly used when the product of this application is in use, they are only for the convenience of describing this application and simplifying the description, and do not indicate or imply that the device or element referred to must have a specific orientation, or be constructed and operated in a specific orientation, and therefore should not be construed as a limitation on this application. In addition, in the description of this application, in order to distinguish different units, the terms "first," "second," etc. are used in this specification, but these are not limited by the manufacturing order, nor should they be construed as indicating or implying relative importance. Their names may differ in the detailed description and claims of this application.

[0029] The vocabulary used in this specification is for illustrative purposes and is not intended to limit the scope of this application. It should also be noted that, unless otherwise expressly specified and limited, the terms "set," "connected," and "linked" should be interpreted broadly. For example, they can refer to a fixed connection, a detachable connection, or an integral connection; they can refer to a mechanical connection, a direct connection, or an indirect connection via an intermediate medium; or they can refer to the internal communication between two components. Those skilled in the art will understand the specific meaning of these terms in this application.

[0030] This application provides an apparatus for magnetic cell sorting, combined with Figures 1 to 4 As shown, the device includes an outer shell, inside which are a liquid inlet 6, a receiving cavity, and a liquid outlet 7 connected sequentially from top to bottom along the fluid flow direction. The receiving cavity contains an upper filter layer 3, a magnetic field amplification layer 4, and a lower filter layer 5 sequentially arranged from top to bottom. In use, an external magnetic field is applied to the area corresponding to the magnetic field amplification layer 4. Under the action of the external magnetic field, the magnetic field amplification layer 4 generates an induced magnetic field, which is superimposed on the external magnetic field, thereby forming an enhanced composite magnetic field in the receiving cavity. This composite magnetic field can more efficiently apply magnetic force to target cells labeled with magnetic particles, achieving precise sorting.

[0031] Furthermore, the magnetic field amplification layer 4 includes a plurality of magnetic matrix particles 41 arranged in a predetermined manner and flow channels 42 formed between the magnetic matrix particles 41. The magnetic matrix particles 41 are composed of balls 411 and a thermoplastic polymer material layer 412 disposed on the outer surface of the balls 411.

[0032] Specifically, the ball 411 can be selected from iron, cobalt, nickel and their alloys. These materials have good magnetic properties and can be rapidly magnetized under the action of an external magnetic field. Preferably, the ball 411 is Q195 carbon steel.

[0033] Preferably, the diameter d1 of the ball 411 is 0.2 to 1.0 mm. This size range can ensure that a reasonable flow channel gap width is maintained while ensuring sufficient magnetic strength.

[0034] Specifically, the thermoplastic polymer material can be selected from polyethylene, polypropylene, polyvinyl chloride, etc. The presence of this thermoplastic polymer material layer creates a stable spacing structure between the balls. This spacing not only effectively prevents direct contact between the balls, reducing magnetic short circuits, but also significantly enhances the internal magnetic field strength, further improving cell sorting efficiency. On the other hand, it also prevents moisture and oxygen from corroding the inner ball layer and prevents direct contact between the inner ball layer and cells, thus preventing cell damage and affecting the sorting effect. In some specific embodiments, the thermoplastic polymer material can be coated onto the surface of the balls using an electrodeposition process. This electrodeposition process allows the thermoplastic polymer material layer to adhere uniformly and tightly to the ball surface, with controllable thickness.

[0035] Preferably, the thickness L of the thermoplastic polymer material layer is 5–30 μm. This range can avoid the situation where the ball surface is not completely covered due to excessive thickness, which may lead to oxidation and corrosion or residual exposed metal sites that cause cell adhesion and increase non-specific adsorption, resulting in an increase in the proportion of contaminated cells in the positive component. On the other hand, it can also avoid the situation where the magnetic matrix particle size is significantly increased due to excessive thickness, which may reduce the width of the flow channel gap, increase the resistance to cell passage, and thus affect the cell recovery rate and increase the shear force, thereby affecting cell activity.

[0036] Furthermore, the magnetic matrix particles 41 are spherical; the arrangement of the magnetic matrix particles 41 is selected from close-packed arrangement or simple cubic arrangement, wherein close-packed arrangement specifically refers to each sphere having 12 nearest neighbor spheres around it, including face-centered cubic close packing (the third layer is aligned with the first layer, forming an ABABAB... periodic arrangement) and hexagonal close packing (the third layer is located above the voids of the first layer, forming an ABCABCABC... arrangement), such as Figure 3 The diagram shown is a schematic representation of the magnetic matrix particle arrangement in one layer of a planar surface in one embodiment of this application. The simple cubic stacking arrangement specifically refers to each sphere having six nearest neighbor spheres (one each in the top, bottom, front, back, left, and right directions), meaning each sphere is located at a vertex of the cube and contacts the midpoint of one of the cube's edges. Figure 4 The diagram shown is a schematic diagram of the magnetic matrix particle arrangement in one of the planar layers of a simple cubic stacked arrangement in another embodiment of this application.

[0037] Furthermore, Figure 5A partially enlarged schematic diagram of one arrangement shows a flow channel 42 formed by adjacent magnetic matrix particles 41. The flow channel 42 is an irregular continuous porous structure, the morphology of which is directly related to the arrangement density and arrangement of the magnetic matrix particles 41. By controlling the arrangement of the magnetic matrix particles 41 and thus the gap width D of the flow channel 42, the flow rate of the cell suspension, the magnetic field gradient distribution, and the probability of cell contact with the magnetic matrix particles can be significantly affected, thereby affecting the sorting effect. Here, the gap width D of the flow channel 42 specifically refers to the length of the distance between the centers of two adjacent magnetic matrix particles minus the diameter of the magnetic matrix particles. When the gap width D of the flow channel is too small, it will lead to an increased flow rate, increasing the shear force of the liquid on the cells and thus damaging the cells. When the gap width D of the flow channel is too large, it will reduce the magnetic field gradient and thus reduce the magnetic sorting efficiency.

[0038] Preferably, the diameter d2 of the magnetic matrix particles 41 is 0.3–0.7 mm;

[0039] Preferably, the filling rate of the magnetic matrix particles 41 (i.e., the proportion of magnetic matrix particles 41 per unit volume) is 40% to 60%.

[0040] Preferably, the gap width D of the flow channel 42 is 0.15 to 0.75 mm. This range can ensure that cells can pass through in a mild fluid environment while maintaining sufficient magnetic field strength to achieve effective sorting.

[0041] In some specific embodiments, a polymer material can be used to bond the magnetic matrix particles to form an integral magnetic field amplification layer 4, and then bond and fix it to the inner wall of the outer shell. The polymer material can be selected from one or more of polyurethane, epoxy resin or acrylate, which have good adhesion, chemical corrosion resistance and biocompatibility.

[0042] Furthermore, the upper filter layer 3 and the lower filter layer 5 are respectively disposed on the side of the magnetic field amplification layer 4 near the liquid inlet 6 and the liquid outlet 7. Both are filter structures with pore channels. The upper filter layer 3 can homogenize the fluid velocity, so that the cell suspension can be evenly distributed in the magnetic field amplification layer 4 area in a short time. The lower filter layer 5 is used to prevent the loss and outflow of magnetic matrix particles 41.

[0043] Preferably, the pore size of the pore channel is 0.25 to 0.5 mm, which can maintain the strength of the filter structure and ensure the uniformity of the fluid.

[0044] In some specific embodiments, the upper and lower filter layers are filter structures formed by sintering medical-grade polypropylene.

[0045] Specifically, the outer shell includes an inlet 6, a receiving cavity, and an outlet 7 that are connected sequentially from top to bottom along the fluid flow direction. The receiving cavity is cylindrical, and the diameters of the inlet 6 and the outlet 7 are smaller than those of the receiving cavity, forming a shape that is thicker in the middle and thinner at both ends. This shape design helps to buffer the flow rate of the cell suspension when it enters the corresponding area of ​​the magnetic field amplification layer 4, avoiding cell damage caused by sudden changes in flow rate. It also helps to distribute the cell suspension evenly in this area, thereby improving the sorting effect.

[0046] Furthermore, the outer casing includes an upper cover 11 and a bottom shell 12 connected to the bottom end of the upper cover 11. The liquid inlet 6 is located at the top of the upper cover 11, and the receiving cavity and the liquid outlet 7 are located inside the bottom shell 12. In some specific embodiments, the upper cover 11 and the bottom shell 12 are connected by ultrasonic welding to achieve a glue-free sealed connection between the two.

[0047] In some specific embodiments, the outer shell is made of medical-grade polycarbonate to avoid toxic side effects on cells.

[0048] Combination Figure 5 To further illustrate the specific application scenario of the device provided in this application embodiment, when the cell magnetic sorting device is working, a peripheral blood mononuclear cell solution containing various cells (the target T cells in the solution have been specifically recognized by antigen and antibody and bound to the nanomagnetic particles) flows in from the inlet 6. After preliminary filtration by the upper filter layer 3, it enters the magnetic field amplification layer 4. Under the action of the enhanced composite magnetic field, the target cells labeled with nanomagnetic particles are adsorbed onto the surface of the magnetic matrix particles 41. Unlabeled cells continue to flow downward with the cell suspension through the flow channel 42 and are finally discharged from the outlet 7. After the sorting is completed, the external magnetic field is removed. Due to the loss of the magnetic field, the adsorbed target cells detach from the magnetic matrix particles 41 under the action of the subsequent rinsing liquid and are discharged from the outlet 7, thereby collecting the purified target cells.

[0049] The specific embodiments of this application have been described in detail above. For those skilled in the art, several improvements and modifications can be made to this application without departing from the principle of this application, and these improvements and modifications also fall within the protection scope of the claims of this application.

Claims

1. A device for magnetic cell sorting, characterized in that, The device includes an outer shell, inside which are a liquid inlet (6), a receiving cavity and a liquid outlet (7) connected sequentially from top to bottom along the fluid flow direction. An upper filter layer (3), a magnetic field amplification layer (4) and a lower filter layer (5) are arranged sequentially from top to bottom in the receiving cavity. The magnetic field amplification layer (4) is composed of a plurality of magnetic matrix particles (41) and flow channels (42) arranged in a set manner. The magnetic matrix particles (41) are composed of balls (411) and a thermoplastic polymer material layer (412) disposed on the outer surface of the balls (411).

2. The device for magnetic cell sorting according to claim 1, characterized in that, The diameter of the ball (411) is 0.2 to 1.0 mm, and / or the ball (411) may be made from iron, cobalt, nickel and their alloys.

3. The device for magnetic cell sorting according to claim 1, characterized in that, The thickness L of the thermoplastic polymer material layer (412) is 5 to 30 μm.

4. The apparatus for magnetic cell sorting according to claim 1, characterized in that, The arrangement of the magnetic matrix particles (41) is selected from close packing or simple cubic packing.

5. The apparatus for magnetic cell sorting according to claim 4, characterized in that, The diameter d2 of the magnetic matrix particles (41) is 0.3 to 0.7 mm.

6. The apparatus for magnetic cell sorting according to claim 5, characterized in that, The magnetic matrix particles (41) have a filling rate of 40% to 60%.

7. The apparatus for magnetic cell sorting according to claim 6, characterized in that, The gap width D of the flow channel (42) is 0.15 to 0.75 mm.

8. The apparatus for magnetic cell sorting according to claim 1, characterized in that, The upper filter layer (3) and the lower filter layer (5) are filter structures with a pore channel range of 0.25 to 0.5 mm.

9. The apparatus for magnetic cell sorting according to claim 1, characterized in that, The cavity is cylindrical, and the diameters of the inlet (6) and outlet (7) are smaller than the cavity. And / or, the outer casing includes an upper cover (11) and a bottom shell (12) connected to the bottom end of the upper cover (11), the liquid inlet (6) is located at the top of the upper cover (11), and the receiving cavity and the liquid outlet (7) are located inside the bottom shell (12).

10. The apparatus for magnetic cell sorting according to claim 9, characterized in that, The upper cover (11) and the bottom shell (12) are connected by ultrasonic welding.

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