Separation system and separation method

Abstract

A separation system according to the present disclosure is a separation system for separating cells, the separation system comprising a filter provided with a plurality of through-holes and a liquid supply device for supplying a liquid containing a plurality of cells to the filter. The liquid supply device changes the liquid supply pressure for supplying the liquid when the liquid is being supplied to the filter.

Description

Separation system and separation method

[0001] The present disclosure relates to a separation system and a separation method.

[0002] For example, Patent Document 1 discloses a method for separating rare cells in a blood sample. In the method described in Patent Document 1, liquid sending conditions are set so that the pressure applied to the filtration is constant, and the blood sample is filtered using a filter.

[0003] JP 2015-87382 A

[0004] In recent years, there has been a demand for efficient cell separation.

[0005] An object of the present disclosure is to provide a separation system and a separation method for efficiently separating cells.

[0006] A separation system according to one aspect of the present disclosure is a separation system for separating cells, comprising: a filter having a plurality of through holes; and a liquid delivery device that delivers a liquid containing a plurality of cells to the filter, wherein the liquid delivery device changes the liquid delivery pressure at which the liquid is delivered while the liquid is being delivered to the filter.

[0007] A separation method according to one aspect of the present disclosure is a method for separating cells, comprising the steps of: feeding a liquid containing a plurality of cells to a filter; separating the plurality of cells using the filter; and changing a liquid feeding pressure at which the liquid is fed while the liquid is being fed to the filter.

[0008] According to the present disclosure, a separation system and a separation method for efficiently separating cells can be provided.

[0009] FIG. 1 is a schematic block diagram showing the main components of an example of a separation system according to a first embodiment of the present disclosure. FIG. 2 is a schematic diagram illustrating an example of a filter attached to a holder. FIG. 3 is a graph showing an example of a change in liquid delivery pressure in the separation system according to the first embodiment of the present disclosure. FIG. 4 is a flowchart showing an example of an operation of the separation system according to the first embodiment of the present disclosure. FIG. 5 is a schematic diagram illustrating cell separation by the separation system according to the first embodiment of the present disclosure. FIG. 6 is a schematic diagram illustrating cell separation by the separation system according to the first embodiment of the present disclosure. FIG. 7 is a schematic block diagram showing the main components of a separation system of Modification 1. FIG. 8 is a schematic block diagram showing the main components of an example of a separation system according to a second embodiment of the present disclosure. FIG. 9 is a graph showing an example of a change in liquid delivery pressure in the separation system according to the second embodiment of the present disclosure. FIG. 10 is a flowchart showing an example of an operation of the separation system according to the second embodiment of the present disclosure. FIG. 11 is a schematic diagram illustrating cell separation by the separation system according to the second embodiment of the present disclosure. FIG. 12 is a schematic diagram illustrating cell separation by the separation system according to the second embodiment of the present disclosure. Fig. 10 is a flowchart showing the operation of the separation system of Modification 2. Fig. 11 is a schematic diagram showing an example of a filter configuration of the separation system of Embodiment 4 according to the present disclosure. Fig. 12 is a graph showing changes in pressure of the liquid after passing through the filter in Examples 1 and 2.

[0010] (Background to the present disclosure) For example, a method for separating cells using a filter is known in which a liquid containing cells is sent to a filter under a constant liquid sending pressure to separate the cells. For example, in this method, the liquid containing cells is sent to the filter under a constant liquid sending pressure, so that cells that are easily deformed pass through the filter and cells that are less easily deformed are captured by the filter.

[0011] However, the above-mentioned method has a problem in that efficient cell separation becomes difficult when the filter becomes clogged. For example, when the liquid is delivered at a constant delivery pressure, cells that are difficult to deform are captured by the filter and accumulate on the filter. This causes the cells that are difficult to deform to clog the through-holes, preventing cells that are easy to deform from passing through the filter. As a result, the efficiency of cell separation decreases.

[0012] Therefore, the present inventors conducted extensive research and discovered a configuration for feeding a liquid containing cells to a filter while changing the liquid feeding pressure, leading to the present disclosure.

[0013] Hereinafter, embodiments of the present disclosure will be described with reference to the accompanying drawings. In each drawing, elements are exaggerated for ease of explanation.

[0014] First Embodiment <Separation System> FIG. 1 is a schematic block diagram showing a main configuration of an example of a separation system 1A according to a first embodiment of the present disclosure.

[0015] As shown in Fig. 1, the separation system 1A includes a container 10, a filter 20, a liquid delivery device 30, and a collection container 40. The container 10, the filter 20, the liquid delivery device 30, and the collection container 40 are connected by a flow path. In this embodiment, the container 10, the filter 20, the liquid delivery device 30, and the collection container 40 are arranged in this order from upstream. For example, these elements are connected by a tube or a pipe. The liquid delivery device 30 may be arranged between the container 10 and the filter 20.

[0016] The container 10 stores a liquid containing a plurality of cells.

[0017] For example, the cells include induced pluripotent stem cells (iPS cells), ES cells, stem cells, mesenchymal stem cells, mononuclear cells, single cells, cell clumps, floating cells, adhesive cells, nerve cells, leukocytes, cells for regenerative medicine, autologous cells, cancer cells, circulating cancer cells (CTCs), HL-60, HELA, eubacteria, fungi, archaea, and fungi such as yeast. Alternatively, "cells" include substances having a membrane containing lipids and proteins, such as liposomes.

[0018] For example, the liquid may be blood, extracellular fluid, electrolyte solution, cell suspension, or cell culture medium.

[0019] The filter 20 separates a plurality of cells.

[0020] FIG. 2 is a schematic diagram illustrating an example of the filter 20 attached to the holder 22. As shown in FIG.

[0021] 2, the filter 20 has a first main surface PS1 and a second main surface PS2 opposite to the first main surface PS1. The filter 20 has a plurality of through holes 21 that communicate between the first main surface PS1 and the second main surface PS2.

[0022] The through holes 21 may be arranged periodically or randomly as viewed from the first main surface PS1 side, and may have a circular, elliptical, rectangular, or polygonal shape as viewed from the first main surface PS1 side.

[0023] The material constituting the filter 20 may be, for example, primarily composed of at least one of a metal, an oxide, and a polymer. In the case of a metal or metal oxide, the material constituting the filter 20 may be, for example, gold, silver, copper, platinum, nickel, palladium, titanium, aluminum, alloys thereof, and oxides thereof. In the case of a polymer, the material constituting the filter 20 may be, for example, polyethylene or polypropylene. In particular, by using a nickel-palladium alloy, titanium, aluminum, oxides thereof, or polyethylene, the amount of eluate is reduced, thereby reducing the impact on cells.

[0024] The outer shape of the filter 20 may be, for example, circular, elliptical, rectangular, or polygonal.

[0025] The size of the through-holes 21 of the filter 20 is designed appropriately depending on the size, shape, properties, elasticity, or quantity of the cells to be separated. The size of the through-holes 21 is, for example, 0.2 μm or more and 200 μm or less. When the through-holes 21 are circular, the size of the through-holes 21 is the diameter. When the through-holes 21 are elliptical, the size of the through-holes 21 is the length in the minor axis direction. When the through-holes 21 are rectangular, the size of the through-holes 21 is the short side. When the through-holes 21 are polygonal, the size of the through-holes 21 is the diameter of the inscribed circle.

[0026] In this embodiment, the size of the multiple through-holes 21 is smaller than the size of the cells.

[0027] The thickness of the filter 20 is, for example, 1 μm or more and 100 μm or less. Preferably, the thickness of the filter 20 is 1 μm or more and 20 μm or less. More preferably, the thickness of the filter 20 is 1 μm or more and 10 μm or less. With such a configuration, the resistance of the liquid passing through the filter 20 can be reduced, and the processing time can be shortened.

[0028] The filter 20 is held by a holder 22. For example, the holder 22 holds the outer edge of the filter 20.

[0029] The holder 22 has a cylindrical shape. The holder 22 has, for example, a cylindrical shape.

[0030] The holder 22 includes a first holder 23 and a second holder 24 connected to the first holder 23. The filter 20 is held by being sandwiched between the first holder 23 and the second holder 24.

[0031] The liquid delivery device 30 delivers a liquid containing a plurality of cells to the filter 20 .

[0032] The liquid delivery device 30 has a mechanism for varying the liquid delivery pressure. The liquid delivery device 30 changes the liquid delivery pressure while delivering liquid to the filter 20. That is, the liquid delivery device 30 changes the liquid delivery pressure while separating cells using the filter 20. For example, the liquid delivery pressure may be measured by providing a pressure gauge near the downstream of the liquid delivery device 30 or by providing one end of a differential pressure gauge.

[0033] For example, the liquid delivery device 30 includes a pump driven by voltage, such as a tube pump, a syringe pump, or a microblower. For example, the control unit changes the liquid delivery pressure by changing the drive voltage of the pump. The control unit may be included in the liquid delivery device 30, or may be included in an external device separate from the liquid delivery device 30.

[0034] The control unit includes, for example, a memory that stores a program and a processing circuit corresponding to a processor such as a CPU (Central Processing Unit). The functions of the control unit may be configured solely by hardware, or may be realized by combining hardware and software. The control unit realizes predetermined functions by reading data and programs stored in the memory and performing various arithmetic operations.

[0035] In this embodiment, the liquid delivery device 30 aspirates a liquid containing a plurality of cells from the container 10 through the filter 20. The liquid delivery device 30 also supplies the aspirated liquid to the collection container 40.

[0036] FIG. 3 is a graph showing an example of changes in liquid delivery pressure in the separation system 1A according to the first embodiment of the present disclosure.

[0037] As shown in FIG. 3 , the liquid delivery device 30 periodically changes the liquid delivery pressure. For example, the liquid delivery device 30 changes the liquid delivery pressure in a range of 100 Pa to 50 kPa. Preferably, the liquid delivery device 30 changes the liquid delivery pressure in a range of 200 Pa to 25 kPa. More preferably, the liquid delivery device 30 changes the liquid delivery pressure in a range of 500 Pa to 10 kPa. By changing the liquid delivery pressure in this range, the filter 20 can be made to easily elastically deform in the liquid delivery direction. This allows the filter 20 to be moved in the liquid delivery direction and in the opposite direction to the liquid delivery direction to peel off cells adhering to the filter 20.

[0038] For example, the liquid delivery device 30 changes the liquid delivery pressure at a cycle of 0.01 seconds or more and 300 seconds or less. Preferably, the liquid delivery device 30 changes the liquid delivery pressure at a cycle of 0.1 seconds or more and 60 seconds or less. More preferably, the liquid delivery device 30 changes the liquid delivery pressure at a cycle of 1 second or more and 30 seconds or less. By changing the liquid delivery pressure at such a cycle, the filter 20 can be periodically elastically deformed in the liquid delivery direction. This makes it possible to suppress adhesion of cells to the filter 20.

[0039] The collection container 40 is a container for collecting the cells and liquid that have passed through the filter 20 .

[0040] <Operation> Next, an example of the operation of the separation system 1A will be described with reference to FIG. 4 and FIGS. 5A to 5C.

[0041] Fig. 4 is a flowchart showing an example of the operation of the separation system 1A according to the first embodiment of the present disclosure. Fig. 5A to Fig. 5C are schematic diagrams for explaining the separation of cells by the separation system 1A according to the first embodiment of the present disclosure.

[0042] As shown in FIG. 4, the separation system 1A performs steps S1 to S4.

[0043] In step S1, the liquid delivery device 30 delivers a liquid containing a plurality of cells to the filter 20. For example, the control unit applies a drive voltage to the pump of the liquid delivery device 30. This causes the liquid containing a plurality of cells to be delivered to the filter 20.

[0044] In step S2, the plurality of cells is separated by the filter 20. Specifically, the filter 20 separates cells that can pass through the through-holes 21 from the plurality of cells.

[0045] As shown in FIG. 5A, the liquid delivery device 30 delivers a liquid containing a plurality of cells 60A and 60B to the filter 20 and separates the cells.

[0046] For example, cell 60A and cell 60B are cells of the same type but have different characteristics. Specifically, cell 60A and cell 60B have different cytoskeleton structures, cell membrane compositions, and the amounts, shapes, positions, and interactions of intracellular organelles. In this embodiment, cell 60A may be referred to as a first cell 60A, and cell 60B may be referred to as a second cell 60B.

[0047] The cytoskeleton has a fibrous structure composed of three types of proteins: actin filaments, intermediate filaments, and microtubules. The stiffness of a cell varies depending on the thickness, density, and orientation of the fibers, as well as the degree of cross-linking between fibers and other intracellular components.

[0048] For example, cells with unidirectional fiber orientation are easily deformed by forces applied from any direction, whereas cells with multidirectional fiber orientation are less likely to deform when forces are applied from any direction.

[0049] For example, older cells have more developed actin filaments than younger cells, with thicker and denser fibers. As a result, older cells are stiffer and less likely to deform than younger cells. The cell membrane is primarily composed of phospholipids and membrane proteins, and its structure is formed as a phospholipid bilayer. This bilayer structure is known to change due to temperature, pressure, or compositional heterogeneity on the front and back of the membrane. This structural change is known to affect the cell's deformability.

[0050] Intracellular organelles include the endoplasmic reticulum, Golgi apparatus, mitochondria, peroxisomes, ribosomes, endosomes, and lipid droplets. The amount, morphology, location, and interactions of these intracellular organelles are known to change due to cell cycle, cell differentiation, nutritional status, energy demand, genetic mutations, viral infection, temperature sensitivity, aging, cellular stress, metabolic state, fluctuations in lipid metabolism, and immune activation. Changes in the amount, morphology, location, and interactions of intracellular organelles are known to alter the intracellular mechanical properties, thereby affecting the cell's susceptibility to deformation.

[0051] Thus, differences in cytoskeleton structure, cell membrane composition, and the amount, shape, location, and interactions of intracellular organelles result in differences in cell stiffness, i.e., the ease with which cells can be deformed.

[0052] 5A, the second cells 60B are older than the first cells 60A and have thicker and denser fibers, i.e., the second cells 60B are less likely to deform under pressure than the first cells 60A.

[0053] 5A, cells 60A and 60B are separated using a filter 20 having a plurality of through-holes 21 smaller than the size of cells 60A and 60B. A liquid delivery device 30 sets a liquid delivery pressure that allows first cells 60A to deform but does not deform second cells 60B. As a result, first cells 60A pass through the through-holes 21 of the filter 20, but second cells 60B do not pass through the through-holes 21 of the filter 20 and are captured on the first main surface PS1 of the filter 20.

[0054] In this way, by applying a liquid delivery pressure to the filter 20, the first cells 60A pass through the through-holes 21 of the filter 20 while being deformed, and are separated.

[0055] In step S3, the liquid delivery pressure is changed while the liquid delivery device 30 delivers the liquid to the filter 20. That is, the liquid delivery device 30 changes the liquid delivery pressure while separating the first cell 60A. For example, the control unit changes the liquid delivery pressure by changing the drive voltage of the pump of the liquid delivery device 30.

[0056] As the liquid delivery pressure changes, the filter 20 elastically deforms in the liquid delivery direction. For example, when the liquid delivery pressure is relatively high, the filter 20 elastically deforms so as to bend convexly in the liquid delivery direction. When the liquid delivery pressure is relatively low, the load on the filter 20 is smaller than when the liquid delivery pressure is relatively high. As a result, a force acts on the filter 20 to return to its original shape, causing the filter 20 to deform in the direction opposite to the liquid delivery direction. In this way, the load on the filter 20 is changed by changing the pressure, and the elastic deformation of the filter 20 is used to move the filter 20. This allows the second cells 60B attached to the filter 20 to be peeled off.

[0057] 5B, when first cells 60A are separated by filter 20, second cells 60B captured by filter 20 adhere to first main surface PS1 of filter 20. Furthermore, when the liquid delivery pressure is relatively high, filter 20 elastically deforms so as to bend convexly in the liquid delivery direction.

[0058] 5C , when the liquid delivery pressure becomes relatively low, the load on filter 20 decreases, causing filter 20 to deform in an attempt to return to its original shape. That is, filter 20 deforms in the direction opposite to the liquid delivery direction. This causes second cells 60B adhering to first main surface PS1 of filter 20 to peel off from filter 20. As a result, first cells 60A can more easily pass through through-holes 21.

[0059] In this embodiment, the liquid delivery device 30 periodically changes the liquid delivery pressure, thereby periodically elastically deforming the filter 20 in the liquid delivery direction. That is, by periodically changing the liquid delivery pressure, the liquid delivery device 30 periodically moves the filter 20 in the liquid delivery direction and the direction opposite to the liquid delivery direction. This makes it possible to suppress adhesion of the second cells 60B to the first main surface PS1 of the filter 20.

[0060] In step S4, the first cells 60A separated by the filter 20 are collected. For example, the first cells 60A and the liquid that have passed through the filter 20 are sent to the collection container 40 by the liquid delivery device 30 and collected in the collection container 40.

[0061] The second cells 60B captured by the filter 20 may also be collected.

[0062] In this way, the separation system 1A performs steps S1 to S4 to separate and recover cells 60A and 60B having different characteristics.

[0063] <Effects> According to the separation system 1A according to the first embodiment, the following effects can be achieved.

[0064] Separation system 1A is a separation system that separates cells 60A and 60B, and includes a filter 20 and a liquid delivery device 30. Filter 20 has a plurality of through-holes 21. Liquid delivery device 30 delivers a liquid containing a plurality of cells 60A and 60B to filter 20. Furthermore, liquid delivery device 30 changes the liquid delivery pressure at which the liquid is delivered to filter 20 while the liquid is being delivered to filter 20.

[0065] This configuration allows efficient separation of cells 60A and 60B. For example, by changing the liquid feed pressure, the filter 20 can be elastically deformed in the liquid feed direction. For example, by changing the liquid feed pressure applied to the filter 20, the filter 20 can be moved in the liquid feed direction and in the opposite direction to the liquid feed direction. This allows cells 60B captured and attached to the filter 20 to be detached, preventing clogging of the filter 20. As a result, cells 60A can more easily pass through the through-holes 21 of the filter 20, shortening the separation time. In addition, the amount of liquid fed can be reduced.

[0066] The liquid delivery device 30 periodically changes the liquid delivery pressure. This configuration allows the filter 20 to periodically elastically deform in the liquid delivery direction. For example, the filter 20 can be periodically vibrated. This makes it easier to detach cells 60B adhering to the filter 20, and allows cells 60A to more easily pass through the through-holes 21 of the filter 20. As a result, the cells 60A and 60B can be separated more efficiently. Furthermore, the pressure loss of the filter 20 relative to the fluid can be reduced, and deformation of the cells can be minimized, allowing the cells 60A and 60B to be separated more accurately.

[0067] The liquid delivery device 30 changes the liquid delivery pressure within a range of 100 Pa to 50 kPa. This configuration allows the filter 20 to easily deform elastically in the liquid delivery direction. This makes it easier to peel off cells 60B adhering to the filter 20 by moving the filter 20 in the liquid delivery direction and the direction opposite to the liquid delivery direction.

[0068] The size of the through-holes 21 is smaller than the size of the cells 60A, 60B. This configuration makes it possible to easily separate cells with different characteristics from the plurality of cells 60A, 60B. For example, if the cytoskeletons of the cells 60A, 60B passing through the through-holes 21 are different, a difference in elastic modulus occurs between the cells, which changes the ease with which they pass through the through-holes 21, making it possible to easily separate cells with different cytoskeletons. This allows cells with different characteristics to be separated without chemically treating the cells, thereby shortening the work time and reducing stress on the cells.

[0069] The filter 20 is mainly composed of at least one of a metal, an oxide, and a polymer. This configuration reduces the amount of elution and the impact on cells, allowing for more efficient separation of the cells 60A and 60B.

[0070] In the present embodiment, an example has been described in which the separation system 1A includes the container 10, the filter 20, the liquid delivery device 30, and the collection container 40, but the present invention is not limited to this. For example, the separation system 1A does not have to include the container 10 and the collection container 40. For example, the container 10 and the collection container 40 may be included in a device separate from the separation system 1A.

[0071] In this embodiment, an example has been described in which the cells 60A and 60B are cells of the same type, but this is not intended to be limiting. For example, the cells 60A and 60B may be cells of different types. Furthermore, in this embodiment, the cytoskeleton has been used as an example of a microscopic feature, but this is not intended to be limiting. For example, the amount, shape, position, and interaction of intracellular organelles such as the nucleus, mitochondria, endoplasmic reticulum, endosome, and Golgi apparatus may also be used.

[0072] In this embodiment, an example has been described in which the size of the through-holes 21 of the filter 20 is smaller than the size of the cells 60A and 60B, but this is not limiting. For example, the size of the through-holes 21 of the filter 20 may be larger than the size of the cells 60A. For example, if the liquid delivery pressure applied to the filter 20 causes the cells 60A to deform and make it difficult for them to pass through the through-holes 21, the liquid delivery pressure can be changed to move the filter 20 in the liquid delivery direction and in the opposite direction to the liquid delivery direction. This makes it easier for the cells 60A to pass through the through-holes 21 of the filter 20.

[0073] In the present embodiment, an example in which the liquid delivery device 30 is disposed between the filter 20 and the collection container 40 has been described, but the present invention is not limited to this. For example, the liquid delivery device 30 may be disposed between the container 10 and the filter 20. Such a configuration widens the range of pressure application, enabling processing in a shorter time.

[0074] In the present embodiment, an example has been described in which the liquid delivery device 30 periodically changes the liquid delivery pressure, but the present invention is not limited to this. For example, the liquid delivery device 30 may randomly change the liquid delivery pressure.

[0075] In this embodiment, an example in which both the separated cells 60A and 60B are recovered has been described, but the present invention is not limited to this. For example, the cells 60A may be recovered without recovering the cells 60B. Alternatively, the cells 60A may not be recovered, but the cells 60B may be recovered.

[0076] <Modification 1> FIG. 6 is a schematic block diagram showing the main configuration of a separation system 1AA according to Modification 1. As shown in FIG.

[0077] 6 , separation system 1AA includes a first collection container 40 for collecting first cells 60A, a second collection container 41 for collecting second cells 60B, and a collection liquid container 50 for storing a collection liquid. Separation system 1AA also includes a first switching valve V1 that switches between a flow path FL1 connecting container 10 and filter 20 and a flow path FL2 connecting filter 20 and second collection container 41. Separation system 1AA also includes a second switching valve V2 that switches between a flow path FL10 connecting liquid delivery device 30 and first collection container 40 and a flow path FL20 connecting liquid delivery device 30 and collection liquid container 50.

[0078] In the separation system 1AA, the recovery liquid stored in the recovery liquid container 50 is sent to the second recovery container 41 via the filter 20, thereby recovering the second cells 60B captured by the filter 20.

[0079] 4, the first switching valve V1 opens the flow path FL1 connecting the container 10 and the filter 20, and closes the flow path FL2 connecting the filter 20 and the second collection container 41. The second switching valve V2 opens the flow path FL10 connecting the liquid delivery device 30 and the first collection container 40, and closes the flow path FL20 connecting the liquid delivery device 30 and the collection liquid container 50.

[0080] 6, the liquid delivery device 30 delivers the liquid containing the cells 60A and 60B stored in the container 10 through the flow path FL1 to the filter 20. The liquid containing the first cells 60A that has passed through the filter 20 passes through the flow path FL10 and is collected in the first collection container 40.

[0081] 4, the first switching valve V1 closes the flow path FL1 connecting the container 10 and the filter 20, and opens the flow path FL2 connecting the filter 20 and the second collection container 41. The second switching valve V2 closes the flow path FL10 connecting the liquid delivery device 30 and the first collection container 40, and opens the flow path FL20 connecting the liquid delivery device 30 and the collection liquid container 50. The liquid delivery device 30 reverses the liquid delivery direction.

[0082] 6 , the liquid delivery device 30 delivers the recovery liquid stored in the recovery liquid container 50 through the flow path FL20 to the filter 20. As a result, the second cells 60B captured on the filter 20 are detached from the filter 20 and are collected in the second collection container 41 via the flow path FL2 together with the recovery liquid.

[0083] The first switching valve V1 and the second switching valve V2 may be controlled by a control unit, or may be manually switched by a user.

[0084] This configuration makes it possible to easily collect the second cells 60B captured by the filter 20. Furthermore, the liquid in which the second cells 60B are cultured can be easily replaced.

[0085] (Embodiment 2) A separation system according to embodiment 2 of the present disclosure will be described. Note that in embodiment 2, components that are the same as or equivalent to those in embodiment 1 will be denoted by the same reference numerals, and descriptions that overlap with embodiment 1 will be omitted.

[0086] Fig. 7 is a schematic block diagram showing a main configuration of an example of a separation system 1B according to a second embodiment of the present disclosure. Fig. 8 is a graph showing an example of a change in liquid delivery pressure in the separation system 1B according to the second embodiment of the present disclosure.

[0087] The second embodiment differs from the first embodiment in that the separation system 1B includes a first collection container 40, a second collection container 41, and a switching valve V10. The second embodiment also differs from the first embodiment in that the liquid delivery device 30 changes the liquid delivery pressure between a first pressure and a second pressure to separate the first cell 60A, the second cell 60B, and the third cell 60C.

[0088] As shown in FIG. 7, the separation system 1B includes a first collection container 40, a second collection container 41, and a switching valve V10.

[0089] First collection container 40 is a container that collects first cells 60A that have passed through filter 20. First collection container 40 is similar to collection container 40 in the first embodiment.

[0090] The second collection container 41 is a container for collecting the second cells 60B that have passed through the filter 20.

[0091] The switching valve V10 switches between a flow path FL11 that connects the filter 20 and the first collection container 40 and a flow path FL12 that connects the filter 20 and the second collection container 41.

[0092] The switching valve V10 may be controlled by a control unit or may be manually switched by a user.

[0093] In this embodiment, the size of the through-holes 21 of the filter 20 is smaller than the size of the first to third cells 60A, 60B, and 60C. Furthermore, the first to third cells 60A, 60B, and 60C are cells of the same type, but each have a different cytoskeletal structure. For example, the second cell 60B is older than the first cell 60A, and the third cell 60C is older than the second cell 60B. That is, the second cell 60B is harder and less deformable than the first cell 60A, and the third cell 60C is harder and less deformable than the second cell 60B.

[0094] As shown in FIG. 8, the liquid delivery device 30 changes the liquid delivery pressure between a first pressure P1 and a second pressure P2 higher than the first pressure P1.

[0095] The first pressure P1 is a pressure at which the first cell 60A is deformed and can pass through the through-holes 21 of the filter 20. At the first pressure P1, the second cell 60B and the third cell 60C do not deform and cannot pass through the through-holes 21 of the filter 20.

[0096] The second pressure P2 is a pressure at which the second cells 60B deform and can pass through the through-holes 21 of the filter 20. At the second pressure P2, the third cells 60C do not deform and cannot pass through the through-holes 21 of the filter 20. For example, the second pressure P2 is 1.1 to 10 times the first pressure P1. Preferably, the second pressure P2 is 1.2 to 5 times the first pressure P1. More preferably, the second pressure P2 is 1.5 to 2 times the first pressure P1.

[0097] <Operation> An example of the operation of the separation system 1B will be described with reference to FIGS. 9, 10A, and 10B.

[0098] 9 is a flowchart showing an example of the operation of the separation system 1B according to the second embodiment of the present disclosure. FIG. 10A and FIG. 10B are schematic diagrams for explaining the separation of cells 60A to 60C by the separation system 1B according to the second embodiment of the present disclosure.

[0099] As shown in FIG. 9, the separation system 1B performs steps S11 to S16.

[0100] In step S11, the liquid delivery device 30 delivers the liquid containing the cells 60A to 60C at a first pressure P1 to the filter 20. At this time, the switching valve V10 opens the flow path FL11 and closes the flow path FL12.

[0101] In step S12, the first cells 60A are separated by the filter 20.

[0102] 10A , while the liquid delivery device 30 delivers liquid at the first pressure P1, the first cell 60A deforms and passes through the through-hole 21 of the filter 20. On the other hand, the second cell 60B and the third cell 60C do not deform and therefore cannot pass through the through-hole 21, and are captured by the filter 20.

[0103] In step S13, the first cells 60A that have passed through the filter 20 are collected by the first collection container 40. The first cells 60A that have passed through the filter 20 are supplied to and collected in the first collection container 40 through the flow path FL11 together with the liquid.

[0104] In step S14, the liquid delivery device 30 changes the liquid delivery pressure from the first pressure P1 to the second pressure P2. For example, after separating and recovering the first cell 60A, the liquid delivery device 30 increases the liquid delivery pressure to the second pressure P2. At this time, the switching valve V10 closes the flow path FL11 and opens the flow path FL12.

[0105] In step S15, the second cells 60B are separated by the filter 20.

[0106] 10B , while the liquid delivery device 30 delivers the liquid at the second pressure P2, the second cell 60B deforms and passes through the through-hole 21 of the filter 20. On the other hand, the third cell 60C does not deform and therefore cannot pass through the through-hole 21, and is captured by the filter 20.

[0107] In step S16, second cells 60B that have passed through filter 20 are collected by second collection container 41. Second cells 60B that have passed through filter 20 are supplied to second collection container 41 together with the liquid through flow path FL12 and collected.

[0108] The third cells 60C captured by the filter 20 may also be collected.

[0109] <Effects> According to the separation system 1B according to the second embodiment, the following effects can be achieved.

[0110] In the separation system 1B, the liquid delivery device 30 changes the liquid delivery pressure between a first pressure P1 and a second pressure P2 higher than the first pressure P1. This configuration allows efficient separation of the cells 60A to 60C. By changing the liquid delivery pressure between the first pressure P1 and the second pressure P2, the cells 60A to 60C having different characteristics can be efficiently separated.

[0111] Separation system 1B includes a first collection container 40, a second collection container 41, flow paths FL11 and FL12, and a switching valve V10. First collection container 40 collects first cells 60A that have passed through filter 20. Second collection container 41 collects second cells 60B that have passed through filter 20. Flow path FL11 supplies a liquid containing first cells 60A that have passed through filter 20 to first collection container 40. Flow path FL12 supplies a liquid containing second cells 60B that have passed through filter 20 to second collection container 41. Switching valve V10 switches between flow paths FL11 and FL12. This configuration allows for easy collection of separated first cells 60A and second cells 60B.

[0112] In this embodiment, the cells 60A to 60C are the same type of cells, but the present invention is not limited to this. For example, the cells 60A to 60C may be different types of cells.

[0113] In the present embodiment, an example has been described in which the size of the through-holes 21 of the filter 20 is smaller than the size of the cells 60A to 60C, but the present invention is not limited to this. For example, the size of the through-holes 21 of the filter 20 may be larger than the size of the cell 60A.

[0114] In the present embodiment, an example has been described in which the separation system 1B includes the first collection container 40 and the second collection container 41, but the present invention is not limited to this. For example, the separation system 1B may include a single collection container. In this case, the collection container may be replaceable.

[0115] In the present embodiment, an example has been described in which the liquid delivery device 30 increases the liquid delivery pressure from the first pressure P1 to the second pressure P2 after separating and collecting the first cell 60A. However, this is not limiting. For example, the liquid delivery device 30 may alternately apply the first pressure P1 and the second pressure P2. For example, while the liquid delivery device 30 is separating the first cell 60A at the first pressure P1, the second cell 60B may be captured and deposited on the filter 20. In this case, the liquid delivery device 30 may change the liquid delivery pressure to the second pressure P2 and pass the second cell 60B captured on the filter 20 through the through-hole 21 of the filter 20. At this time, the switching valve V10 may open the flow path FL12 and close the flow path FL11, and collect the second cell 60B in the second collection container 41.

[0116] In this embodiment, the third cells 60C may be collected by sending the collection liquid from the collection liquid container 50 to the filter 20, as in the separation system 1AA of Modification 1 shown in FIG.

[0117] (Embodiment 3) A separation system according to embodiment 3 of the present disclosure will be described. Note that in embodiment 3, components that are the same as or equivalent to those in embodiment 1 will be denoted by the same reference numerals, and descriptions that overlap with embodiment 1 will be omitted.

[0118] FIG. 10 is a schematic block diagram illustrating a main configuration of an example of a separation system 1C according to the third embodiment of the present disclosure.

[0119] The third embodiment differs from the first embodiment in that a separation system 1C includes a detection device 31 and an output device 32.

[0120] As shown in FIG. 10, the separation system 1C includes a detection device 31 and an output device 32.

[0121] The detection device 31 detects information related to the pressure applied to the filter 20. The information related to the pressure applied to the filter 20 is information that changes in relation to the liquid delivery pressure. In this embodiment, the information related to the pressure applied to the filter 20 is a drive voltage that drives a pump included in the liquid delivery device 30. The detection device 31 is a voltmeter that detects the drive voltage of the pump of the liquid delivery device 30.

[0122] The output device 32 outputs information relating to the state of the filter 20. Information relating to the state of the filter 20 includes information on the clogging state of the filter 20, the timing of replacing the filter 20, damage to the filter 20, and the like.

[0123] The output device 32 outputs information about the state of the filter 20 based on the information detected by the detection device 31 .

[0124] The output device 32 is, for example, a display, a speaker, or an LED. The display displays information about the state of the filter 20 as visual information. The speaker displays information about the state of the filter 20 as audio information. The LED displays information about the state of the filter 20 by, for example, lighting up a red LED.

[0125] The detection device 31 and the output device 32 are controlled by a control unit.

[0126] FIG. 12 is a graph showing an example of a change in voltage detected by the detection device 31. In FIG.

[0127] 12, when the filter 20 becomes clogged, the pressure around the filter 20 increases. When the pressure around the filter 20 increases, the drive voltage of the pump of the liquid delivery device 30 also increases. In other words, the pressure around the filter 20 and the drive voltage are proportional to each other.

[0128] Therefore, the clogging state of the filter 20 can be known based on the driving voltage of the pump of the liquid delivery device 30.

[0129] The control unit causes the output device 32 to output information relating to the state of the filter 20 based on the drive voltage detected by the detection device 31. For example, when the drive voltage detected by the detection device 31 becomes equal to or greater than a predetermined threshold value VS1, the control unit generates information relating to the state of the filter 20 and causes the output device 32 to output the information relating to the state of the filter 20.

[0130] <Operation> An example of the operation of the separation system 1C will be described with reference to FIG.

[0131] FIG. 13 is a flowchart showing an example of the operation of the separation system 1C according to the third embodiment of the present disclosure.

[0132] 13, the separation system 1C performs steps S21 to S23 in addition to steps S1 to S4 of embodiment 1. Specifically, the separation system 1C performs steps S21 to S23 after steps S1 to S3.

[0133] In step S21, the detection device 31 detects information about the pressure applied to the filter 20. In this embodiment, the detection device 31 detects the drive voltage of the pump of the liquid delivery device 30.

[0134] In step S22, the output device 32 outputs information about the state of the filter 20 based on information about the pressure acting on the filter 20. In this embodiment, when the drive voltage detected by the detection device 31 becomes equal to or greater than the threshold value VS1, the control unit causes the output device 32 to output a warning that the filter 20 is clogged or information suggesting filter replacement.

[0135] <Effects> According to the separation system 1C according to the third embodiment, the following effects can be achieved.

[0136] The separation system 1C includes a detection device 31 that detects information related to the pressure applied to the filter 20. With this configuration, it is possible to detect information related to the pressure applied to the filter 20. For example, based on the information related to the pressure applied to the filter 20, it is possible to obtain information related to the state of the filter, such as the clogging state of the filter 20 and the timing for replacing the filter 20.

[0137] The liquid delivery device 30 includes a pump that is driven by a voltage, and the detection device 31 detects the voltage. With this configuration, information about the state of the filter can be easily obtained based on the voltage of the pump.

[0138] The separation system 1C includes an output device 32 that outputs information. The output device 32 outputs information about the state of the filter 20 based on the information detected by the detection device 31. With this configuration, the user can easily know the clogging state of the filter 20 and the replacement timing.

[0139] In this embodiment, an example in which the detection device 31 is a voltmeter has been described, but this is not limiting. For example, the detection device 31 may be a flow meter. In this case, the information regarding the pressure applied to the filter 20 is the flow rate of the liquid flowing through the filter 20. The detection device 31 may also be an ammeter. In this case, the information regarding the pressure applied to the filter 20 is the driving current of the pump of the liquid delivery device 30.

[0140] The information about the pressure applied to the filter 20 may also be a resistance change, a transit time difference, or a temperature difference. The detection device 31 may be a pressure gauge, a differential pressure gauge, or a flow meter.

[0141] In the present embodiment, an example has been described in which the separation system 1C includes the output device 32, but the present invention is not limited to this. For example, the separation system 1C may not include the output device 32. Furthermore, the separation system 1C may control the liquid delivery device 30 based on information regarding the pressure applied to the filter 20.

[0142] <Modification 2> FIG. 14 is a flowchart showing the operation of the separation system of Modification 2.

[0143] As shown in FIG. 14, in the separation system of the second modification, steps S21, S24 and S25 are performed in addition to steps S1 to S4 of the first embodiment.

[0144] In step S21, the detection device 31 detects the drive voltage of the liquid delivery device 30. Specifically, the detection device 31 detects the drive voltage of the pump of the liquid delivery device 30.

[0145] In step S24, the control unit determines whether the drive voltage detected by the detection device 31 is equal to or greater than the threshold value VS1.

[0146] If the drive voltage is equal to or greater than the threshold value VS1, the flow proceeds to step S25. If the drive voltage is less than the threshold value VS1, the flow proceeds to step S4.

[0147] In step S25, the liquid delivery device 30 stops delivering the liquid.

[0148] With this configuration, the liquid delivery by the liquid delivery device 30 can be stopped based on the voltage that drives the liquid delivery device 30. This makes it possible to prevent the filter 20 from becoming clogged and being damaged.

[0149] In the second modification, an example in which the liquid delivery device 30 stops delivering liquid in step S25 has been described, but this is not limiting. For example, in step S25, the liquid delivery device 30 may change the liquid delivery direction. For example, the liquid delivery device 30 may deliver liquid in the opposite direction to detach cells adhering to the filter 20.

[0150] (Embodiment 4) A separation system according to embodiment 4 of the present disclosure will be described. Note that in embodiment 4, components that are the same as or equivalent to those in embodiment 1 will be denoted by the same reference numerals, and descriptions that overlap with embodiment 1 will be omitted.

[0151] FIG. 15 is a schematic diagram illustrating an example of a filter configuration of a separation system according to a fourth embodiment of the present disclosure.

[0152] The fourth embodiment differs from the first embodiment in that the separation system includes two filters 20A and 20B.

[0153] As shown in FIG. 15, the separation system includes a first filter 20A and a second filter 20B.

[0154] The first filter 20A has a plurality of through-holes 21A. The first filter 20A is disposed near the inlet. The inlet is an inlet through which the liquid containing cells flows into the holder 22. For example, the inlet is provided at an end of the holder 22.

[0155] The second filter 20B has a plurality of through holes 21B. The second filter 20B is disposed farther from the inlet than the first filter 20A. That is, the second filter 20B is disposed downstream of the first filter 20A. The second filter 20B is disposed with a distance L1 between it and the first filter 20A.

[0156] For example, the distance L1 between the first filter 20A and the second filter 20B is 1 mm or more and 200 mm or less. Preferably, the distance L1 is 10 mm or more and 100 mm or less. More preferably, the distance L1 is 20 mm or more and 50 mm or less.

[0157] The through holes 21B of the second filter 20B are smaller than the through holes 21A of the first filter 20A. For example, the size of the through holes 21B is 0.1 to 0.9 times the size of the through holes 21A. Preferably, the size of the through holes 21B is 0.2 to 0.8 times the size of the through holes 21A. More preferably, the size of the through holes 21B is 0.3 to 0.5 times the size of the through holes 21A.

[0158] Between the first filter 20A and the second filter 20B, a region R1 is provided, which reduces the pressure change of the liquid that has passed through the first filter 20A.

[0159] FIG. 16 is a graph showing the change in pressure of the liquid after passing through the filters 20, 20A, and 20B in Examples 1 and 2.

[0160] Example 1 is a separation system having the filter configuration of the first embodiment, and Example 2 is a separation system having the filter configuration of the fourth embodiment.

[0161] As shown in FIG. 16, in Example 2, the pressure change after passing through the filter is more gradual than in Example 1.

[0162] <Effects> The separation system according to the fourth embodiment can provide the following effects.

[0163] In the separation system of the fourth embodiment, the filters include a first filter 20A located near the inlet and a second filter 20B located farther from the inlet than the first filter 20A. This configuration allows the pressure change of the liquid after passing through the filters to be gradual, thereby reducing damage to the cells.

[0164] The through-holes 21A of the first filter 20A are smaller than the through-holes 21B of the second filter 20B. With this configuration, cells that have passed through the through-holes 21A can pass through the through-holes 21B at low pressure due to the effect of the first filter 20A, so that the cells can be separated based on the difference in cell size.

[0165] The distance L1 between the first filter 20A and the second filter 20B is 1 mm or more and 200 mm or less. This configuration makes it possible to more gently change the pressure of the liquid after passing through the filters, thereby further reducing damage to the cells.

[0166] In the present embodiment, the through-holes 21A of the first filter 20A are smaller than the through-holes 21B of the second filter 20B. However, the present invention is not limited to this. For example, the size of the through-holes 21A may be the same as the size of the through-holes 21B.

[0167] As described above, the above embodiments have been described as examples of the technology disclosed in this application. However, the technology in this disclosure is not limited to these embodiments and can be applied to embodiments in which modifications, substitutions, additions, omissions, etc. are made as appropriate. Furthermore, these general and specific aspects may be realized by an apparatus, a system, a method, a computer program, a computer-readable storage medium, or a combination thereof.

[0168] As used herein, terms such as "first," "second," etc. are used for descriptive purposes only and should not be understood as expressing or implying relative importance or ranking of technical features. Features qualified as "first" and "second" expressly or imply the inclusion of one or more of that feature.

[0169] Although the present disclosure has been fully described in connection with the preferred embodiments with reference to the accompanying drawings, various changes and modifications will be apparent to those skilled in the art, and such changes and modifications are to be understood as being included within the scope of the present disclosure as defined by the appended claims unless they depart therefrom.

[0170] (Summary of embodiment) (1) A separation system according to one aspect of the present disclosure is a separation system for separating cells, comprising a filter having a plurality of through-holes and a liquid delivery device that delivers a liquid containing a plurality of cells to the filter, and the liquid delivery device changes the liquid delivery pressure at which the liquid is delivered while the liquid is being delivered to the filter.

[0171] (2) In the separation system of (1), the liquid delivery device may periodically change the liquid delivery pressure.

[0172] (3) In the separation system of (1) or (2), the liquid delivery device may change the liquid delivery pressure between a first pressure and a second pressure higher than the first pressure.

[0173] (4) In the separation system of any one of (1) to (3), the liquid delivery device may change the liquid delivery pressure in a range of 100 Pa or more and 50 kPa or less.

[0174] (5) The separation system of any one of (1) to (4) may further include a detection device that detects information regarding the pressure applied to the filter.

[0175] (6) In the separation system of (5), the liquid delivery device may include a pump driven by a voltage, and the detection device may detect the voltage.

[0176] (7) The separation system of (5) or (6) may further include an output device that outputs information, and the output device may output information regarding the state of the filter based on the information detected by the detection device.

[0177] (8) In any one of the separation systems (1) to (7), the filters may include a first filter close to the inlet and a second filter positioned farther from the inlet than the first filter.

[0178] (9) In the separation system of (8), the through-holes of the first filter may be smaller than the through-holes of the second filter.

[0179] (10) Any one of the separation systems (1) to (9) may further include a first collection container that collects first cells that have passed through the filter, a second collection container that collects second cells that have passed through the filter, a first flow path that supplies a liquid containing the first cells that have passed through the filter to the first collection container, a second flow path that supplies a liquid containing the second cells that have passed through the filter to the second collection container, and a switching valve that switches between the first flow path and the second flow path.

[0180] (11) In the separation system of any one of (1) to (10), the size of the plurality of through-holes may be smaller than the size of the plurality of cells.

[0181] (12) In the separation system according to any one of (1) to (11), the filter may be mainly composed of at least one of a metal, a metal oxide, and a polymer.

[0182] (13) A separation method according to one aspect of the present disclosure is a method for separating cells, comprising the steps of: feeding a liquid containing a plurality of cells to a filter; separating the plurality of cells using the filter; and varying a feeding pressure for feeding the liquid while feeding the liquid to the filter.

[0183] (14) In the separation method of (13), the step of changing the liquid sending pressure may include periodically changing the liquid sending pressure.

[0184] (15) In the separation method of (13) or (14), the step of changing the liquid supply pressure may include changing the liquid supply pressure between a first pressure and a second pressure higher than the first pressure.

[0185] (16) In any one of the separation methods (13) to (15), the method may further include a step of detecting information regarding the pressure applied to the filter.

[0186] (17) The separation method of (16) may further include a step of outputting information about the filter based on the detected information.

[0187] (18) In any one of the separation methods (13) to (17), the filter may include a first filter close to the inlet and a second filter positioned farther from the inlet than the first filter.

[0188] (19) In the separation system of (18), the size of the through-holes of the first filter may be smaller than the size of the through-holes of the second filter.

[0189] (20) In the separation method according to any one of (13) to (19), the size of the plurality of through-holes may be smaller than the size of the plurality of cells.

[0190] The separation systems and methods of the present disclosure are useful for cell separation applications.

[0191] 1A, 1AA, 1B, 1C Separation system 10 Container 20 Filter 21 Through-hole 22 Holder 23 First holder 24 Second holder 30 Liquid delivery device 31 Detection device 32 Output device 40 Collection container (first collection container) 41 Collection container (second collection container) 50 Collection liquid container 60A Cell (first cell) 60B Cell (second cell) 60C Cell (third cell)

Claims

1. A separation system for separating cells, comprising: a filter having a plurality of through holes; and a liquid delivery device that delivers a liquid containing a plurality of cells to the filter, wherein the liquid delivery device changes the liquid delivery pressure at which the liquid is delivered while the liquid is being delivered to the filter.

2. The separation system according to claim 1, wherein the liquid delivery device periodically changes the liquid delivery pressure.

3. The separation system according to claim 1 or 2, wherein the liquid delivery device changes the liquid delivery pressure between a first pressure and a second pressure higher than the first pressure.

4. The separation system according to any one of claims 1 to 3, wherein the liquid delivery device changes the liquid delivery pressure in the range of 100 Pa or more and 50 kPa or less.

5. The separation system according to any one of claims 1 to 4, further comprising a detection device for detecting information relating to the pressure applied to the filter.

6. The separation system according to claim 5, wherein the liquid delivery device includes a pump driven by a voltage, and the detection device detects the voltage.

7. The separation system according to claim 5 or 6, further comprising an output device that outputs information, wherein the output device outputs information about the state of the filter based on the information detected by the detection device.

8. A separation system according to any one of claims 1 to 7, wherein the filters include a first filter located closer to the inlet and a second filter located farther from the inlet than the first filter.

9. The separation system according to claim 8, wherein the through holes of the first filter are smaller than the through holes of the second filter.

10. The separation system according to any one of claims 1 to 9, further comprising: a first collection container that collects first cells that have passed through the filter; a second collection container that collects second cells that have passed through the filter; a first flow path that supplies a liquid containing the first cells that have passed through the filter to the first collection container; a second flow path that supplies a liquid containing the second cells that have passed through the filter to the second collection container; and a switching valve that switches between the first flow path and the second flow path.

11. The separation system according to any one of claims 1 to 10, wherein the size of the plurality of through-holes is smaller than the size of the plurality of cells.

12. The separation system according to any one of claims 1 to 11, wherein the filter is primarily composed of at least one of a metal, a metal oxide, and a polymer.

13. A separation method for separating cells, comprising the steps of: feeding a liquid containing a plurality of cells to a filter; separating the plurality of cells using the filter; and changing a liquid feeding pressure for feeding the liquid while feeding the liquid to the filter.

14. The separation method according to claim 13, wherein the step of changing the liquid supply pressure comprises periodically changing the liquid supply pressure.

15. The separation method according to claim 13 or 14, wherein the step of changing the liquid supply pressure comprises changing the liquid supply pressure between a first pressure and a second pressure higher than the first pressure.

16. The separation method according to any one of claims 13 to 15, further comprising the step of detecting information relating to the pressure on the filter.

17. The separation method of claim 16, further comprising the step of outputting information about the filter based on the detected information.

18. A separation method according to any one of claims 13 to 17, wherein the filters include a first filter located closer to the inlet and a second filter located farther from the inlet than the first filter.

19. The separation method according to claim 18, wherein the size of the through holes of the first filter is smaller than the size of the through holes of the second filter.

20. The separation method according to any one of claims 13 to 19, wherein the size of the plurality of through-holes is smaller than the size of the plurality of cells.

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