Cell Culture Systems

Abstract

To provide a cell culture system that can precisely control the inflow and discharge of a medium from a medium bag to a culture bag.SOLUTION: A cell culture system comprises: a plurality of culture bags made of flexible packaging material, each having a plurality of ports; a first culture medium container containing a culture medium to be transferred to the plurality of culture bags; a second culture medium container containing the culture medium transferred from the plurality of culture bags; and a control unit controlling the feeding of the culture medium. A tubular member is connected to each of the ports, and the culture bags are connected to the first culture medium container and the second culture medium container with the tubular member. One liquid feeding means is disposed in each tubular member connecting each branch portion of the tubular member with the port of the plurality of culture bags. The control unit controls the operation of each liquid feeding means to control the transfer of the culture medium in the first culture medium container, the plurality of culture bags, and the second culture medium container.SELECTED DRAWING: Figure 15

Description

[Technical Field]

[0001] The present invention relates to a cell culture technique, and more particularly to a technique for controlling the delivery of a culture medium to a culture bag used for culturing cells. [Background technology]

[0002] In recent years, there has been a demand for efficient mass cultivation of cells and tissues in an artificial environment in the fields of pharmaceutical production, gene therapy, regenerative medicine, immunotherapy, and the like. In this situation, various techniques for automatically culturing cells in a closed system using culture bags made of soft packaging materials are being researched in order to mass-cultivate cells, and the development of cell culture systems to realize these techniques is underway.

[0003] Here, in the cell culture system, it is desirable to perform liquid transfer control to transfer an appropriate amount of culture medium to the culture bag at an appropriate timing. To control the supply of culture medium, for example, a culture medium bag for supplying the culture medium to the culture bag is connected with a tube, and a pump is provided in the tube and operated to control the supply of culture medium from one culture medium bag to another. Specifically, in the cell culture system described in Patent Document 1, a culture medium bag and multiple culture bags are connected by tubes, a pump is attached to the tubes, and the pump controls the transfer of culture medium from the culture medium bag to the culture bag. [Prior art documents] [Patent documents]

[0004] [Patent Document 1] Patent No. 5892216 [Patent Document 2] International Publication No. 2016 / 190312 Brochure [Patent Document 3] International Publication No. 2016 / 190314 Brochure Summary of the Invention [Problem to be solved by the invention]

[0005] The cell culture system described in Patent Document 1 uses only one pump, thereby eliminating the need for synchronization when multiple pumps are connected in series, and aims to reduce the work of attaching the pump to the tube. On the other hand, when multiple pumps are connected in series, it is possible to synchronize the pumps by controlling their operation in detail. In particular, by forming multiple ports in the culture bag, providing pumps on each of the tubes connected to those ports, and controlling these pumps simultaneously, it becomes possible to precisely control the inflow and outflow of culture medium into the culture bag.

[0006] Therefore, the inventors developed a cell culture system in which a culture bag with multiple ports is connected in a circular shape to a culture medium bag via tubes, and pumps are placed on the tubes connected to each port, and the operation of these pumps is controlled to control the supply of culture medium to the culture bag. Such a cell culture system allows for precise control of the inflow and outflow of culture medium from the culture medium bag to the culture bag, so that, for example, it is possible to fill the culture medium bag with the amount of culture medium required from the start to the end of the culture, and then automatically culture cells by sending this culture medium to the culture bag.

[0007] In addition, in a cell culture system, it is desirable to grasp the state of the culture bag and perform optimal fluid transfer control for that state. For example, in the past, a weight sensor was used to measure the combined weight of the culture bag and the holding plate that holds it, and the weight of the culture medium was calculated based on this weight to control the supply of culture medium to the culture bag.

[0008] In contrast, if the condition of the culture bag is determined by checking the liquid thickness of the culture medium filled in the culture bag (the width of the gap between the upper and lower films of the culture bag) and controlling the liquid supply according to that condition, it becomes possible to perform a wider variety of controls in cell culture. For example, when using a culture bag with many recesses formed in the culture section to culture spheres (spheroids, aggregates) or organoids in the recesses, it is possible to prevent the spheres from moving between the recesses by adjusting the liquid thickness of the culture medium to a level that prevents the spheres from moving, and then pumping the culture medium. Therefore, the inventors used a length measuring sensor to detect the thickness of the culture medium filled in the culture bag, and by controlling the operation of the pump based on the detection information, it became possible to control the supply of culture medium as well as the thickness of the culture medium.

[0009] Measurements using weight sensors typically use load cells, which require the weight of the entire culture bag or holding plate to be applied to a minute point on the load cell, making them difficult to fix and vulnerable to impacts. Weight sensors also have temperature-sensitive characteristics, such as low temperatures, and require uniform installation in multiple locations, making installation difficult. Furthermore, when piping such as tubes is used, the weight sensor may come into contact with other housings through the tubes, which can apply tension to the tubes and result in erroneous measurements.

[0010] Here, an example of a technology related to the control of the supply of culture medium to a culture bag is the culture device described in Patent Document 2. In this culture device, a distance sensor is placed on the spacer, making it possible to detect a state in which the distance between the holding plate and the spacer has exceeded a certain level. However, this culture device does not detect the liquid thickness of the culture medium in the culture bag, and therefore does not control the liquid thickness of the culture medium. In addition, in the culture device described in Patent Document 3, the weight of the culture bag and its holder is detected using a weight detection unit. However, this culture device also does not detect the liquid thickness of the culture medium in the culture bag, and does not control the liquid thickness of the culture medium.

[0011] In addition, in a cell culture system, it is desirable to maintain the medium in the culture bag in a uniform state in order to obtain cells or spheres of uniform size. Therefore, the inventors have made the above-mentioned cell culture system rotatable, thereby rotating the culture bag to agitate the culture medium and maintaining the culture medium in the culture bag in a uniform state.

[0012] According to such a cell culture system, when adherent cells are attached to the culture section of a culture bag and cultured, the adherent cells are attached to the lower surface of the culture bag, the culture bag is then inverted upside down to attach the adherent cells to the original upper surface (new lower surface) of the culture bag, and the culture bag is then inverted upside down as appropriate for culturing, thereby doubling the culture area. In addition, after controlling the liquid thickness of the culture medium to a size that prevents the spheres from moving, the culture bag can be turned upside down, allowing the culture medium to be discharged while the spheres are retained in the recesses formed in the culture section of the culture bag.

[0013] Furthermore, in order to enable mass cultivation of cells using such a cell culture system, it is desirable to increase the culture area by using multiple culture bags. Therefore, the inventors connected multiple culture bags in parallel and connected them to a common culture medium bag, and installed pumps in each tube connected to each culture bag.By controlling the operation of these pumps, it became possible to individually control the supply of culture medium to each culture bag.

[0014] The present invention has been made in view of the above circumstances, and aims to provide a cell culture system that can precisely control the inflow and outflow of culture medium from a culture medium bag to a culture bag. [Means for solving the problem]

[0015] In order to achieve the above-mentioned object, the cell culture system of the present invention is a cell culture system comprising a culture bag made of a soft packaging material having a plurality of ports, a culture medium container for storing culture medium to be transferred to the culture bag, and a control unit for controlling the supply of the culture medium, wherein a tubular member is connected to each of the plurality of ports, and the culture bag and the culture medium container are connected in a ring shape via the tubular member, a first liquid supply means is disposed in the tubular member connected to a first port of the plurality of ports, and a second liquid supply means is disposed in a tubular member connected to a second port of the plurality of ports, and the control unit controls the operation of at least the first liquid supply means or the second liquid supply means to circulate the culture medium in the culture bag and the culture medium container. [Effects of the Invention]

[0016] According to the present invention, it is possible to provide a cell culture system that can precisely control the inflow and outflow of culture medium from a culture medium bag to a culture bag. [Brief explanation of the drawings]

[0017] [Figure 1] 1 is an explanatory diagram showing the configuration of a cell culture system according to a first embodiment of the present invention. [Figure 2] 1A and 1B are schematic diagrams showing an outline of a culture bag used in the cell culture system of each embodiment of the present invention, where FIG. 1A is a plan view and FIG. 1B is a front view. [Figure 3] 1 is a schematic diagram showing a part of a culture section provided with various recesses in a culture bag used in a cell culture system according to each embodiment of the present invention. FIG. [Figure 4] FIG. 10 is an explanatory diagram showing the configuration of a cell culture system according to a second embodiment of the present invention. [Figure 5] FIG. 10 is a front view showing an outline of a culture bag storage unit used in a cell culture system according to a second embodiment of the present invention, showing a state in which a culture medium is sealed in the culture bag. [Figure 6] FIG. 10 is a left side view showing an outline of a culture bag storage unit used in a cell culture system according to a second embodiment of the present invention, showing a state in which a culture medium is sealed in the culture bag. [Figure 7] FIG. 10 is a plan view showing an outline of a culture bag storage unit used in a cell culture system according to a second embodiment of the present invention. [Figure 8] FIG. 10 is a front view showing an outline of a culture bag storage unit used in a cell culture system according to a second embodiment of the present invention, showing a state in which no culture medium is sealed in the culture bag. [Figure 9] FIG. 10 is a left side view showing an outline of a culture bag storage unit used in a cell culture system according to a second embodiment of the present invention, showing a state in which no culture medium is sealed in the culture bag. [Figure 10] FIG. 10 is an explanatory diagram showing the configuration of a cell culture system according to a third embodiment of the present invention. [Figure 11] FIG. 10 is an explanatory diagram showing the configuration of a cell culture system according to a fourth embodiment of the present invention. [Figure 12] FIG. 10 is an explanatory diagram showing the configuration of a cell culture system according to a fifth embodiment of the present invention. [Figure 13] FIG. 10 is an explanatory diagram showing the configuration of a cell culture system according to a sixth embodiment of the present invention. [Figure 14] FIG. 10 is an explanatory diagram showing the configuration of a cell culture system according to a seventh embodiment of the present invention. [Figure 15] FIG. 13 is an explanatory diagram showing the configuration of a cell culture system according to an eighth embodiment of the present invention. DETAILED DESCRIPTION OF THE INVENTION

[0018] Hereinafter, embodiments of the cell culture system of the present invention will be described in detail, but the present invention is not limited to the specific contents of the following embodiments.

[0019] [First embodiment] First, a first embodiment of a cell culture system according to the present invention will be described with reference to FIGS. The cell culture system of this embodiment is a cell culture system comprising a culture bag made of a soft packaging material having a plurality of ports, a culture medium container for storing culture medium to be transferred to the culture bag, and a control unit for controlling the supply of the culture medium, wherein a tubular member is connected to each of the plurality of ports, and the culture bag and the culture medium container are connected in a ring shape via the tubular member, a first liquid supply means is disposed in the tubular member connected to a first port of the plurality of ports, and a second liquid supply means is disposed in the tubular member connected to a second port of the plurality of ports, and the control unit controls the operation of at least the first liquid supply means or the second liquid supply means to circulate the culture medium in the culture bag and the culture medium container.

[0020] Specifically, as shown in Figure 1, for example, a culture bag 10 having two ports 13 and a culture medium bag 40 are connected in a ring shape by a tubular member 14 such as a tube, and a liquid delivery means 30 such as a pump is arranged in each of the tubular members 14 connected to the two ports 13, and the operation of these liquid delivery means 30 is controlled by a control unit 60. In this embodiment, a culture bag 10 is accommodated in the culture bag accommodation section 20, and a single culture unit 50 using one culture bag 10 is formed by combining this culture bag accommodation section 20, the culture medium bag 40, the tubular member 14, and the liquid delivery means 30, etc.

[0021] The culture bag 10 is made of a soft packaging material, and can be formed by bonding the peripheral portions of a bottom film 11 (lower film) and a top film 12 (upper film) together by heat sealing or the like, as shown in Fig. 2. The culture bag 10 is also provided with a plurality of ports 13 (two in Fig. 2). Tubular members are connected to these ports 13, and a liquid supply means disposed on the tubular members is used to supply the culture medium, such as by injecting the culture medium into the culture bag 10 or discharging the culture medium from the culture bag 10, via the ports 13. The width of the gap (liquid thickness) between the bottom film 11 and the top film 12 changes depending on the amount of culture medium poured into the culture bag 10.

[0022] When two ports are provided in the culture bag 10, it is preferable to arrange the first port and the second port point-symmetrically with respect to the center of the culture section at both ends of the culture bag 10 so that the culture medium in the culture bag 10 can be sufficiently exchanged. The culture section is the bottom surface in contact with the culture medium in the culture bag 10, and the center of the culture section refers to the center of the horizontal surface of the culture section. The horizontal surface of the culture section is also a surface parallel to the upper end surface of the culture section.

[0023] The surface of the bottom film 11 in the culture bag 10 is used as a culture area for culturing cells or spheres. As shown in FIG. 3, the culture area of the bottom film 11 has a plurality of recesses formed therein for accommodating culture objects such as cells or spheres. By forming such recesses, spheres can be suitably formed and cultured. The arrangement of the plurality of recesses in the culture area 11 can be, for example, a staggered or lattice pattern.

[0024] 2, the bottom film 11 of the culture bag 10 is flat, and the top film 12 bulges upward as the culture medium flows in. By configuring the culture bag 10 in this way, the bottom surfaces of all recesses can be brought into contact with the mounting surface, and the thickness of the culture medium inside the culture bag 10 can be made roughly uniform. This makes it possible to reduce differences in the proliferation of the culture medium and variations in the number of cells when seeding the cells.

[0025] The culture bag 10 used in the cell culture system of this embodiment is not limited to the shape shown in Figure 2(b), and can also be formed by bonding a bottom side film 11 and a top side film 12 of the same shape together by heat sealing or the like. 2(a), the shape of the culture bag 10 near the port of the culture section is inclined relative to the port, which makes it easier for the culture medium to flow to the port. However, the shape of the culture section is not limited to this shape, and the culture section may be rectangular, for example.

[0026] The shape of the recess formed in the culture portion of the bottom film 11 can be a cone or pyramid such as a square pyramid as shown in FIG. 3(a), or a hemisphere or curved shape as shown in FIG. 3(b). It is also preferable that the shape of the recesses formed in the culture portion of the bottom film 11 is such that at least a portion of the sidewall is formed substantially vertically, as shown in Figure 3(c). In these recesses, the vertical length of the substantially vertical portion of the sidewall is longer than half the maximum diameter of the culture object. Furthermore, the shape of the region of the recess where the sidewall is substantially vertical can be, for example, cylindrical or rectangular prism-like. By making the recess in the culture bag 10 in this shape, when the culture medium is sent into the culture bag 10 from the port 13, even if the flow rate is 1 ml / min or more, it is possible to prevent the culture object from jumping out of the recess and moving to another recess.

[0027] When the culture object is a sphere, the depth of the recess is preferably 50 to 500 μm. This is because if the depth of the recess is greater than 500 μm, it may be difficult to sufficiently replace the culture medium in the recess even when the culture medium is sent to the culture bag 10, and a bag with a recess deeper than 500 μm is difficult to process. In addition, the size of a sphere is about 50 μm to 100 μm at the smallest and about 200 μm to 300 μm at the largest.

[0028] The shape of the bottom of the well is not particularly limited, but if the culture object is a sphere, it is preferable that it be conical, hemispherical, or rounded, in order to facilitate aggregation of single cells and facilitate the formation of spheres.

[0029] When the culture object is a single cell, the depth of the well is preferably 5 to 50 μm, because the size of a single cell is about 6 μm to 15 μm, with most being about 10 μm.

[0030] The shape of the opening of the well is not particularly limited and may be circular or rectangular, such as square. The width of the opening of the well can be appropriately set according to the size of the culture object. For example, when the culture object is a sphere, the lower limit of the diameter of the circle or inscribed circle of the opening of the well may be 60 μm or more, 70 μm or more, 80 μm or more, 90 μm or more, 100 μm or more, 110 μm or more, 120 μm or more, 150 μm or more, etc. Furthermore, the upper limit of the diameter of the circle or inscribed circle of the opening of the well may be 1 mm or less, 900 μm or less, 800 μm or less, 700 μm or less, 500 μm or less, etc. Furthermore, when the culture object is a single cell, the lower limit of the diameter of the circle or inscribed circle of the opening of the well may be 5 μm or more, 6 μm or more, 8 μm or more, etc. Furthermore, the upper limit of the diameter of the circle or inscribed circle of the opening of the well may be 50 μm or less, 40 μm or less, 30 μm or less, etc.

[0031] In this specification, the term "cell" includes not only single cells and spheres, but also organoids and tissues formed from these.

[0032] The culture bag 10 used in the cell culture system of this embodiment can be preferably made of polyolefin resins such as polyethylene and polypropylene. Examples of suitable materials include polyethylene, copolymers of ethylene and α-olefins, copolymers of ethylene and vinyl acetate, and ionomers using ethylene, acrylic acid, or methacrylic acid copolymers with metal ions. Other suitable materials include polyolefins, styrene-based elastomers, polyester-based thermoplastic elastomers, silicone-based thermoplastic elastomers, and silicone resins. Other suitable materials include silicone rubber, soft vinyl chloride resin, polybutadiene resin, ethylene-vinyl acetate copolymer, chlorinated polyethylene resin, polyurethane-based thermoplastic elastomers, polyester-based thermoplastic elastomers, silicone-based thermoplastic elastomers, styrene-based elastomers such as SBS (styrene-butadiene-styrene), SIS (styrene-isoprene-styrene), SEBS (styrene-ethylene-butylene-styrene), and SEPS (styrene-ethylene-propylene-styrene), polyolefin resins, and fluorine-based resins.

[0033] Furthermore, it is preferable to apply a low-adhesion surface treatment coating to prevent spheres or single cells from adhering to the culture portion of the bottom film 11. Specifically, it is preferable to apply a cell adhesion inhibitor (cell low-adhesion treatment agent) to the culture portion. Examples of cell adhesion inhibitors that can be used include phospholipid polymers, polyvinyl alcohol derivatives, phospholipid-polymer complexes, polyhydroxyethyl methacrylate, polyvinyl alcohol, agarose, chitosan, polyethylene glycol, and albumin. These may also be used in combination.

[0034] The material of the port 13 in the culture bag 10 may be, for example, a thermoplastic resin such as polyethylene, polypropylene, vinyl chloride, polystyrene elastomer, or FEP.

[0035] Furthermore, examples of materials that can be used for the tubular member 14 include silicone resin, soft vinyl chloride resin, polybutadiene resin, ethylene-vinyl acetate copolymer, polyurethane-based thermoplastic elastomer, polyester-based thermoplastic elastomer, silicone-based thermoplastic elastomer, styrene-based elastomer such as SBS (styrene-butadiene-styrene), SIS (styrene-isoprene-styrene), SEBS (styrene-ethylene-butylene-styrene), SEPS (styrene-ethylene-propylene-styrene), polyolefin resin, and fluorine-based resin.

[0036] The culture bag storage section 20 includes a stand on which the culture bag 10 is placed, a pressing member that uniforms the liquid thickness in the culture bag 10 by pressing the culture bag 10 against the stand, a top plate portion arranged opposite the pressing member, and a support mechanism that supports the pressing member so that it can move vertically relative to the top plate portion. Such a culture bag storage section will be described in detail in a second embodiment to be described later.

[0037] Furthermore, it is also preferable that the cell culture system of this embodiment be configured to include a culture bag storage section 20 having a stand on which the culture bag 10 is placed, a pressing member that presses the culture bag 10 against the stand to make the liquid thickness in the culture bag 10 uniform, a top plate section arranged opposite the pressing member, and a support mechanism that supports the pressing member so that it can move vertically relative to the top plate section, a culture unit 50 having a culture medium container storage section (not shown) that stores the culture medium bag 40, and a storage section (not shown) that stores each liquid delivery means 30 and a tubular member 14 that connects the culture bag 10 and the culture medium bag 40. By configuring the cell culture system of this embodiment in this way, it is possible to suitably control the supply of the culture medium in the culture bag 10.

[0038] The liquid delivery means 30 is disposed in a tubular member 14 that connects the culture bag 10 and the medium bag 40. As the liquid delivery means 30, it is preferable to use a pump such as a tube pump that can deliver liquid at a low speed with high precision. Such a pump can suitably control the filling of the culture medium from the culture medium bag 40 into the culture bag 10 and the discharge of the culture medium from the culture bag 10 into the culture medium bag 40 with a simple configuration.

[0039] As the liquid delivery means 30, a tube pump and an electromagnetic valve such as a pinch valve or a needle-type flow control valve may be used in combination, a diaphragm pump and an electromagnetic valve may be used in combination, or a liquid delivery means using a head difference (dead weight) or pressurization may be used in combination.

[0040] The culture medium bag 40 is used as a culture medium supply bag that circulates and supplies the culture medium to the culture bag 10. In this embodiment, the culture medium container is not limited to the culture medium bag 40 made of a soft packaging material, and a rigid container can also be used as the culture medium container.

[0041] The control unit 60 (control unit) is not particularly limited as long as it can control the operation of the liquid delivery means 30 based on preset information, and can be configured to include an input / output unit 61 (input / output unit), a PLC (programmable logic controller) 62, a relay unit 63 (relay), an operation unit 64 (touch panel, etc.), a power supply unit 65 (stabilized power supply, etc.), and a circuit breaker unit 66 (breaker). Such a configuration may also be realized by a microcomputer or computer.

[0042] The input / output unit 61 is connected by wiring cords to each of the two liquid delivery means 30. Desired control contents are preprogrammed and stored in the PLC 62. Then, based on instruction information from the PLC 62, the operation of these liquid delivery means 30 is controlled. For example, the PLC 62 in the control unit 60 can store first information corresponding to a first state in which the liquid thickness of the culture medium is zero, calculate second information by adding a predetermined amount of change to the first information, and control the operation of the liquid delivery means 30 based on the second information to control the liquid thickness of the culture medium to the second state corresponding to the second information. The first information and second information can be voltage, current, transistor output, liquid thickness of the culture medium, etc.

[0043] According to the cell culture system of this embodiment, the inflow and outflow of culture medium from the culture medium bag 40 to the culture bag 10 can be precisely controlled, so that, for example, the culture medium required from the start to the end of the culture can be filled into the culture medium bag, and then this culture medium can be sent to the culture bag 10, thereby enabling automatic cell culture.

[0044] [Second embodiment] Next, a second embodiment of the cell culture system according to the present invention will be described with reference to FIGS. The cell culture system of this embodiment differs from the cell culture system of the first embodiment in that it includes a detection unit that measures changes that occur in response to changes in the amount of culture medium in the culture bag, and controls the operation of at least the first liquid-transfer means or the second liquid-transfer means based on detection information input from the detection unit. Furthermore, except for the points described below, the other configurations can be the same as those of the first embodiment.

[0045] The change occurring in response to a change in the amount of culture medium in the culture bag is preferably a displacement of the position of a member, and a length measuring sensor is preferably used as the detection unit. Specifically, as shown in Figure 4, the cell culture system of this embodiment is equipped with a length measuring sensor 25 in the culture bag storage section 20a, and is configured to be able to control the operation of the liquid delivery means 30 based on detection information from this length measuring sensor 25.

[0046] Various types of sensors, such as electromagnetic induction type, laser type, ultrasonic type, and magnetic type, can be used as the length measuring sensor 25. There are also analog (linear) type and switch type sensors, but it is preferable to use an analog type sensor that is a type that measures constantly. Furthermore, when using an electromagnetic induction type length measuring sensor, it is preferable to attach a metal member to the measurement position, as this improves the sensitivity of the sensor. However, when using a laser type or ultrasonic type length measuring sensor, measurements can be performed without attaching a metal member to the measurement position.

[0047] Specifically, when an analog proximity sensor (electromagnetic induction type, measurement distance 1 to 6 mm, MDA-C5 manufactured by Sensortec Co., Ltd.) is used as the length measuring sensor 25, the output voltage (or output current) is roughly proportional to the measurement distance, and within the measurement distance range, a change in liquid thickness (liquid thickness of the culture medium in the culture bag) of 0.01 mm can be detected for every 0.01 V of output voltage. For example, if the horizontal surface area of the culture area is 50 cm 2 When using a culture bag with a thickness of 0.01 mm, the volume of culture medium is 0.05 ml. In this case, if the liquid delivery rate is 2 ml / min, the culture medium can be delivered in 1.5 seconds, and the liquid thickness can be increased or decreased by 0.01 mm. Also, if the liquid delivery rate is 0.5 ml / min, the culture medium can be delivered in 6 seconds, and the liquid thickness can be increased or decreased by 0.01 mm.

[0048] Furthermore, by using an analog distance sensor (electromagnetic induction type, measuring distance 0 to 12 mm, manufactured by Fuji Electric Co., Ltd., PE2-LA10D) as the length measuring sensor 25, it is possible to control a liquid thickness up to a larger value of, for example, about 8 mm.

[0049] When the liquid thickness is measured directly using the length measuring sensor 25, for example, the length measuring sensor 25 can be placed on a stand on which the culture bag is placed, and a metal member can be placed on the pressing member, and the distance to the metal member can be measured using the length measuring sensor 25. In this case, the liquid thickness can be obtained by multiplying the output voltage of the length measuring sensor by a predetermined coefficient and performing correction by subtracting the thickness of the film or the like.

[0050] As will be described later, it is also possible to place a length measuring sensor 25 on the top plate that supports the pressing member, and place a metal member on the pressing member, and use the length measuring sensor 25 to measure the distance to the metal member. In this case, the liquid thickness can be obtained by multiplying the output voltage of the length measuring sensor 25 by a predetermined coefficient to calculate the distance from the length measuring sensor 25 to the metal member, and then subtracting the calculated distance, the thickness of the metal member, the height of the pressing member, and the thickness of the film from the distance from the top surface of the stand on which the culture bag is placed to the bottom surface of the length measuring sensor 25.

[0051] Furthermore, the change occurring in response to the change in the amount of culture medium in the culture bag can be detected as a change in weight of the culture bag storage section 20a on which the culture bag 10 is placed, and a weight sensor can be used as the detection section. Then, the control section 60 can calculate the liquid thickness of the culture medium based on the change in weight, and the operation of the liquid delivery means can be controlled based on this liquid thickness. In other words, the weight at the time of measurement is subtracted from the weight at the time when the liquid thickness of the culture bag 10 is zero, converted into volume, and then this is divided by the area of the horizontal surface of the culture section to calculate the liquid thickness, and the operation of the liquid delivery means can be controlled based on this liquid thickness.

[0052] 5 to 9, the culture bag storage section 20a in this embodiment has top plate support parts 22 erected at the four corners of a stand 21, and a top plate part 23 fixed to the top plate support parts 22. Furthermore, guide pins 273 are provided on the periphery of the top plate part 23, and pass through guide holes provided in an attachment part 272 of the pressing member 27, so that the pressing member 27 is provided to be movable vertically along the guide pins 273. Furthermore, the top panel portion 23 is provided with a top panel opening 231, through which the inside of the culture bag receiving portion 20a can be seen.

[0053] Furthermore, permanent magnets 26 are provided at the four corners of the top plate portion 23 as biasing means for biasing the pressing member 27 downward, and permanent magnets 2721 are provided as biasing means at positions on the mounting portion 272 corresponding to the permanent magnets 26. These permanent magnets are arranged with their polarities facing each other, and the repulsive force acting between the permanent magnets 26 and 2721 allows the pressing portion 271 of the pressing member 27 to move vertically while pressing vertically against the top plate side film 12 of the culture bag 10. The biasing means 73 is not limited to a permanent magnet. Alternatively, the biasing means 73 may be omitted, and the pressing member 27 may be moved up and down relative to the top plate side film 12 by its own weight.

[0054] The pressing member 27 is sized so that the bottom surface of the pressing portion 271 is positioned inside the culture section of the culture bag 10. That is, the shape of the horizontal surface of the pressing portion 271 is the same as the shape of the horizontal surface of the culture section of the culture bag 10, and the pressing portion 271 presses only within the region of the culture section. In this way, the pressing member 27 presses the culture bag 10, thereby suppressing the occurrence of undulations in the top plate-side film 12. As a result, while the culture medium is being injected or discharged, the top plate-side film 12 is kept parallel to the mounting surface of the stand 21, and the liquid thickness of the culture medium in the culture bag 10 becomes uniform.

[0055] The pressing member 27 may be made of, for example, a synthetic resin such as polycarbonate, or may be made of glass. In addition, it is preferable that the pressing member 27 is partially or entirely transparent so that the progress of the culture and the state of the culture object can be checked.

[0056] When the culture bag 10 is filled with the culture medium, the pressing member 27 is pushed up by the culture bag 10, as shown in FIGS. On the other hand, when the culture medium is discharged from the culture bag 10 by the liquid supply means 30 arranged on the tubular member 14 via the tubular member 14 connected to the port 13 of the culture bag 10, the liquid thickness of the culture medium in the culture bag 10 decreases accordingly, and the pressing member 27 descends accordingly.

[0057] 8 and 9, the culture bag 10 is pressed by the pressing member 27 until the upper and lower films are in contact with each other (the plurality of recesses formed in the bottom film 11 are closed by the top film 12). At this time, the liquid thickness of the culture medium in the culture bag 10 becomes zero.

[0058] The sensor support part 24 is fixed to the periphery of the top plate part 23, and has the length measurement sensor 25 attached to its tip. In addition, a metal member 2722 is attached to the attachment part 272 of the pressing member 27 so as to face the length measurement sensor 25. Note that the arrangement of the sensor support part 24 and the length measurement sensor 25 is not limited to this, and they can be attached at any position with respect to the top plate part 23 as long as the distance to the metal member 2722 can be measured.

[0059] The distance to the metal member 2722 is measured by the length measuring sensor 25, and the liquid thickness of the culture medium in the culture bag 10 is calculated, thereby making it possible to detect the liquid thickness. For example, the liquid thickness can be obtained by multiplying the output voltage of the length measuring sensor 25 by a predetermined coefficient to calculate the distance from the length measuring sensor 25 to the metal member 2722, and then subtracting the calculated distance, the thickness of the metal member 2722, the height of the pressing member, and the thicknesses of the upper and lower films from the distance from the upper surface of the frame 21 to the lower surface of the length measuring sensor 25.

[0060] The operation of the liquid delivery means 30 is controlled by the control unit 60 based on the detection information from the length measurement sensor 25. At this time, the control unit 60 calculates the liquid thickness of the culture medium in the culture bag 10 based on the output voltage (or output current) input (transmitted) from the length measurement sensor 25, and controls the start and stop of the operation of the liquid delivery means 30, the rotation speed, etc. based on the liquid thickness, thereby making it possible to control the liquid thickness to a desired size.

[0061] The control unit 60 (control unit) is not particularly limited as long as it can control the operation of the liquid delivery means 30 based on the detection information from the length measurement sensor 25, and the same one as in the first embodiment can be used. After discharging the culture medium in the culture bag 10 and storing the output voltage corresponding to a state in which the liquid thickness is zero by the control unit 60, the operation of the liquid delivery means 30 such as a pump can be controlled based on the detection information from the length measurement sensor 25 so that the culture medium in the culture bag 10 has the desired liquid thickness, and the culture medium can be supplied to the culture bag 10.

[0062] In this embodiment, the culture bag storage section 20a can also be configured by arranging the length measuring sensor 25 on the stand 21 on which the culture bag 10 is placed and by arranging a metal member 2722 on the pressing member 27. Also, a weight sensor can be used to detect the weight of the culture bag 10 including the culture medium and output the detected weight to the control unit 60, and the control unit 60 can calculate the liquid thickness of the culture medium based on the weight of the culture medium and control the operation of the liquid sending means 30.

[0063] Here, there are three types of control for the medium supply in the cell culture system: full volume exchange, half volume exchange, and circulating supply. In a full exchange, all of the culture medium in the culture bag 10 is discharged, the culture bag 10 is emptied, and then the culture bag 10 is filled with culture medium again, and the liquid thickness of the culture medium in the culture bag 10 is returned to its original liquid thickness. In half-exchange, half of the culture medium in the culture bag 10 is discharged, and then the culture bag 10 is filled with culture medium again, returning the liquid thickness of the culture medium in the culture bag 10 to its original liquid thickness. In the circulating liquid transfer, the liquid is continuously transferred while the liquid thickness of the culture medium in the culture bag 10 is maintained within a certain range. Full and half volume exchanges can be performed by first discharging the medium and then supplying it until the medium reaches a certain liquid thickness, and therefore can be controlled relatively easily.

[0064] However, it is difficult to operate the circulating liquid feed stably, and it is necessary to perform special control on the liquid feed means 30. For example, if the first liquid delivery means and the second liquid delivery means are operated simultaneously and the liquid delivery speeds are exactly the same, it is theoretically possible to perform circulatory liquid delivery while maintaining the liquid thickness. However, since there is generally variation in the liquid delivery speed of pumps, simply operating them simultaneously will result in a change in the liquid thickness due to the error. Therefore, if the discharge pump is faster than the inlet pump, the culture medium in the culture bag 10 will be emptied, and if the discharge pump is slower than the inlet pump, the culture medium in the culture bag 10 will continue to increase, which may damage the bag.

[0065] Therefore, in the cell culture system of this embodiment, the culture medium delivery rates by the first and second fluid delivery means are controlled to different rates. Then, of the rates of delivery of culture medium by the first and second fluid delivery means, the fluid delivery rate by the slower fluid delivery means is kept constant, and the stop and start of the operation of the faster fluid delivery means is controlled based on input information from the detection unit so that the liquid thickness of the culture medium in the culture bag 10 is within a certain range, thereby enabling stable operation of circulatory fluid delivery.

[0066] Specifically, when starting the circulating liquid supply, first, with the culture medium in the culture bag 10 empty, the pump on the discharge side is stopped, and the pump on the injection side is operated at a speed V, and the culture medium begins to be filled so that the liquid thickness of the culture medium in the culture bag 10 becomes, for example, a maximum liquid thickness of 3d. Next, when the liquid thickness of the medium in the culture bag 10 reaches 2d, operate the pump on the discharge side at a speed V'. Keep the pump on the injection side operating at speed V. At this time, set V < V' so that the speed of the pump on the discharge side is faster than the speed of the pump on the injection side.

[0067] Next, when the liquid thickness of the medium in the culture bag 10 reaches d, stop the pump on the discharge side. Keep the pump on the injection side operating at speed V. As a result, the liquid thickness of the medium in the culture bag 10 increases toward reaching the maximum liquid thickness of 3d again. And by repeating these operations, it becomes possible to keep the liquid thickness of the medium in the culture bag 10 within a certain range.

[0068] Conversely, it is also possible to make the speed of the pump on the injection side faster than the speed of the pump on the discharge side and control the stop and start of the operation. Specifically, first, fill the culture bag 10 with the medium so that the liquid thickness reaches 3d. With the pump on the injection side stopped, operate the pump on the discharge side at speed V and start discharging the medium so that the liquid thickness of the medium in the culture bag 10 reaches d. Next, when the liquid thickness of the medium in the culture bag 10 reaches d, operate the pump on the injection side at a speed V'. Keep the pump on the discharge side operating at speed V. At this time, set V < V' so that the speed of the pump on the injection side is faster than the speed of the pump on the discharge side.

[0069] Next, when the liquid thickness of the medium in the culture bag 10 reaches 2d, stop the pump on the injection side. Keep the pump on the discharge side operating at speed V. As a result, the liquid thickness of the medium in the culture bag 10 decreases toward reaching d again. And by repeating these operations, it becomes possible to keep the liquid thickness of the medium in the culture bag 10 within a certain range.

[0070] Note that the pump on the injection side is operated at a speed V', and the pump on the discharge side is operated at a speed V, with V < V'. The speed of the pump on the injection side is made faster than the speed of the pump on the discharge side, and each is stopped when the liquid thickness of the culture medium in the culture bag 10 reaches a certain value based on the input information from the detection unit. This is also possible. However, with such a configuration, the operation start (pump motor ON) and stop (pump motor OFF) of the pump are frequently repeated. Although control is possible to some extent, damage is caused to the motor for driving the pump. Therefore, it is more preferable to control as described above.

[0071] According to such a cell culture system of the present embodiment, since the liquid thickness of the culture medium in the culture bag can be grasped, it is possible to grasp the remaining amount of the culture medium in the culture bag. Also, the timing when the upper film and the lower film of the culture bag come into contact can be grasped. Thereby, it is possible to prevent the state where there is no culture medium in the culture bag. It is also possible to detect a liquid feeding error due to a pump failure or inappropriate installation.

[0072] Furthermore, by controlling the operation of the liquid feeding means, it is also possible to suitably perform the circulating liquid feeding of the culture medium in the cell culture system. Also, the width of the gap between the upper film of the culture bag and the lower film having a plurality of recesses can be controlled to a size (a gap of about 50 μm) where the spheres cannot move, thereby preventing the movement of the spheres between the recesses.

[0073] [Third Embodiment] Next, a third embodiment of the cell culture system according to the present invention will be described with reference to FIG. 10. 10, the cell culture system of this embodiment is characterized by including a housing 70 that stores the culture unit 50, a rod-shaped member 71 that stands upright from the side wall of the housing 70, a drive unit 72 that rotates the housing 70 along the central axis of the rod-shaped member 71, and a support mechanism 73 that rotatably supports the housing 70. Except for the points described below, the other configurations can be the same as those of the first or second embodiment.

[0074] In this embodiment, it is preferable to use a tube pump as the liquid delivery means 30, and it is also preferable to arrange the liquid delivery means 30 in each tubular member 14 connected to multiple ports 13 provided in the culture bag 10.

[0075] By disposing the liquid supply means 30 in this manner, even when the culture bag 10 is pressed by a pressing member, the culture medium can be prevented from flowing back into the culture medium bag 40 due to the pressure of the pressing member. Furthermore, even if air bubbles are generated in the tubular member 14 connected to the culture bag 10, by operating the liquid delivery means 30, these air bubbles can be transferred to the culture medium bag 40 and discharged. Furthermore, even if the culture bag 10 is rotated or otherwise changed in position, it is not affected by the difference in head pressure, and sudden pressure fluctuations within the culture bag 10 can be prevented, thereby eliminating any adverse effects on the cells.

[0076] Furthermore, in this embodiment, the housing 70 fixes and holds the culture bag storage section 20, the liquid delivery means 30, the tubular member 14, and the culture medium bag 40 in the culture unit 50, and the arrangement of these in the housing 70 is not particularly limited, nor is the method of fixing them limited. The rod-shaped member 71 is a rotation axis for rotating the housing 70, and may be formed separately from the housing 70 and fixed to the housing 70, or may be formed integrally with the housing 70. The rod-shaped member 71 may be formed in a cylindrical shape, and the wiring cord connected to the input / output unit 61 of the control unit 60 may be connected to the liquid delivery means 30 and the length measuring sensor 25 through its interior.

[0077] The driving unit 72 rotates the rod-shaped member 71 along its central axis, thereby rotating the housing 70. The driving unit 72 is connected to the input / output unit 61 of the control unit 60 via a wiring cord, and can be controlled by the control unit 60. Alternatively, another control unit may be used to control the driving unit 72. The support mechanism 73 supports the rod-shaped member 71, thereby supporting the housing 70 in a rotatable manner.

[0078] According to the cell culture system of this embodiment, it is possible to transfer culture medium to the culture bag 10 even when the culture bag 10 is rotated to make the culture section vertical or when the culture bag 10 is turned upside down. Furthermore, for example, when adhering and culturing adherent cells to the culture section of culture bag 10, the culture area can be doubled by adhering the adherent cells to the lower surface of culture bag 10, then inverting culture bag 10 upside down to adhering the adherent cells to the original upper surface (new lower surface) of culture bag 10, and then appropriately inverting culture bag 10 upside down for culturing. Furthermore, after controlling the liquid thickness of the culture medium to a size that prevents the spheres from moving, the culture bag 10 is turned upside down, allowing the spheres to be retained in the recesses formed in the culture section of the culture bag 10, and dead cells and culture medium can be discharged, or the entire amount or half of the culture medium can be replaced in an appropriate manner.

[0079] [Fourth embodiment] Next, a fourth embodiment of the cell culture system according to the present invention will be described with reference to FIG. 11, the cell culture system of this embodiment is characterized by including a second housing 70a that stores (stacked) a plurality of culture units 50 (50-1, 50-2, 50-3), a second rod-shaped member 71a that stands from the side wall of the second housing 70a, a second drive unit 72a that rotates the second housing 70a along the central axis of the second rod-shaped member 71a, and a second support mechanism 73a that rotatably supports the second housing 70a. Except for the points described below, the other configurations can be the same as those of the third embodiment.

[0080] The housing 70a fixes and holds the culture bag accommodating sections 20, the liquid sending means 30, the tubular members 14, and the culture medium bags 40 in the plurality of culture units 50, respectively. The rod-shaped member 71a is a rotation axis for rotating the housing 70a, and may be formed separately from the housing 70a and fixed to the housing 70a, or may be formed integrally with the housing 70a. The rod-shaped member 71a may be formed in a cylindrical shape, and the wiring cord connected to the input / output unit 61 of the control unit 60 may be connected to the liquid delivery means 30 and the length measuring sensor 25 through its interior.

[0081] The driving unit 72a rotates the rod-shaped member 71a along its central axis, thereby rotating the housing 70a. The driving unit 72a is connected to the input / output unit 61 of the control unit 60 via a wiring cord, and can be controlled by the control unit 60. Alternatively, another control unit may be used to control the driving unit 72a. The support mechanism 73a supports the rod-shaped member 71a, thereby supporting the housing 70a in a rotatable manner.

[0082] According to the cell culture system of this embodiment, in addition to the effects of the third embodiment, the culture area can be increased by using multiple culture bags 10, and the inflow and outflow of culture medium into each culture bag 10 can be precisely controlled, making it possible to culture large quantities of cells.

[0083] [Fifth embodiment] Next, a fifth embodiment of the cell culture system according to the present invention will be described with reference to FIG. The cell culture system of this embodiment is characterized in that a plurality of culture bags are connected in parallel to one culture medium container (culture medium bag 40) by tubular members, and one fluid supply means is provided on each tubular member connecting each branched portion of the tubular member to the ports of the plurality of culture bags. Furthermore, except for the points described below, the other configurations can be the same as those of the cell culture system of the first embodiment.

[0084] 12, each culture bag 10 is provided with two ports 13. Tubular members 14 are connected to these ports 13, and each tubular member 14 is connected to another culture bag 10 and a culture medium bag 40. In addition, the tubular member 14 is partially branched and connected to the culture bag 10 and the culture medium bag 40.

[0085] A liquid delivery means 30 is provided in each of the tubular members 14 on both sides of each port of the culture bag 10. In this case, one liquid delivery means 30 is provided in each of the tubular members 14 that connect a certain branched portion of the tubular member 14 to the port 13 of the culture bag 10. It is preferable to use a tube pump as these liquid delivery means 30. Furthermore, the input / output unit 61 in the control unit 60 is connected to each of the liquid delivery means 30 by a wiring cord. Then, the operation of these liquid delivery means 30 is controlled based on instruction information from the PLC 62. These liquid delivery means 30 can be controlled individually.

[0086] It is also preferable that the cell culture system of this embodiment includes a second culture unit 50b (a multiple culture unit in which multiple culture bags are connected to a common culture medium container) having a base on which the culture bag 10 is placed, a pressing member that presses the culture bag 10 against the base to make the liquid thickness in the culture bag 10 uniform, a top plate portion arranged facing the pressing member, and a support mechanism that supports the pressing member so that it can move vertically relative to the top plate portion, a culture medium container holding portion (not shown) that holds the culture medium bag 40, and a holding portion (not shown) that holds each liquid delivery means 30 and a tubular member 14 that connects the multiple culture bags 10 and the culture medium bag 40. By configuring the cell culture system of this embodiment in this way, it becomes possible to suitably control the supply of culture medium in each culture bag 10.

[0087] Here, when connecting multiple culture bags 10 to a common culture medium bag 40, if a pump is placed on only one of the tubular members 14 connected to a culture bag 10 having two ports, or if solenoid valves are placed on both of the tubular members 14, the culture medium will flow more easily through the branched tubular members 14, which creates the problem that the culture medium cannot be uniformly delivered to each culture bag 10. In contrast, according to the cell culture system of this embodiment, a pump is provided as a liquid delivery means 30 in each of the tubular members 14 on both sides of each port of the culture bag 10, and the operation of these liquid delivery means 30 can be controlled, making it possible to uniformly deliver culture medium to multiple culture bags 10.

[0088] Furthermore, according to the cell culture system of this embodiment, even when a culture medium bag 40 common to multiple culture bags 10 is used, the operation of each liquid delivery means 30 can be controlled individually, making it possible to finely control the delivery of culture medium to each of the multiple culture bags 10 separately. Furthermore, in this way, since the culture can be suitably carried out using a plurality of culture bags 10, it becomes possible to culture a large amount of cells.

[0089] [Sixth embodiment] Next, a sixth embodiment of the cell culture system according to the present invention will be described with reference to FIG. The cell culture system of this embodiment, like the second embodiment, is characterized in that it includes a detection unit that measures changes that occur in response to changes in the amount of culture medium in the culture bag, and controls the operation of each fluid delivery means based on detection information input from the detection unit, and differs from the cell culture system of the fifth embodiment in that, except for the points described below, the other configurations can be the same as those of the fifth embodiment.

[0090] 13, the input / output unit 61 in the control unit 60 is connected to each of the liquid delivery means 30 by a wiring cord. The input / output unit 61 is also connected to each of the length measurement sensors 25 by a wiring cord. Then, based on the detection information from each of the length measurement sensors 25, the control unit 60 can individually control the operation of each of the liquid delivery means 30 (provided for the same culture bag 10) corresponding to each of the length measurement sensors 25.

[0091] For example, the control unit 60 can calculate the liquid thickness of the culture medium in the culture bag 10 based on the output voltage input from the length measuring sensor 25, and control the start and stop of the operation of the liquid delivery means 30 and the rotation speed, etc. based on the liquid thickness, thereby controlling the liquid thickness to the desired size.

[0092] It is also preferable that the cell culture system of this embodiment comprises a stand on which the culture bag 10 is placed, a pressing member that presses the culture bag 10 against the stand to make the liquid thickness in the culture bag 10 uniform, a top plate portion arranged facing the pressing member, and a support mechanism that supports the pressing member so that it can move vertically relative to the top plate portion, and also comprises a third culture unit 50c (a multiple culture unit having a length measuring sensor in which a plurality of culture bags are connected to a common culture medium container) that has a plurality of culture bag storage sections 20a equipped with a length measuring sensor, a culture medium container storage section (not shown) that stores culture medium bags 40, and a storage section (not shown) that stores each liquid delivery means 30 and a tubular member 14 that connects the plurality of culture bags 10 to the culture medium bag 40.

[0093] By configuring the cell culture system of this embodiment in this manner, in addition to the effects of the cell culture system of the fifth embodiment, it is possible to more precisely control the supply of culture medium in the culture bag 10, and it is also possible to control the liquid thickness of the culture medium in the culture bag 10.

[0094] [Seventh embodiment] Next, a seventh embodiment of the cell culture system according to the present invention will be described with reference to FIG. 14, the cell culture system of this embodiment is characterized by including a third housing 70b that stores multiple culture units (50b, 50c), a third rod-shaped member 71b standing from the side wall of the third housing 70b, a third drive unit 72b that rotates the third housing 70b along the central axis of the third rod-shaped member 71b, and a third support mechanism 73b that rotatably supports the third housing 70b. Except for the points described below, the other configurations can be the same as those of the fifth or sixth embodiment.

[0095] The housing 70b fixes and holds the culture bag storage section 20, the liquid delivery means 30, the tubular member 14, and the culture medium bag 40 in the multiple culture units (50b, 50c). The rod-shaped member 71b is a rotation axis for rotating the housing 70b, and may be formed separately from the housing 70b and fixed to the housing 70b, or may be formed integrally with the housing 70b. The rod-shaped member 71b may be formed in a cylindrical shape, and the wiring cord connected to the input / output unit 61 of the control unit 60 may be connected to the liquid delivery means 30 and the length measuring sensor 25 through its interior.

[0096] The driving unit 72b rotates the rod-shaped member 71b along its central axis, thereby rotating the housing 70b. The driving unit 72b is connected to the input / output unit 61 of the control unit 60 via a wiring cord, and can be controlled by the control unit 60. Alternatively, another control unit may be used to control the driving unit 72b. The support mechanism 73b supports the rod-shaped member 71b, thereby supporting the housing 70b in a rotatable manner.

[0097] According to the cell culture system of this embodiment, when a culture medium bag 40 common to multiple culture bags 10 is used, it is possible to properly deliver culture medium to each culture bag 10 even when the culture bags 10 are rotated to make the culture section vertical or are turned upside down.

[0098] [Eighth embodiment] Next, an eighth embodiment of the cell culture system according to the present invention will be described with reference to FIG. The cell culture system of this embodiment is a cell culture system comprising a plurality of culture bags made of soft packaging material, each having a plurality of ports, a first culture medium container for accommodating culture medium to be transferred to the plurality of culture bags, a second culture medium container for accommodating culture medium transferred from the plurality of culture bags, and a control unit for controlling the transfer of culture medium, wherein a tubular member is connected to each of the plurality of ports, and the plurality of culture bags are connected to the first culture medium container and the second culture medium container by the tubular member, and one liquid transfer means is disposed on each of the tubular members connecting each branch portion of the tubular member to the ports of the plurality of culture bags, and the control unit controls the operation of each liquid transfer means to control the transfer of culture medium in the plurality of culture bags, the first culture medium container, and the second culture medium container.

[0099] That is, in the cell culture system of this embodiment, the culture medium is not circulated but is sent from the first culture medium container 41 to each culture bag 10 and from each culture bag 10 to the second culture medium container 42. The cell culture system of this embodiment also makes it possible to precisely and appropriately control the liquid thickness of the culture medium in the culture bag 10, similar to the cell culture system of the sixth embodiment.

[0100] Moreover, the length measuring sensor 25 can be omitted from the culture bag storage unit 20a in the cell culture system of this embodiment. In such a cell culture system, it is possible to suitably control the supply of the culture medium in the culture bag 10, as in the cell culture system of the fifth embodiment.

[0101] Furthermore, the cell culture system of this embodiment can be combined with the cell culture system of the seventh embodiment, so that the culture medium can be properly delivered to each culture bag 10 even when multiple culture bags 10 are rotated to make the culture section vertical or are turned upside down.

[0102] In addition, for the culture bag used in the cell culture system of this embodiment, it is also preferable that the top plate side film 12 has a top plate protrusion on the inside of the culture bag 10, the width of the top plate protrusion is smaller than the width of the opening of the recess formed in the bottom plate side film 11, and the height of the top plate protrusion is smaller than the minimum diameter of the culture object, so that when the top plate protrusion comes into contact with part of the upper end surface of the culture section, a gap is formed between the upper end surface and the underside of the top plate side film 12.

[0103] By configuring the culture bag used in the cell culture system of this embodiment in this way, the top plate protrusion can be brought into contact with a portion of the top surface, forming a gap between the top surface and the top plate that is smaller than the size of the culture object. This allows liquid such as culture medium to pass through the gap while preventing the culture object, such as spheres, from moving.

[0104] When the culture object is a sphere, the culture device of this embodiment can be used in processes such as forming the sphere, culturing and growing the cells while maintaining the sphere state, and inducing differentiation in the sphere state. It can also be used in methods such as forming spheres using differentiated cells obtained from iPS cells, ES cells, or other stem cells that have already undergone a differentiation induction process, or in methods in which differentiated cells are frozen and stored, and then thawed again to form spheres.

[0105] As described above, according to the cell culture system of the embodiment of the present invention, the inflow and outflow of the culture medium from the culture medium container to the culture bag can be precisely controlled. In addition, by knowing the liquid thickness of the culture medium in the culture bag, it is possible to know the remaining amount of culture medium in the culture bag and the timing when the upper and lower films of the culture bag come into contact, making it possible to prevent the culture bag from running out of culture medium and to detect fluid transfer errors caused by pump failure or improper installation.

[0106] Furthermore, it is possible to transfer culture medium to the culture bag even when the culture section is rotated to make it vertical or when it is turned upside down. For example, when adhering cells to the culture section of the culture bag and culturing them, it is possible to double the culture area by appropriately turning the culture bag upside down. In addition, by using multiple culture bags, the culture area can be increased and the inflow and outflow of culture medium into each culture bag can be precisely controlled, making it possible to culture large quantities of cells.

[0107] The present invention is not limited to the above-described embodiments, and various modifications can be made within the scope of the present invention. For example, the culture bag may be provided with three or more ports, and pumps may be provided on tubular members connected to the ports, and these pumps may be controlled based on information from a detector. Alternatively, the arrangement of the pressing member and length measuring sensor in the culture bag storage unit may be changed. [Industrial Applicability]

[0108] The present invention can be suitably used when efficiently producing large quantities of uniformly sized spheres, organoids, etc. [Explanation of symbols]

[0109] 10 culture bags 11, 11a, 11b Bottom film 111, 111a, 111b recess 12 Top panel film 13 ports 14 Tubular member 20, 20a Culture bag storage section 21 Mounting stand 22 Top plate support 23 Top plate 231 Top panel opening 24 Sensor support part 25 Length measuring sensor 26 Permanent Magnets 27 Pressing member 271 Pressing part 272 Mounting part 2721 Permanent magnets 2722 Metal parts 273 Guide Pin 30 Liquid delivery means 40,41,42 Culture medium bag 50, 50a, 50b, 50c, 50d Culture Unit 60 Control Unit 61 Input / output section 62 PLC 63 Relay section 64 Operation section 65 Power supply section 66 Wiring breaker 70, 70a, 70b housing 71, 71a, 71b Rod-shaped members 72, 72a, 72b Drive unit 73,73a,73b Support mechanism

Claims

1. A cell culture system including a plurality of culture bags made of a soft packaging material each having a plurality of ports, a first culture medium container for accommodating culture media to be transferred to the plurality of culture bags, a second culture medium container for accommodating culture media transferred from the plurality of culture bags, and a control unit for controlling the transfer of the culture media, a tubular member is connected to each of the plurality of ports, and the plurality of culture bags are connected to the first culture medium container and the second culture medium container by the tubular member; a tubular member connecting each branched portion of the tubular member to the ports of the plurality of culture bags, each of which is provided with a liquid supply means; the control unit controls the operation of each first fluid supply means disposed in a tubular member connecting the first culture medium container and the plurality of culture bags, and each second fluid supply means disposed in a tubular member connecting the second culture medium container and the plurality of culture bags, thereby controlling the transfer of culture medium between the first culture medium container, the plurality of culture bags, and the second culture medium container; a detection unit for measuring a change occurring in response to a change in the amount of culture medium in the culture bag; controlling the operations of the first liquid-transfer means and the second liquid-transfer means based on the detection information input from the detection unit, and controlling the liquid-transfer rates of the culture medium by the first liquid-transfer means and the second liquid-transfer means to be different; The liquid transfer speed of the slower liquid transfer means out of the liquid transfer speeds of the culture medium by the first liquid transfer means and the second liquid transfer means is kept constant, and the stop and start of the operation of the liquid transfer means with a faster speed is controlled based on input information from the detection unit so that the liquid thickness of the culture medium in the culture bag is within a certain range. A cell culture system characterized by:

2. 2. The cell culture system according to claim 1, wherein the liquid delivery means is a tube pump.

3. 2. The cell culture system according to claim 1, wherein the change occurring in response to the change in the amount of culture medium is a displacement of the position of a member, and the detection unit is a length measurement sensor.

4. 2. The cell culture system according to claim 1, wherein the change occurring in response to the change in the amount of culture medium is a change in weight of a storage section in which the culture bag is placed, and the detection section is a weight sensor.

5. The control unit calculates the liquid thickness of the culture medium in the culture bag, and controls the operation of at least the first liquid supply means or the second liquid supply means based on the liquid thickness. The cell culture system according to claim 1 .

6. a culture bag accommodating section including a stand on which the culture bag is placed, a pressing member that presses the culture bag against the stand to make the liquid thickness in the culture bag uniform, a top plate portion disposed facing the pressing member, and a support mechanism that supports the pressing member so as to be vertically movable relative to the top plate portion; a culture unit having a culture medium container accommodating section that accommodates the first culture medium container and the second culture medium container; and an accommodating section that accommodates the first liquid supply means, the second liquid supply means, and tubular members that connect the culture bag to the first culture medium container and the second culture medium container. The cell culture system according to any one of claims 1 to 5.

7. 7. The cell culture system according to claim 6, wherein a length measuring sensor is disposed on the top plate portion, and the length measuring sensor measures the distance to the metal member disposed on the pressing member.

8. The culture system includes a second housing that stores a plurality of the culture units, a second rod-shaped member that stands upright from a side wall of the second housing, a second drive unit that rotates the second housing along the central axis of the second rod-shaped member, and a second support mechanism that rotatably supports the second housing. The cell culture system according to claim 7 .

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