Cell Culture Systems

Abstract

This cell culture system (10) comprises: a reactor (12); a distribution channel (16) through which a medium flows in and out of the reactor (12); a waste liquid channel (18) through which the medium is discharged from the distribution channel (16); and a waste liquid container (62) capable of storing the medium transported via the waste liquid channel (18). The waste liquid channel (18) has a temporary storage part (64) capable of temporarily storing the medium. The waste liquid container (62) is located below the temporary storage part (64) in the direction of gravity. The temporary storage part (64), which is located above the distribution channel (16) in the direction of gravity, temporarily stores the medium discharged from the distribution channel (16) and allows the medium to flow out toward the waste liquid container (62).

Description

[Technical Field]

[0001] The present invention relates to a cell culture system that cultures cells in a reactor by flowing a culture medium into and out of the reactor. [Background technology]

[0002] In regenerative medicine, cells from a living body are collected, cultured, and then administered to a patient. For cell culture processing, a cell culture system using a cell culture vessel (reactor) with hollow fibers inside a case is used, as disclosed in, for example, Japanese Patent Application Laid-Open No. 2017-143775. In the cell culture system, cells are seeded inside the hollow fibers of the reactor, and then a culture medium is pumped into the reactor via a distribution channel to culture the cells. The culture medium that flows out of the reactor during culture is discharged into a waste liquid collection container (waste liquid section). Summary of the Invention

[0003] In this type of cell culture system, the waste liquid section, which is made up of a medical bag or the like, is suspended from a stand and positioned above the reactor in the direction of gravity. This allows the cell culture system to apply positive pressure to the reactor and the distribution channel through the culture medium flowing into the waste liquid section, thereby preventing excess air (air bubbles) from entering the reactor.

[0004] However, when a large amount of culture medium is discharged into the waste liquid section over a long period of time, if a small-capacity medical bag is used, the waste liquid section must be replaced frequently, which places a heavy burden on the operator. Even if the waste liquid section is configured using a large tank or the like, the operator must still remove the waste liquid section that has accumulated a large amount of culture medium above the reactor in the direction of gravity, which also places a heavy burden on the operator.

[0005] The present invention has been made in consideration of the above-mentioned problems, and aims to provide a cell culture system that can appropriately apply positive pressure to the reactor and circulation path, and can reduce the workload of replacing and removing the waste liquid section.

[0006] In order to achieve the above-mentioned object, one aspect of the present invention is a cell culture system comprising: a reactor for culturing cells based on the flow of a culture medium; a flow path through which the culture medium flows into and out of the reactor; a waste liquid path connected to the flow path and for discharging the culture medium from the flow path; and a waste liquid container connected to the waste liquid path and capable of storing the culture medium that has passed through the waste liquid path, wherein the waste liquid path has a temporary storage section capable of temporarily storing the culture medium, the waste liquid container is located below the temporary storage section in the direction of gravity, and the temporary storage section is located above the flow path in the direction of gravity, and temporarily stores the culture medium discharged from the flow path and discharges the culture medium toward the waste liquid container.

[0007] The above-described cell culture system can appropriately apply positive pressure to the reactor and the flow path, and can reduce the workload involved in replacing and removing the waste liquid section. [Brief explanation of the drawings]

[0008] [Figure 1] 1 is a perspective view showing the overall configuration of a cell culture system according to one embodiment of the present invention. [Figure 2] FIG. 10 is a circuit diagram showing a flow path between a culture medium reservoir and a reactor, and a flow path control mechanism. [Figure 3] FIG. 2 is an explanatory diagram schematically illustrating a waste liquid path and a waste liquid section. [Figure 4] 4A and 4B are perspective and cross-sectional views of a temporary reservoir according to a first modified example. [Figure 5] FIG. 10 is an explanatory diagram schematically showing a waste liquid path and a waste liquid section according to a second modified example. DETAILED DESCRIPTION OF THE INVENTION

[0009] DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS The present invention will now be described in detail with reference to preferred embodiments thereof and the accompanying drawings.

[0010] As shown in Fig. 1, a cell culture system 10 according to one embodiment of the present invention is configured as a stationary device installed in a sterile room or the like, and performs a culture process for culturing living cells in regenerative medicine. To this end, the cell culture system 10 includes a reactor 12, which is a cell culture container. The cell culture system 10 performs cell culture over a long period of time by supplying culture medium and oxygen to the reactor 12 while discharging lactic acid, carbon dioxide, and the like (including unused culture medium and oxygen) generated during cell culture from the reactor 12.

[0011] The biological cells are not particularly limited, but examples thereof include cells contained in blood (T cells, etc.) and stem cells (ES cells, iPS cells, mesenchymal stem cells, etc.). The medium may also be selected appropriately depending on the biological cells, and examples thereof include those prepared by adding various amino acids, vitamins, serum, etc. to a balanced salt solution (BSS) as a base solution.

[0012] In addition to the reactor 12, the cell culture system 10 has a culture medium reservoir 14 that stores the culture medium, a flow path 16 provided between the reactor 12 and the culture medium reservoir 14, a waste liquid path 18 that discharges the culture medium from the flow path 16, and a waste liquid part 20 that stores the culture medium flowing through the waste liquid path 18. The cell culture system 10 according to this embodiment is provided with a plurality of reactors 12 (five in FIG. 1 ) to improve the efficiency of the culture process. That is, the cell culture system 10 is configured to circulate the culture medium through each of the plurality of reactors 12 and culture cells in each reactor 12, thereby obtaining several times the number of cells compared to culture using a single reactor 12, without significantly changing the culture period.

[0013] A hard (or soft) tank capable of storing a large amount of culture medium is used as the culture medium reservoir 14 in order to supply the culture medium to each reactor 12. For example, the tank preferably has a volume of about 5 L to 30 L, which reduces the workload of frequently replacing the culture medium reservoir 14 during the culture treatment. Note that a flexible medical bag or the like may also be used as the culture medium reservoir 14.

[0014] The distribution path 16 is composed of a plurality of tubes 22 (only the tubes 22 connected to the culture medium reservoir 14 are shown in FIG. 1). Each tube 22 is connected to the culture medium reservoir 14 and several medical bags (not shown), and is also connected to each reactor 12. As a result, the cell culture system 10 supplies and discharges the culture medium from the culture medium reservoir 14 and the liquids (cell liquid, washing liquid, detachment liquid, etc.) from each medical bag to and from the reactor 12 via each tube 22.

[0015] The cell solution is a liquid containing cells to be seeded (cultured) in the reactor 12. The cleaning solution is a liquid used when priming the reactor 12 and the flow path 16. Examples of this cleaning solution include buffer solutions such as PBS (Phosphate Buffered Salts) and TBS (Tris-Buffered Saline), or physiological saline. The detachment solution is a liquid that detaches cells cultured by a culture process. Examples of the detachment solution that can be used include trypsin and EDTA solution.

[0016] When the cell culture system 10 is constructed, the flow path 16 is set to pass through a flow path control mechanism 24. The flow path control mechanism 24 includes a first housing 26 that houses a portion of the flow path 16. The flow path control mechanism 24 also includes, in the first housing 26, a plurality of clamps 28 that open and close predetermined tubes 22, a plurality of pumps 30 that circulate the liquid in the tubes 22, and a control unit 32 that controls the operation of each clamp 28 and each pump 30 (see FIG. 2). That is, the flow path control mechanism 24 selectively switches the tubes 22 through which the liquid flows by opening and closing each clamp 28, while circulating the liquid in the flow path 16 under the operation of the pump 30.

[0017] In addition to the plurality of tubes 22, the distribution path 16 may also include a cassette (not shown) having a plurality of liquid flow paths and to which some of the tubes 22 are connected. In this case, the cassette is placed in a clamp 28 of the flow path control mechanism 24 when set in the first housing 26, and the clamp 28 opens, closes, switches, etc. the flow paths in the cassette.

[0018] To ensure a large culture area, each reactor 12 connected to the flow path 16 preferably has a structure including, for example, hollow fibers 34. Specifically, each reactor 12 includes a plurality of hollow fibers 34 (for example, 10,000 or more) and a case 36 that houses the plurality of hollow fibers 34 along the axial direction.

[0019] Each hollow fiber 34 has a lumen (not shown) that penetrates along the extension direction, and cells are seeded on the inner circumferential surface that constitutes the lumen. Each hollow fiber 34 also has a plurality of pores (not shown) that communicate between the outside and the lumen, and each pore is impermeable to cells and proteins but permeable to solutions and low-molecular-weight substances. Therefore, culture medium, predetermined gas components, etc. are supplied to the cells on the inner circumferential surface of the hollow fiber 34 through the pores. Hereinafter, a configuration in which liquid is circulated mainly through the lumen of the hollow fiber 34 will be referred to as an IC (intra capillary), and a configuration in which liquid is circulated mainly outside the hollow fiber 34 will be referred to as an EC (extra capillary).

[0020] The material constituting the hollow fibers 34 is not particularly limited, and examples thereof include polymeric materials such as polyolefin resins such as polypropylene and polyethylene, polysulfone, polyethersulfone, polyacrylonitrile, polytetrafluoroethylene, polystyrene, polymethyl methacrylate, cellulose acetate, cellulose triacetate, and regenerated cellulose.

[0021] The case 36 is cylindrical and rigid. The case 36 includes a first IC terminal 36a, a second IC terminal 36b, a first EC terminal 36c, and a second EC terminal 36d, which are connected to the tubes 22. The first IC terminal 36a is provided at one axial end of the case 36 and is connected to the lumen of the hollow fibers 34. The second IC terminal 36b is provided at the other axial end of the case 36 and is connected to the lumen of the hollow fibers 34. The first EC terminal 36c is provided near the other end of a side surface of the case 36 and is connected to the space outside the hollow fibers 34 within the case 36. The second EC terminal 36d is provided near one end of a side surface of the case 36 and is connected to the space outside the hollow fibers 34 within the case 36.

[0022] Hereinafter, with reference to FIG. 2, the flow path 16 between one reactor 12 and the culture medium reservoir 14, and the configuration of the flow path control mechanism 24 will be specifically described.

[0023] The flow path 16 has a culture medium delivery route 40 connected to the culture medium reservoir 14, and an IC route 42 (internal route) and an EC route 44 (external route) branching from the culture medium delivery route 40. The IC route 42 is a route for supplying liquid to the lumen of the hollow fibers 34, and liquids such as cleaning liquid, cell liquid, culture medium, and detachment liquid circulate through it. The EC route 44 is a route for supplying liquid to the outside of the hollow fibers 34 (inside the case 36), and liquids such as cleaning liquid, culture medium, and detachment liquid circulate through it.

[0024] The culture medium delivery route 40 is provided with a first clamp 40a that opens or closes the supply of culture medium from the culture medium reservoir 14.

[0025] The IC route 42 has an IC circulation circuit 42a capable of circulating liquid between the reactor 12 and the IC route 42, and an IC supply circuit 42b capable of distributing liquid from the culture medium delivery route 40 to the IC circulation circuit 42a. The IC circulation circuit 42a is provided with an IC circulation pump 30a for circulating the liquid. The IC supply circuit 42b is provided with an IC supply pump 30b for distributing liquid from the culture medium delivery route 40 to the IC circulation circuit 42a. Although not shown, in addition to the culture medium reservoir 14, tubes 22 connected to each medical bag storing a cleaning solution, a cell solution, a detachment solution, etc. are connected to the IC supply circuit 42b.

[0026] The IC circulation circuit 42a is connected to the first IC terminal 36a and the second IC terminal 36b of the reactor 12. Therefore, the liquid circulating through the IC circulation circuit 42a flows through the lumen of the hollow fibers 34 under the operation of the IC circulation pump 30a.

[0027] An IC waste liquid circuit 46 is connected to the IC circulation circuit 42a downstream of the reactor 12. The IC waste liquid circuit 46 constitutes part of the waste liquid path 18 and is connected to a confluence route 50 of the waste liquid path 18. The IC waste liquid circuit 46 is provided with a second clamp 46a that opens or closes the discharge of liquid from the IC circulation circuit 42a.

[0028] On the other hand, the EC route 44 has an EC circulation circuit 44a that can circulate liquid between the reactor 12 and the EC route 44, and an EC supply circuit 44b that can circulate liquid from the culture medium delivery route 40 to the EC circulation circuit 44a. The EC circulation circuit 44a is provided with an EC circulation pump 30c for circulating the liquid. The EC supply circuit 44b is provided with an EC supply pump 30d that circulates liquid from the culture medium delivery route 40 to the EC circulation circuit 44a. Although not shown, in addition to the culture medium reservoir 14, the EC supply circuit 44b is also connected to tubes 22 that lead to medical bags that store a cleaning solution, a stripping solution, etc.

[0029] The EC circulation circuit 44a is connected to the first EC terminal 36c and the second EC terminal 36d of the reactor 12. Therefore, the liquid circulating through the EC circulation circuit 44a flows through the case 36 under the operation of the EC circulation pump 30c. A gas exchanger 52 is provided upstream of the reactor 12 in the EC circulation circuit 44a. The gas exchanger 52 has the function of discharging carbon dioxide mixed in the culture medium while mixing predetermined gas components (nitrogen N2: 75%, oxygen O2: 20%, carbon dioxide CO2: 5%) into the culture medium. The structure of the gas exchanger 52 is not particularly limited, and a gas exchanger having a plurality of hollow fibers inside a case, similar to the reactor 12, can be used.

[0030] An EC waste liquid circuit 48 is connected to the EC circulation circuit 44a downstream of the reactor 12. The EC waste liquid circuit 48 constitutes a part of the waste liquid path 18 and is connected to a confluence route 50 of the waste liquid path 18. The EC waste liquid circuit 48 is provided with a third clamp 48a that opens or closes the discharge of liquid from the EC circulation circuit 44a.

[0031] Furthermore, as described above, when a plurality of (five) reactors 12 are provided, the cell culture system 10 may be configured to include a plurality of IC circulation circuits 42a and EC circulation circuits 44a corresponding to each reactor 12. That is, another IC circulation circuit and an EC circulation circuit (not shown) that circulate liquid to another reactor 12 are connected in parallel to a branch point X between the IC supply pump 30b and the IC circulation circuit 42a, and to a branch point Y between the EC supply pump 30d and the EC circulation circuit 44a.

[0032] Returning to FIG. 1 , the cell culture system 10 has a second housing 54 that houses each reactor 12, located adjacent to the first housing 26 that constitutes the flow path control mechanism 24. The second housing 54 has the function of maintaining the temperature of the housing chamber of each reactor 12 at 37°C. In other words, by using a second housing 54 that is different from the first housing 26 of the flow path control mechanism 24, the cell culture system 10 can easily create an environment suitable for cell culture in each reactor 12. Note that the cell culture system 10 is not limited to a configuration in which each reactor 12 and the flow path 16 are housed in multiple housings, and may be housed in a single housing.

[0033] The second housing 54 may be configured to house a part of the flow path control mechanism 24 (such as the clamp 28 and the pump 30) therein. For example, the second clamp 46a on the IC waste liquid circuit 46 and the third clamp 48a on the EC waste liquid circuit 48 are provided inside the second housing 54. Furthermore, the second housing 54 is preferably configured to rotatably fix each reactor 12 in the direction of gravity, horizontally, or around the axis of the case 36. This allows air inside each reactor 12 to be easily discharged from the case 36.

[0034] Furthermore, the cell culture system 10 includes an installation stand 56 on which the first housing 26 and the second housing 54 are installed. The installation stand 56 has a top plate 58 on which the first housing 26 and the second housing 54 are placed, and this top plate 58 is supported at a predetermined height (approximately 50 cm to 150 cm) by the side walls of the installation stand 56 or the like. The culture medium reservoir 14 is accommodated in a culture medium storage chamber 60 of the installation stand 56, which is provided below the top plate 58.

[0035] The waste liquid path 18 of the cell culture system 10 is connected to the above-mentioned distribution path 16 and also to the waste liquid section 20, thereby discharging liquids such as culture medium and cleaning liquid from the distribution path 16 to the waste liquid section 20. The waste liquid path 18 has an IC waste liquid circuit 46 of the IC route 42, an EC waste liquid circuit 48 of the EC route 44, and a confluence route 50 (see FIG. 2). The confluence route 50 of the waste liquid path 18 is provided so as to extend from inside the second housing 54 (or the first housing 26) to the outside.

[0036] 1 and 3, the waste liquid section 20 is connected to the most downstream part of the waste liquid path 18 and has a waste liquid container 62 capable of storing the culture medium that has passed through the waste liquid path 18. The waste liquid path 18 also has a temporary storage section 64 upstream of the waste liquid container 62 that is capable of temporarily storing the culture medium.

[0037] A hard (or soft) tank with a large capacity is used as the waste liquid container 62 in order to store the culture medium used in each reactor 12. For example, the tank preferably has a capacity of about 5 L to 30 L. This reduces the workload of frequently replacing the waste liquid container 62. Alternatively, a flexible medical bag or the like may be used as the waste liquid container 62.

[0038] The waste liquid container 62 is stored in a waste liquid storage container 61 of the installation stand 56, which is provided below the top plate 58. In other words, the waste liquid container 62 is provided lower in the direction of gravity than the temporary storage unit 64. Furthermore, the waste liquid container 62 according to this embodiment is located lower in the direction of gravity than the second housing 54 that houses the first housing 26 and each reactor 12. Note that, although FIG. 1 illustrates a state in which the waste liquid container 62 is exposed from the waste liquid storage container 61, the waste liquid storage container 61 may be configured to seal the waste liquid container 62.

[0039] On the other hand, the temporary storage section 64 temporarily stores the liquid discharged from the flow path 16 and then discharges it toward the waste liquid container 62. Therefore, the volume of the temporary storage section 64 is sufficiently smaller than the volume of the waste liquid container 62. The temporary storage section 64 is, for example, hung from a stand 66 fixed to the installation table 56, and is located higher in the direction of gravity than the first housing 26 and the second housing 54 that houses the multiple reactors 12. In other words, the temporary storage section 64 is disposed higher in the direction of gravity than the reactors 12 and the flow path 16. The height of the temporary storage section 64 is not particularly limited, but may be set in the range of 150 cm to 180 cm, for example, and may be set to have a predetermined difference (in the range of 10 cm to 80 cm) from the heights of the reactors 12 and the flow path 16.

[0040] The waste liquid path 18 (confluence route 50) has an upstream line 70 provided between the distribution path 16 and the temporary storage section 64 via the IC waste liquid circuit 46 and the EC waste liquid circuit 48, and a downstream line 72 provided between the temporary storage section 64 and the waste liquid container 62. The upstream line 70 and the downstream line 72 are configured by a tube 73 having a flow path therein. The upstream line 70 extends upward from the second housing 54 in the direction of gravity and is connected to the lower part of the temporary storage section 64. The downstream line 72 extends downward from the temporary storage section 64 in the direction of gravity and is connected to the upper part of the waste liquid container 62.

[0041] A flexible medical bag is applied to the temporary storage section 64. The temporary storage section 64 has a sealed section 74 that seals the outer peripheries of two sheets that make up the medical bag, and a storage space 76 is provided inside the sealed section 74 and between the two sheets. The upstream line 70 and the downstream line 72 are connected to the sealed section 74 (hereinafter referred to as the lower sealed section 74a) on the lower side of the temporary storage section 64. The temporary storage section 64 may also be made of a hard container.

[0042] A partition 82 is provided inside the temporary storage section 64, separating the lower side of the storage space 76 into a first storage section 78 and a second storage section 80. The partition 82 is connected to the lower seal section 74a and extends upward in the direction of gravity from the lower seal section 74a. The partition 82 is formed, for example, by sealing two sheets that constitute the medical bag. Alternatively, the partition 82 may be formed by sandwiching a plate member having a predetermined thickness in a direction perpendicular to the surfaces of the two sheets between the two sheets and welding the edges of the plate member to each sheet.

[0043] A communication section 84 (part of the storage space 76) that communicates the first storage section 78 and the second storage section 80 is provided inside the temporary storage section 64 above the partition wall 82 in the direction of gravity. That is, the storage space 76 is composed of the communication section 84 above in the direction of gravity, and the first storage section 78 and the second storage section 80 that are adjacent to each other in the lateral direction of the partition wall 82 (the direction perpendicular to the direction of gravity) below the communication section 84 in the direction of gravity. The first storage section 78 is connected to the flow path of the upstream line 70 that is fixed to the lower seal section 74a. The second storage section 80 is connected to the flow path of the downstream line 72 that is fixed to the lower seal section 74a.

[0044] Therefore, the liquid that flows from the upstream line 70 into the temporary storage section 64 is first stored in the first storage section 78, and when the first storage section 78 is filled, the liquid flows over the partition wall 82 into the second storage section 80. The liquid that flows into the second storage section 80 flows out into the downstream line 72 (outside the temporary storage section 64).

[0045] First reservoir 78 is configured to apply an appropriate pressure (positive pressure) to reactor 12 and flow path 16 through the culture medium stored therein. For example, the volume of first reservoir 78 is preferably set to a range of approximately 0.5 to 3 times the volume of second reservoir 80. The actual volume of first reservoir 78 is preferably set to a range of 50 cc to 300 cc, for example.

[0046] Furthermore, the temporary storage section 64 has an atmosphere release section 86 that applies atmospheric pressure to the liquid that has flowed into the first storage section 78. In this embodiment, the atmosphere release section 86 is configured with a vent mechanism 88 that allows gas to pass through but blocks liquid from passing through. This allows the vent mechanism 88 to apply atmospheric pressure to the liquid in the first storage section 78 without causing the liquid that has flowed into the storage space 76 to leak to the outside. Note that the atmosphere release section 86 is not limited to the vent mechanism 88, and may be configured with an opening that simply opens the upper side of the temporary storage section 64 in the direction of gravity.

[0047] As shown in Fig. 3, the waste liquid unit 20 may be connected to a plurality of cell culture systems 10. For example, by branching an upstream line 70 of the waste liquid path 18, the temporary storage unit 64 is connected to the distribution path 16 of the first cell culture system 10A (see the solid line in Fig. 3) and the distribution path 16 of the second cell culture system 10B (see the two-dot chain line in Fig. 3). Therefore, the waste liquid unit 20 temporarily stores in one temporary storage unit 64 both the liquid flowing out from the distribution path 16 of the first cell culture system 10A and the liquid flowing out from the distribution path 16 of the second cell culture system 10B. The liquid stored in the temporary storage unit 64 is then discharged to one or more waste liquid containers 62 via a downstream line 72.

[0048] The cell culture system 10 according to this embodiment is basically configured as described above, and its operation will be described below.

[0049] 1, in the cell culture system 10, before a culture process is performed, an operator sets the multiple reactors 12 in the second housing 54 and sets the flow path 16 in the flow path control mechanism 24. The operator also places the culture medium reservoir 14 in the culture medium storage container 60 of the installation table 56, installs a waste liquid container 62 in the waste liquid storage container 61 of the installation table 56, and hangs the temporary reservoir 64 on the stand 66. As a result, the flow path 16 shown in FIG. 2 is established between the culture medium reservoir 14 and each reactor 12, and the temporary reservoir 64 is positioned above the flow path 16 in the direction of gravity.

[0050] After the above setup, the cell culture system 10 sequentially performs the following steps in the culture process: priming, medium replacement, seeding, culture, detachment, and recovery. In the priming step, a cleaning solution stored in a medical bag (not shown) is supplied to each reactor 12 through the flow path 16, and air is removed from the reactors 12 and the flow path 16. In the medium replacement step, medium is supplied from the medium reservoir 14 to each reactor 12 through the primed flow path 16, filling the inside and outside of the hollow fibers 34 with medium. In the seeding step, cell liquid stored in a medical bag (not shown) is supplied into the hollow fibers 34 of each reactor 12 through the IC route 42, and cells are seeded on the inner surface of the hollow fibers 34.

[0051] As shown in FIG. 2, in the culture step, the cell culture system 10 supplies the culture medium from the culture medium reservoir 14 into the hollow fibers 34 through both the IC route 42 and the EC route 44, and cultures cells in the hollow fibers 34. At this time, the gas exchanger 52 discharges carbon dioxide from the culture medium and supplies oxygen to the culture medium. The culture step is carried out for a longer period (e.g., several days) than other steps, so that cells gradually grow on the inner surface of the hollow fibers 34. Note that the cell culture system 10 may be configured to supply the culture medium to the reactor 12 via the EC route 44 without passing through the IC supply circuit 42b. The culture medium that flows into the reactor 12 via the EC route 44 is supplied to the cells by seeping from the outside to the inside of the hollow fibers 34.

[0052] During the culturing process, the culture medium circulating through the IC circulation circuit 42a flows into the IC waste liquid circuit 46 when the second clamp 46a is opened. The culture medium circulating through the EC circulation circuit 44a flows into the EC waste liquid circuit 48 when the third clamp 48a is opened. This allows the culture medium to flow through the waste liquid path 18. The culture medium in the IC waste liquid circuit 46 and the EC waste liquid circuit 48 flows into the upstream line 70 of the confluence route 50, and moves outside the second housing 54. This culture medium flows upward in the direction of gravity via the upstream line 70 and flows into the first reservoir 78 of the temporary reservoir 64.

[0053] 3, temporary storage section 64 continues to store the culture medium in first storage section 78 until the culture medium exceeds partition wall 82. When the culture medium in first storage section 78 exceeds partition wall 82, the culture medium flows over partition wall 82 (through communication section 84) into second storage section 80. The culture medium that has moved to second storage section 80 flows into downstream line 72 fixed to the lower part of second storage section 80. In other words, when the amount of culture medium flowing into temporary storage section 64 exceeds a certain amount, the culture medium is automatically discharged into downstream line 72 below.

[0054] The culture medium discharged into the downstream line 72 flows downward in the direction of gravity and flows into the waste liquid container 62 installed below the installation stand 56. The waste liquid container 62 has a capacity large enough to store the culture medium, which significantly reduces the number of times the waste liquid container 62 needs to be replaced during the culture process.

[0055] The temporary storage section 64, which is disposed above the flow path 16 and the reactor 12 in the direction of gravity, can apply positive pressure to the flow path 16 via the first storage section 78 and the culture medium in the upstream line 70. Therefore, in the EC circulation circuit 44a, the positive pressure of the culture medium can suppress the inflow of excess air into the gas exchanger 52, and the air and the culture medium can be stably mixed. As a result, the cell culture system 10 significantly suppresses the inflow of air bubbles into the flow path 16.

[0056] Furthermore, the atmosphere release part 86 provided in the temporary storage part 64 can apply positive pressure to the distribution path 16 even when there is little culture medium by applying atmospheric pressure to the culture medium in the first storage part 78 and the upstream line 70. In particular, the atmosphere release part 86 employs a vent mechanism 88, which makes it possible to prevent the culture medium from leaking from the temporary storage part 64.

[0057] In the detachment step after the culture step, the cell culture system 10 introduces a detachment solution stored in a medical bag (not shown) into the hollow fibers 34 of the reactor 12 via the IC route 42, and detaches the cultured (grown) cells. In the recovery step after the detachment step, the cell culture system 10 supplies a culture medium to the IC route 42, thereby causing the cells detached in the detachment step to flow out of the reactor 12 and move to a recovery bag (not shown).

[0058] Through the above steps, the cell culture system 10 can effectively store the cells cultured in the reactor 12 in the collection bag. In particular, the cell culture system 10 can stably apply positive pressure to the distribution channel 16, and even when a large amount of culture medium is used, the workload on the operator can be reduced by reducing the need to replace or remove the waste liquid container 62.

[0059] The present invention is not limited to the above-described embodiment, and various modifications are possible within the spirit and scope of the invention. For example, the cell culture system 10 may be configured to perform the culture process using a single reactor 12, rather than using multiple reactors 12. If the number of cultured cells in the culture process is to be increased, a large reactor 12 may be used.

[0060] 4A and 4B, the temporary storage section 90 may be provided in the upper part of the second housing 54 (or the first housing 26: see FIG. 1) that houses the plurality of reactors 12. Even in this case, the temporary storage section 90 is disposed above the reactors 12 and the flow path 16 in the direction of gravity. For example, the temporary storage section 90 is formed by a small and hard container 92, and is configured so that its volume does not change within the second housing 54 (so that the flexible bag does not compress other mechanisms).

[0061] Furthermore, no partition wall 82 (see FIG. 1) is provided inside the container 92, and a storage space 92a is formed that is surrounded by the inner surface of the container 92. The upstream line 70 and downstream line 72 of the waste liquid path 18 are each connected to the lower end of the container 92 of the temporary storage unit 90. The upstream line 70 extends upward in the direction of gravity from the connection point with the distribution path 16 (see FIG. 2). Meanwhile, the downstream line 72 temporarily heads upward in the direction of gravity from the lower part of the container 92 inside the second housing 54, and is exposed to the outside of the second housing 54. Then, outside the second housing 54, the downstream line 72 extends downward in the direction of gravity and is connected to a waste liquid container 62 (see FIG. 1) that is installed lower than the second housing 54 in the direction of gravity.

[0062] Furthermore, an atmosphere release part 86 is provided at the upper end of the container 92 of the temporary storage part 90, which applies atmospheric pressure to the culture medium that has flowed into the storage space 92a. The atmosphere release part 86 is configured with a vent mechanism 88 that allows gas to pass through but blocks liquid from passing through. Note that the atmosphere release part 86 may also be configured by simply opening the upper end of the container 92.

[0063] The temporary storage section 90 configured as described above can also achieve the same effect as the temporary storage section 64. That is, the culture medium that flows into the temporary storage section 90 from the flow path 16 via the upstream line 70 is temporarily stored in the storage space 92a of the container 92. Then, atmospheric pressure is applied to the culture medium in the container 92 from the atmosphere vent 86, so that positive pressure can be applied to the flow path 16 through the culture medium in the upstream line 70. In particular, the temporary storage section 90 provided in the second housing 54 can prevent inconveniences such as an operator accidentally dropping the temporary storage section 90.

[0064] Furthermore, the culture medium in the storage space 92a is guided to the downstream line 72 by the siphon effect. The culture medium first flows upward in the direction of gravity in the downstream line 72, and then is guided downward in the direction of gravity to be stored in the waste liquid container 62. As a result, the culture medium is temporarily stored in the temporary storage section 90, and then smoothly discharged into the waste liquid container 62, preventing a large amount of culture medium from accumulating in the container 92.

[0065] 5 , the cell culture system 10 may employ a temporary reservoir 94 without a partition 82, and may include a sensor 96 for detecting the weight or liquid level of the temporary reservoir 94 and a valve 98 for opening and closing the downstream line 72. The sensor 96 and the valve 98 are connected to the control unit 32 of the cell culture system 10 via wired or wireless communication. The control unit 32 normally closes the valve 98, monitors the amount of liquid flowing into the temporary reservoir 94 based on the detection information of the sensor 96, and opens the valve 98 when the amount of liquid exceeds a predetermined threshold. Even in this case, the cell culture system 10 can stably apply a positive pressure to the distribution channel 16.

[0066] The technical ideas and effects that can be understood from the above-described embodiments will be described below.

[0067] One aspect of the present invention is a cell culture system 10 comprising a reactor 12 for culturing cells based on the flow of culture medium, a flow path 16 for flowing the culture medium into and out of the reactor 12, a waste liquid path 18 connected to the flow path 16 and for discharging the culture medium from the flow path 16, and a waste liquid container 62 connected to the waste liquid path 18 and capable of storing the culture medium that has passed through the waste liquid path 18, wherein the waste liquid path 18 has temporary storage sections 64, 90, 94 capable of temporarily storing the culture medium, the waste liquid container 62 is located below the temporary storage sections 64, 90, 94 in the direction of gravity, and the temporary storage sections 64, 90, 94 are located above the flow path 16 in the direction of gravity, and temporarily store the culture medium discharged from the flow path 16 and discharge the culture medium toward the waste liquid container 62.

[0068] As described above, in the cell culture system 10, the temporary storage units 64, 90, 94 are located above the flow path 16 in the direction of gravity, so that it is possible to appropriately apply positive pressure to the reactor 12 and the flow path 16 via the culture medium in the temporary storage units 64, 90, 94. This allows the cell culture system 10 to suppress the inflow of excess air into the reactor 12 and the flow path 16. Furthermore, since the waste liquid container 62 is located below the temporary storage units 64, 90, 94 in the direction of gravity, the cell culture system 10 can reduce the workload involved in replacing, removing, etc. the waste liquid container 62.

[0069] Furthermore, the waste liquid container 62 is provided below the installation position of the reactor 12 in the direction of gravity. This makes it easier for the operator of the cell culture system 10 to replace or remove the waste liquid container 62.

[0070] Further, inside temporary storage units 64, 94, there are provided a first storage unit 78, a second storage unit 80, a partition wall 82 separating first storage unit 78 and second storage unit 80 from each other, and a communication unit 84 communicating first storage unit 78 and second storage unit 80 above partition wall 82 in the direction of gravity, and waste liquid path 18 includes an upstream line 70 communicating between distribution path 16 and first storage unit 78 and a downstream line 72 communicating between second storage unit 80 and waste liquid container 62, and downstream line 72 extends downward in the direction of gravity from second storage unit 80. This allows the culture medium stored in first storage unit 78 to apply an appropriate positive pressure to reactor 12 and distribution path 16.

[0071] Furthermore, the first storage section 78 and the second storage section 80 are located adjacent to each other in a direction perpendicular to the direction of gravity, and the volume of the first storage section 78 is larger than the volume of the second storage section 80. This allows the temporary storage section 64 to apply a large positive pressure from the first storage section 78 to the reactor 12 and the flow path 16.

[0072] Furthermore, the temporary storage section 90 has an atmosphere release section 86 that applies atmospheric pressure to the culture medium that has flowed into the temporary storage section 90, and the atmosphere release section 86 is configured with a vent mechanism 88 that is permeable to gas but not to liquid. As a result, even in the cell culture system 10 configured with the atmosphere release section 86, leakage of the culture medium from the temporary storage section 90 is suppressed.

[0073] The waste liquid path 18 also includes an upstream line 70 that communicates between the distribution path 16 and the temporary storage section 90, and a downstream line 72 that communicates between the temporary storage section 90 and the waste liquid container 62, and the downstream line 72 first heads upward in the direction of gravity from the bottom of the temporary storage section 90 and then heads downward in the direction of gravity. Even in this case, the cell culture system 10 can discharge the culture medium from the temporary storage section 90 to the waste liquid container 62 while applying positive pressure to the reactor 12 and the distribution path 16 by the culture medium stored in the temporary storage section 90.

[0074] Furthermore, the temporary storage units 64, 94 are disposed outside the housing (second housing 54) that houses the reactor 12. This allows the operator to easily set up the temporary storage units 64, 94 when preparing the cell culture system 10. Furthermore, the operator can visually check the culture medium in the external temporary storage units 64, 94 and recognize the waste liquid state of the culture medium.

[0075] Moreover, the temporary reservoir 90 is disposed inside a housing (second housing 54) that houses the reactor 12. This allows the cell culture system 10 to avoid inconveniences such as an operator accidentally dropping the temporary reservoir 90.

[0076] Furthermore, the flow path 16 has a circulation circuit (EC circulation circuit 44a) that circulates the culture medium between the reactor 12 and the circulation circuit, and a supply circuit (EC supply circuit 44b) that supplies the culture medium to the circulation circuit, and the circulation circuit has a gas exchanger 52 that mixes a gas with the culture medium upstream of the reactor 12 in the direction of culture medium flow, and a waste liquid path 18 is connected downstream of the reactor 12 in the direction of culture medium flow. This makes it possible for the cell culture system 10 to apply an appropriate positive pressure to the circulation circuit having the gas exchanger 52 via the temporary storage units 64, 90, 94 and the culture medium in the waste liquid path 18, and to suppress the inflow of excessive gas into the gas exchanger 52.

[0077] Furthermore, a plurality of reactors 12 are provided, and the culture medium that has flowed through the plurality of reactors 12 is discharged collectively into the waste liquid path 18 and the waste liquid container 62. The cell culture system 10 can improve the efficiency of cell culture by using the plurality of reactors 12. Furthermore, by using the plurality of reactors 12, the cell culture system 10 can stably discharge the culture medium through the waste liquid container 62 and temporary storage units 64, 90, 94, even if a large amount of culture medium is flowed.

Claims

1. a reactor for culturing cells based on the flow of a culture medium; A flow path for introducing and discharging the culture medium into and from the reactor; a waste liquid path connected to the distribution path and discharging the culture medium from the distribution path; A cell culture system comprising: a waste liquid container connected to the waste liquid path and capable of storing the culture medium that has passed through the waste liquid path; the waste liquid path has a temporary storage section capable of temporarily storing the culture medium, the waste liquid container is located below the temporary storage section in the direction of gravity, the temporary storage section is provided above the flow path in the direction of gravity, and temporarily stores the culture medium discharged from the flow path and allows the culture medium to flow out toward the waste liquid container; the temporary storage section is provided therein with a first storage section, a second storage section, a partition wall separating the first storage section and the second storage section from each other, and a communication section communicating the first storage section and the second storage section above the partition wall in the direction of gravity, the waste liquid path includes an upstream line communicating between the distribution path and the first reservoir, and a downstream line communicating between the second reservoir and the waste liquid container, The downstream line extends downward in the direction of gravity from the second storage section. Cell culture system.

2. The cell culture system according to claim 1, The waste liquid container is provided below the installation location of the reactor in the direction of gravity. Cell culture system.

3. The cell culture system according to claim 1, The first storage section and the second storage section are located adjacent to each other in a direction perpendicular to the direction of gravity, and the volume of the first storage section is larger than the volume of the second storage section. Cell culture system.

4. The cell culture system according to any one of claims 1 to 3, The temporary storage section is disposed outside a housing that houses the reactor. Cell culture system.

5. The cell culture system according to any one of claims 1 to 4, The temporary storage section is disposed inside a housing that houses the reactor. Cell culture system.

6. The cell culture system according to any one of claims 1 to 5, The distribution channel is a circulation circuit for circulating the culture medium between the reactor and the circulation circuit; a supply circuit for supplying the culture medium to the circulation circuit, The circulation circuit has a gas exchanger that mixes a gas with the culture medium, located upstream of the reactor in the direction of flow of the culture medium, and the waste liquid path is connected downstream of the reactor in the direction of flow of the culture medium. Cell culture system.

7. The cell culture system according to any one of claims 1 to 6, The reactor is provided in plurality, The culture medium that has flowed through the plurality of reactors is discharged collectively into the waste liquid path and the waste liquid container. Cell culture system.

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