Cell stripping system and cell culture system

Abstract

The invention relates to a cell dissection system and a cell culture system. Provided is a cell dissecting system, comprising: a magnetic scraper for dissecting cells attached to a culture surface; a magnetic field applying unit configured to apply a magnetic field to the blade; and a blade position control unit configured to control a position of the blade by controlling the magnetic field application unit, in which the blade is configured to move while rotating in a state in which the magnetic field is not applied, in a state in which the magnetic field is applied by the magnetic field application means, the blade is moved by the blade position control means without rotating the blade, thereby peeling the adhered cells.

Description

Technical Field

[0001] The invention relates to a cell detachment system and a cell culture system. Background Art

[0002] In the culture of adherent cells, there is a step called cell stripping. Usually, when stripping, a method is adopted in which the cell-to-cell attachment is weakened by an enzyme and then the cells are physically stripped by using a scraper or the like. In addition, cell culture needs to be carried out in a clean area to prevent bacteria and other sample cells from entering. In fact, a variety of systems have been developed in which, in a clean area, cells are sown in a culture container, and a culture medium is supplied to and discharged from the culture container. In order to strip cells in such a system, a scraper needs to be inserted into the container and the culture surface is scraped. However, the cells, the scraper, and the drive system that holds and operates the scraper exist in the same space, so that even for the drive system, a clean area needs to be prepared. This is a factor that hinders cost reduction.

[0003] To solve this problem, as disclosed in Japanese Patent Application Laid-Open No. 2004-129558, a system has been developed in which a triangular prism scraper made of a magnetic substance is installed in a closed container, and the scraper is driven from the outside of the closed container using magnetic force, thereby peeling off cells by the scraper while keeping the inside of the container clean.

[0004] In Japanese Patent Application Laid-Open No. 2004-129558, a simple culture system is implemented, wherein only a scraper in the shape of a triangular prism is installed in a culture container. However, there is a concern that when the scraper is only installed in the culture container, the scraper may contact the cells in culture and peel off the cells. For example, in Japanese Patent Application Laid-Open No. 2004-129558, the scraper is not fixed in the container under normal conditions, and it is expected that the scraper moves in the container due to the water flow or the tilt of the container when the culture medium is replaced, and the cells in culture are peeled off by the culture surface contact portion of the scraper while the scraper moves. Summary of the invention

[0005] The present invention is proposed in view of the above problems. An object of the present invention is to provide a cell detachment system in which a scraper is prevented from contacting cells during cell culture, or friction is suppressed even when the scraper contacts cells, thereby reducing damage to cells during culture, and also enabling the scraper to detach cells while maintaining a closed system.

[0006] The present invention adopts the following structure. That is, a cell peeling system is provided, comprising: a scraper having magnetism for peeling off cells attached to a culture surface; a magnetic field applying unit configured to apply a magnetic field to the scraper; and a scraper position control unit configured to control the position of the scraper by controlling the magnetic field applying unit, wherein the scraper is configured to move while rotating in a state where no magnetic field is applied, and in a state where a magnetic field is applied by the magnetic field applying unit, the scraper does not rotate but is moved by the scraper position control unit to peel off the attached cells.

[0007] Furthermore, according to one aspect of the present invention, there is provided a cell culture system, comprising: the above-mentioned cell detachment system; and a culture container comprising a culture surface.

[0008] In addition, according to one aspect of the present invention, a cell peeling system is provided, comprising: a scraper having magnetism for peeling off cells attached to a culture surface; a magnetic field applying unit configured to apply a magnetic field to the scraper; a scraper position control unit configured to control the position of the scraper by controlling the magnetic field applying unit; and a scraper holding portion configured to hold the scraper at a position where it is not in contact with the culture surface.

[0009] Furthermore, according to one aspect of the present invention, there is provided a cell culture system, comprising: the above-mentioned cell detachment system; and a culture container comprising a culture surface.

[0010] Further features of the present invention will become apparent from the following description of exemplary embodiments with reference to the attached drawings. BRIEF DESCRIPTION OF THE DRAWINGS

[0011] Figure 1A is a perspective view showing an outline of the overall configuration of an example of a cell culture system including a cell detachment system according to the present invention, Figure 1B It is shown in Figure 1A A perspective view showing a state in which cells attached to a culture surface are peeled off by a scraper in the example of the cell culture system shown.

[0012] Figure 2 It is a perspective view showing how the spherical scraper in the present invention is installed on a culture surface having a concave curved surface.

[0013] Figure 3A , Figure 3B and Figure 3C Each is a view showing an example of a cylindrical or cylindrical scraper in the present invention, wherein, Figure 3A is a perspective view showing a cylindrical scraper made of a magnetic substance, Figure 3B is a perspective view showing a cylindrical scraper using a magnetic substance in the center portion and a non-magnetic substance in the outer peripheral portion, Figure 3Cis a perspective view showing a cylindrical scraper made of a magnetic substance.

[0014] Figure 4A is a schematic diagram showing how a scraper of the prior art moves due to an external force while in contact with cells, Figure 4B is a schematic diagram showing how the scraper of the present invention moves due to an external force while in contact with cells, Figure 4C is a schematic diagram showing how the scraper of the present invention peels off cells.

[0015] Figure 5A , Figure 5B , Figure 5C and Figure 5D Each is a schematic diagram showing an example of a cell detachment system according to the present invention, wherein a scraper holding portion is installed in a culture container, wherein Figure 5A is a view showing a state during cell culture in a cell detachment system in which a scraper holding portion has an inclined surface, Figure 5B is a view showing how to move a scraper to a culture surface after cell culture in a cell detachment system, Figure 5C is a view showing a state during cell culture in a cell detachment system in which a scraper is held by magnetic attraction, Figure 5D is a view showing how to move a scraper to a culture surface after cell culture in a cell detachment system.

[0016] Fig. 6A and Figure 6B are views showing an example of a cell detachment system according to the present invention, wherein a scraper holder is installed outside a culture container, wherein: Fig. 6A is a schematic diagram showing the state during cell culture, Figure 6B is a schematic diagram showing how to move the scraper to the culture surface after cell culture.

[0017] Figure 7 : is a schematic diagram showing an example of the overall configuration of a cell culture system including a cell detachment system according to the first embodiment.

[0018] Fig. 8A , Figure 8B and Figure 8C are schematic diagrams showing an example of the overall configuration of a cell culture system including a cell detachment system according to a second embodiment, wherein: Fig. 8A is a view showing a state before the scraper is moved to the culture surface, Figure 8B is a view showing how to move the scraper to the culture surface, Figure 8C is a view showing a state where the movement of the scraper to the culture surface has been completed.

[0019] Fig.9A , Fig. 9B and Fig. 9C are schematic diagrams showing an example of the overall configuration of a cell culture system including a cell detachment system according to a second embodiment, wherein: Fig.9A is a view showing a state before the scraper is moved to the culture surface, Fig. 9B is a diagram showing how to move the scraper to the culture surface, Fig. 9C This is a diagram showing a state where the movement of the scraper to the culture surface has been completed.

[0020] Fig. 10A , Fig. 10B and Fig. 10C are schematic diagrams showing an example of the overall configuration of a cell culture system including a cell detachment system according to a third embodiment, wherein: Fig. 10A is a view showing a state before the scraper is moved to the culture surface, Fig. 10B is a view showing how to move the scraper to the culture surface, Fig. 10C This is a diagram showing a state where the movement of the scraper to the culture surface has been completed.

[0021] Fig.11A , Fig. 11B and Fig. 11C are schematic diagrams showing an example of the overall configuration of a cell culture system including a cell detachment system according to a fourth embodiment, wherein: Fig.11A is a view showing a state before the scraper is moved to the culture surface, Fig. 11B is a diagram showing how to move the scraper to the culture surface, Fig. 11C This is a diagram showing a state where the movement of the scraper to the culture surface has been completed.

[0022] Fig.12 is a block diagram showing a configuration example of a cell detachment system according to the present invention.

[0023] Fig.13 is a block diagram showing a hardware configuration example of the information processing unit in the present invention. DETAILED DESCRIPTION

[0024] According to the present invention, the cell peeling system includes: a magnetic scraper for peeling cells attached to a culture surface; a magnetic field applying unit, configured to apply a magnetic field to the scraper; and a scraper position control unit, configured to control the position of the scraper by controlling the magnetic field applying unit, wherein the scraper is configured to move while rotating in a state where no magnetic field is applied, and in a state where a magnetic field is applied by the magnetic field applying unit, the scraper does not rotate but is moved by the scraper position control unit to peel off the attached cells.

[0025] Exemplary embodiments of the present invention are described below with reference to the accompanying drawings. In the accompanying drawings, the same parts are represented by the same figure numerals in principle, and the description is omitted. The term "external force" used in this specification refers to a force other than magnetic force, and examples thereof include gravity, inertial force, and water flow applied when the culture container is moved. The term "sample solution, etc." used in this specification refers to culture medium, enzyme solution, phosphate buffered saline (PBS), cell suspension, etc. In the following description, adherent cells are described as an example, but the cell detachment system and cell culture system according to the present invention can also be applied to cells other than adherent cells.

[0026] (System Structure)

[0027] First, refer to Figure 1A and Figure 1B A cell detachment system according to the present invention and a cell culture system comprising the cell detachment system are described. Figure 1A 1 is a perspective view showing an overview of the overall configuration of an example of a cell culture system including a cell peeling system according to the present invention. The culture surface 101 has a surface to which cells can attach and to which cells can proliferate. The culture container 102 includes the culture surface 101 and can contain a liquid. The scraper 103 has magnetism and is used to peel off cells attached to the culture surface. The scraper 103 slides in the direction indicated by the arrow. The magnetic field applying unit 104 applies a magnetic field to the scraper. The scraper position control unit 105 controls the magnetic field applying unit to control the position of the scraper 103 in the culture container 102. The scraper 103 moves while rotating in a state where no magnetic field is applied. In a state where a magnetic field is applied by the magnetic field applying unit 104, the scraper 103 does not rotate but is moved by the scraper position control unit 105, thereby peeling off cells attached to the culture surface 101.

[0028] On the wall surface of the culture container 102, there is a supply and discharge port 106 inserted into the culture container 102 to allow the culture medium and the like to be put in and taken out. The upper surface of the culture container 102 remains closed and is connected to the outside via the supply and discharge port 106. A plurality of supply and discharge ports 106 may be provided, each connected to a corresponding tube (not shown) to be connected to a storage container (not shown) of a sample solution and the like (e.g., culture medium, PBS) via the tube under sterile conditions. It is also possible to create a closed system by providing a wide opening instead of the upper surface of the culture container, injecting the sample solution and the like from the wide opening, and then tightly closing the lid. However, from the viewpoint of preventing foreign matter from entering from the outside during the injection process, as shown in FIG. Figure 1A As shown, it is preferred to provide supply and discharge ports and narrow the openings. An incubator (not shown) may be prepared as a cell culture unit, and the culture container 102 may be installed therein.

[0029] Figure 1B1 is a perspective view showing a state in which cells attached to a culture surface 101 are peeled off by a scraper 103 in a cell culture system including a cell peeling system according to the present invention. Figure 1B In the figure, the shaded area on the culture surface 101 indicates that cells are attached thereto. The scraper 103 to which the magnetic field is applied by the magnetic field applying unit 104 is controlled by the scraper position controlling unit 105 to move on the culture surface 101 while sliding on the culture surface, thereby being able to peel off the cells attached to the culture surface 101.

[0030] (Cultivation Container)

[0031] The cell culture system according to the invention may comprise the cell detachment system according to the invention and a culture container comprising a culture surface.

[0032] Culture surface is a culture substrate that can be attached to and proliferate for cells, and culture surface preferably adopts a transparent material that is easy to observe. Preferably, transparent resins, glass, etc. that are easy to process and observe are used as the material of culture surface. When using resin, polystyrene, polycarbonate, acrylic acid, etc. can be used. Material is not limited to transparent material, as long as the material does not block the light path when observing, but magnetic material (such as some types of metal) is inappropriate. Culture surface can form a part or all of a culture vessel, or can be provided separately from a culture vessel. From the perspective of the cost and ease of preparation, it is preferred that culture surface is the material identical with the culture vessel and culture surface forms a part of the culture vessel.

[0033] The culture surface is preferably subjected to a surface treatment such as a plasma treatment so that the cells can easily attach only to the culture surface.

[0034] The culture container is a container for holding a culture medium for culturing cells. It is preferred to use a transparent resin, glass, etc. that is easy to process and observe as the material of the culture container. In the case of using a resin, polystyrene, polycarbonate, acrylic acid, etc. can be used. The material is not limited to a transparent material as long as the material does not block the light path during observation, but a magnetic material (such as some types of metal) is not suitable. From the perspective of being able to prevent foreign matter from entering from the outside and ensuring a predetermined temperature and a predetermined humidity, the culture container is preferably a closed system.

[0035] (scraper)

[0036] The cell detachment system according to the present invention comprises a scraper having magnetism for detaching cells attached to a culture surface.

[0037] The scraper can move while rotating in a state where a magnetic field is not applied, and the scraper does not rotate but can be moved by a scraper position control unit to peel off the attached cells when a magnetic field is applied by a magnetic field applying unit. The term "rotation" used herein refers to rotation caused by an external force, including not only continuous rotation but also intermittent rotation. Specifically, the term "rotation" includes moving while rotating a certain distance, then moving while sliding without rotating, and moving again while rotating. The scraper can move while rotating in a state where a magnetic field is not applied, so that the scraper can rotate on the culture surface when an external force is applied to the culture container during cell culture. Therefore, the friction force applied by the scraper to the cells in culture can be suppressed. In this case, damage to the cells in culture can be reduced, and a low-cost simple cell peeling system can be realized. In addition, in a state where a magnetic field is applied by a magnetic field applying unit, the scraper does not rotate but can be moved by a scraper position control unit, so that a magnetic field can be applied to peel off the cells when the cells are peeled.

[0038] The shape of the scraper is not limited, as long as the scraper can rotate on the culture surface, and its examples include polyhedron (such as regular polyhedron, truncated polyhedron, etc.), spherical, ellipsoidal, cylindrical and cylindrical. The example of polyhedron includes a regular polyhedron with more than 20 surfaces and its truncated polyhedron. Wherein, the shape of the scraper is preferably a shape in which the scraper surface in contact with the culture surface is a curved surface, or the scraper surface in contact with the culture surface is only formed by a curved surface, such as spherical, ellipsoidal, cylindrical and cylindrical. When using a scraper with such a shape, it is assumed that the scraper is arranged so that the scraper surface as a curved surface contacts the culture surface. In the present invention, the situation in which the scraper is cylindrical includes the situation in which the scraper is a regular cylinder and an ellipsoidal cylinder. Furthermore, in the present invention, in the case where the scraper is in the shape of an ellipsoid, assuming that the x-axis, y-axis, and z-axis lengths of the ellipsoid are represented by "a", "b", and "c", respectively, the ellipsoid can be set so that a / b, b / c, and c / a are all equal to or greater than 1 / 3 and equal to or less than 3. In the present invention, in the case where the scraper is in the shape of an elliptical cylinder, assuming that the ratio of the short diameter and the long diameter of the elliptical surface is set to (short diameter): (long diameter) = a:b and "a" is 1, the scraper can be set to have a shape in which "b" is greater than 1 and equal to or less than 3.

[0039] In the case where the culture surface is a flat surface, for example, when the shape of the scraper is cylindrical or cylindrical, the contact area between the culture surface and the scraper is a straight line. When the cells are peeled off, the scraper moves while maintaining the contact area, so that the cells on the culture surface area through which the contact area passes can be peeled off. The contact area of ​​the scraper relative to the culture surface is preferably a line or a plane.

[0040] Furthermore, when the culture surface in the present invention includes a concave surface, the shape of the scraper is preferably spherical or ellipsoidal. The advantage of the culture surface being a concave curved substrate is that the culture surface area per unit volume of the culture container can be increased compared to the case where the culture surface is a flat surface substrate. Figure 2 1 is a stereogram showing how the spherical scraper 103 is installed on the culture surface 101 having a concave surface in the present invention. When the culture surface is a concave surface or a tubular surface, a scraper having a spherical or ellipsoidal shape that conforms to the concave surface or the tubular surface can be used. For example, when a concave curved substrate having a curvature of 1 mm in diameter is set as the culture surface, in the case of culturing cells on the culture surface, it is preferred to use a scraper having a sphere of 1 mm in diameter or a scraper having an ellipsoidal shape of 1 mm in any one of the major axis, minor axis or depth (x-axis, y-axis or z-axis).

[0041] The scraper contains a magnetic material, and in particular, preferably contains a metal with strong magnetism. Resin materials mixed with magnetic particles can also be used. It is also effective to use different components to form the center and periphery of the scraper. For example, it is also effective to use a magnetic metal as the center and to cover the periphery with a non-magnetic buffer rubber component. Covering with a rubber component also allows the use of magnetic metals that are easy to corrode, and its buffering properties increase the adhesion of the scraper during peeling, thereby improving the peeling performance. In addition, a hollow scraper can be used. The case where the scraper in the present invention is cylindrical includes the case where the scraper is a hollow right cylinder or elliptical cylinder. The case where the scraper is spherical or ellipsoidal includes the case where the scraper is a hollow sphere or a hollow ellipsoid. By using a hollow scraper, the peeled cells and the extruded liquid enter the hollow space during cell peeling, and the scraper can move smoothly without obstruction of its path. In addition, the scraper can be made lighter in weight, and the damage to the cells in culture can be further reduced when the scraper contacts the cells in culture.

[0042] exist FIG. 3A to FIG. 3C , an example of a cylindrical or drum-shaped scraper is shown. Figure 3A is a perspective view showing a cylindrical scraper made of a magnetic substance. Figure 3B is a perspective view showing a cylindrical scraper using a magnetic substance in a central portion and a non-magnetic substance in an outer peripheral portion. Figure 3C is a perspective view showing a cylindrical scraper made of a magnetic material. FIG. 3A to FIG. 3C , the portion indicated by “m” represents a magnetic substance, the portion indicated by “n” represents a non-magnetic substance, and the portion indicated by “h” represents a hollow.

[0043] (Magnetic field applying unit)

[0044] The magnetic field applying unit applies a magnetic field to the scraper. For the magnetic field applying unit, a magnet can be used, and a permanent magnet or an electromagnet can be used. As the permanent magnet, a ferrite magnet, a neodymium magnet, a samarium cobalt magnet, an alnico magnet, etc. can be used. When a permanent magnet is used as the magnetic field applying unit, there is a case where the magnetic field is applied to the scraper by the magnetic field applying unit even in the culture that does not involve cell peeling. In this case, the scraper is fixed during cell culture, thereby facilitating the reduction of damage to the cells.

[0045] (Scraper position control unit)

[0046] The scraper position control unit controls the position of the scraper by controlling the magnetic field application unit. The scraper position control unit enables the scraper to slide on the culture surface. When the culture surface is a flat surface, it is preferred to move the magnetic field application unit in a two-dimensional direction along the culture surface. When the length of the scraper is sufficient to cover the width of the culture surface, a single-axis stage can be used as the scraper position control unit. In contrast, when the length of the scraper is shorter than the width of the culture surface, a double-axis stage is preferably used as the scraper position control unit.

[0047] (Movement of scraper)

[0048] FIG. 4A to FIG. 4C Each shows how the scraper comes into contact with the cells. FIG. 4A to FIG. 4C 1 shows a cell 100. When a member such as a scraper is not fixedly mounted on the culture surface, the scraper moves on the culture surface due to an external force (eg, gravity or water flow) when the culture medium is replaced during culture or the like. Figure 4A 1 is a view showing the state of the cells 100 when an angular prior art scraper 403 that cannot rotate on the culture surface is moved on the culture surface 101 by an external force in a state where no magnetism is applied. The scraper 403 takes the form of dragging on the culture surface 101 and is prone to peeling off the cells 100 at an unexpected time (for example, during culture). Figure 4B 1 is a view showing the state of the cells 100 when the scraper 103 of the present invention moves on the culture surface 101 in a state where no magnetism is applied. The scraper 103 moves on the culture surface 101 while rotating, so the friction force is reduced, thereby Figure 4A The difference is the amount of cells 100 that can inhibit accidental detachment.

[0049] Figure 4C 1 is a diagram showing the state of cells 100 when the scraper 103 of the present invention moves on the culture surface 101 in a state where magnetism is applied. The scraper 103 is fixed in the magnetic pole direction by the magnetic field applying unit 104, so that the scraper takes the form of dragging on the culture surface 101, so that the cells 100 can be peeled off.

[0050] In this way, the scraper in the present invention moves on the culture surface while rotating without being applied with a magnetic field and can reduce damage to cells during the culture process even when the scraper is placed on the culture surface, while the scraper can peel off the cells by applying magnetism during peeling.

[0051] As a method of controlling the position of the scraper, the magnetic field applying unit may be fixed to the scraper position controlling unit, and the scraper position controlling unit may be moved to move the magnetic field applying unit, thereby moving the scraper to which the magnetic field is applied.

[0052] (Scraper holding unit and scraper moving unit)

[0053] The cell peeling system according to the present invention is a cell peeling system comprising: a scraper having magnetism for peeling off cells attached to a culture surface; a magnetic field applying unit for applying a magnetic field to the scraper; and a scraper position control unit for controlling the position of the scraper by controlling the magnetic field applying unit, wherein the scraper moves while rotating in a state where no magnetic field is applied, and the scraper does not rotate but is moved by the scraper position control unit in a state where a magnetic field is applied by the magnetic field applying unit to peel off the attached cells; and the cell peeling system can be configured to also include a scraper holding portion that holds the scraper in a position where it does not contact the culture surface.

[0054] In addition, the cell detachment system according to the present invention can be a cell detachment system comprising: a scraper having magnetism for detaching cells attached to a culture surface; a magnetic field applying unit for applying a magnetic field to the scraper; a scraper position control unit for controlling the position of the scraper by controlling the magnetic field applying unit; and a scraper holding portion for holding the scraper at a position where it does not contact the culture surface.

[0055] Furthermore, the cell detachment system according to the present invention may be a cell detachment system including a scraper moving unit that moves the scraper held by the scraper holding portion to the culture surface.

[0056] It is important to take appropriate measures to prevent the scraper from damaging the cells in culture. As such a measure, it is effective to provide a scraper holding portion for holding the scraper at a position different from the culture surface during cell culture. Examples of the form of the scraper holding portion include a form in which the scraper holding portion is included inside the culture container and a form in which the scraper holding portion is included outside the culture container. As a form in which the scraper holding portion is included inside the culture container, for example, a space and an inclined surface may be provided in the culture container to hold the scraper, or a magnet may be used to hold the scraper. Examples of the form in which the scraper holding portion is included outside the culture container include a form in which a box-shaped scraper holding portion is provided outside the culture container. When the cell detachment system includes a scraper holding portion, it is preferred to provide a scraper moving unit that moves the scraper held by the scraper holding portion to the culture surface. For example, the scraper moving unit may be provided using a magnet, or the scraper moving unit may be provided using a cover, a valve, or the like that can be opened and closed. Refer to the following FIG. 5A to FIG. 5D , Fig. 6A and Figure 6B The scraper holding part and the scraper moving unit are described. The forms of the scraper holding part and the scraper moving unit are not limited to the forms described below. Any form can be used as long as a scraper holding part that holds the scraper at a position that does not contact the culture surface is provided and the scraper can be moved to the culture surface by the scraper moving unit.

[0057] FIG. 5A to FIG. 5D 1 and 2 are schematic diagrams each showing an example of a cell detachment system in which a scraper holding portion 107 is installed in a culture container 102 . Figure 5A 1 is a view showing a state during cell culture in a cell detachment system in which a scraper holding portion 107 has an inclined surface. During the culture of cells 100, the scraper 103 is prevented from moving to the culture surface 101 by the scraper holding portion 107 having an inclined surface. This also prevents the cells 100 in culture from being detached. At the time of detachment, a scraper moving unit 108 is used to move the scraper 103 to the culture surface 101. As the scraper moving unit 108, a movable stage or the like can be used, and when the cell detachment system includes an imaging device, the scraper moving unit 108 is preferably arranged so as not to hinder the imaging device from imaging. Figure 5B 1 is a view showing how to move the scraper 103 to the culture surface 101 in the above-mentioned cell detachment system after cell culture. In the case of this form, when the scraper 103 has a shape that allows the scraper 103 to move while rotating in a state where no magnetic field is applied, the scraper 103 can be moved to the culture surface 101 only by moving the scraper moving unit 108 to tilt the culture container 102.

[0058] Furthermore, a magnet may be used as the scraper moving unit 108 . Figure 5C1 is a view showing a state during cell culture in the cell detachment system, in which the scraper 103 is held by being attracted by a magnet. During the culture, the scraper 103 is fixed in the scraper holding portion 107 by being attracted by the scraper moving unit 108 which is a magnet. Figure 5D 1 is a schematic diagram showing how to move the scraper 103 to the culture surface 101 in the above-mentioned cell detachment system after cell culturing. At the time of cell detachment, the scraper 103 can be moved to the culture surface 101 by releasing the magnetic force of the scraper moving unit 108.

[0059] Fig. 6A and Figure 6B Each is a view showing an example of a cell detachment system in which a scraper holding portion 107 is installed outside the culture container 102 . Fig. 6A 1 is a schematic diagram showing a state during cell culture in the above-mentioned cell detachment system. The culture container 102 and the scraper holding portion 107 holding the scraper 103 are connected to each other via a tube 111. The above-mentioned configuration enables the scraper 103 to detach cells while maintaining a closed system. The tube 111 is provided with a valve serving as a scraper moving unit 108, and closing the scraper moving unit 108 during the culture process can keep the scraper 103 in the scraper holding portion 107.

[0060] Figure 6B 1 is a schematic diagram showing how to move the scraper 103 to the culture surface 101 after cell culture. Fig. 6A As shown, the scraper 103 is fixed in the scraper holder 107, thereby preventing the cells 100 from being peeled off during the culture process. At the time of peeling, the scraper 103 can be moved to the culture surface 101 by opening the scraper moving unit 108 to allow the scraper 103 to pass through the inside of the tube 111. It is also possible to connect the supply port to the scraper holder 107 and inject some sample solution or the like through these supply ports. The method of moving the scraper 103 in the tube 111 can be achieved by allowing the culture medium, PBS or other liquids to flow together, or the scraper 103 can be simply moved in the tube by gravity. When the scraper 103 has a shape that allows the scraper 103 to rotate on the surface, the scraper 103 is not easily stuck when passing through the inside of the tube 111.

[0061] (Camera device)

[0062] The cell peeling system according to the present invention may include a camera that acquires information about the state of cells attached to the culture surface. Examples of the camera include a CMOS camera and a CCD camera. By using the camera, information about the state of cells attached to the culture surface (e.g., the area of ​​the colony included in the culture surface) can be acquired, and it can be determined whether the desired amount of cells has been cultured. It is also effective to add a process of checking the attachment area of ​​the cells from the image acquired by the camera and moving the scraper to the attachment area to peel off the cells in the desired area.

[0063] (Information Processing Unit)

[0064] The cell detachment system according to the present invention may include an information processing unit. In particular, the cell detachment system according to the present invention may include an information processing unit configured to cause the scraper moving unit to move the scraper from the scraper holding portion to the culture surface based on information acquired by the camera. Fig.12 1 is a block diagram showing a configuration example of a cell peeling system according to the present invention, which includes an imaging device 110 and an information processing unit 113. The information processing unit 113 can process information related to the state of cells attached to the culture surface, particularly, for example, information related to the state of cells attached to the culture surface acquired by the imaging device 110. In addition, for example, the information processing unit 113 can control a scraper position control unit 105 for moving the scraper 103 at the time of cell peeling. In addition, for example, when the magnetic field applying unit 104 is an electromagnet or the like, the information processing unit 113 can turn on and off a switch. In addition, for example, the information processing unit 113 can issue an instruction to move the scraper 103 from the scraper holding portion to the culture surface by using the scraper moving unit 108 at the time of cell peeling. For example, the above-mentioned instruction can be issued after determining whether a desired amount of cells have been cultured based on the colony area or other information included in the culture surface acquired by the imaging device 110. The information processing unit 113 can also determine whether the desired amount of cells has been cultured based on, for example, information on the area of ​​colonies included in the culture surface that has been acquired by the camera 110, and issue an instruction to continue the culture or an instruction to end the culture and shift to cell peeling. In the case of cells with a stable proliferation rate, it is also possible to use the information processing unit 113 to pre-set that cell peeling starts after a specified culture time period has passed, and issue an instruction to perform cell peeling after the culture time period has passed. In addition, for example, an instruction can be issued to perform a process for peeling cells in a desired area by checking the attachment area of ​​the cells from the image acquired by the camera 110 and moving the scraper 103 to the attachment area using the scraper position control unit 105.

[0065] Fig.13201 is a block diagram showing an example of a hardware configuration of the information processing unit 113 in the present invention. The information processing unit 113 has the function of a computer. For example, the information processing unit 113 can be configured integrally with, for example, a desktop personal computer (PC), a laptop PC, a tablet PC, or a smart phone. In order to realize the function as a computer that performs arithmetic operations and storage, the information processing unit 113 includes a central processing unit (CPU) 200, a random access memory (RAM) 201, a read-only memory (ROM) 202, and a hard disk drive (HDD) 203. The information processing unit 113 also includes a communication interface (I / F) 204, a display device 205, and an input device 206. The CPU 200, the RAM 201, the ROM 202, the HDD 203, the communication I / F 204, the display device 205, and the input device 206 are connected to each other via a bus 207. The display device 205 and the input device 206 can be connected to the bus 207 via a drive device (not shown) for driving these devices.

[0066] exist Fig.13 , the various components constituting the information processing unit 113 are illustrated as an integrated device, but some functions of these components may be implemented by external devices. For example, the display device 205 and the input device 206 may be external devices different from the components implementing the functions of the computer including, for example, the CPU 200.

[0067] The CPU 200 performs predetermined operations according to programs stored in, for example, the RAM 201 and the HDD 203, and also has a function of controlling the various components of the information processing unit 113. The RAM 201 is constructed of a volatile storage medium, and provides a temporary storage area required for the operation of the CPU 200. The ROM 202 is constructed of a nonvolatile storage medium, and stores necessary information such as programs to be used for the operation of the information processing unit 113. The HDD 203 is formed of a nonvolatile storage medium, and is a storage device that stores information on cell states and the like.

[0068] The communication I / F 204 is a communication interface based on a standard such as Wi-Fi (trademark) or 4G, and is a module for communicating with other devices. The display device 205 is, for example, a liquid crystal display or an organic light emitting diode (OLED) display, and is used to display, for example, dynamic images, static images, characters, etc. The input device 206 is, for example, a button, a touch panel, a keyboard, or a pointing device, and is used by the user to operate the information processing unit 113. The display device 205 and the input device 206 can be integrally formed as a touch panel.

[0069] Fig.13The hardware configuration shown in is an example, and devices other than the devices shown may be added, or some of the devices shown may be omitted. In addition, some devices may be replaced by other devices having the same functions. In addition, some functions may be provided by other devices via a network, and the functions to be implemented may be shared and implemented by multiple devices. For example, HDD 203 may be replaced by a solid-state drive (SSD) using semiconductor elements such as flash memory, or may be replaced by cloud storage.

[0070] The CPU 200 realizes various functions by loading a program stored in the ROM 202 or the like into the RAM 201 and executing the program. The CPU 200 also controls the display device 205. The CPU 200 also controls the HDD 203.

[0071] (Cell Culture System)

[0072] The cell culture system according to the present invention is a cell culture system comprising: the cell detachment system of the present invention; and a culture container comprising a culture surface. In addition, the cell culture system according to the present invention may include a cell culture unit. Examples of cell culture units include a heat preservation unit that maintains the entire cell culture system at 37°C, a unit that maintains the entire cell culture system under a CO2 atmosphere, and a unit that maintains the entire cell culture system at 95% humidity. Specifically, an incubator in which the entire culture container is maintained at a temperature of 37°C, a CO2 concentration of 5%, and a humidity of 95% is taken as an example. In the following embodiments, cells are sown and cultured using tubes through supply and discharge ports, but cells can also be sown by using a pipette.

[0073] By using the above system, it is possible to provide a cell detachment system that can minimize physical damage to cells during cell culture and also enable a scraper to detach cells while maintaining a closed system.

[0074] [First embodiment]

[0075] Now, a cell detachment system and a cell culture system including the cell detachment system according to a first embodiment will be described. In the drawings, the same components are denoted by the same reference numerals in principle, and description is omitted.

[0076] Figure 71 is a schematic diagram showing an example of the overall configuration of a cell culture system including a cell detachment system according to the first embodiment. In this system, a plurality of tubular supply and discharge ports 106 made of polystyrene are arranged at the edge of a culture container 102, which is a rectangular parallelepiped closed container made of polystyrene. In addition, a membrane filter 109 through which a gas such as CO2 can pass is arranged in the upper surface. The upper surface of the culture container 102 remains closed except for a portion where the membrane filter 109 is arranged and a portion where the supply and discharge ports 106 are arranged, and is connected to the outside via the supply and discharge ports 106. In addition, the lower surface of the inner side of the culture container 102 is a culture surface 101, and a surface treatment is performed to allow cell attachment. In the culture container 102, a cylindrical metal rod made of SUS440 with a diameter of 1 mm and a length of 5 mm is arranged as a scraper 103. Below the culture container 102, an electromagnet is arranged as a magnetic field applying unit 104, and the magnetic field applying unit 104 is arranged in a scraper position control unit 105 as a biaxial automatic stage. Furthermore, in the central portion below the culture container 102 , a CMOS camera is installed as an imaging device 110 that allows observation of the culture surface 101 .

[0077] The actual cell culture and detachment sequence using this system is described below.

[0078] (Initial setting)

[0079] Three supply and exhaust ports 106 are prepared, and a silicone tube (not shown) is attached to each of the supply and exhaust ports.

[0080] The first supply and discharge port 106 can be connected to a container (not shown) for a culture medium, a container (not shown) for an enzyme solution during stripping, a container (not shown) for PBS, and a container (not shown) for a cell suspension mixed with cells before culture. The second supply and discharge port 106 can be connected to a waste container (not shown) for accumulating waste liquid. The third supply and discharge port 106 can be connected to a recovery container (not shown) for recovering a cell suspension containing stripped cells. The silicone tube is provided with a joint (not shown) for connecting to each container. The silicone tube to be connected to the first supply and discharge port 106 is branched to correspond to the type of sample solution, etc., and the opening and closing of these branches can be switched by a valve. The entire culture container 102 connected with these silicone tubes is sterilized with gamma rays.

[0081] In the case of starting cell culture, each silicone tube is connected to each container containing sample solution, etc., waste container and recovery container. In this case, it is necessary to connect them aseptically. After connection, each tube is connected to a tube pump (not shown).

[0082] Liquid is fed from each container such as the sample solution to the culture container 102 and liquid is fed from the culture container 102 to the waste container and the recovery container by each tube pump. As an actuator for feeding liquid, a tube pump is used in the first embodiment, but any actuator can be used instead of the tube pump as long as the actuator can realize a closed system. Finally, the entire cell culture system except for each container such as the sample solution is installed in an incubator (not shown), and the interior of the incubator is maintained at a temperature of 37° C., a CO2 concentration of 5%, and a humidity of 95%.

[0083] (Cell Culture)

[0084] First, a cell suspension as a mixed solution of culture medium, extracellular matrix and cells to be cultured is fed into the culture container 102 via the supply and discharge port 106. The feeding amount of the cell suspension is sufficient to immerse the cell suspension in the entire area of ​​the culture surface 101. Afterwards, the culture container 102 is stored in an incubator for a certain period of time. Sometimes the culture medium is replaced. In this case, after the culture medium in the culture container 102 is moved to a waste container, a new culture medium is moved into the culture container. The frequency of the culture medium replacement is preferably set according to the cells to be cultured. The replacement of the culture medium and the preservation in the incubator continue until the cells have proliferated by the required amount. By using the camera 110, it can be determined whether the desired amount of cells has been cultured according to the area of ​​the bacterium colonies included in the culture surface. This determination can also be automatically performed by providing an information processing unit (not shown). A CMOS camera is used as the camera 110, but a CCD camera can also be used.

[0085] (Cell detachment)

[0086] After the desired amount of cells have been cultured, the cells are peeled off. The timing of cell peeling is determined based on the image of the cells attached to the culture surface 101 that has been acquired by the camera 110. It is effective to set the timing when the area of ​​the colonies included in the culture surface becomes equal to or greater than the predetermined area as the peeling timing. This determination can also be made automatically by providing an information processing unit (not shown), and an instruction to transfer to cell peeling can also be issued. In the case of cells with a stable proliferation rate, the peeling timing can be set using a specified culture time period. In the case where an information processing unit is provided, it can also be set so that it is automatically transferred to cell peeling after a specified culture time period has passed.

[0087] First, after the culture medium in the culture container 102 is moved to the waste container via the supply and discharge port 106, PBS is fed into the culture container 102 via the supply and discharge port 106 until the culture surface 101 is immersed. Then, after the PBS in the culture container 102 is moved to the waste container via the supply and discharge port 106, an enzyme solution such as trypsin is fed into the culture container 102 via the supply and discharge port 106 until the culture surface 101 is immersed, while the culture container 102 is maintained in an incubator at 37° C. for a certain period of time. Thereafter, the enzyme solution in the culture container 102 is moved to the waste container via the supply and discharge port 106, and then the culture medium is fed into the culture container 102 via the supply and discharge port 106 until the culture surface 101 is immersed.

[0088] Thereafter, the magnetic field applying unit 104, which is an electromagnet, is turned on and driven by the scraper position control unit 105, which is a two-dimensional stage, so that the magnetic field applying unit 104 passes twice over the entire area below the culture surface 101. The scraper 103 can move freely on the culture surface 101 before the start of culture, so the scraper 103 is not always present in the vicinity of the magnetic field applying unit 104 when the culture starts, but the scraper 103 is reliably fixed by the magnetic force of the magnetic field applying unit 104 at the first pass. Therefore, the two passes ensure that the scraper 103 passes over the entire area of ​​the culture surface 101 at least once, so that almost all cells attached to the culture surface 101 can be peeled off.

[0089] It is also effective to add a process of checking the cell attachment area from the image acquired by the imaging device 110 and moving the scraper 103 to the attachment area to peel off the cells in a desired area.

[0090] This processing may be performed manually or automatically by using an information processing unit.

[0091] (Cell Recycling)

[0092] In the culture container 102, there is a suspension of a culture medium and detached cells. The tube pump is operated to move the cell suspension to the recovery container via the supply and discharge port 106. In this case, the cell suspension can be recovered more efficiently by tilting the culture container 102 so that the side where the supply and discharge port 106 is installed becomes lower, which is therefore effective.

[0093] As described above, according to the first embodiment, a cell peeling system can be provided, comprising: a scraper having magnetism for peeling off cells attached to a culture surface; a magnetic field applying unit for applying a magnetic field to the scraper; and a scraper position control unit for controlling the position of the scraper by controlling the magnetic field applying unit, wherein the scraper moves while rotating when no magnetic field is applied, and when a magnetic field is applied by the magnetic field applying unit, the scraper does not rotate but is moved by the scraper position control unit to peel off the attached cells; and a cell culture system comprising the above-mentioned cell peeling system can also be provided.

[0094] The first embodiment has been verified by using iPS cells, but most types of cells can be used as long as the cells are adherent cells. In this case, the types of culture media and reagents and the timing of detachment can be appropriately changed according to the characteristics of each cell and the protocol to be implemented.

[0095] [Second embodiment]

[0096] Now, a cell detachment system and a cell culture system including the cell detachment system according to a second embodiment are described. In the drawings, the same components are denoted by the same reference numerals in principle, and description is omitted.

[0097] FIG. 8A to FIG. 8C 1 and 2 are schematic diagrams each showing an example of an overall configuration of a cell culture system including a cell detachment system according to the second embodiment. Fig. 8A 1 is a view showing a state presented before the scraper 103 is moved to the culture surface 101 .

[0098] In this system, a plurality of tubular supply and discharge ports 106 made of polystyrene are arranged at the edge of a culture container 102, which is a closed container made of polystyrene. In addition, a membrane filter 109 through which gases such as CO2 can pass is arranged in the upper surface. In addition, the lower surface of the inner side of the culture container 102 is a culture surface 101, and a surface treatment is performed to allow cell attachment. In the culture container 102, a scraper holder 107 is arranged at a position that does not contact the culture surface 101, and the scraper holder has an inclined wall for preventing the scraper 103 from easily moving to the culture surface 101.

[0099] In the scraper holding portion 107 in the culture container 102, a cylindrical metal rod made of SUS440 with a diameter of 1 mm and a length of 5 mm is installed as the scraper 103. Outside the culture container 102, a magnetic field applying unit 104 as an electromagnet is arranged, and the magnetic field applying unit 104 is installed in a scraper position control unit 105 as a two-axis automatic stage. In addition, in the central part below the culture surface 101 of the culture container 102, a CMOS camera is installed as an imaging device 110 that allows observation of the culture surface 101. Below the culture container 102, a scraper moving unit 108 as a movable stage is installed.

[0100] The steps in the “(Initial Setting)”, “(Cell Culture)” and “(Cell Recovery)” sections of this system are the same as those in the first embodiment, so their description is omitted and only the steps in the “(Cell Peeling)” section are described below.

[0101] (Cell detachment)

[0102] After the desired amount of cells have been cultured, the cells are peeled off. The timing of cell peeling is determined based on the image of the cells attached to the culture surface 101 that has been acquired by the camera 110. It is effective to set the timing when the area of ​​the colonies included in the culture surface becomes equal to or greater than the predetermined area as the peeling timing. This determination can also be made automatically by providing an information processing unit (not shown), and an instruction to transfer to cell peeling can also be issued. In the case of cells with a stable proliferation rate, the peeling timing can be set using a specified culture time period. In the case where an information processing unit is provided, it can also be set so that it is automatically transferred to cell peeling after a specified culture time period has passed.

[0103] First, after the culture medium in the culture container 102 is moved to the waste container via the supply and discharge port 106, PBS is fed into the culture container 102 via the supply and discharge port 106 until the culture surface 101 is immersed. Then, after the PBS in the culture container 102 is moved to the waste container via the supply and discharge port 106, an enzyme solution such as trypsin is fed into the culture container 102 via the supply and discharge port 106 until the culture surface 101 is immersed, while the culture container 102 is maintained in an incubator at 37° C. for a certain period of time. Thereafter, the enzyme solution in the culture container 102 is moved to the waste container via the supply and discharge port 106, and then the culture medium is fed into the culture container 102 via the supply and discharge port 106 until the culture surface 101 is immersed.

[0104] Then, if Figure 8BAs shown, the scraper moving unit 108 is driven to tilt the culture container 102, thereby moving the scraper 103 to the culture surface 101 side, and then returning the culture container 102 to the position as shown in FIG. Figure 8C The driving of the scraper moving unit 108 may be performed manually, or may be performed automatically by using an information processing unit.

[0105] Thereafter, in the same manner as in the first embodiment, the magnetic field applying unit 104 as an electromagnet is turned on, and driven by driving the scraper position control unit 105 as a two-dimensional stage so that the magnetic field applying unit 104 passes twice over the entire area below the culture surface 101. In the same manner as in the first embodiment, the scraper 103 is reliably fixed by the magnetic force of the electromagnet at the time of the first pass. Therefore, the two passes of the magnetic field applying unit 104 ensure that the scraper 103 passes over the entire area of ​​the culture surface 101 at least once, thereby being able to peel off almost all the cells attached to the culture surface 101.

[0106] In another example, 9A to 9C As shown, a cell culture system may be provided, which includes a cell detachment system using an electromagnet as a scraper moving unit 108 . 9A to 9C 1 and 2 are schematic diagrams each showing an example of an overall configuration of a cell culture system including a cell detachment system according to the second embodiment. Fig.9A 1 is a view showing a state presented before the scraper 103 is moved to the culture surface 101 .

[0107] In this system, a plurality of tubular supply and discharge ports 106 made of polystyrene are arranged at the edge of a culture container 102, which is a closed container made of polystyrene. In addition, a membrane filter 109 through which gases such as CO2 can pass is arranged in the upper surface. In addition, the lower surface of the inner side of the culture container 102 is a culture surface 101, and a surface treatment is performed to allow cell attachment. In the culture container 102, a scraper holding portion 107 including an inclined wall and holding a scraper 103 is arranged at a position not in contact with the culture surface 101, and a scraper moving unit 108 as an electromagnet is arranged near the scraper holding portion 107.

[0108] In the culture container 102, a cylindrical metal rod made of SUS440 with a diameter of 1 mm and a length of 5 mm is installed in a scraper holding portion 107 as a scraper 103. Outside the culture container 102, a magnetic field applying unit 104 as an electromagnet is arranged, and the magnetic field applying unit 104 is installed in a scraper position control unit 105 as a two-axis automatic stage. In addition, in the central part below the culture surface 101 of the culture container 102, a CMOS camera is installed as an imaging device 110 that allows observation of the culture surface 101.

[0109] Fig.9A The structure shown is Fig. 8A The configuration shown is the same except that an electromagnet is installed as a scraper moving unit 108 near a scraper holding portion 107 provided with an inclined surface toward the culture surface 101, and the scraper 103 is held by being attracted to the wall surface of the scraper holding portion 107 by magnetic force.

[0110] Fig. 9B 1 is a view showing how to move the scraper 103 to the culture surface 101 . Fig. 9C 101 is a diagram showing a state where the movement of the scraper 103 to the culture surface 101 is completed. When the scraper moving unit 108 is disconnected, the scraper 103 moves from the inclined surface of the scraper holding portion 107 to the culture surface 101. Then, according to Figure 8C In the same sequence as in the illustrated configuration, the scraper 103 is moved by the magnetic field applying unit 104 moved by the scraper position control unit 105, thereby enabling the cells to be peeled off. The switching on and off of the scraper moving unit 108 may be performed manually, or may be performed automatically by using an information processing unit.

[0111] As described above, in the second embodiment, the scraper holding portion is set at a position that does not contact the culture surface in the culture container, and a scraper moving unit that moves the scraper held by the scraper holding portion to the culture surface is provided. Therefore, a cell peeling system that can reduce physical damage to cells during cell culture and can also peel cells while maintaining a closed system can be provided, and a cell culture system including the above-mentioned cell peeling system can also be provided. In addition, at this time, when the scraper can move while rotating in a state where no magnetic field is applied, the scraper can be easily moved to the culture surface.

[0112] The second embodiment has been verified by using iPS cells, but most types of cells can be used as long as the cells are adherent cells. In this case, the types of culture media and reagents and the timing of detachment can be appropriately changed according to the characteristics of each cell and the protocol to be implemented.

[0113] [Third embodiment]

[0114] Now, a cell detachment system and a cell culture system including the cell detachment system according to a third embodiment are described. In the drawings, the same components are denoted by the same reference numerals in principle, and description is omitted.

[0115] FIG. 10A to FIG. 10C 1 and 2 are schematic diagrams each showing an example of an overall configuration of a cell culture system including a cell detachment system according to the third embodiment. Fig. 10A1 is a view showing a state presented before the scraper 103 is moved to the culture surface 101 .

[0116] In this system, a plurality of tubular supply and discharge ports 106 made of polystyrene are arranged at the edge of a culture container 102, which is a closed container made of polystyrene. In addition, a membrane filter 109 through which gases such as CO2 can pass is arranged in the upper surface. In addition, the lower surface of the inner side of the culture container 102 is a culture surface 101, and a surface treatment is performed to allow cell attachment. On the outer side of the culture container 102, a scraper holder 107 made of polystyrene and connected to the culture container 102 via a tube 111 is prepared. The scraper holder 107 is also provided with a supply port 112, and can be connected to each container such as a sample solution via a silicone tube. The tube 111 is provided with a valve as a scraper moving unit 108. In the scraper holder 107, a cylindrical metal rod made of SUS440 with a diameter of 1 mm and a length of 5 mm is arranged as a scraper 103. A magnetic field applying unit 104 as an electromagnet is arranged outside the culture container 102, and the magnetic field applying unit 104 is installed in a scraper position control unit 105 as a two-axis automatic stage. In addition, in the central part below the culture surface 101 of the culture container 102, a CMOS camera is installed as an imaging device 110 that allows observation of the culture surface 101.

[0117] The steps in the “(Initial Setting)”, “(Cell Culture)” and “(Cell Recovery)” sections of this system are the same as those in the first embodiment, so their description is omitted and only the steps in the “(Cell Peeling)” section are described below.

[0118] (Cell detachment)

[0119] After confirming that the desired amount of cells have been cultured, the cells are peeled off. The timing of cell peeling is determined based on the image of the cells attached to the culture surface 101 that has been acquired by the camera 110. It is effective to set the time when the area of ​​the colonies included in the culture surface becomes equal to or greater than the predetermined area as the peeling timing. This determination can also be made automatically by providing an information processing unit (not shown), and an instruction to transfer to cell peeling can also be issued. In the case of cells with a stable proliferation rate, the peeling timing can be set using a specified culture time period. In the case where an information processing unit is provided, it can also be set so that it automatically transfers to cell peeling after a specified culture time period has passed.

[0120] First, after the culture medium in the culture container 102 is moved to the waste container via the supply and discharge port 106, PBS is fed into the culture container 102 via the supply and discharge port 106 until the culture surface 101 is immersed. Then, after the PBS in the culture container 102 is moved to the waste container via the supply and discharge port 106, an enzyme solution such as trypsin is fed into the culture container 102 via the supply and discharge port 106 until the culture surface 101 is immersed, while the culture container 102 is maintained in an incubator at 37° C. for a certain period of time. Thereafter, the enzyme solution in the culture container 102 is moved to the waste container via the supply and discharge port 106, and then the culture medium is fed into the culture container 102 via the supply and discharge port 106 until the culture surface 101 is immersed.

[0121] Then, the valve serving as the scraper moving unit 108 is released, and the scraper 103 moves to the culture surface 101 by passing through the inside of the tube 111 . Fig. 10B 1 is a view showing how to move the scraper to the culture surface. The driving of the scraper moving unit 108 may be performed manually, or may be performed automatically by using an information processing unit. Fig. 10C It is a view showing a state where the movement of the scraper to the culture surface has been completed. In the case of moving the scraper 103, it is also effective to inject the culture medium into the scraper holding portion 107 via the supply port 112, release the scraper moving unit 108, and then move the scraper 103 using the force of flowing the culture medium.

[0122] Thereafter, in the same manner as in the first embodiment, the magnetic field applying unit 104 as an electromagnet is turned on, and driven by driving the scraper position control unit 105 as a two-dimensional stage so that the magnetic field applying unit 104 passes twice over the entire area below the culture surface 101. In the same manner as in the first embodiment, the scraper 103 is reliably fixed by the magnetic force of the electromagnet at the time of the first pass. Therefore, the two passes of the magnetic field applying unit 104 ensure that the scraper 103 passes over the entire area of ​​the culture surface 101 at least once, thereby being able to peel off almost all the cells attached to the culture surface 101.

[0123] As described above, in the third embodiment, a scraper holding portion and a scraper moving unit that moves the scraper held by the scraper holding portion to the culture surface are provided outside the culture container. Therefore, a cell peeling system that reduces physical damage to cells during cell culture and can peel cells while maintaining a closed system can be provided, and a cell culture system including the above-mentioned cell peeling system can also be provided. In addition, at this time, when the scraper can move while rotating in a state where no magnetic field is applied, the scraper is not easily stuck in the tube and can smoothly pass through the tube.

[0124] The third embodiment has been verified by using iPS cells, but most types of cells can be used as long as the cells are adherent cells. In this case, the types of culture media and reagents and the timing of detachment can be appropriately changed according to the characteristics of each cell and the protocol to be implemented.

[0125] [Fourth embodiment]

[0126] Now, a cell detachment system and a cell culture system including the cell detachment system according to a fourth embodiment are described. In the drawings, the same components are denoted by the same reference numerals in principle, and description is omitted.

[0127] FIG. 11A to FIG. 11C 1 and 2 are schematic diagrams each showing an example of an overall configuration of a cell culture system including a cell detachment system according to a fourth embodiment. Fig.11A 1 is a diagram showing a state before the scraper 103 is moved to the culture surface 101 . Fig. 11B 1 is a view showing how to move the scraper 103 to the culture surface 101 . Fig. 11C 101 is a view showing a state where the movement of the scraper 103 to the culture surface 101 has been completed. Fig.11A and Fig. 11C The figures in the lead-out boxes of are detailed figures showing the culture surface 101 and the scraper 103 installed on the culture surface 101, respectively.

[0128] This system is almost the same as that in the third embodiment, except that a plurality of rows of concave curved substrates are provided as the culture surface 101 and scrapers 103 each having a spherical shape are used.

[0129] The culture surface 101 is in the form of concave curved substrates each having a diameter of φ1 mm arranged parallel to each other and formed as a part of a culture container 102 made of polystyrene.

[0130] As each scraper 103, a sphere made of SUS440 with a diameter of φ1 mm is used. When a magnetic field is applied to the scraper 103 during peeling, the scraper 103 is in close contact with the culture surface 101 and is moved in this state, thereby scraping off cells attached to the culture surface 101.

[0131] The steps of the subsections “(Initial Setting)”, “(Cell Culture)”, “(Cell Recovery)” and “(Cell Detachment)” in this system are the same as those in the third embodiment, and thus description is omitted.

[0132] As described above, in the fourth embodiment, a scraper holding portion and a scraper moving unit that moves the scraper held by the scraper holding portion to the culture surface are arranged outside the culture container. In addition, substrates each having a concave curved surface shape are installed as the culture surface, and scrapers each having a spherical shape are provided. Therefore, a cell peeling system that reduces physical damage to cells during cell culture and is also capable of peeling cells while maintaining a closed system can be provided, and a cell culture system including the above-mentioned cell peeling system can also be provided. In addition, each scraper is spherical, so it is easy to move from the scraper holding portion to the culture surface in the tube.

[0133] The fourth embodiment has been verified by using iPS cells, but most types of cells can be used as long as the cells are adherent cells. In this case, the types of culture media and reagents and the timing of detachment can be appropriately changed according to the characteristics of each cell and the protocol to be implemented.

[0134] A cell detachment system can be provided which reduces physical damage to cells during cell culture and enables a scraper to detach cells while maintaining a closed system.

[0135] Other embodiments

[0136] The embodiments of the present invention may also be implemented in the following manner: a computer of a system or device, the computer reads and executes computer executable instructions (e.g., one or more programs) recorded on a storage medium (which may also be more completely referred to as a "non-transitory computer-readable storage medium") to perform the functions in one or more of the above embodiments, and / or the computer includes one or more circuits (e.g., application-specific integrated circuits (ASICs)) for performing the functions in one or more of the above embodiments; and a method implemented by a computer of a system or device, by, for example, reading and executing computer executable instructions from a storage medium to implement the functions in one or more of the above embodiments, and / or controlling one or more circuits to implement the functions in one or more of the above embodiments. The computer may include one or more processors (e.g., a central processing unit (CPU), a microprocessing unit (MPU)), and may include a network composed of separate computers or separate processors to read and execute computer executable instructions. Computer executable instructions may be provided to the computer, for example, from a network or a storage medium. The storage medium may include, for example, a hard disk, a random access memory (RAM), a read-only memory (ROM), a memory of a distributed computing system, an optical disk (such as a compact disc (CD), a digital versatile disc (DVD), or a Blu-ray disc (BD)) TM ), flash memory devices, memory cards, etc.

[0137] While the present invention has been described with reference to exemplary embodiments, it is to be understood that the invention is not limited to the disclosed exemplary embodiments. The scope of the following claims is to be accorded the broadest interpretation so as to encompass all modifications and equivalent structures and functions.

Claims

1. A cell detachment system, include: A magnetic scraper for detaching cells attached to the culture surface; a magnetic field applying unit configured to apply a magnetic field to the scraper; and a scraper position control unit configured to control the position of the scraper by controlling the magnetic field applying unit, The scraper is configured to move while rotating when no magnetic field is applied, and to peel off attached cells by moving the scraper without rotating when a magnetic field is applied by a magnetic field applying unit.

2. The cell detachment system according to claim 1, in, The shape of the scraper includes one of a spherical shape, an ellipsoidal shape, a cylindrical shape or a cylindrical shape. 3 . The cell detachment system according to claim 1 , further comprising a scraper holding portion configured to hold the scraper in a position not in contact with the culture surface. 4 . The cell detachment system according to claim 3 , further comprising a scraper moving unit configured to move the scraper held by the scraper holding portion to the culture surface. 5 . The cell detachment system according to claim 4 , further comprising an imaging device configured to acquire information on the state of the cells attached to the culture surface. 6 . The cell detachment system according to claim 5 , further comprising an information processing unit configured to cause the scraper moving unit to move the scraper from the scraper holding portion to the culture surface based on the information acquired by the imaging device.

7. The cell detachment system according to claim 1, in, The culture surface includes a concave curved surface, and the shape of the scraper includes a spherical shape or an ellipsoidal shape.

8. The cell detachment system according to claim 1, in, The cells include adherent cells.

9. A cell culture system, include: The cell detachment system according to any one of claims 1 to 8; and A culture vessel comprising a culture surface.

10. The cell culture system according to claim 9, further comprising a cell culture unit.

11. A cell detachment system, include: A magnetic scraper for detaching cells attached to the culture surface; a magnetic field applying unit configured to apply a magnetic field to the scraper; a scraper position control unit configured to control the position of the scraper by controlling the magnetic field applying unit; and The scraper holding part is configured to hold the scraper at a position where it does not contact the culture surface. 12 . The cell detachment system according to claim 11 , further comprising a scraper moving unit configured to move the scraper held by the scraper holding portion to the culture surface. 13 . The cell detachment system according to claim 12 , further comprising an imaging device configured to acquire information on the state of cells attached to the culture surface. 14 . The cell detachment system according to claim 13 , further comprising an information processing unit configured to cause the scraper moving unit to move the scraper from the scraper holding portion to the culture surface based on information acquired by the imaging device.

15. The cell detachment system according to claim 11, in, The cells include adherent cells.

16. A cell culture system, include: The cell detachment system according to any one of claims 11 to 15; and A culture vessel comprising a culture surface. The cell culture system according to claim 16 , further comprising a cell culture unit.

Images