Cell Culture Systems

Abstract

To provide an automated culture device using a single-use, closed-system flow path for culturing cells or tissues, for achieving reduced manufacturing cost and higher device integration.SOLUTION: There is provided a cell culture system comprising an automatic culture device and an information processing device. The automatic culturing apparatus comprises multiple types of closed flow channels that can be installed and removed, and multiple culturing apparatuses. The information processing device comprises: an input device that receives as input at least one data selected from a group consisting of patient identifier data, transplant method data, cell type data, required cell number data, and treatment plan data; an arithmetic device that selects, on the basis of the input data, a cell culture method, a culture apparatus, and a closed system flow path to be used from options for the cell culture method, the culture apparatus, and the closed system flow path; and an output device that outputs the number of the closed system channel to be used and the number of the culture apparatus to be used.SELECTED DRAWING: Figure 6

Description

[Technical Field]

[0001] The present invention relates to a cell culture system. [Background technology]

[0002] Regenerative medicine, which uses regenerative tissues manufactured from cells to restore the functions of organs, is expected to be a cure for diseases for which there were no conventional treatments. Clinical applications are rapidly increasing for a variety of target tissues, including skin, cornea, esophagus, heart, bone, and cartilage. In the regenerative tissue manufacturing process, stem cells are isolated from biological samples taken from the patient or others, and the isolated stem cells are then proliferated and organized. This process is carried out in a cell processing facility (CPF) in accordance with Good Manufacturing Practices (GMP), which are the standards for manufacturing and quality control of pharmaceuticals, etc. The process is carried out in accordance with Standard Operating Procedures (SOPs) that meet Good Manufacturing Practices.

[0003] Operating a CPF requires significant costs and personnel with specialized culture techniques. Additionally, the regenerative tissue manufacturing process is primarily manual, limiting the amount of regenerative tissue that can be produced. Low productivity and high manufacturing costs are hindering the widespread adoption of regenerative medicine. Automation of the culture process, which is particularly labor-intensive and costly, is anticipated, which would reduce labor and costs in the manufacturing of regenerative tissue and enable mass production of regenerative tissue.

[0004] Furthermore, among products already on the market and those under development, there are more regenerative tissues intended for autologous transplantation than allogeneic transplantation. Autologous transplantation, which uses the patient's own cells, is expected to have good treatment outcomes due to a low risk of rejection, and there is thought to be a high demand for it from the perspective of improving patients' quality of life. Since regenerative tissue is produced for each patient in autologous transplantation, it is possible to produce a wide variety of regenerative tissues in small quantities. In this case, when manufacturing using an automated culture device, it is preferable to produce enough tissue for one patient per automated culture device to avoid the risk of biological contamination. On the other hand, when manufacturing cells for multiple patients, it is also important that the device is small and highly integratable. Therefore, it is important that the automated culture device for autologous transplantation has low manufacturing costs and high integratability.

[0005] There are two main types of automatic culture equipment: open-system automatic culture equipment and closed-system automatic culture equipment. In open-system automatic culture equipment, open-system culture vessels, such as culture vessels with lids that are commonly used in manual culture and culture plates, are automatically handled in a closed space equipped with an articulated robot or the like. This closed space can be decontaminated using decontamination gases or the like. On the other hand, closed-system automatic culture equipment automatically handles a closed-system flow path with a closed space. In a closed-system flow path, the closed-system culture vessel is constantly connected by flow path tubes or the like, and the interior is sterilized in advance by gamma ray sterilization or the like.

[0006] When comparing open-system automated culture equipment with closed-system automated culture equipment, with the open-system automated culture equipment, after regenerative tissue is produced, the open-system culture vessel is removed from the open-system automated culture equipment and moved to the next process. However, the transfer of the open-system culture vessel carries the risk of biological contamination due to medium leakage, etc. With the closed-system automated culture equipment, after regenerative tissue is produced, the closed-system culture vessel is removed from the closed-system automated culture equipment and moved to the next process. However, since medium leakage does not occur when transferring the closed-system culture vessel, the risk of biological contamination is lower than with open-system automated culture equipment that uses open-system culture vessels. Furthermore, open-system automated culture equipment is often large because it requires the installation of articulated robots and other devices within a closed space. Therefore, if multiple open-system automated culture equipment are to be used in a stacked configuration, a large CPF room is required to accommodate them. With closed-system automated culture equipment, only the inside of the closed-system flow path is kept sterile, allowing the equipment to be made smaller. Therefore, using multiple closed-system automated culture equipment in a stacked configuration is not as effective as open-system automated culture equipment. It is easier than placing

[0007] As an example of a closed-system automatic culture device, a device has been disclosed that has a single-layer closed-system culture vessel and is constantly connected to a flow path for supplying or discharging a culture medium or the like (see Patent Document 1). When feeding a culture medium or the like, a valve installed outside the closed-system flow path is adjusted and a pump is operated. The closed-system flow path is designed for single-use to avoid biological contamination. It is also possible to use multiple devices in an integrated state. Another closed-system automatic culture device has been disclosed that has multiple types of flow paths, branches, etc. within a closed-system culture vessel (see Patent Document 2). With this device, by selecting appropriate ones from multiple types of flow paths and branches, it is possible to carry out an automatic culture protocol according to the selected ones. [Prior art documents] [Patent documents]

[0008] [Patent Document 1] Japanese Patent Application Laid-Open No. 2007-312668 [Patent Document 2] US2019 / 0330584 Summary of the Invention [Problem to be solved by the invention]

[0009] An object of the present invention is to provide a novel cell culture system. [Means for solving the problem]

[0010] One embodiment of the present invention is a cell culture system including an automated culture device and an information processing device. The automated culture device includes multiple types of installable and removable closed flow paths and multiple culture devices. The information processing device includes an input device that receives as input at least one data selected from the group consisting of patient identifier data, transplantation method data, cell type data, required cell number data, and treatment plan data. An arithmetic unit that selects a cell culture method, culture device, and closed flow path to be used from options for the cell culture method, culture device, and closed flow path based on the input data. The cell culture system includes an output device that outputs the number of the closed flow path to be used and the number of the culture device to be used. The closed flow path may include a valve for opening and closing a flow path tube, a pump for delivering fluid or gas through the flow path tube, a culture medium container for holding fresh culture medium, and a culture supernatant container for holding fresh culture supernatant. One closed flow path may be installed for multiple culture devices. The output device may output a work procedure protocol for the closed flow path to be used. The output device may output a work procedure protocol for the selected incubation device. [Effects of the Invention]

[0011] According to the automatic culture device of the present invention, by using a culture program that operates multiple culture devices using a highly versatile closed flow path, the integration of the automatic culture device can be improved, and as a result, manufacturing costs can be reduced. [Brief explanation of the drawings]

[0012] [Figure 1] FIG. 1 is a diagram showing the configuration of a culture device in one embodiment of the present invention. [Figure 2] FIG. 1 is a diagram showing a state in which a closed flow path is installed in a culture device in one embodiment of the present invention. [Figure 3] FIG. 1 is a diagram showing four types of closed flow channels installed in a culture device in one embodiment of the present invention. [Figure 4] FIG. 1 is a diagram showing a configuration in which a plurality of culture devices are stacked in one embodiment of the present invention. [Figure 5] FIG. 1 is a diagram showing a state in which a closed flow path is provided for each culture device in a configuration in which a plurality of culture devices are stacked in one embodiment of the present invention. [Figure 6] FIG. 10 is a diagram showing a state in one embodiment of the present invention in which a plurality of closed-system automatic culture devices are stacked, with one closed-system flow path being installed for every two and three culture devices. [Figure 7-1] FIG. 1 is a diagram showing a closed flow path installed in a plurality of culture devices in one embodiment of the present invention. [Figure 7-2] FIG. 1 is a diagram showing a closed flow path installed in a plurality of culture devices in one embodiment of the present invention. [Figure 8] FIG. 2 is a diagram showing culture information relating to a culture device used for culture and a closed-system flow path in one embodiment of the present invention. [Figure 9] FIG. 2 is a diagram showing the flow of culture information regarding a culture apparatus and a closed-system flow path used for culture in one embodiment of the present invention. [Figure 10] FIG. 10 is a diagram showing an example of a screen of a culture program in one embodiment of the present invention. [Figure 11] FIG. 2 is a diagram showing a control mechanism of the automatic culture device in one embodiment of the present invention. [Figure 12] FIG. 1 is a diagram showing a flow during operation of a closed-system automatic culture device in one embodiment of the present invention. [Figure 13]FIG. 10 is a diagram showing the configuration of a culture device in another embodiment of the present invention, in which there is no refrigerator and the culture medium container is installed on the side of the device at room temperature. [Figure 14] This is a diagram showing the configuration of a culture device in another embodiment of the present invention, in which a refrigerator and a flow path mechanism unit are not provided and the culture medium container is installed on the side of the device at room temperature. DETAILED DESCRIPTION OF THE INVENTION

[0013] Preferred embodiments of the present invention will be described in detail below with reference to the accompanying drawings, but the present invention is not necessarily limited thereto. The objects, features, advantages, and concepts of the present invention will be apparent to those skilled in the art from the description in this specification, and those skilled in the art will be able to easily reproduce the present invention from the description in this specification. The embodiments and specific examples of the invention described below show preferred embodiments of the present invention and are presented for illustrative or explanatory purposes, and are not intended to limit the present invention thereto. It will be apparent to those skilled in the art that various changes and modifications can be made based on the description in this specification within the spirit and scope of the present invention disclosed herein.

[0014] The cell culture system of the present invention includes an information processing device and an automatic culture device.

[0015] The automated culture device includes multiple types of installable and removable closed flow paths and multiple culture devices. The information processing device includes an input device that receives as input at least one data selected from the group consisting of patient identifier data, transplantation method data, cell type data, required cell number data, and treatment plan data, a computing device that identifies a cell culture method, culture device, and closed flow path to be used from options for the cell culture method, culture device, and closed flow path based on the input data, and an output device that outputs the number of the closed flow path to be used and the number of the culture device to be used.

[0016] The closed flow path used for culturing includes a closed culture vessel, a medium vessel, a culture supernatant vessel, etc. Various closed flow paths with different types and numbers of closed culture vessels can be used according to the cell culture method corresponding to the cell type to be cultured, the cell type after culture, and the morphology of the regenerated tissue, etc. In other words, one automatic culture device can be used to culture cells using multiple types of closed flow paths with different cell culture methods.

[0017] The information processing device includes an input device, a calculation device, and an output device. To perform a cell culture method using multiple types of closed flow paths for multiple culture devices according to each closed flow path, the information processing device uses a program for operating the multiple culture devices. This program has an electronic tag that can link each culture device and each closed flow path to the cell culture method. The calculation device determines the cell culture method, culture device, and closed flow path to be used from at least one data selected from the group consisting of patient identifier data, transplant method data, cell type data, required cell number data, and treatment plan data received by the input device. The output device outputs a work procedure manual according to the cell culture method to be used.

[0018] In this way, by automatically selecting from multiple culture devices and multiple types of closed flow paths to be used for culturing cells, it is possible to improve the integration of cell culture systems and reduce manufacturing costs.

[0019] Below, with reference to the drawings, we will explain in detail an embodiment of a cell culture system that automatically cultures cells using a closed flow path. However, the cell culture system of the present invention is not limited to the following embodiment, and a person skilled in the art can perform various measurements.

[0020] <Embodiment I> In this embodiment I, a cell culture system that uses a closed flow channel and performs automatic culture will be described with reference to FIGS.

[0021] Each culture device comprises an incubator 101, which is a space for culturing cells at a culture temperature of 37°C, a flow path mechanism unit 102 with a liquid transfer function, and a refrigeration unit 103 (Fig. 1). The flow path mechanism unit 102 comprises a pump 104 and an electromagnetic valve 105. The refrigeration unit 103 stores medium containers containing medium and culture supernatant containers for collecting culture supernatant. These containers can be shaped, for example, in the form of bottles or bags.

[0022] FIG. 2 shows the state in which a closed flow path is installed inside the culture device of FIG. 1. A closed-system culture vessel 201 for culturing cells is installed inside the incubator 101. The number of closed-system culture vessels 201 may be one or more. The closed-system culture vessel 201 is constantly connected to a medium container 202, a culture supernatant container, and the like in the refrigerated section 103 via a flow path tube 203. Because the flow path tube 203 is routed between the inside and outside of the incubator 101, a notch 106 is provided in the incubator 101 to provide a space through which the flow path tube 203 passes. This notch 106 may be provided in either the door or the main body of the incubator 101. The notch 106 is airtight so that the interior of the incubator 101 can be maintained at an appropriate temperature, CO2 concentration, and humidity with the flow path tube 203 installed. For example, the flow path tube 203 may be made of a plastic material, and the gap of the notch 106 may be filled so that it can be crushed. Furthermore, in the notch 106 with the flow path tube 203 passing through, the gap may be filled with, for example, a plastic jig. Note that even when the flow path tube 203 is not installed, the gap of the notch 106 may be filled with a plastic jig that fits into the gap of the notch 106. Thereby, when the inside of the incubator 101 is decontaminated with a decontamination gas such as hydrogen peroxide, the inside of the incubator 101 can be sealed even when the flow path tube 203 is not installed.

[0023] Next, an embodiment of a closed flow channel will be described with reference to Figure 3. In this embodiment, four types of closed flow channels can be used with one automatic culture device, but the types and number of closed flow channels are not limited to this, and closed flow channels with partially modified configurations can also be used. Depending on the type of closed flow channel, the cell culture method that can be implemented and the culture protocol used for culture also differ.

[0024] Figure 3(A) shows a flow channel that enables adherent culture of stem cells, such as iPS cells and mesenchymal stem cells. This configuration allows for both the discharge of culture supernatant and the supply of fresh culture medium during culture medium exchange. Adjusting the height of the outlet pipe within the closed-system culture vessel 301 allows for medium exchange at any desired rate. The incubator 101 is equipped with a closed-system culture vessel 301 with an external filter. The closed-system culture vessel 301 with an external filter has a filter 302 for gas exchange between the gas phase within the incubator 101 and the gas phase within the closed-system culture vessel 301. The refrigeration unit 103 is equipped with a medium container 306 containing fresh medium and a culture supernatant container 307. All components are constantly connected by flow channel tubing 303. A pump 304 externally provides driving force for the flow of medium and other fluids through the flow channel tubing 303, and a solenoid valve 305 selects the direction of the fluid delivery, thereby delivering the medium and other fluids. The solenoid valve 305 is closed during culture. Although a closed culture vessel 301 with an external filter is used, a closed culture vessel 308 with a gas-permeable membrane as shown in Figure 3(B) may also be used. As a culture protocol, culture may be performed without using a culture supernatant vessel 307, with only the supply of fresh medium.

[0025] Figure 3(B) shows an example of a flow path that enables suspension culture of T cells. The culture protocol involves only supplying the culture medium. A common culture medium container 306 is provided for two closed-system culture vessels 308. The closed-system culture vessel 308 with a gas-permeable membrane is installed in the incubator 101. The closed-system culture vessel 308 with a gas-permeable membrane has a gas-permeable membrane 309 for gas exchange between the gas phase in the incubator 101 and the gas phase in the closed-system culture vessel 308 with a gas-permeable membrane. A culture medium container 306 containing fresh culture medium is installed in the refrigeration unit 103. When transferring liquid, two solenoid valves 305 determine the liquid transfer direction. The solenoid valve 305 for the closed-system culture vessel 308 with a gas-permeable membrane to which liquid is to be transferred is opened, and the solenoid valve 305 for the closed-system culture vessel 308 with a gas-permeable membrane to which liquid is not to be transferred is closed. The solenoid valve 305 is closed during culture. In Figure 3(B), the closed culture vessel 308 with a gas-permeable membrane has gas-permeable membranes 309 in two places, the lid and the bottom, but only one may be provided. Although the closed culture vessel 308 with a gas-permeable membrane is used, the closed culture vessel 301 with an external filter used in Figure 3(A) may also be used.

[0026] Figure 3(C) shows an example of a flow channel that enables suspension and adherent culture of T cells. The culture protocol involves only supplying the medium. Two closed culture vessels 308 each have a medium container 306 containing fresh medium. In other words, two closed flow channels are used. In this figure, the closed culture vessel 308 with gas-permeable membrane has gas-permeable membranes 309 in two places, the lid and the bottom, but only one can be used. Although a closed culture vessel 308 with gas-permeable membrane is used, the closed culture vessel 301 with an external filter used in Figure 3(A) can also be used.

[0027] Figure 3(D) shows a flow channel that enables, for example, adherent culture of stem cells such as iPS cells and mesenchymal stem cells, and suspension culture of T cells. The culture protocol involves perfusion culture, which maintains a constant volume of fluid while simultaneously supplying new medium and discharging old medium. A closed-system culture vessel 310 for perfusion culture is installed in the incubator 101. The closed-system culture vessel 310 for perfusion culture has a gas-permeable membrane 309 for gas exchange between the gas phase in the incubator 101 and the gas phase in the closed-system culture vessel 310 for perfusion culture. A medium container 306 containing fresh medium and a culture supernatant container 307 are installed in the refrigeration section 103. Note that, while the closed-system culture vessel 308 with a gas-permeable membrane has a single gas-permeable membrane 309 at the top in this figure, it may also have a gas-permeable membrane at the bottom or both.

[0028] Next, the case where the culture devices are used in a stacked state will be described with reference to FIGS.

[0029] FIG. 4 shows examples of stacking two and three culture devices. The number of stacked devices can be selected by the user depending on the purpose, etc. FIG. 5 shows one culture device performing culture using one closed flow path. FIG. 6(A) shows two culture devices performing culture using one closed flow path. FIG. 6(B) shows three culture devices performing culture using one closed flow path. Although not shown here, four or more culture devices may also be used to perform culture using one closed flow path. In this way, the automatic culture device disclosed herein can connect multiple culture devices with one closed flow path for culture. This allows culture in any number of culture vessels, enabling mass production of cells.

[0030] The closed flow path includes closed-system culture vessels 201, culture medium vessels, and culture supernatant vessels corresponding to the number of culture devices. The solenoid valve 105 is opened and closed to switch the liquid supply direction to the culture vessel to be supplied, and then the pump 104 is operated to supply the liquid to each culture vessel. The culture medium vessels are installed in each refrigerator, and the solenoid valve 105 is opened and closed to switch the liquid supply direction to the culture medium vessel to be supplied, and then the pump 104 is operated to supply the liquid from the culture medium vessel. If the culture is to be continued for a long period of time, new culture medium may be installed using a sterile connection, as the culture medium has an expiration date. As a large amount of culture medium is required for culture, it may be installed separately in each refrigerator.

[0031] Next, another embodiment of the automatic culture device used in a stacked state will be described with reference to Fig. 7. Fig. 7 shows an automatic culture device used in a stacked state of two devices and a stacked state of three devices. The culturing device shown in the figure is different depending on the type of closed flow path. The culturing protocol that can be implemented varies depending on the type of closed flow path. Therefore, the user can select a closed flow path and also select a culturing method according to the selected closed flow path.

[0032] Figures 7(A) and 7(B) show automated culture devices for culturing stem cells, such as iPS cells and mesenchymal stem cells. Figure 7(A) shows an automated culture device using two stacked culture devices. Figure 7(B) shows an automated culture device using three stacked culture devices. The culture protocol allows for both the discharge of culture supernatant and the supply of new culture medium during culture medium replacement. Adjusting the height of the exhaust pipe inside the closed-system culture vessel 301 allows for medium replacement at any desired rate. The incubator 101 is equipped with a closed-system culture vessel 301 with an external filter. The refrigeration section 103 is equipped with a culture medium container 306. While Figure 7(A) shows two culture medium containers 306 and two culture supernatant containers, the number of culture medium containers 306 may be one or three depending on the required amount of medium. The same applies to the culture supernatant container. While the closed-system culture vessel 301 with an external filter is used, a closed-system culture vessel 308 with a gas-permeable membrane may also be used. The culture protocol may involve only supplying new culture medium without using a culture supernatant container. This applies to the automated culture device in Figure 7(B). Figures 7(C) and 7(D) show, as an example, flow paths for performing suspension culture of T cells. Figure 7(C) shows an automated culture device using two stacked culture devices. Figure 7(D) shows an automated culture device using three stacked culture devices. The culture protocol involves only supplying the culture medium. A closed-system culture vessel 308 with a gas-permeable membrane is installed in the incubator 101. A culture medium container 306 is installed in the refrigeration section 103. Note that, although two culture medium containers 306 are installed in Figure 7(C), the number of culture medium containers 306 may be one or three depending on the required amount of culture medium. Although a closed-system culture vessel 308 with a gas-permeable membrane is used, a closed-system culture vessel 301 with an external filter may also be used. This applies to the automated culture device in Figure 7(D).

[0033] When culturing cells from multiple patients, it is preferable to install integrated culture devices in one cell preparation room. Each culture device can culture cells from different patients using its own culture program. The culture program varies depending on the cell type to be cultured, and the culture program The program schedule varies depending on the treatment schedule, such as transplantation, which differs for each patient. The type and number of culture vessels used in each closed flow path differ depending on the cell type to be cultured and the transplant form. Therefore, it is necessary to select the appropriate culture program, closed flow path, and culture device for each patient from the multiple available options.

[0034] FIG. 8 explains the culture information related to the multiple culture devices and multiple closed flow paths used for culture, which is handled by the culture program possessed by the information processing device. The table in FIG. 8 shows the generation and acquisition of culture information generated for each process. FIG. 9 also shows the flow of culture information related to the multiple culture devices and multiple closed flow paths used for culture, and the relationship between the culture program that handles the culture information and the culture information. The information processing device comprises a processing unit consisting of a CPU (Central Processing Unit), a memory unit, a display device, The input device and the arithmetic unit are provided in the CPU, and the display device corresponds to the display device.

[0035] Starting with a patient's visit to the hospital, information regarding the cells to be manufactured is generated as the treatment method is decided. The output device outputs a work procedure protocol regarding the extraction of the closed-system flow path to be used and the identification of the selected culture device, and the output device displays the protocol on the display device. When each item is determined and input into the input device, the arithmetic device limits the options for subsequent selection based on the decision, and the output device outputs the options to the display device, which displays the options. For example, when the type of cell to be manufactured is determined and input into the input device, the arithmetic device limits the types of culture vessels that can be used for culture to those corresponding to the cell type, and the output device outputs the options to the display device, which displays the options. Furthermore, when the type, number, and transplant form of cells to be manufactured are determined and input into the input device, the arithmetic device limits the manufacturing method according to those conditions.

[0036] When determining the manufacturing method, first, the type of closed-system flow path to be used is determined and entered into the input device. This is determined according to the type and number of culture vessels to be used. Next, the culture device to be used is determined and entered into the input device. Furthermore, the culture period varies depending on the cells to be produced, which means that the operating period of the automated culture device also varies. Culture schedules also vary for each patient. Increasing the utilization rate of the device and producing as many patient cells as possible reduces manufacturing costs, so the culture device to be used may be selected to maximize utilization rate. When the closed-system flow path to be used is removed from the warehouse, the lot number of the closed-system flow path to be removed is displayed in the work procedure protocol on the display device. An electronic tag or two-dimensional barcode containing lot number information is attached to the closed-system flow path in advance, and it is confirmed that the lot number displayed in the work procedure manual matches the information on the electronic tag or two-dimensional barcode of the selected closed-system flow path. When the closed-system flow path to be used is installed in the culture device, the device number of the culture device to be used is displayed in the work procedure protocol on the display device. An electronic tag and a two-dimensional barcode containing the device number are attached to the culture device in advance, and it is confirmed that the lot number displayed in the work procedure manual matches the information on the electronic tag or two-dimensional barcode of the selected culture device. This method prevents mistakes when selecting the closed flow path and culture device to be used. After production, the quality evaluation results are used to determine whether to use the produced cells for therapy.

[0037] Figure 10 shows an example of a display screen for a work procedure protocol driven by a culture program. Figure 10(A) is a screen for selecting a cell type when selecting a treatment method. In this example, cancer immunotherapy is selected as the treatment method before selecting a cell type, so only cell types that can be used for cancer immunotherapy are displayed as options for cell type selection, from which a choice can be made. Cell types that are not used for cancer immunotherapy are not displayed, thereby avoiding incorrect selection. Figure 10(B) is a screen for selecting a culture vessel type when selecting a culture method. Since suspension cells are selected as the culture method before selecting a culture vessel type, only types of culture vessels that can be used for suspension cells are displayed as options for culture vessel selection, from which a choice can be made. Figure 10(C) shows a similar selection form. Figure 10(D) shows the selection form. This is the screen that displays all the information.

[0038] FIG. 11 is a block diagram illustrating the functional configuration of an automatic culture device including a culture vessel 1101. This is an overall configuration diagram in which each component controlled by a control device 1102 possessed by an information processing device is connected to each other and to an incubator 1103. The aforementioned closed-system culture vessel 1101 is installed inside the incubator 1103. The control device 1102 is connected to a temperature control unit 1104 for controlling the temperature of the incubator 1103, a gas concentration control unit 1106 having a gas supply unit 1105 for controlling the gas concentration inside the incubator 1103, a pump 1107 installed in a closed-system flow path circuit for automatically supplying the culture medium inside the closed-system culture vessel 1101, and a CO₂ / O₂ sensor 1108. The control device 1102 is provided in the CPU of a typical computer. The control device 1102 runs various programs stored in a memory unit on the CPU, which functions as a processing unit. This controls the culture environment in the incubator 1103 through the temperature adjustment unit 1104, gas supply unit 1105, pump 1107, CO2 / O2 sensor 1108, gas concentration adjustment unit 1106, temperature sensor 1110, medium container 306 / culture supernatant container 1111, and enables a predetermined culture process to be carried out in the closed-system culture vessel 1101. Gas exchange within the closed-system culture vessel 1101 is achieved between the incubator 1103 and the closed-system culture vessel 1101 by a gas-permeable membrane installed in the closed-system culture vessel 1101. The gas concentration adjustment unit 1106 may be directly connected to the closed-system culture vessel 1101. The temperature adjustment unit 1104, gas concentration adjustment unit 1106, and CO2 / O2 sensor 1108 may also be connected to the closed-system culture vessel 1101. In this configuration, gas is supplied directly into the closed-system culture vessel 1101.

[0039] Figure 12 shows the procedure for producing regenerated tissue using the automated culture device with the above functions.

[0040] <Step S1: Installing the flow path> The automatic culture device is started. Information about the multiple culture devices to be used, such as the device number, is recorded in the work procedure protocol. An electronic tag or two-dimensional barcode is pre-installed on the culture device, and it is preferable to confirm in advance that the electronic tag or two-dimensional barcode matches the information recorded in the work procedure protocol, such as the device number, by reading it with a barcode reader or the like. The operator presses the start switch on the operating section of the control device to start it. The inside of the device has been disinfected or sterilized in advance to create a clean environment.

[0041] Next, the operator checks the operation screen on the control unit's display to ensure that the internal environment of the automatic culture device is appropriate. For example, the operator checks that the incubator temperature is 37°C. These values are not limited; for example, the temperature can be selected from a range of 0°C to 45°C. The automatic culture schedule is determined in advance by the culture program, and this information is reflected in the automatic culture device. The automatic culture schedule also includes conditions such as the date and time, volume of fluid, etc. for cell seeding, medium change, culture supernatant collection, tissue collection for testing, and tissue collection for transplantation.

[0042] A closed flow path including a closed culture vessel is installed in the culture device beforehand. When the closed flow path is removed from the warehouse, the electronic tag or 2D barcode installed in the closed flow path beforehand is read using a barcode reader or similar device to confirm that it matches the information such as the device number recorded in the work procedure protocol. The closed flow path consists of a closed culture vessel, a medium container containing fresh medium, a closed culture vessel with a gas-permeable membrane for collecting the culture supernatant after culture, and the flow path tubing that connects them. The configuration depends on the type of closed flow path selected. When cells are seeded manually, the seeding is performed in advance in a safety cabinet, and the closed culture vessel containing the cell suspension is installed in the device. When cells are seeded automatically using an automatic culture device, a cell suspension prepared to a specified concentration is placed in a cell bottle in a safety cabinet beforehand, and the closed culture vessel containing it is installed in the device. Automatic culture start After the start of the process, the automatic culture device transfers the cell suspension from the cell bottle to each closed culture vessel to seed the cells.

[0043] <Step S2: Cell culture> The closed culture vessel is left stationary and cultured for the specified time. The temperature is maintained at 37°C using an incubator. The air inside the vessel is constantly agitated by a fan to ensure uniform temperature distribution. To improve production safety, a particle counter or viable cell counting device may be installed in the vessel to monitor cleanliness. When using a closed culture vessel equipped with a gas-permeable membrane or filter, gas exchange occurs between the gas phase inside the incubator and the gas phase inside the closed culture vessel via the gas-permeable membrane or filter. When using a closed culture vessel equipped with an air supply channel, gas exchange occurs by supplying a specific gas directly into the closed culture vessel. Gas exchange is performed several times a day during the culture period. For example, air containing 5% CO2 is used as the supplied gas. The gas is delivered from a gas cylinder while the flow rate is controlled by a gas flow meter. It is then supplied to each closed culture vessel via a humidifying bottle to saturate the water vapor. After being supplied to the closed culture vessel, unnecessary gas is discharged outside the flow path via a filter, preferably one with a pore size of 0.22 μm or more.

[0044] <Step S3: Medium exchange> Culture medium changes are performed every few days during the culture period. Medium addition can simply involve adding fresh medium. Culture medium stored at 4°C in a refrigerator is transferred to a preheating bottle and preheated to 37°C. The temperature can also be raised to 37°C while transferring the medium at a slow transfer rate. When changing the medium, first discharge the old medium from the closed culture vessel. After discharge, quickly supply new medium into the closed culture vessel. The old medium is finally discharged into a culture supernatant container. If necessary, collect the culture supernatant from the culture supernatant container and evaluate the cell growth state by medium component analysis. The medium can also be changed by pushing out new medium while the old medium is still inside.

[0045] <Step S4: Check if it is the day before the scheduled transplant date> It is checked whether it is the day before the planned transplantation date, and if it is not the day before, return to step S2 to continue culturing. If it is the day before, proceed to the next step.

[0046] <Step S5: Collection of tissue for testing> When multiple closed culture vessels are being cultured simultaneously, some of the closed culture vessels may be collected for testing. The door of the culture device is opened, and the flow path tubing of the closed culture vessel to be tested is aseptically cut using heat welding or other means to remove the closed culture vessel. The removed closed culture vessel is transported to a safety cabinet or outside the CPF, and testing is carried out promptly. For example, the cell count, viability, and expression of specific proteins of the biological sample are evaluated.

[0047] <Step S6: Cultivation and medium change immediately before transplantation> Cultivation is carried out by the same procedure as in step S3, and immediately before step S7, the medium is replaced by the same procedure as in step S4.

[0048] <Step S7: Collection and transportation of transplant tissue> If the evaluation in step S5 determines that the condition is suitable for transplantation, the biological sample is collected and used for regenerative medicine treatment. As in step S5, the closed culture vessel is aseptically separated from the closed flow path and removed from the incubator. If necessary, it is transported to a safety cabinet and processed.

[0049] In the shipping room, the closed culture vessels are placed in short-distance or long-distance transport containers. By using heat storage materials, airtight containers, packaging, etc., the effects of temperature, pressure, shock, etc. are minimized throughout the entire transport process. In this state, the transport containers are taken out of the CPF and transported to the operating room by vehicle, train, airplane, hand-carrying, etc. as needed. Before treatment in the operating room, an acceptance inspection is conducted as needed. In the case of short-distance transport, it is expected that the state of the cells will remain almost unchanged from immediately before transport, so it is up to the operator to decide not to perform this step.

[0050] <Step S8: Transplantation> After arriving at the operating room, the regenerated tissue is removed from the closed culture vessel. When opened, the outside of the closed culture vessel may be contaminated with organisms such as bacteria or particles, so it is opened aseptically to maintain the cleanliness of the inside of the closed culture vessel.

[0051] Finally, the closed flow path used for culturing is removed. Next, the inside of the device is decontaminated with decontamination gas or wiped and disinfected with ethanol or the like using appropriate procedures to clean it. The various software programs for the automatic culturing device are shut down, and the operation of the automatic culturing device is terminated.

[0052] According to a preferred embodiment of the automatic culture device including the closed culture vessel configured as described above, by using a highly versatile closed flow path for multiple culture devices and using a culture program to operate multiple devices, the integration of the culture devices can be improved, and as a result, manufacturing costs can be reduced.

[0053] <Embodiment II> A method using an automatic culture device with a different configuration from that of Example 1 will be described with reference to FIG.

[0054] This embodiment does not have the refrigeration unit that is provided in embodiment I. The medium container and culture supernatant container are placed on the side of the device, which is kept at room temperature. In Figure 13, a medium container 1301 is placed.

[0055] The closed flow path can be installed in the same manner as in embodiment I. The culture devices can be integrated by stacking or other methods. Cells can be cultured using one closed flow path for multiple culture devices. This makes it possible to culture a large amount of cells. In addition, the culture program shown in embodiment 1 can be used to operate multiple culture devices.

[0056] <Embodiment III> A method using an automatic culture device with a different configuration from that of embodiment I will be described with reference to FIG.

[0057] This example does not have the refrigeration unit that is provided in embodiment I. The flow path mechanism unit 102 in embodiment I is integrated with the incubator 101. The medium container, culture supernatant container, etc. are placed on the side of the device, which is kept at room temperature. In Figure 14, a medium container 1401 is placed.

[0058] It is possible to install a closed flow path similar to that of embodiment I. The culture devices can be integrated by stacking them, etc. It is possible to culture cells using one closed flow path for multiple culture devices. This makes it possible to culture a large amount of cells. In addition, since multiple culture devices are operated, the culture program shown in embodiment I can be used. [Explanation of symbols]

[0059] 101 Incubator 102 Flow path mechanism 103 Refrigerated section 104 Pump 105···Solenoid valve 106 Notch 107...Installation stand 108···Humidification tray 201...Closed culture container 202 Culture medium container 203 Flow path tube 204···Filter 301···Closed culture vessel with external filter 302···Filter 303 Flow path tube 304 Pump 305···Solenoid valve 306 Culture medium container 307...Culture supernatant container 308....Closed culture vessel with gas-permeable membrane 309 Gas-permeable membrane 310...Closed culture vessel for perfusion culture 1101...Closed culture container 1102 Control device 1103 Incubator 1104...Temperature control section 1105 Gas supply unit 1106 Gas concentration adjusting unit 1107 Pump 1108···CO2·O2 sensor 1109...Display device 1110 Temperature Sensor 1111···Culture medium container·Culture supernatant container 1301 Culture medium container 1401 Culture medium container

Claims

1. A cell culture system comprising an automatic culture device and an information processing device, The automatic culture device is A plurality of types of closed flow paths that can be installed and removed; A plurality of culture devices; Equipped with the plurality of types of closed flow paths are configured so that different culture methods can be performed using the culture devices connected to each closed flow path, The information processing device includes: an input device that receives data regarding the cell type and / or data regarding the required number of cells as input; a computing device that selects the cell culture method, the culture device, and the closed flow path to be used from options for the cell culture method, the culture device, and the closed flow path based on the input data; and an output device that outputs the selected option for the closed system flow path and the selected option for the culture device; Equipped with Cell culture system.

2. The closed flow path is 2. The cell culture system according to claim 1, comprising: a valve for opening and closing a flow path tube; a pump for supplying a fluid or gas through the flow path tube; a medium container for holding fresh medium; and a culture supernatant container for holding a culture supernatant after culture.

3. The cell culture system according to claim 1 or 2, wherein the output device outputs a work procedure manual for the closed flow path to be used.

4. The cell culture system according to any one of claims 1 to 3, wherein the output device outputs an operating procedure manual for the selected culture device.

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