A cell separation apparatus and continuous separation method

Abstract

The application discloses a cell separation device, which comprises a centrifugal chamber and a rotating driving part for driving the rotation of the centrifugal chamber, a joint assembly is arranged on the centrifugal chamber, a liquid inlet interface, a waste outlet interface and a suction interface are connected to the joint assembly, a flow guide seat is arranged in the centrifugal chamber, a liquid inlet channel, a waste outlet channel and a suction channel are arranged on the flow guide seat, the liquid inlet interface is communicated with the liquid inlet channel through a first connecting pipe, the waste outlet interface is communicated with the waste outlet channel through a second connecting pipe, the suction interface is communicated with the suction channel through a third connecting pipe, and the suction channel extends to the end of the centrifugal chamber. A continuous separation method comprises the following steps: a) filling a buffer solution; b) filling a liquid containing target cells; c) cell layering; d) cell separation; or d') target cell washing. The application has the advantages of simple structure, compactness, continuous centrifugal separation, and the ability to process a larger volume of liquid.

Description

Technical Field

[0001] This invention relates to centrifugation equipment and methods, and more particularly to a cell separation equipment and continuous separation method. Background Technology

[0002] The cell therapy market is growing rapidly. Cell separation from blood is a fundamental technology, and the application of automation, disposable closed systems, separation efficiency, and recovery rates are key considerations. Existing cell separation equipment uses the same flow channel for the feed solution entering the separation chamber and for the cells exiting the chamber, making continuous centrifugation impossible. Furthermore, to accommodate large volumes of feed solution input, the separation chamber needs to be large, increasing costs and requiring more frequent solution changes. Summary of the Invention

[0003] The technical problem to be solved by the present invention is to overcome the shortcomings of the prior art and provide a cell separation device with a simple and compact structure that can realize continuous centrifugal separation and process large volumes of liquid.

[0004] The present invention further provides a continuous separation method for the above-mentioned cell separation device.

[0005] To solve the above-mentioned technical problems, the present invention adopts the following technical solution:

[0006] A cell separation device includes a centrifuge chamber and a rotary drive for driving the centrifuge chamber to rotate. The centrifuge chamber is provided with a connector assembly, which is connected to a liquid inlet, a waste outlet, and a suction outlet. The centrifuge chamber is provided with a flow guide seat, which is provided with a liquid inlet channel, a waste outlet channel, and a suction channel. The liquid inlet is connected to the liquid inlet channel through a first connecting pipe, the waste outlet is connected to the waste outlet channel through a second connecting pipe, and the suction outlet is connected to the suction channel through a third connecting pipe. The suction channel extends to the end of the centrifuge chamber.

[0007] As a further improvement to the above technical solution: the axis of the first connecting tube coincides with the rotation axis of the centrifuge chamber, the third connecting tube is coaxially sleeved on the outer periphery of the first connecting tube with a gap between them, and the second connecting tube is coaxially sleeved on the outer periphery of the third connecting tube with a gap between them.

[0008] As a further improvement to the above technical solution: the liquid inlet channel is located on the side of the guide seat near the rotary drive component, the waste discharge channel is located on the side of the guide seat near the connector assembly, and the suction channel is located between the liquid inlet channel and the waste discharge channel.

[0009] As a further improvement to the above technical solution: the liquid inlet channel, the waste discharge channel and the suction channel are all provided with multiple channels and are evenly arranged around the rotation axis of the centrifuge chamber.

[0010] As a further improvement to the above technical solution: a sealing element is provided between the first connecting pipe and the connector assembly, between the second connecting pipe and the connector assembly, and between the third connecting pipe and the connector assembly.

[0011] As a further improvement to the above technical solution: the centrifuge chamber is provided with a conical part on at least one side.

[0012] As a further improvement to the above technical solution: the liquid inlet is connected to an input pump, the inlet of the input pump is connected to a raw liquid bag, a replacement liquid bag and a spare interface through on-off valves, the waste discharge interface is equipped with a first liquid detection component, the waste discharge interface is connected to a waste liquid bag through on-off valves, the suction interface is equipped with a suction pump, a second liquid detection component and a color detection component, and the outlet of the suction pump is connected to a harvest bag, a separation liquid bag, a buffer solution bag and a waste liquid bag through on-off valves.

[0013] A continuous separation method for the above-mentioned cell separation device includes the following steps:

[0014] a) Adding buffer solution: Add buffer solution into the centrifuge chamber through the third connecting tube and the suction channel at the suction port until the waste discharge port detects that the chamber is full of liquid;

[0015] b) Adding liquid containing target cells: The liquid inlet interface adds liquid containing target cells into the centrifuge chamber through the first connecting pipe and the liquid inlet channel. At the same time, the rotating drive drives the centrifuge chamber to rotate, and the cells gather at the end of the centrifuge chamber. The supernatant is discharged to the waste outlet through the waste outlet channel and the second connecting pipe.

[0016] c) Cell stratification: After all the liquid containing the target cells has been added, the rotation speed of the rotating drive increases to cause the cells to stratify in the centrifuge chamber;

[0017] d) Cell separation: The suction port aspirates waste cells from the end of the centrifuge chamber into the waste liquid bag through the third connecting tube and suction channel. After the waste cells are aspirated, the port switches to the harvest bag to harvest the target cells until the target cells are harvested. At the same time as the suction port aspirates cells, the liquid inlet port replenishes buffer or gas into the centrifuge chamber.

[0018] Alternatively, dˊ) Target cell washing: The suction port aspirates the waste cells at the end of the centrifuge chamber into the waste liquid bag. After the waste cells are aspirated, the suction port adds buffer solution into the centrifuge chamber through the third connecting tube and suction channel to wash the target cells. The waste discharge port collects the waste liquid.

[0019] As a further improvement to the above technical solution:

[0020] In step b, the suction port adds the separation liquid into the centrifuge chamber;

[0021] Step dˊ is followed by:

[0022] e) Concentration: The rotation speed of the rotary drive increases, causing the target cells to aggregate. The suction port aspirates the target cells from the centrifuge chamber into the harvest bag. At the same time, the suction port aspirates the target cells and the liquid inlet port replenishes the centrifuge chamber with buffer solution or gas.

[0023] As a further improvement to the above technical solution:

[0024] The discarded cells are red blood cells, and the target cells are white blood cells. In step c, the red blood cells are located near the end of the centrifuge chamber, and the white blood cells are located near the rotation axis of the centrifuge chamber.

[0025] Compared with the prior art, the advantages of the present invention are as follows: The cell separation device disclosed in the present invention is provided with a liquid inlet, a waste outlet, and a suction outlet on the connector assembly, and a liquid inlet channel, a waste outlet channel, and a suction channel on the guide seat. The liquid inlet, waste outlet, and suction outlet are connected to the liquid inlet channel, waste outlet channel, and suction channel respectively through a first connecting pipe, a second connecting pipe, and a third connecting pipe. The liquid inlet, waste outlet, and suction processes in the centrifuge chamber can be carried out simultaneously to achieve continuous centrifugation separation. After the cell culture is completed, the liquid volume will be relatively large. The cell separation device can adapt to the input of a large volume of liquid. For example, a 30ml centrifuge chamber can adapt to the input of 50L of liquid containing target cells, and users can reduce the liquid change during the centrifugation process.

[0026] The continuous separation method of the cell separation device disclosed in this invention realizes continuous centrifugation separation and can adapt to the input of large volume liquids. For example, a 30ml centrifuge chamber can adapt to the input of 50L of liquid containing target cells. Users can reduce the liquid change during the centrifugation process and flexibly select the centrifugation, washing or concentration process according to the specific needs of the user. Attached Figure Description

[0027] Figure 1 This is a schematic diagram of the cell separation device of the present invention.

[0028] Figure 2 This is a schematic diagram of the centrifuge chamber in this invention.

[0029] Figure 3 yes Figure 2 A magnified view of a portion of the image.

[0030] The labels in the diagram represent: 1. Centrifuge chamber; 11. Quick-release connector; 2. Rotary drive component; 3. Connector assembly; 4. Liquid inlet; 41. Input pump; 5. Waste outlet; 51. First liquid detection component; 6. Suction interface; 61. Suction pump; 62. Second liquid detection component; 63. Color detection component; 7. Flow guide; 71. Liquid inlet channel; 72. Waste outlet channel; 73. Suction channel; 74. Suction tube; 81. First connecting tube; 82. Second connecting tube; 83. Third connecting tube; 84. Seal; 90. On / off valve; 91. Stock solution bag; 92. Replacement bag; 93. Spare interface; 94. Harvest bag; 95. Separation solution bag; 96. Buffer solution bag; 97. Waste solution bag; 98. Bubble sensor; 100. Cell. Detailed Implementation

[0031] The present invention will be further described in detail below with reference to the accompanying drawings and specific embodiments.

[0032] Example 1

[0033] Figures 1 to 3 An embodiment of the cell separation device of the present invention is shown. The cell separation device of this embodiment includes a centrifuge chamber 1 and a rotary drive 2 for driving the centrifuge chamber 1 to rotate. The centrifuge chamber 1 is provided with a connector assembly 3, which is connected to a liquid inlet 4, a waste outlet 5 and a suction port 6. The centrifuge chamber 1 is provided with a flow guide seat 7, which is provided with a liquid inlet channel 71, a waste outlet channel 72 and a suction port 73. The liquid inlet 4 is connected to the liquid inlet channel 71 through a first connecting pipe 81. The waste outlet 5 is connected to the waste outlet channel 72 through a second connecting pipe 82. The suction port 6 is connected to the suction port 73 through a third connecting pipe 83. The suction port 73 extends to the end of the centrifuge chamber 1. Preferably, a bearing is provided between the connector assembly 3 and the centrifuge chamber 1, so that the connector assembly 3 can remain stationary when the centrifuge chamber 1 rotates, thus preventing the pipes connected to the connector assembly 3 from becoming entangled. The centrifuge chamber 1 is installed on the centrifuge chamber 1 via a quick-release connector 11 (e.g., a snap-fit ​​connector, a threaded connector, etc.), which facilitates the disassembly, assembly, and replacement of the centrifuge chamber 1. The rotary drive component 2 and the connector assembly 3 can be arranged opposite each other on both sides of the centrifuge chamber 1, resulting in a compact structure and reasonable layout. The rotary drive component 2 can be, for example, a motor. A suction tube 74 can be installed in the suction channel 73, extending to the end of the centrifuge chamber 1. Of course, in other embodiments, the suction tube 74 can also be an integral structure with the suction channel 73.

[0034] The cell separation device has an inlet port 4, a waste outlet port 5, and a suction port 6 on the connector assembly 3, and an inlet channel 71, a waste outlet channel 72, and a suction channel 73 on the guide seat 7. The inlet port 4, waste outlet port 5, and suction port 6 are connected to the inlet channel 71, waste outlet channel 72, and suction channel 73 through the first connecting pipe 81, the second connecting pipe 82, and the third connecting pipe 83, respectively. The inlet, waste outlet, and suction processes in the centrifuge chamber 1 can be carried out simultaneously to achieve continuous centrifugation separation. After the cell culture is completed, the liquid volume will be relatively large. This cell separation device can adapt to the input of large volumes of liquid. For example, the 30ml volume centrifuge chamber 1 can adapt to the input of 50L of liquid containing target cells, and users can reduce the liquid change during the centrifugation process.

[0035] Furthermore, in this embodiment, the axis of the first connecting pipe 81 coincides with the rotation axis of the centrifuge chamber 1, the third connecting pipe 83 is coaxially sleeved on the outer periphery of the first connecting pipe 81 with a gap between them, and the second connecting pipe 82 is coaxially sleeved on the outer periphery of the third connecting pipe 83 with a gap between them. The liquid inlet 4 can be directly connected to the liquid inlet channel 71 through the first connecting pipe 81, the waste discharge 5 can be connected to the waste discharge channel 72 through the gap between the second connecting pipe 82 and the third connecting pipe 83, and the suction 6 can be connected to the suction channel 73 through the gap between the third connecting pipe 83 and the first connecting pipe 81. The overall structure is compact and occupies little space; when rotating with the centrifuge chamber 1, it helps to maintain dynamic force balance and make the equipment work stably.

[0036] Furthermore, in this embodiment, the liquid inlet channel 71 is located on the side of the guide seat 7 near the rotary drive component 2, the waste discharge channel 72 is located on the side of the guide seat 7 near the connector assembly 3, and the suction channel 73 is located between the liquid inlet channel 71 and the waste discharge channel 72, which is conducive to making various liquids and cells flow along the set path and ensuring the cell separation effect.

[0037] In a preferred embodiment, the liquid inlet channel 71, waste outlet channel 72, and suction channel 73 are all provided in multiples and are evenly arranged circumferentially around the rotation axis of the centrifuge chamber 1. Since the axes of the first connecting pipe 81, the second connecting pipe 82, and the third connecting pipe 83 all coincide with the rotation axis of the centrifuge chamber 1, the even arrangement of the liquid inlet channel 71, waste outlet channel 72, and suction channel 73 around the rotation axis is beneficial to maintain the uniform distribution of various liquids and cells in the centrifuge chamber 1, while ensuring that liquids and cells in all directions can be discharged or suctioned, thus guaranteeing the separation effect.

[0038] Furthermore, the liquid inlet 4 and the suction port 6 are located on the side of the connector assembly 3 away from the centrifuge chamber 1, while the waste discharge port 5 is located on the side of the connector assembly 3 close to the centrifuge chamber 1. This helps to shorten the overall length of the first connecting pipe 81, the second connecting pipe 82 and the third connecting pipe 83, and reduce the axial dimensions of the equipment.

[0039] In a preferred embodiment, a sealing element 84 (such as a commonly used sealing ring) is provided between the first connecting pipe 81 and the connector assembly 3, between the second connecting pipe 82 and the connector assembly 3, and between the third connecting pipe 83 and the connector assembly 3. This helps to maintain the sealing and reliability of the equipment and prevent leakage.

[0040] In a preferred embodiment, the centrifuge chamber 1 has conical portions on opposite sides. Of course, in other embodiments, the centrifuge chamber 1 can also adopt a single conical structure, and a counterweight can be set on the side opposite to the conical portion. Correspondingly, the suction channel 73 extends to the tip of the conical portion, which facilitates the aggregation of target cells in the conical portion, makes suction convenient, and ensures the cell separation effect.

[0041] Furthermore, in this embodiment, the inlet port 4 is connected to an input pump 41. The inlet of the input pump 41 is connected to a raw liquid bag 91, a replacement liquid bag 92, and a spare port 93 via an on / off valve 90 (preferably a clamp valve, but other valves that can control the on / off of the pipeline). The waste discharge port 5 is equipped with a first liquid detection component 51 and is connected to a waste liquid bag 97 via an on / off valve 90. The suction port 6 is equipped with a suction pump 61, a second liquid detection component 62, and a color detection component 63. The outlet of the suction pump 61 is connected to a harvest bag 94, a separation liquid bag 95, a buffer solution bag 96, and a waste liquid bag 97 via an on / off valve 90. The stock solution bag 91 is used to hold the liquid containing the target cells, and the replacement bag 92 can be used to hold the buffer solution (such as physiological saline). The spare interface 93 can be used to connect the gas supply component, or to connect the stock solution bag 91 and the replacement bag 92 in an emergency. The first liquid detection component 51 and the second liquid detection component 62 can be, for example, bubble sensors, which can detect the liquid in the pipeline and, together with the color detection component 63, detect red blood cells and white blood cells, thereby switching the operating status of the equipment in a timely manner and improving the automation and intelligence of the equipment. Preferably, the spare interface 93 and each bag are also equipped with bubble sensors. The waste liquid bag 97 is used to collect waste liquid, discarded cells, etc. The harvest bag 94 is used to collect the target cells, the separation liquid bag 95 is used to hold the separation liquid, and the buffer solution bag 96 is used to hold the buffer solution.

[0042] Example 2

[0043] An embodiment of the continuous separation method of the above-mentioned cell separation equipment includes the following steps:

[0044] a) Adding buffer solution: The suction port 6 adds buffer solution into the centrifuge chamber 1 through the third connecting tube 83 and the suction channel 73 until the waste port 5 detects that it is full. Specifically, the buffer solution in the centrifuge chamber 1 flows out to the waste port 5 through the waste channel 72 and the second connecting tube 82, and the first liquid detection component 51 detects the buffer solution. The introduction of buffer solution from the suction port 6 helps to remove the air in the centrifuge chamber 1 and ensures the subsequent cell separation effect.

[0045] b) Adding liquid containing target cells 100: The liquid inlet 4 adds liquid containing target cells 100 into the centrifuge chamber 1 through the first connecting pipe 81 and the liquid inlet channel 71. At the same time, the rotating drive 2 drives the centrifuge chamber 1 to rotate. The cells 100 gather at the end of the centrifuge chamber 1. The supernatant is discharged to the waste outlet 5 through the waste outlet channel 72 and the second connecting pipe 82.

[0046] c) Cell 100 stratification: After all the liquid containing the target cells 100 has been added, the rotation speed of the rotating drive 2 is increased to cause the cells 100 in the centrifuge chamber 1 to stratify.

[0047] d) Cell 100 separation: The suction port 6 suctions the waste cells at the end of the centrifuge chamber 1 to the waste liquid bag 97 through the third connecting tube 83 and the suction channel 73. After the waste cells are suctioned, the port switches to the harvest bag 94 to harvest the target cells until the target cells are harvested. At the same time as the suction port 6 suctions the cells, the liquid inlet port 4 replenishes the centrifuge chamber 1 with buffer or gas.

[0048] Alternatively, dˊ) Target cell washing: Aspiration port 6 aspirates waste cells from the end of centrifuge chamber 1 into waste liquid bag 97. After the waste cells are aspirated, aspiration port 6 adds buffer solution to centrifuge chamber 1 through third connecting tube 83 and aspiration channel 73 to wash the target cells. Waste liquid is collected by waste discharge port 5.

[0049] This continuous separation method enables continuous centrifugation and can accommodate the input of larger volumes of liquid. For example, a 30ml centrifuge chamber 1 can accommodate 50L of liquid containing target cells. Users can reduce the need for liquid changes during centrifugation and can flexibly choose between centrifugation, washing, or concentration processes according to their specific needs.

[0050] Furthermore, in this embodiment,

[0051] In step b, the aspiration port 6 adds separation fluid into the centrifuge chamber 1, which can effectively improve cell separation efficiency and reduce the loss of target cells;

[0052] Step dˊ is followed by:

[0053] e) Concentration: The increased rotation speed of the rotary drive 2 causes the target cells to aggregate. The suction port 6 aspirates the target cells from the centrifuge chamber 1 into the harvest bag 94. Simultaneously, the inlet port 4 replenishes the centrifuge chamber 1 with buffer solution or gas, which further promotes cell aggregation. The suction port 6 can then aspirate a higher concentration of target cells. During concentration, the rotation speed should be controlled to avoid excessively high speeds that could cause cell clumping, hindering cell recovery.

[0054] In a preferred embodiment, the waste cells are red blood cells and the target cells are white blood cells. In step c, the red blood cells are near the end of the centrifuge chamber 1, and the white blood cells are near the rotation axis or guide seat 7 of the centrifuge chamber 1.

[0055] While the present invention has been disclosed above with reference to preferred embodiments, it is not intended to limit the invention. Any person skilled in the art can make many possible variations and modifications to the technical solutions of the present invention, or modify them into equivalent embodiments, without departing from the scope of the present invention. Therefore, any simple modifications, equivalent changes, and modifications made to the above embodiments based on the technical essence of the present invention, without departing from the scope of the present invention, should fall within the protection scope of the present invention.

Claims

1. A cell separation device, comprising a centrifuge chamber (1) and a rotary drive (2) for driving the centrifuge chamber (1) to rotate, characterized in that: The centrifuge chamber (1) is provided with a connector assembly (3), which is connected to an inlet port (4), a waste outlet port (5), and a suction port (6). The centrifuge chamber (1) is provided with a flow guide seat (7), which is provided with an inlet channel (71), a waste outlet channel (72), and a suction channel (73). The inlet port (4) is connected to the inlet channel (71) via a first connecting pipe (81). The waste outlet port (5) is connected to the waste outlet channel (72) via a second connecting pipe (82). The suction port (6) is connected to the suction channel (73) via a third connecting pipe (83). The suction channel (73) extends to the centrifuge chamber. At the end of the centrifuge chamber (1), the axis of the first connecting pipe (81) coincides with the rotation axis of the centrifuge chamber (1), the third connecting pipe (83) is coaxially sleeved on the outer periphery of the first connecting pipe (81) and there is a gap between them, the second connecting pipe (82) is coaxially sleeved on the outer periphery of the third connecting pipe (83) and there is a gap between them, the liquid inlet channel (71) is located on the side of the guide seat (7) near the rotary drive (2), the waste discharge channel (72) is located on the side of the guide seat (7) near the connector assembly (3), and the suction channel (73) is located between the liquid inlet channel (71) and the waste discharge channel (72).

2. The cell separation device according to claim 1, characterized in that: The liquid inlet channel (71), the waste discharge channel (72), and the suction channel (73) are all provided with multiple channels and are evenly arranged circumferentially along the rotation axis of the centrifuge chamber (1).

3. The cell separation device according to claim 1, characterized in that: A sealing element (84) is provided between the first connecting pipe (81) and the connector assembly (3), between the second connecting pipe (82) and the connector assembly (3), and between the third connecting pipe (83) and the connector assembly (3).

4. The cell separation apparatus according to any one of claims 1 to 3, characterized in that: The centrifuge chamber (1) has a conical section on at least one side.

5. The cell separation apparatus according to any one of claims 1 to 3, characterized in that: The inlet port (4) is connected to an input pump (41). The inlet of the input pump (41) is connected to a raw liquid bag (91), a replacement liquid bag (92), and a spare port (93) via a shut-off valve (90). The waste outlet port (5) is equipped with a first liquid detection component (51). The waste outlet port (5) is connected to a waste liquid bag (97) via a shut-off valve (90). The suction port (6) is equipped with a suction pump (61), a second liquid detection component (62), and a color detection component (63). The outlet of the suction pump (61) is connected to a harvest bag (94), a separation liquid bag (95), a buffer solution bag (96), and a waste liquid bag (97) via a shut-off valve (90).

6. A continuous separation method for the cell separation apparatus of claim 5, characterized in that: Including the following steps: a) Add buffer solution: Add buffer solution into the centrifuge chamber (1) through the third connecting tube (83) and the suction channel (73) via the suction port (6) until the waste port (5) detects that it is full; b) Adding liquid containing target cells: The liquid inlet (4) adds liquid containing target cells into the centrifuge chamber (1) through the first connecting tube (81) and the liquid inlet channel (71). At the same time, the rotating drive (2) drives the centrifuge chamber (1) to rotate. The cells (100) gather at the end of the centrifuge chamber (1). The supernatant is discharged to the waste outlet (5) through the waste outlet channel (72) and the second connecting tube (82). c) Cell (100) stratification: After all the liquid containing the target cells has been added, the rotation speed of the rotating drive (2) is increased to cause the cells (100) in the centrifuge chamber (1) to stratify. d) Cell (100) separation: The suction port (6) sucks the waste cells at the end of the centrifuge chamber (1) into the waste liquid bag (97) through the third connecting tube (83) and the suction channel (73). After the waste cells are sucked out, the port switches to the harvest bag (94) to harvest the target cells until the target cells are harvested. At the same time as the suction port (6) sucks out the cells, the liquid inlet port (4) replenishes the centrifuge chamber (1) with buffer or gas. Or, dˊ) Target cell washing: The suction port (6) suctions the waste cells at the end of the centrifuge chamber (1) into the waste liquid bag (97). After the waste cells are suctioned, the suction port (6) adds buffer solution to the centrifuge chamber (1) through the third connecting tube (83) and the suction channel (73) to wash the target cells. The waste discharge port (5) collects the waste liquid.

7. The continuous separation method according to claim 6, characterized in that: In step b), the suction port (6) adds the separation liquid into the centrifuge chamber (1); Step d) is followed by: e) Concentration: The rotation speed of the rotating drive (2) increases, causing the target cells to aggregate. The suction port (6) suctions the target cells in the centrifuge chamber (1) to the harvest bag (94). At the same time as the suction port (6) suctions the target cells, the liquid inlet port (4) replenishes the centrifuge chamber (1) with buffer or gas.

8. The continuous separation method according to claim 6, characterized in that: The discarded cells are red blood cells, and the target cells are white blood cells. In step c), the red blood cells are near the end of the centrifuge chamber (1), and the white blood cells are near the rotation axis of the centrifuge chamber (1).

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