WO2024106996A1

WO2024106996A1 - Rare cell isolation device using centrifugal force

Info

Publication number: WO2024106996A1
Application number: PCT/KR2023/018531
Authority: WO
Inventors: 이정민; 김승훈; 김종만
Original assignee: 주식회사 씨티셀즈
Priority date: 2022-11-18
Filing date: 2023-11-17
Publication date: 2024-05-23
Also published as: KR20240073374A

Abstract

A rare cell isolation device using centrifugal force according to the present invention comprises: a rotating disc part; an inlet part which is arranged on one side of the disc part and rotates together therewith, and in which whole blood, a lytic material that lyses red blood cells, and microbeads that are combined with rare cells having nuclei in the whole blood are injected and mixed; an isolation part which is arranged in the disc part to be connected adjacent to the inlet part and isolates, by density difference using centrifugal force, only the rare cells combined with the microbeads from the mixed solution of the whole blood, the lytic material, and the microbeads, by passing the rare cells through a density gradient material, the mixed solution being provided from the inlet part; and a collection part which is arranged in the disc part to be connected adjacent to the isolation part and collects the rare cells having passed through the density gradient material. By this configuration, only nucleated red blood cells can be isolated by density difference using centrifugal force after red blood cells in blood are lysed, thus enabling high-purity isolation of rare cells.

Description

Rare cell separation device using centrifugal force

The present invention relates to a device for separating rare cells using centrifugal force. More specifically, it can effectively improve the detection rate of rare cells by increasing the binding force between rare cells with nuclei in the blood and microbeads and the density difference caused by centrifugal force. This is about a rare cell separation device using centrifugal force.

Cancer is one of the leading causes of death worldwide, along with cardiovascular disease. In particular, the death rate due to cancer has hardly decreased over the past 50 years, so the probability of cancer occurring if one survives to the average life expectancy is approximately 34%, or 1 in 3 people. Accordingly, various studies are being conducted to reduce the mortality rate from cancer by diagnosing cancer early and increasing the cancer treatment rate.

As one of these early cancer diagnosis methods, a method of selectively recognizing circulating cancer cells existing in the blood using microbeads has been recently applied. Microbeads bind to cancer cells in the blood and use their high-density properties to separate cancer cells from the blood. Meanwhile, as the most important step in the method of isolating rare cells such as cancer cells in the blood for detection is the step of binding microbeads to cancer cells in the blood, various studies have been continuously conducted in recent years to increase the cell detection rate. There is a trend.

The purpose of the present invention is to provide a rare cell separation device using centrifugal force that can improve the detection rate of rare cells by increasing the binding force between rare cells with nuclei in the blood and microbeads.

A rare cell separation device using centrifugal force according to the present invention to achieve the above object includes a rotating disc unit, provided on one side of the disc unit and rotated together with the disc unit, and contains whole blood, a lytic substance that dissolves red blood cells, and a substance in the whole blood. An injection unit where microbeads bound to rare cells with nuclei are injected and mixed, the disk unit is adjacent to and connected to the injection unit, and the microbeads are supplied from the mixture of whole blood, lysate, and microbeads provided from the injection unit. A separation unit that separates only the rare cells bound to beads by passing a density gradient material through a density difference caused by centrifugal force, and the disk unit is provided adjacent to the separation unit and connected to the rare cells that have passed through the density gradient material. It includes a collection unit that collects.

Additionally, the rare cells may include nucleated red blood cells or cancer cells.

In addition, the disk portion includes a disk-shaped disk body and rotates about a rotation center, and the injection portion, separation portion, and collection portion are arranged to be sequentially adjacent to each other in a circular arc in the radial direction from the rotation center of the disk body. You can.

Additionally, the injection unit may include a blood chamber into which the whole blood, dissolved material, and microbeads are injected, adjacent to the rotation center of the disc unit.

In addition, the injection unit may include a blood chamber into which the whole blood and the lysate are injected to lyse the red blood cells from the whole blood, a collection chamber for collecting the residue dissolved by the lysate from the blood chamber, and Once the residue is collected, it may include a bead chamber for injecting the microbeads into the blood chamber.

Additionally, a valve is provided between the injection unit and the separation unit to selectively open or close the movement of the mixed liquid.

In addition, the separation unit includes a separation chamber provided adjacent to the injection unit to allow the mixed liquid to flow in between the injection unit and the valve, and the separation chamber is located further than the injection unit in a radial direction from the rotation center of the disk unit. It can be arranged in a remote location.

In addition, the lysate contains RBC lysis buffer (Red Blood cell Lysis buffer) that lyses red blood cells, and the microbeads may have a density higher than that of the whole blood and lower than that of the nucleated red blood cells.

Additionally, the microbeads may have a density of 1.13 to 1.17 g/mL.

In addition, the collection unit includes a collection chamber provided adjacent to the separation chamber at a position radially away from the rotation center of the disk unit so as to be connected to the separation chamber and a separation passage therebetween, and the rotation center of the disk unit. The density gradient material may be filled in the collection chamber connected to the separation passage from the rear end of the separation chamber in the radial direction.

A rare cell separation device using centrifugal force according to another preferred aspect of the present invention is provided with a rotating disc unit, the disc unit adjacent to the rotation center of the disc unit, and rotated together with the disc unit, and dissolves whole blood and red blood cells. An injection unit through which the material and microbeads bound to rare cells with nuclei in the whole blood are injected, and an injection unit provided in the disc unit so as to be adjacent to and connected to the injection unit at a position radially away from the rotation center of the disc unit, wherein the injection unit is provided. A separation unit that separates only the rare cells bound to the microbeads from the whole blood, lysate, and microbeads provided from the unit by density difference by centrifugal force, and a separation unit located at a position radially away from the rotation center of the disc unit. It is provided in the disk unit to be connected adjacently, and is provided in the disk unit to be adjacent to and connected to the separation unit, and includes a collection unit that collects the rare cells separated from the separation unit.

The rare cells may include nucleated red blood cells or cancer cells.

In addition, the disk unit includes a disc-shaped disk body and rotates around a rotation center, and can reciprocate within a predetermined rotation angle range.

Additionally, the injection unit may include a blood chamber into which the whole blood, dissolved material, and microbeads are injected.

Additionally, a valve is provided between the injection unit and the separation unit to selectively open or close the movement of the mixture of whole blood, dissolved material, and microbeads.

Additionally, the separation unit may include a separation chamber provided adjacent to the injection unit to allow the mixture of whole blood, dissolved material, and microbeads to flow in between the injection unit and the valve.

Additionally, the microbeads may have a density of 1.13 to 1.17 g/mL.

In addition, the collection unit includes a collection chamber connected to the separation chamber and the separation passage, and is connected to the collection chamber connected to the separation passage from the rear end of the separation chamber in the radial direction from the rotation center of the disk unit. The density gradient material may be filled.

According to the present invention having the above configuration, first, by lysing cells in whole blood except nucleated red blood cells, which are rare cells to be detected, the binding force between nucleated red blood cells and microbeads can be increased.

Second, the nucleated red blood cells can be separated and detected by density difference after dissolving red blood cells with similar density components to nucleated red blood cells in whole blood with a lytic substance, thereby improving the detection rate of high-purity rare cells.

Third, nucleated red blood cells and microbeads can be separated from blood by passing through a density gradient material due to the density difference in a combined state, thereby improving the separation and detection rate of rare cells.

Fourth, as it becomes unnecessary to stop the rotation of the disc portion when opening and closing the valve for combining existing rare cells and microbeads, a configuration to prevent blood backflow in the existing blood chamber becomes unnecessary. As a result, the mixing power between rare cells and microbeads in the blood chamber can be improved, and cell layer separation due to the combination between rare cells and microbeads in the blood chamber becomes more accurate due to the centrifugal force of the rotating disk unit. .

Fifth, the detection rate of rare cells such as nucleated red blood cells and cancer cells in the blood can be increased, thereby improving medical technology and improving public health.

Figure 1 is a diagram schematically showing a rare cell separation device using centrifugal force in a preferred embodiment of the present invention.

FIG. 2 is a schematic cross-sectional view taken along line II-II shown in FIG. 1.

FIG. 3 is a schematic configuration diagram to explain the separation operation of the rare cell separation device using centrifugal force according to an embodiment shown in FIG. 1.

Figure 4 is a diagram schematically showing a rare cell separation device using centrifugal force in another preferred embodiment of the present invention. and,

Figure 5 is a schematic configuration diagram to explain the separation operation of a rare cell separation device using centrifugal force according to another embodiment shown in Figure 4.

Hereinafter, a preferred embodiment of the present invention will be described with reference to the attached drawings. However, the spirit of the present invention is not limited to such embodiments, and the spirit of the present invention may be proposed differently by adding, changing, or deleting components constituting the embodiments, but this is also included in the spirit of the invention. It will happen.

Referring to Figure 1, according to a preferred embodiment of the present invention, a rare cell separation device (1) using centrifugal force includes a disk unit (10), an injection unit (20), a separation unit (30), and a collection unit ( 40).

For reference, the rare cell separation device 1 using centrifugal force described in the present invention is for separating and detecting rare cells in the blood, and the rare cells described in this embodiment are fetal cells that entered the mother's blood. Here, the fetal cells contained in the mother's blood are red blood cells with a nucleus and are referred to as nucleated red blood cells (NRBC) (N). Meanwhile, the rare cells described in the present invention are described as nucleated red blood cells (N) in the blood, but are not limited to this and various embodiments such as cancer cells in the blood are possible.

The disk unit 10 includes a rotating disk-shaped disk body 11. This disk unit 10 is provided with a rotation drive means (not shown) for rotating the disk body 11, and the disk body 11 is connected to the rotation center 12 by connection with the rotation drive means (not shown). It rotates around . Here, the disk unit 10 can be rotated 360 degrees around the rotation center 12, or can be rotated back and forth within a predetermined rotation angle.

Meanwhile, the disc body 11 of the disc unit 10 is provided with first and

second passages

13 and 14 to provide a movement path for fluid, including blood, and these first and second passages 13 ) The configuration of 14 will be described in more detail later along with the configuration of the injection unit 20, separation unit 30, and collection unit 40.

The injection unit 20 is provided on one side of the disc unit 10 and rotates together with the disc unit 10, and whole blood, dissolved substances (L), and microbeads (M) are injected. Here, whole blood (B), dissolved material (L), and microbeads (M) injected into the injection unit 20 may be injected in a mixed state or may be injected individually. Here, the whole blood (B) is the mother's blood, and the lytic substance (L) dissolves unnecessary cells, including red blood cells, contained in the whole blood (B). The microbeads (M) are combined with nucleated red blood cells (N) in whole blood (B) that have not been dissolved by the lysing agent (L).

In this example, it is exemplified that the lysate (L) uses RBC lysis buffer (Red Blood cell Lysis buffer), which lyses red blood cells by osmotic pressure. As this RBC lysis buffer does not dissolve nucleated red blood cells (N), which are red blood cells with nuclei, the nucleated red blood cells (N), which are rare cells contained in whole blood (B), do not dissolve but bind to the microbeads (M). do.

Microbeads (M) are a type of antibody that binds to nucleated red blood cells (N), which are antigens, through an antigen-antibody reaction. Additionally, in this example, the microbeads (M) are illustrated to include CD71 microbeads (M), but this is not a limitation. These microbeads (M) are high-density materials, and their density increases by combining with nucleated red blood cells (N), which have the highest density in whole blood (B).

Meanwhile, the injection unit 20 may include a blood chamber 21 provided relatively adjacent to the rotation center 12 of the disc body 11. Whole blood (B), dissolved material (L), and microbeads (M) are injected into the blood chamber 21 and mixed. At this time, the blood chamber 21 is provided with a first flow path 13 in the disc body 11 for injection of whole blood (B), dissolved material (L), and microbeads (M).

The separation unit 30 is provided to be connected to the disk unit 10 adjacent to the injection unit 20, and the whole blood (B), dissolved material (L), and microbeads (M) provided from the injection unit 20 are mixed. Only nucleated red blood cells (N), which are rare cells combined with microbeads (M), are separated from the mixed solution by density difference using centrifugal force. Here, the separation part 30 is provided adjacent to the injection part 20, and with respect to the rotation center 12 of the disk body 11, the injection part 20 moves in a radial arc from the rotation center 12. ) and a separation chamber 31 located further away from the blood chamber 11.

The separation chamber 31 may be connected to the blood chamber 21 of the injection unit 20 with a blood flow path 22 therebetween. A valve 23 that can be opened and closed may be provided in the blood flow path 22, and the mixed whole blood (B) is dissolved in the blood chamber 21 of the injection unit 20 by opening or closing the valve 23. A fluid containing the substance (L) and the microbeads (M) may flow into the separation chamber 31.

Meanwhile, as the disk body 11 of the disk unit 10 rotates, whole blood (B), dissolved material (L), and microbeads (M) within the blood chamber 21 are rotated by the disk body 11. are mixed together. At this time, the disk body 11 is reciprocally rotated at a predetermined angle around the rotation center 12, thereby mixing the whole blood (B), dissolved material (L), and microbeads (M) in the blood chamber 21. The whole blood (B), dissolved material (L), and microbeads (M) mixed in the blood chamber 21 are moved in the radial direction of the disc body 11 by the centrifugal force generated by the rotation of the disc body 11. By moving to , it flows into the separation chamber 31.

Among the mixture of whole blood (B), lysate (L), and microbeads (M) introduced into the separation chamber 31 in this way, relatively high-density microbeads (M) and nucleated red blood cells (N) are separated from the disc body by centrifugal force. As it continues to move in the radial direction of (11), it is separated from the whole blood (B) due to the density difference. That is, in whole blood (B), cells other than the nucleated red blood cells (N) bound to the microbeads (M) are separated from the nucleated red blood cells (N) and microbeads (M) in a dissolved state by the lysate (L). .

A density gradient material (DGM) is provided at the rear end of the separation chamber 31 based on a direction gradually moving away from the rotation center 12 of the disk body 11 in the radial direction. Here, the density gradient material (D) may be a material having a density of approximately 1.13 to 1.17 g/mL, which is higher than the density of water, in order to separate only the nucleated red blood cells (N) bound to the microbeads (M) from blood. More preferably, the density gradient material (D) is made of a material having a density range that is higher than that of whole blood (B) and lower than the density of nucleated red blood cells (N) bound to microbeads (M), but is prepared as a fluid. good night.

As the nucleated red blood cells (N) combined with the microbeads (M) moving in the radial direction due to the centrifugal force of the disc body 11 pass through this density gradient material (D), the nucleated red blood cells (N) are separated from the blood. .

For reference, whole blood (B) that does not pass through the density gradient material (D) due to centrifugal force in the separation chamber (31), and cells included in the whole blood (B) are transferred to the second flow path (14) provided in the disc body (11). ) can be discharged through.

The collection unit 40 is connected to the disc unit 10 adjacent to the separation unit 30 and collects nucleated red blood cells (N), which are rare cells that have passed through the density gradient material (D). The collection unit 40 is provided adjacent to the separation unit 30 in the radial direction from the rotation center 12 of the disk body 11, and is connected to the separation chamber 31 and the separation passage 32 of the separation unit 30. It includes a collection chamber 41 connected to it. This collection chamber 41 is provided adjacent to the arc of the disc body 11, so that it is spaced farther away from the rotation center 12 of the disc body 11 than the blood chamber 21 and the separation chamber 31 described above. It is provided in location.

Meanwhile, since the separation passage 32 between the separation chamber 31 and the collection chamber 41 is in an open state, the density gradient material D provided at the rear end of the separation chamber 31 is in the collection chamber 41. It is filled to the inside. That is, the density gradient material D filled from the collection chamber 41 to the rear end of the separation chamber 31 is in a fluid state due to centrifugal force caused by the rotation of the disk body 11.

Meanwhile, as shown in FIG. 2, a jaw 42 may be provided in the separation passage 32 provided between the separation chamber 31 and the collection chamber 41. Here, the chin 42 is provided in the shape of a kind of triangular protrusion, so that blood in the separation chamber 31 can be blocked from flowing into the collection chamber 41.

As described above, the blood chamber 21, the separation chamber 31, and the collection chamber 41 are sequentially arranged to be adjacent to each other in a radial arc from the rotation center 12 of the disc body 11. Each of these blood chambers 21, separation chambers 31 and collection chambers 41 all have an internal cross-sectional shape that narrows obliquely in a direction radially away from the center of rotation 12. Therefore, the inclined shape of the blood chamber 21, separation chamber 31, and collection chamber 41 guides the components in the blood so that they can be easily separated by the density difference caused by the centrifugal force caused by the rotation of the disc body 11. can do.

The rare cell separation operation of the rare cell separation device 1 using centrifugal force according to a preferred embodiment of the present invention having the above configuration will be described with reference to FIGS. 1 and 3 as follows.

1 and 3, the blood chamber 21 of the injection unit 20, the separation chamber 31 of the separation unit 30, and the collection unit ( The collection chambers 41 of 40 are provided adjacent to each other. More preferably, as shown in FIG. 3, the disc body 11 can be rotated back and forth within a predetermined angle range while standing vertically, and the blood chamber 21 and the separation chamber are formed based on the rotation center 12. (31) and the collection chamber (41) are arranged to be gradually spaced apart in the radial direction.

When whole blood (B), lysate (L), and microbeads (M) flow into the blood chamber 21, whole blood (B), lysate (L), and microbeads ( M) are mutually mixed. At this time, the lytic substance (L) dissolves cells other than the nucleated red blood cells (N) in the whole blood (B), so that the nucleated red blood cells (N) and the microbeads (M) are easily combined with each other. For reference, the lysate (L) dissolves red blood cells with a density similar to that of the nucleated red blood cells (N), thereby separating the red blood cells from the whole blood (B) together with the nucleated red blood cells (N) by the density difference in the separation chamber 31. to block collection into the collection chamber 41.

The mixed whole blood (B), lysate (L), and microbeads (M) flow into the separation chamber (31) connected to the blood flow path (22) by opening the valve (23), and within the separation chamber (31) Nucleated red blood cells (N) combined with relatively high-density microbeads (M) are moved to the rear end of the separation chamber 31 by centrifugal force. At this time, a density gradient material (D) is provided at the rear end of the separation chamber 31, so that the nucleated red blood cells (N) combined with the high-density microbeads (M) pass through the density gradient material (D) and enter the adjacent collection chamber. It is gathered as (41). Here, a jaw 42 is provided in the separation passage 32 between the separation chamber 31 and the collection chamber 41, so that blood in the separation chamber 31 does not flow into the collection chamber 41. Additionally, blood in the separation chamber 31 that is not collected in the collection chamber 41 may be discharged through the second flow path 14.

In this way, the user collects the nucleated red blood cells (N) that pass through the density gradient material (D) and are collected in the collection chamber 41 by the density difference caused by centrifugal force, thereby allowing the user to separate and test the nucleated red blood cells (N) in the blood. You can do it.

Referring to Figures 4 and 5, a rare cell separation device 100 using centrifugal force according to another preferred embodiment of the present invention is schematically shown.

As shown in FIGS. 4 and 5, the rare cell separation device 100 using centrifugal force according to another embodiment includes a disk unit 110, an injection unit 120, a separation unit 130, and a collection unit 140. Includes.

Like the above-described embodiment, the disk unit 110 includes a disk body 111 that is provided in a disk shape and rotates about the rotation center 112. Since this disk unit 110 is similar to the above-described embodiment, detailed description will be omitted.

The injection unit 120 is provided on one side of the disc unit 110 and rotates with the disc unit 110, and whole blood (B), dissolved material (L), and microbeads (M) are injected. This injection unit 120 includes a blood chamber 121, a collection chamber 122, and a bead chamber 123.

Whole blood (B) and dissolved material (L) are injected into the blood chamber 121. Such whole blood (B) and dissolved material (L) may be injected into the blood chamber 121 through the blood supply passage 113 provided in the disc body 111. The whole blood (B) and the lysed material (L) injected into the blood chamber 121 are mixed with each other by the reciprocating rotation movement of the disc body 111, so that unnecessary cells such as red blood cells in the whole blood (B) are mixed with the lysed material (L). dissolved by

The collection chamber 122 collects the remainder of the whole blood (B) dissolved by the dissolved substance (L) from the blood chamber (121). That is, the collection chamber 122 collects residues including red blood cells, which are unnecessary cells dissolved by the dissolved substance (L) in the whole blood (B), and is connected to the blood chamber 121 through the collection passage 124. . The residue collected in the collection chamber 122 may be discharged through the residue discharge passage 115.

The bead chamber 123 injects high-density microbeads (M) into the blood chamber 121 after the residue is collected from the blood chamber 121 into the collection chamber 122. This bead chamber 123 can receive microbeads (M) injected through the bead supply passage 116 provided in the disk body 111. By connecting the bead chamber 123 to the blood chamber 121 across the bead passage 125, residues in the whole blood (B) are removed and only the nucleated red blood cells (N) remain inside the blood chamber 121. Micro beads (M) are supplied. As a result, the nucleated red blood cells (N) have better binding force with the microbeads (M) inside the blood chamber 121.

Among the blood chamber 121, collection chamber 122, and bead chamber 123 described above, the bead chamber 123 is provided closest to the rotation center 112 of the disc body 111, and the bead chamber 123 A blood chamber 121 is provided adjacent to the radial direction away from the rotation center 112. Additionally, the collection chamber 122 may be provided to adjoin the blood chamber 121 in the circumferential direction. The blood chamber 121, the collection chamber 122, and the bead chamber 123 may all have a cross-sectional shape in which the internal space becomes narrower as the radial distance from the rotation center 112 of the disc body 111 increases. there is. Therefore, the inclined shapes of the blood chamber 121, the collection chamber 122, and the bead chamber 123 are guided so that the blood components can be easily separated by the density difference due to the centrifugal force caused by the rotation of the disc body 111. can do.

The separation unit 130 is provided adjacent to the injection unit 120 on the disc unit 110, and is provided to collect nucleated red blood cells, which are rare cells combined with microbeads (M), from the blood provided from the blood chamber 121 of the injection unit 120. Only (N) is separated by density difference due to centrifugal force. To this end, the separation unit 130 includes a separation chamber 131 and a density gradient material D (see FIG. 5) provided at the rear end of the separation chamber 131, as in one embodiment. This density gradient material (D) has a density range that is higher in density than whole blood (B) and lower in density than nucleated red blood cells (N), thereby providing a density boundary between whole blood (B) and nucleated red blood cells (N).

Inside the separation chamber 131, the nucleated red blood cells (N) combined with the microbeads (M) move away in the radial direction of the disk body 111 by centrifugal force and pass through the density gradient material (D). Blood other than the nucleated red blood cells (N) bound to the microbeads (M) in the whole blood (B) can be discharged through the discharge passage 114 provided in the disc body 111.

The collection unit 140 is provided adjacent to the separation unit 130 on the disk unit 110 and collects nucleated red blood cells (N), which are rare cells that have passed through the density gradient material (D). This collection unit 140 includes a separation chamber 131 provided in the disk body 111 and a collection chamber 141 provided adjacent to the separation passage 132 with the separation channel 132 in between. Here, although not shown in detail, as in one embodiment, the separation passage 132 is provided with a jaw 42 (see FIG. 2), so that blood, excluding the nucleated red blood cells (N) bound to the microbeads (M), is collected into the collection chamber ( 141), the inflow can be blocked.

These separation chambers 131 and collection chambers 141 are provided adjacent to each other in a direction radially away from the rotation center 112 with respect to the disk body 111, and have an inclined shape so that the internal space in the radial direction is gradually narrowed. By having , it is possible to guide the collection of density differences in blood by centrifugal force. Additionally, the collection chamber 141 is located above the separation chamber 131 with respect to the disk body 111, and the separation passage 132 between the collection chamber 141 and the separation chamber 131 is open. Therefore, the density gradient material D, which is a fluid, may be filled from the collection chamber 141 to the rear end of the separation chamber 131.

The separation operation of the rare cell separation device 100 using centrifugal force according to another embodiment of the present invention having the above configuration will be described with reference to FIGS. 4 and 5 as follows.

4 and 5, whole blood (B) and lysate (L) are first injected into the blood chamber 121, and unnecessary cells such as red blood cells in the whole blood (B) are dissolved by the lysate (L). At this time, whole blood (B) and dissolved material (L) may be supplied through the blood supply channel 113. The residue in the dissolved whole blood (B) is collected into the collection chamber 122 through the collection passage 124, and the residue collected in the collection chamber 122 can be discharged through the residue discharge passage 115. there is.

With the residue in the blood chamber 121 removed, microbeads (M) are introduced into the blood chamber 121 from the bead chamber 123 through the bead passage 125. Here, the bead chamber 123 can receive microbeads (M) through the bead supply passage 116 and provide them to the blood chamber 121. When microbeads (M) are supplied into the blood chamber 121, nucleated red blood cells (N) that are not dissolved in whole blood (B) are combined with the microbeads (M). At this time, as the disk body 111 reciprocates within a predetermined rotation angle, bonding between the microbeads (M) and the nucleated red blood cells (N) within the blood chamber 121 is easier.

Whole blood (B) containing nucleated red blood cells (N) combined with microbeads (M) flows into the separation chamber 131 connected through the blood flow path 126. The separation chamber 131 separates the nucleated red blood cells (N) bound to the relatively high-density microbeads (M) by centrifugal force caused by the rotation of the disk body 111. Here, the nucleated red blood cells (N) combined with the microbeads (M) are moved in the radial direction away from the rotation center 112 of the disc body 111 by centrifugal force, thereby passing through the density gradient material (D) and producing whole blood (B). ) can be separated from.

The nucleated red blood cells (N) combined with the separated microbeads (M) flow into the collection chamber 141 through the separation passage 132 and are finally collected. For reference, the separation passage 132 is provided with a jaw 142 to prevent whole blood B from flowing into the separation passage 132.

Meanwhile, as shown in FIG. 5,

valves

124a, 125a, and 126a are provided in the collection flow path 124, the bead flow path 125, and the blood flow path 126, respectively, so that the collection flow path 124 and the beads The flow path 125 and the blood flow path 126 can be selectively opened and closed, but this is not a limitation.

As described above, although the present invention has been described with reference to preferred embodiments, those skilled in the art may make various modifications and changes to the present invention without departing from the spirit and scope of the present invention as set forth in the claims below. You will understand that you can do it.

Included in the text.

Claims

a rotating disk unit;

An injection unit provided on one side of the disc unit and rotated with the disc unit, where whole blood, a lytic substance that dissolves red blood cells, and microbeads combined with rare cells with nuclei in the whole blood are injected and mixed;

The disk unit is provided to be adjacent to and connected to the injection unit, and from the mixture of whole blood, dissolved material and microbeads provided from the injection unit, only the rare cells combined with the microbeads are infused with a density gradient material through a density difference due to centrifugal force. A separation part that passes through and separates; and

A collection unit provided in the disk unit adjacent to and connected to the separation unit to collect the rare cells that have passed through the density gradient material;

Rare cell separation device using centrifugal force including.
According to paragraph 1,

A rare cell separation device using centrifugal force, wherein the rare cells include nucleated red blood cells or cancer cells.
According to paragraph 1,

The disk unit includes a disk-shaped disk body and rotates around a rotation center,

A rare cell separation device using centrifugal force wherein the injection unit, separation unit, and collection unit are sequentially adjacent to each other in a circular arc in a radial direction from the rotation center of the disc body.
According to paragraph 1,

The injection part,

A rare cell separation device using centrifugal force in which a blood chamber into which the whole blood, dissolved material, and microbeads are injected is provided adjacent to the rotation center of the disc unit.
According to paragraph 1,

The injection part,

a blood chamber into which the whole blood and lysate are injected to lyse the red blood cells from the whole blood;

a collection chamber for collecting residue dissolved by the dissolved material from the blood chamber; and

a bead chamber for injecting the microbeads into the blood chamber when the residue is collected from the blood chamber;

Rare cell separation device using centrifugal force including.
According to paragraph 1,

A rare cell separation device using centrifugal force where a valve is provided between the injection unit and the separation unit to selectively open or close the movement of the mixed solution.
According to paragraph 1,

The separation unit includes a separation chamber provided adjacent to the injection unit so that the mixed solution flows in between the injection unit and the valve,

The separation chamber is a rare cell separation device using centrifugal force, wherein the separation chamber is provided at a position farther away from the injection unit in a radial direction from the rotation center of the disc unit.
According to paragraph 2,

The lysate includes RBC lysis buffer (Red Blood cell Lysis buffer) that lyses red blood cells,

A rare cell separation device using centrifugal force wherein the microbeads have a density higher than the density of the whole blood and lower than the density of the nucleated red blood cells.
According to clause 8,

The microbeads are a rare cell separation device using centrifugal force having a density of 1.13 to 1.17 g/mL.
In clause 7,

The collection unit includes a collection chamber provided adjacent to the separation chamber at a position radially away from the rotation center of the disk unit so as to be connected to the separation chamber with a separation passage therebetween,

A rare cell separation device using centrifugal force in which the density gradient material is filled into the collection chamber connected to the separation passage from the rear end of the separation chamber based on the radial direction from the rotation center of the disk unit.
a rotating disk unit;

An injection unit provided in the disc unit adjacent to the rotation center of the disc unit and rotated together with the disc unit, into which whole blood, a lytic substance that dissolves red blood cells, and microbeads combined with rare cells having a nucleus in the whole blood are injected;

The disc unit is provided to be adjacent to and connected to the injection unit at a position radially away from the rotation center of the disc unit, and the rare cells combined with the microbeads from the whole blood, lysed material, and microbeads provided from the injection unit. A separation unit that separates the bays by density difference due to centrifugal force; and

The disk portion is provided to be adjacent to and connected to the separation portion at a position radially away from the rotation center of the disk portion, and the disk portion is provided to be adjacent to and connected to the separation portion, and the rare cells separated from the separation portion are provided. a collection unit that collects;

Rare cell separation device using centrifugal force including.
According to clause 11,

A rare cell separation device using centrifugal force, wherein the rare cells include nucleated red blood cells or cancer cells.
According to clause 11,

A rare cell separation device using centrifugal force where the disk unit includes a disk-shaped disk body and rotates around a rotation center, reciprocating within a predetermined rotation angle range.
According to paragraph 1,

The injection part,

A rare cell separation device using centrifugal force including a blood chamber into which the whole blood, lysate, and microbeads are injected.
According to clause 11,

The injection part,

a blood chamber into which the whole blood and lysate are injected to lyse the red blood cells from the whole blood;

a collection chamber for collecting residue dissolved by the dissolved material from the blood chamber; and

a bead chamber for injecting the microbeads into the blood chamber when the residue is collected from the blood chamber;

Rare cell separation device using centrifugal force including.
According to clause 11,

A rare cell separation device using centrifugal force, wherein a valve is provided between the injection unit and the separation unit to selectively open or close the movement of the mixture of whole blood, lysate, and microbeads.
According to clause 11,

The separation unit is a rare cell separation device using centrifugal force including a separation chamber provided adjacent to the injection unit so that the mixture of whole blood, lysate, and microbeads flows in between the injection unit and the valve.
According to clause 12,

The lysate includes RBC lysis buffer (Red Blood cell Lysis buffer) that lyses red blood cells,

A rare cell separation device using centrifugal force wherein the microbeads have a density higher than the density of the whole blood and lower than the density of the nucleated red blood cells.
According to clause 18,

The microbeads are a rare cell separation device using centrifugal force having a density of 1.13 to 1.17 g/mL.
According to clause 17,

The collection unit includes a collection chamber connected to the separation chamber with a separation passage therebetween,

A rare cell separation device using centrifugal force in which the density gradient material is filled into the collection chamber connected to the separation passage from the rear end of the separation chamber based on the radial direction from the rotation center of the disk unit.