WO2024159554A1

WO2024159554A1 - Magnetic bead cell sorting device for vitiligo

Info

Publication number: WO2024159554A1
Application number: PCT/CN2023/075437
Authority: WO
Inventors: 李春英; 郭伟楠; 李舒丽; 郭森; 易秀莉; 陈健儒
Original assignee: 中国人民解放军空军军医大学
Priority date: 2023-02-02
Filing date: 2023-02-10
Publication date: 2024-08-08
Also published as: CN116286600A

Abstract

The present invention belongs to the technical field of medical instruments, and in particular relates to a magnetic bead cell sorting device for vitiligo. The device comprises a base, a tube rack, and a support rod, wherein a magnetic attraction assembly is further rotatably connected to the support rod; the magnetic attraction assembly comprises a rotating arm, a support arm, a limiting plate, a rotating column, and a sliding block; the rotating column is provided with an ascending channel and a descending channel on the circumference; a rectangular slot is formed in the limiting plate, the sliding block is slidable in the rectangular slot, and the end of the sliding block facing the rotating column is rotatably connected to a limiting block; the limiting block is slidable along the ascending channel and the descending channel; a second electromagnet is mounted at the end of the sliding block away from the rotating column; a power-on strip configured to power a second electromagnet circuit to be on or off is provided in the descending channel; when the limiting block slides along the descending channel, the second electromagnet is powered on; and when the limiting block slides along the ascending channel, the second electromagnet is powered off. It is possible to solve the problems in existing magnetic bead cell sorting devices, such as the binding effect of magnetic beads to cells being poor, and the magnetic beads being attached to a side wall and thus being inconvenient for the subsequent transfer.

Description

A vitiligo cell magnetic bead sorting device

Technical Field

The invention belongs to the technical field of medical devices, and in particular relates to a vitiligo cell magnetic bead sorting device.

Background Art

Vitiligo in humans is a depigmented skin disease caused by the specific killing of melanocytes by endogenous CD8+T cells that can recognize melanocyte-specific antigens. Previous studies have found that melanoma cells and melanocytes have common antigens, and melanoma immunotherapy can activate CD8+T cells that recognize the common antigens of the two, leading to vitiligo in some melanoma patients receiving immunotherapy. In addition, Treg cells can induce CD8+T cell disability, and the loss of Treg cells can induce the onset of vitiligo. When conducting animal model-related experiments, cell sorting experiments are required.

In cell sorting experiments, the most commonly used method is magnetic bead sorting, which is based on the specific binding of cell surface antigens to magnetic beads coupled with antibodies, so that the cells are labeled. When the labeled cells pass through the sorting column (there is a stable magnetic field around the sorting column), the cells with magnetic beads remain on the column, and the cells without magnetic beads flow away, thereby achieving the sorting of target cells. The key point of using magnetic beads to sort cells is high-quality cell labeling. There are two labeling methods, including direct labeling and indirect labeling. Direct labeling is to label the target cells, which is a faster and more specific magnetic labeling method. At present, there are a variety of MACS direct-labeled microbeads for sorting human, mouse, rat and non-human primate cells. Indirect labeling is to label non-target cells, which requires the combined use of monoclonal or polyclonal antibodies and microbeads. Unbound antibodies, biological, biotinylated antibodies or fluorescein-labeled antibodies can all be used as primary antibodies to label cells.

When this method is used for labeling, the magnetic beads are generally in a static state. The magnetic beads are relatively easy to bind to cells close to the tube wall, and the cells located on the axis line of the tube are not easily labeled by the magnetic beads. There is a problem of poor binding effect between the magnetic beads and the cells. In addition, the magnetic beads are easily attached to the side wall of the tube, which makes it inconvenient to transfer the magnetic beads to the subsequent experimental cup.

Summary of the invention

In view of this, the purpose of the present invention is to provide a vitiligo cell magnetic bead sorting device to solve the problems that the existing magnetic bead sorting device has poor binding effect between magnetic beads and cells during magnetic bead sorting, and the magnetic beads are easily attached to the side walls, making it inconvenient to transfer them to the subsequent cup body.

The present invention solves the above technical problems by the following technical means:

A vitiligo cell magnetic bead sorting device comprises a base, a tube rack and a support rod, wherein the tube rack and the support rod are fixed on the base, a centrifuge tube containing magnetic beads and cell fluid is installed on the tube rack, the support rod is located directly below the centrifuge tube, a first electromagnet is fixedly connected to the top of the support rod, a first gear is fixedly connected to the middle of the support rod, and a magnetic attraction assembly is rotatably connected to the support rod;

The magnetic attraction assembly includes a rotating arm, a supporting arm, a limiting plate, a rotating column and a slider. The rotating arm is rotatably connected to the support rod, the supporting arm is fixedly connected to the rotating arm, the limiting plate is fixedly connected to the supporting arm, the rotating column is rotatably connected to the supporting arm, the bottom of the rotating shaft of the rotating column is fixedly connected to a second gear, an ascending channel and a descending channel are provided on the circumference of the rotating column, the ascending channel and the descending channel are both spiral-shaped, the ascending channel and the descending channel are connected end to end to form a closed track, a rectangular groove is provided on the limiting plate, the slider is located in the rectangular groove and slides, the slider is rotatably connected to a limiting block toward one end of the rotating column, the limiting block can slide along the ascending channel and the descending channel, a second electromagnet is installed on the end of the slider away from the rotating column, a clutch gear is also provided on the rotating arm, and the clutch gear can mesh with the first gear and the second gear at the same time;

A power belt for providing on-off of the second electromagnet circuit is arranged in the descending channel. When the limit block slides along the descending channel, the second electromagnet is powered on, and when the limit block slides along the ascending channel, the second electromagnet is powered off.

On the basis of the above technical solution, the present invention also makes the following improvements:

Furthermore, a connecting plate is fixedly connected to the bottom of the limit plate, and the connecting plate is connected to the rotating shaft bearing of the rotating column. The connecting plate can fix the limit plate on the rotating column, prevent the limit plate from being away from the rotating column, and prevent the slider from being separated from the rotating column, which has better stability.

Furthermore, the rotating arm is hollow, a third gear is arranged in the rotating arm, a motor for driving the third gear to rotate is arranged on the rotating arm, and a fourth gear meshing with the third gear is arranged on the support rod. The gears rotate, and the third gear revolves along the fourth gear, thereby being able to bring the rotating arm to revolve around the support rod.

Furthermore, the tube rack includes a frame body and a rotating disk, the frame body is fixedly connected to the base, the rotating disk is rotatably connected to the frame body, and the rotating disk is provided with a plurality of through holes for installing centrifuge tubes, and the axis center line of each through hole is at an equal distance from the axis center line of the rotating disk. A plurality of centrifuge tubes can be installed on the rotating disk at one time, and a centrifuge tube can be replaced by rotating the rotating disk for magnetic bead separation operation, which is more efficient.

Furthermore, the apertures of the through holes on the rotating disk in the clockwise direction become larger in sequence. Through holes with different apertures are provided to facilitate installation of centrifuge tubes of different sizes.

Furthermore, the magnetic force of the first electromagnet is greater than the magnetic force of the second electromagnet, so as to prevent the second electromagnet from sucking away the magnetic beads at the bottom of the centrifuge tube.

Beneficial effects of the present invention:

1. When the clutch gear is separated from the first gear and the second gear, the limit block is placed in the descending channel, the magnetic assembly revolves along the support rod, the second electromagnet moves in a circle around the centrifuge tube, and the magnetic beads in the centrifuge tube move with the second electromagnet. The magnetic beads move in the centrifuge tube, and the binding effect between the magnetic beads and the cells is better;

2. When the clutch gear is meshed with the first gear and the second gear, the magnetic assembly revolves along the support rod. Under the action of the clutch gear, the second gear can rotate along the axis, thereby driving the rotating column to rotate. The limit block slides up and down along the limit plate driven by the rotating column. When the limit block descends along the descending channel, the second electromagnet is energized, and the second electromagnet can drive the magnetic beads on the upper layer of the centrifuge tube to move downward. Under the joint action of the first electromagnet and the second electromagnet, the magnetic beads on the side wall of the centrifuge tube can be concentrated to the bottom of the centrifuge tube, which is convenient for subsequent operations.

BRIEF DESCRIPTION OF THE DRAWINGS

The present invention can be further illustrated by means of non-limiting examples given in the accompanying drawings;

FIG1 is a schematic diagram of the three-dimensional structure of a sorting device in an embodiment of the present invention;

FIG2 is a front view of a sorting device according to an embodiment of the present invention;

FIG3 is a schematic diagram of a partial three-dimensional structure of a magnetic attraction assembly according to an embodiment of the present invention;

FIG4 is a schematic diagram of a circuit of a second electromagnet and a power supply belt in an embodiment of the present invention;

FIG5 is a schematic diagram of the three-dimensional structure of a limiting plate in an embodiment of the present invention;

6 is a schematic diagram of the internal structure of the rotating arm in an embodiment of the present invention;

7 is a schematic diagram of the three-dimensional structure of the pipe rack in an embodiment of the present invention;

Among them, 1-base, 2-tube rack, 21-frame, 22-rotating disk, 23-through hole, 3-support rod, 31-first electromagnet, 32-first gear, 33-fourth gear, 4-magnetic attraction component, 41-rotating arm, 411-third gear, 412-motor, 42-support arm, 43-limiting plate, 431-connecting plate, 44-rotating column, 45-slider, 46-second gear, 47-limiting block, 48-second electromagnet, 49-clutch gear.

DETAILED DESCRIPTION

The present invention will be described in detail below with reference to the accompanying drawings and specific embodiments:

As shown in Figures 1 to 7

A vitiligo cell magnetic bead sorting device comprises a base 1, a tube rack 2 and a support rod 3, wherein the tube rack 2 and the support rod 3 are fixed on the base 1, a centrifuge tube containing magnetic beads and cell fluid is installed on the tube rack 2, the support rod 3 is located directly below the centrifuge tube, a first electromagnet 31 is fixedly connected to the top of the support rod 3, the magnetic force of the first electromagnet 31 can be freely controlled, when the clutch gear 49 is meshed with the first gear 32 and the second gear 46, the switch of the first electromagnet 31 is turned on, and it is closed at other times, the middle part of the support rod 3 is fixedly connected to the first gear 32, the support rod 3 is also rotatably connected to a magnetic attraction component 4, and the end of the magnetic attraction component 4 is located below the first gear 32;

The magnetic attraction assembly 4 includes a rotating arm 41, a supporting arm 42, a limiting plate 43, a rotating column 44 and a slider 45. The rotating arm 41 is rotatably connected to the support rod 3, and the rotating arm 41 can rotate along the circumference of the support rod 3. The supporting arm 42 is fixedly connected to the rotating arm 41, the limiting plate 43 is fixedly connected to the supporting arm 42, the limiting plate 43 is vertically arranged, the rotating column 44 is rotatably connected to the supporting arm 42, the limiting plate 43 and the rotating column 44 are parallel to each other, the rotating column 44 includes a rotating shaft and a swivel, the swivel is fixed on the rotating shaft, and the bottom of the rotating shaft of the rotating column 44 is fixedly connected with a second gear 46. An ascending channel and a descending channel are opened on the circumference of the rotating column 44, and the ascending channel and the descending channel are both spiral-shaped. The ascending channel and the descending channel are staggered with each other, and the ascending channel and the descending channel are connected end to end to form a closed track. The end and end connection of the ascending channel and the descending channel are smoothly connected to facilitate the passage of the limiting block 47. A rectangular groove is provided on the positioning plate 43, and a slider 45 is located in the rectangular groove and slides. The slider 45 is rotated toward one end of the rotating column 44 to connect with a limiting block 47, and the limiting block 47 can slide along the ascending channel and the descending channel. A second electromagnet 48 is installed on the end of the slider 45 away from the rotating column 44. A clutch gear 49 is also provided on the rotating arm 41. The clutch gear 49 can mesh with the first gear 32 and the second gear 46 at the same time. The clutch gear 49 includes a gear body and a telescopic rod. The gear body is rotatably connected to the telescopic rod, and the telescopic rod is fixedly connected to the rotating arm 41. When the telescopic rod is extended, the clutch gear 49 meshes with the first gear 32 and the second gear 46. When the telescopic rod is shortened, the clutch gear 49 is separated from the first gear 32 and the second gear 46.

A power-on belt for providing on and off circuit for the second electromagnet 48 is provided in the descending channel. The power-on belt is made of conductive material. The ends of the two wires on the second electromagnet 48 for powering on are embedded in the two sides where the limit block 47 contacts the rotating column 44. A battery is provided on the second electromagnet 48. One wire of the second electromagnet 48 is connected in series with the battery. When the limit block 47 slides along the descending channel, the two wires contact the power-on belt, and the two wires of the second electromagnet 48 form a closed path. The second electromagnet 48 is powered on. When the limit block 47 slides along the ascending channel, the two wires do not contact the power-on belt, and the second electromagnet 48 is powered off.

Specifically, a connecting plate 431 is fixedly connected to the bottom of the limiting plate 43, and the connecting plate 431 is connected to the shaft bearing of the rotating column 44. The connecting plate 431 restricts the limiting plate 43 and the rotating column 44 together, preventing the limiting plate 43 from being away from the rotating column 44, and preventing the slider 45 from being separated from the rotating column 44, so as to provide better stability.

Specifically, the rotating arm 41 is hollow, a third gear 411 is provided in the rotating arm 41, a motor 412 for driving the third gear 411 to rotate is provided on the rotating arm 41, and a fourth gear 33 meshing with the third gear 411 is provided on the support rod 3. The motor 412 can drive the third gear 411 to rotate, and the third gear 411 rotates along the fourth gear 33, thereby driving the rotating arm 41 to rotate, and then driving the entire magnetic attraction assembly 4 to rotate, and the magnetic attraction assembly 4 can automatically rotate in a circle along the support rod 3, that is, revolve.

Specifically, the tube rack 2 includes a frame body 21 and a rotating disk 22, the frame body 21 is fixedly connected to the base 1, the rotating disk 22 is rotatably connected to the frame body 21, and the rotating disk 22 is provided with a plurality of through holes 23 for installing centrifuge tubes, and the axis center line of each through hole 23 is equidistant from the axis center line of the rotating disk 22. Multiple centrifuge tubes can be installed on the rotating disk 22 at one time, and the next centrifuge tube can be replaced by rotating the rotating disk 22 for magnetic bead separation operation, which is more efficient.

Specifically, the apertures of the through holes 23 on the rotating disk 22 become larger in a clockwise direction. The through holes 23 with different apertures are provided to facilitate installation of centrifuge tubes of different sizes.

Specifically, the magnetic force of the first electromagnet 31 is greater than the magnetic force of the second electromagnet 48. The magnetic force of the second electromagnet 48 is smaller than the magnetic force of the first electromagnet 31. No matter where the second electromagnet 48 is located, the magnetic beads in the centrifuge tube are more strongly affected by the magnetic force of the first electromagnet 31, thereby preventing the second electromagnet from sucking away the magnetic beads at the bottom of the centrifuge tube.

The method of using the present invention is as follows: install the centrifuge tube on the tube rack 2, first separate the clutch gear 49 from the first gear 32 and the second gear 46, place the limit block 47 in the descending channel, the magnetic assembly revolves along the support rod 3, the second electromagnet 48 moves in a circle around the centrifuge tube with the rotating arm 41, and the magnetic beads in the centrifuge tube move with the second electromagnet 48. The movement of the magnetic beads in the centrifuge tube improves the binding effect between the magnetic beads and the cells, and then the clutch gear 49 is meshed with the first gear 32 and the second gear 46, and the magnetic assembly revolves along the support rod 3. Under the action of the clutch gear 49, the second gear 46 can rotate along the axis, thereby driving the rotating column 44 to rotate, and the limit block 47 slides up or down along the limit plate 43 under the action of the ascending channel or the descending channel on the rotating column 44. When the limit block 47 descends along the descending channel, the second electromagnet 48 is energized, and the second electromagnet 48 can drive the magnetic beads in the upper solution in the centrifuge tube to move downward. Under the joint action of the first electromagnet 31 and the second electromagnet 48, the magnetic beads on the side wall of the centrifuge tube can be concentrated to the bottom of the centrifuge tube, which is convenient for subsequent operations.

The above embodiments are only used to illustrate the technical solutions of the present invention and are not intended to limit the present invention. Although the present invention is described in detail with reference to the preferred embodiments, a person skilled in the art should understand that the technical solutions of the present invention can be modified or replaced by equivalents without departing from the purpose and scope of the technical solutions of the present invention, which should be included in the scope of the claims of the present invention. The techniques, shapes, and structural parts not described in detail in the present invention are all well-known technologies.

Claims

A vitiligo cell magnetic bead sorting device comprises a base (1), a tube rack (2) and a support rod (3), wherein the tube rack (2) and the support rod (3) are fixed on the base (1), a centrifuge tube containing magnetic beads and cell fluid is installed on the tube rack (2), and the support rod (3) is located directly below the centrifuge tube, characterized in that: a first electromagnet (31) is fixedly connected to the top of the support rod (3), a first gear (32) is fixedly connected to the middle of the support rod (3), and a magnetic attraction component (4) is also rotatably connected to the support rod (3);

The magnetic attraction assembly (4) comprises a rotating arm (41), a supporting arm (42), a limiting plate (43), a rotating column (44) and a sliding block (45); the rotating arm (41) is rotatably connected to the support rod (3); the supporting arm (42) is fixedly connected to the rotating arm (41); the limiting plate (43) is fixedly connected to the supporting arm (42); the rotating column (44) is rotatably connected to the supporting arm (42); a second gear (46) is fixedly connected to the bottom of the rotating shaft of the rotating column (44); an ascending channel and a descending channel are provided on the circumference of the rotating column (44); the ascending channel and the descending channel are both spiral-shaped. The ascending channel and the descending channel are connected end to end to form a closed track, the limiting plate (43) is provided with a rectangular groove, the slider (45) is located in the rectangular groove and slides, the slider (45) is rotated toward one end of the rotating column (44) to connect the limiting block (47), the limiting block (47) can slide along the ascending channel and the descending channel, the slider (45) is installed with a second electromagnet (48) at one end away from the rotating column (44), and the rotating arm (41) is also provided with a clutch gear (49), and the clutch gear (49) can mesh with the first gear (32) and the second gear (46) at the same time;

A power belt for providing on-off switching for the circuit of the second electromagnet (48) is provided in the descending channel; when the limit block (47) slides along the descending channel, the second electromagnet (48) is powered on; when the limit block (47) slides along the ascending channel, the second electromagnet (48) is powered off.
The vitiligo cell magnetic bead sorting device according to claim 1 is characterized in that a connecting plate (431) is fixedly connected to the bottom of the limiting plate (43), and the connecting plate (431) is connected to the rotating shaft bearing of the rotating column (44).
The vitiligo cell magnetic bead sorting device according to claim 2 is characterized in that: the rotating arm (41) is hollow, a third gear (411) is provided inside the rotating arm (41), and the rotating arm The third gear (41) is provided with a motor (412) for driving the third gear (411) to rotate, and the support rod (3) is provided with a fourth gear (33) meshing with the third gear (411).
A vitiligo cell magnetic bead sorting device according to claim 3, characterized in that: the tube rack (2) comprises a frame body (21) and a rotating disk (22), the frame body (21) is fixedly connected to the base (1), the rotating disk (22) is rotatably connected to the frame body (21), and the rotating disk (22) is provided with a plurality of through holes (23) for installing centrifuge tubes, and the axis center line of each of the through holes (23) is equidistant from the axis center line of the rotating disk (22).
The vitiligo cell magnetic bead sorting device according to claim 4 is characterized in that the apertures of the through holes (23) on the rotating disk (22) gradually increase in a clockwise direction.
The vitiligo cell magnetic bead sorting device according to claim 1 is characterized in that the magnetic force of the first electromagnet (31) is greater than the magnetic force of the second electromagnet (48).