Cell state estimation system

Abstract

Provided is a cell state estimation system capable of accurately determining the state of a cell to be cultured. The cell state estimation system is provided with: an appearance information acquisition unit that acquires appearance information of a second cell different from a first cell to be cultured, and that provides a culture environment to the second cell from at least some of the culture media that provide the culture environment to the first cell; a physical property measurement unit that measures at least one physical property for at least one of the second cells and the medium in which an environment for culturing the second cells is provided; and an estimation reference information unit that stores estimation reference information referenced for estimating the state of at least one of the first cell and the culture medium, the estimation of the state of at least one of the first cell and the culture medium being based on at least one of the result acquired by the appearance information acquisition unit and the result measured by the physical property measurement unit.

Description

Technical Field

[0001] The present invention relates to a system for estimating the state of cells to be cultured. Background Art

[0002] A system is known that determines the state of cells to be cultured based on an image obtained by imaging the cells (for example, see Patent Document 1). This system images cells cultured in a cell culture apparatus and determines the state.

[0003] Prior Art Literature Patent Literature Patent Document 1: International Publication No. WO2014 / 203322 Summary of the invention Problems to be solved by the invention The cells are immersed in the culture medium in the cell culture tank. The culture medium is a medium that provides a culture environment and is an indispensable substance for culturing cells. The culture medium often becomes turbid during the culture process, and the cells cannot be clearly photographed in the presence of turbid culture medium. Therefore, it is difficult to accurately determine the state of the cells.

[0004] The present invention has been made in view of the above-mentioned problems, and an object of the present invention is to provide a system capable of accurately determining the state of cells to be cultured.

[0005] Technical solutions to solve problems The cell state estimation system of the present invention is characterized by comprising: an appearance information acquisition unit that acquires appearance information of a second cell different from a first cell to be cultured, wherein at least a portion of a culture medium that provides a culture environment for the first cell provides a culture environment for the second cell; a physical property measuring unit configured to measure at least one physical property of at least one of a culture medium providing a culture environment for the second cells and the second cells; and An estimation reference information unit stores estimation reference information for reference in estimating a state of at least one of the first cell and the culture medium, wherein the estimation of the state of at least one of the first cell and the culture medium is based on at least one of a result obtained by the appearance information acquisition unit and a result measured by the physical property measurement unit.

[0006] Effects of the Invention The state of cells to be cultured can be accurately determined. BRIEF DESCRIPTION OF THE DRAWINGS

[0007] Figure 1 This is a block diagram showing an example of a cell state estimating system according to the present embodiment.

[0008] Figure 2: is a functional block diagram showing the structure of the control device 330 of the cell state estimation system according to the present embodiment.

[0009] Figure 3 3 is a flowchart showing the product cell state determination process performed by the control device 330 .

[0010] Figure 4 This is a table showing an example of the estimation state space table.

[0011] Figure 5 It is a schematic plan view showing the holding member 510 according to the present embodiment.

[0012] Figure 6 It is a cross-sectional view taken along the line AA′ of the holding member 510 according to the present embodiment.

[0013] Figure 7 It is a schematic plan view showing a holding member 710 according to a modified example of the present embodiment. DETAILED DESCRIPTION

[0014] <<<<Overview of this embodiment>>>> <<First embodiment>> According to a first embodiment, there is provided a cell state estimating system comprising: an appearance information acquisition unit (e.g., the camera 310 described later, the captured image acquisition unit 340 described later, etc.) that acquires appearance information of a second cell (e.g., the sensor cell described later, etc.) different from a first cell (e.g., the product cell described later, etc.) that is a culture target, and at least a portion of the culture medium that provides a culture environment for the first cell provides a culture environment for the second cell; a physical property measuring unit (e.g., the analytical measurement device 320, the analytical measurement result acquisition unit 350, etc.) for measuring at least one physical property of at least one of a culture medium providing a culture environment for the second cells and the second cells; and An estimation reference information unit (e.g., the state estimation unit 360 described later, a state space table for estimation, etc.) stores estimation reference information for reference in order to estimate the state of at least one of the first cell and the culture medium, wherein the estimation of the state of at least one of the first cell and the culture medium is based on at least one of the result obtained by the appearance information acquisition unit and the result measured by the physical property measurement unit.

[0015] The cell state estimation system includes an appearance information acquisition unit, a physical property measurement unit, and an estimation reference information unit.

[0016] The appearance information acquisition unit acquires appearance information of a second cell different from a first cell that is a culture object. The first cell is a culture object. The second cell is different from the first cell. It should be noted that the second cell may also be a culture object.

[0017] The culture medium is a medium that provides a culture environment for cells. At least a portion of the culture medium that provides a culture environment for the first cell is provided as a culture environment for the second cell. Thus, at least a portion of the culture medium is shared by the first cell and the second cell. It should be noted that the culture medium provided to the first cell may also be provided as a culture environment for the second cell.

[0018] The physical property measuring unit measures at least one physical property of at least one of the culture medium providing a culture environment for the second cells and the second cells. The physical property related to both the culture medium providing a culture environment for the second cells and the second cells may be measured, or the physical property related to either one of them may be measured.

[0019] The estimation reference information unit stores estimation reference information. The estimation reference information is information referenced to estimate the state of at least one of the first cell and the culture medium. The estimation reference information unit may estimate both the state of the first cell and the state of the culture medium, or may estimate only the state of one of them.

[0020] The estimation is performed based on at least one of the result acquired by the appearance information acquisition unit and the result measured by the physical property measurement unit. The estimation may be performed based on both results or based on either result.

[0021] The appearance information acquisition unit acquires appearance information of a second cell different from the first cell. That is, the appearance information acquisition unit does not directly acquire appearance information of the first cell. In addition, the physical property measurement unit measures physical properties related to the culture medium that provides a culture environment for the second cell and physical properties related to the second cell. That is, the physical property measurement unit does not directly acquire physical properties related to the culture medium that provides a culture environment for the first cell and the first cell.

[0022] That is, the second cell is preferably a cell that functions as a sensor for estimating the state of the first cell.

[0023] Furthermore, the second cell is preferably arranged at a position separated from the first cell.

[0024] The estimation reference information unit estimates the state of the first cell and the state of the culture medium based on the appearance information of the second cell, the physical properties related to the culture medium providing the culture environment of the second cell, and the physical properties related to the second cell. That is, the estimation reference information unit indirectly determines the state of the first cell based on the state of the second cell. With such a structure, even if the culture environment of the first cell is not suitable for observation, measurement, etc., the state of the first cell can be accurately determined and judged by using the state of the second cell.

[0025] In addition, the culture environment of the first cells will not be contaminated by observation, measurement, etc., and the culture environment of the first cells can be maintained clean.

[0026] <<Second embodiment>> Regarding the second embodiment, in the first embodiment, further comprising: at least one first tank (for example, the cell culture tank 100 described later), which contains the first cells immersed in a culture medium; at least one second tank (for example, the simulated culture tank 110 described later), which contains the second cells immersed in a culture medium; and The communication portion (for example, the pipe 200B, the pipe 200C, the organ cell tank 120, etc. described later) connects the first tank and the second tank so that the culture medium can flow between the first tank and the second tank.

[0027] <<Third Embodiment>> Regarding the third embodiment, in the second embodiment, further comprising: At least one third tank (eg, the organ cell tank 120 described below) contains third cells (eg, feeder cells described below) for culturing the first cells, and the culture medium can flow.

[0028] The culture medium preferably flows in the first tank, the second tank, and the third tank.

[0029] <<Fourth embodiment>> Regarding the fourth embodiment, in the first embodiment, further comprising: A control unit (eg, the estimation device 300, the control device 330, the pump 210, etc., which will be described later) that determines the flow rate of the culture medium based on the reference result of the estimation reference information.

[0030] <<Fifth embodiment>> Regarding the fifth embodiment, in the first embodiment, further comprising: A control unit (eg, the estimation device 300 or the control device 330 described later) that determines a timing for replacing the culture medium based on a reference result of the estimation reference information.

[0031] <<Sixth embodiment>> Regarding the sixth embodiment, in the first embodiment, further comprising: A control unit (for example, the estimation device 300 or the control device 330 described later) that determines a timing for adding nutrients to the culture medium based on a reference result of the estimation reference information.

[0032] <<<<Details of this embodiment>>>> The cell culture estimation system 10 according to the present embodiment maintains the culture environment of the product cells clean, and estimates the growth degree of the product cells based on the growth degree of the sensor cells and the state of the culture medium providing the culture environment of the sensor cells. The cell culture estimation system 10 controls the state of the liquid culture medium based on the estimation result of the liquid culture medium, for example, by adding nutrients required for the liquid culture medium. The cell culture estimation system 10 estimates the growth degree of the product cells at that moment, or predicts the growth degree of the product cells after that moment (in the future).

[0033] <<<Object of the cell culture estimation system 10>>> The target objects of the cell culture estimation system 10 include product cells, sensor cells (canary cells), culture media (liquid culture media), feeder cells, and the like.

[0034] <<Product Cell>> Product cells are culture objects. For example, product cells are preferably cells that constitute edible meat, etc. Product cells are not limited to edible cells, as long as they can be cultivated. Product cells are cultivated by culture. The degree of cultivation of product cells, health status, etc. are also referred to as the status of product cells.

[0035] <<Sensor cells (canary cells) (substitute for sensory cells)>> Sensor cells are cells used to estimate the state of product cells. In the cell culture estimation system 10, the state of product cells is not directly acquired, but acquired (estimated) indirectly through sensor cells.

[0036] The sensor cells are preferably non-proliferative cells. For example, nerve cells, sertoli cells, etc. can be used as sensor cells. In addition, the sensor cells can be the same cells as the product cells. For example, muscle cells, liver cells, etc. can be used as sensor cells.

[0037] <<Culture medium (liquid culture medium)>> The culture medium is supplied to both the product cells and the sensor cells. The culture medium flows between the product cells and the sensor cells. The culture medium is shared by both the product cells and the sensor cells. The liquid culture medium is also called a culture solution.

[0038] <<Feeder Cells>> Feeder cells are cells used to grow product cells. For example, feeder cells can be cells of the lung, intestine, brain, muscle, skin, gonad, stomach, etc. in addition to liver and kidney. Feeder cells are used to grow product cells or improve the quality of product cells through a combination of these organs. Specifically, various endocrine factors (hormones, cytokines) produced by feeder cells are given to product cells to grow product cells.

[0039] Hereinafter, embodiments will be described based on the drawings. Figure 1 This is a block diagram showing an example of a cell state estimating system according to the present embodiment. Figure 2 : is a functional block diagram showing the structure of the control device 330 of the cell state estimation system according to the present embodiment.

[0040] <<<Configuration of cell culture estimation system 10>>> The cell culture estimation system 10 mainly includes a cell culture tank 100 , a simulated culture tank 110 , an organ cell tank 120 , a buffer tank 130 , a pump 210 , and an estimation device 300 .

[0041] <<Cell culture tank 100>> The cell culture tank 100 is a tank for culturing product cells. A culture medium is stored in the cell culture tank 100. The product cells are kept immersed in the culture medium.

[0042] The cell culture tank 100 has a certain shape and size. The cell culture tank 100 has, for example, a rectangular parallelepiped shape. The cell culture tank 100 only needs to have a size sufficient to grow the product cells. The cell culture tank 100 only needs to have a size that can accommodate the product cells in a manner that the product cells are not removed during the growth process until they reach the final growth state.

[0043] The cell culture tank 100 is formed of a material that allows visual confirmation of the interior. The material of the cell culture tank 100 may be appropriately determined according to the culture environment of the product cells, etc. The cell culture tank 100 is formed of, for example, glass, plastic, or other resins, etc. The cell culture tank 100 allows visual confirmation of the culture state of the product cells, such as the amount of culture medium. It should be noted that the cell culture tank 100 may be partially made of metal, etc., as long as it has a window that allows visual confirmation.

[0044] The cell culture tank 100 has a cover (not shown). The cell culture tank 100 is sealed by the cover. The cell culture tank 100 is kept sealed by the cover during the cultivation of product cells. The cell culture tank 100 can be kept clean by being sealed.

[0045] The cell culture tank 100 stores a medium in an amount required for culturing the product cells. The medium can flow in the cell culture tank 100. The product cells are completely immersed in the medium in the cell culture tank 100. In the cell culture tank 100, the surface of the product cells is kept in contact with the medium.

[0046] The cell culture tank 100 has a holding member (not shown) for holding the product cells, a locking member (not shown), etc. The product cells are held or locked without contacting the side walls (not shown) of the cell culture tank 100. In the cell culture tank 100, the product cells can be smoothly cultured.

[0047] <Pipe 200A and Pipe 200B> The pipe 200A and the pipe 200B are connected to the cell culture tank 100 .

[0048] The cell culture tank 100 has an opening (not shown) for the pipe 200A. The cell culture tank 100 is connected to the pipe 200A. The cell culture tank 100 is connected to the buffer tank 130 through the pipe 200A. The culture medium flows from the buffer tank 130. The culture medium flows through the pump 210 described later. The position and size of the opening of the pipe 200A can be appropriately determined.

[0049] The cell culture tank 100 has an opening (not shown) for the pipe 200B. The pipe 200B is connected to the cell culture tank 100. The cell culture tank 100 is connected to the organ cell tank 120 described later through the pipe 200B. The culture medium flows out to the organ cell tank 120. The position and size of the opening of the pipe 200B can be appropriately determined.

[0050] <Flow of Culture Medium in Cell Culture Tank 100 > The culture medium flows in from the buffer tank 130, flows in the cell culture tank 100, and flows out to the organ cell tank 120. The culture medium flows in through the pipe 200A and flows out through the pipe 200B. The culture medium always flows in the cell culture tank 100. The culture medium is smoothly supplied to the cell culture tank 100, providing a culture environment suitable for the culture of product cells.

[0051] <<Simulated culture tank 110>> The sensor cells are housed in the simulated culture tank 110. The simulated culture tank 110 is a tank for growing and observing the sensor cells. The culture medium is stored in the simulated culture tank 110. The sensor cells are kept immersed in the culture medium in the same manner as the product cells.

[0052] The simulated culture tank 110 has a certain shape and size. The simulated culture tank 110 has, for example, a rectangular parallelepiped shape. The simulated culture tank 110 only needs to have a size sufficient for growing and observing the sensor cells. The simulated culture tank 110 only needs to have a size that can accommodate the sensor cells in a manner that the sensor cells are not removed during the growth process until they reach the final growth state of the product cells.

[0053] The simulated culture tank 110 is formed of a material that allows observation of the interior. The material of the simulated culture tank 110 may be appropriately determined according to the cultivation environment of the sensor cells, the observation environment, etc. The simulated culture tank 110 is formed of, for example, glass, plastic, or other resins. Specifically, the simulated culture tank 110 may be configured so that the state of the sensor cells can be photographed using the camera 310 described later.

[0054] The simulated culture tank 110 stores a culture medium in an amount required for growing and maintaining the sensor cells. The culture medium can flow in the simulated culture tank 110. The sensor cells are completely immersed in the culture medium in the simulated culture tank 110. The surfaces of the sensor cells are kept in contact with the culture medium in the simulated culture tank 110.

[0055] Simulated culture tank 110 Figure 5 The sensor cell is shown with a holding member 510. The sensor cell is held at a fixed position by the holding member 510. By holding the sensor cell at a fixed position, the fixed part of the sensor cell can be stably photographed by the camera 310.

[0056] use Figure 5 and Figure 6 The holding member 510 will be described. Figure 5 It is a schematic plan view showing the holding member 510 . Figure 6 It is along Figure 5 A cross-sectional view of the retaining member 510 taken along line AA′ is shown.

[0057] The holding member 510 includes a frame member 520, a predetermined holding area 522, a rod member 524, etc. The frame member 520 has a generally annular shape. The predetermined holding area 522 is an area formed by being surrounded by the frame member 520. The predetermined holding area 522 has a generally circular shape. The predetermined holding area 522 can be defined according to the size and shape of the frame member 520. The rod member 524 has an elongated shape. The rod member 524 has a first end and a second end in the length direction. The frame member 520 is connected to the first end. The second end is held on a wall surface (not shown) of the simulated culture tank 110, etc. The holding member 510 can be positioned at a fixed position of the simulated culture tank 110. The frame member 520 is held in the simulated culture tank 110 in such a manner that the predetermined holding area 522 is opposite to the camera 310. Details will be described later.

[0058] The frame member 520 is a member that surrounds and holds all of the predetermined area 522. The frame member 520 is made of a material that does not have cytotoxicity. The frame member 520 is at least partially covered by the cell group 530 of the sensor cell, whereby the cell group 530 is held in the frame member 520. Therefore, the material of the frame member 520 is preferably a substrate having cell adhesion. By making a substrate having cell adhesion, the holding force can be improved. The material of the frame member 520 can also be a substrate that does not have cell adhesion. The substrate that can be used can be, for example, a polymer material or synthetic resin material such as polyethylene, polypropylene, polyester, polyamide, polyimide, polyurethane, polycarbonate, polypeptide, etc., an inorganic compound such as stainless steel, apatite, calcium phosphate, or a natural material such as a sponge, cellulose, soy protein, polysaccharides, gelatin, egg white, mycelium, and decellularized tissue. From the viewpoint of cell adhesion, the substrate is preferably a polyimide or a sponge.

[0059] exist Figure 6 In the embodiment, the cross-sectional shape of the frame member 520 is generally annular. The cross-sectional shape of the frame member 520 may also be polygonal, generally circular, or generally elliptical. The cross-sectional diameter of the frame member 520 (the maximum width of the cross-sectional length of the frame member orthogonal to the plane holding the predetermined area) is not particularly limited. The lower limit of the cross-sectional diameter of the frame member 520 is, for example, 0.1 μ m or more, 1 μ m or more, 3 μ m or more, 10 μ The upper limit of the cross-sectional diameter of the frame member 520 is, for example, 10 mm or less, 1 mm or less, 100 mm or less, or 100 mm or less. μ m or less.

[0060] The predetermined holding region 522 is a region formed by the inner portion of the frame member 520, and is a region capable of holding a cell group 530 formed by a plurality of sensor cells. The predetermined holding region 522 may be a region surrounded by the frame member 520, and is preferably a region that is a space inside the frame member 520. The shape of the predetermined holding region 522 is generally circular, but may also be polygonal or generally elliptical.

[0061] The area of ​​the predetermined region 522 is such that the frame member 520 can be immersed in the culture medium. Specifically, for example, 1,000,000 μ m 2 Below, preferably 500 to 600000 μ m 2 , more preferably 1000 to 600000 μ m 2 , particularly preferably 1000 to 6000 μ m 2 By setting the planned holding region 522 to such a range, the cell group 530 can be held at a high occupancy rate in the planned holding region.

[0062] The rod member 524 is a long rod-shaped member. The rod member 524 has a first end 525 and a second end 526 in the length direction. The frame member 520 is connected to the first end 525. The second end 526 is held (held) by a holding portion (not shown) having a holding mechanism provided on the wall surface of the simulated culture tank 110. The rod member 524 is formed of the same base material as the frame member 520, but can also be formed of other base materials. The rod member 524 is integrally formed with the frame member 520. The rod member 524 can also be separately formed from the frame member 520. The shape of the rod member 524 is not particularly limited as long as it can be held.

[0063] The holding mechanism has a drive motor for moving in the X direction, a drive motor for moving in the Y direction, a drive motor for moving in the Z direction, a drive motor for tilting, etc. The control device 330 sends a control signal to control the movement amount of the drive motor for moving in the X direction, the drive motor for moving in the Y direction, the drive motor for moving in the Z direction, the tilt of the drive motor for tilting, etc.

[0064] The control device 330 controls the movement and inclination of the rod member 524 (holding member 510) held by the holding portion in the XYZ direction. By performing such control, the rod member 524 (holding member 510) can be easily operated. That is, the cell group 530 held by the holding member 510 can be easily moved. Thus, it is possible to adjust the holding area 522 to face the camera 310, so that a specific part of the cell group 530 can be easily photographed by the camera 310.

[0065] The cell group 530 occupies at least a portion of the predetermined holding area 522 and is a collection of membrane-like sensor cells held by the frame member 520. Figure 5 In the embodiment, the cell group 530 occupies the entirety of the predetermined holding region 522, but the cell group 530 may also occupy a portion of the predetermined holding region 522. The cell group 530 preferably occupies 50% or more of the area of ​​the predetermined holding region 522, more preferably 80% or more, further preferably 90% or more, and most preferably occupies the entirety of the predetermined holding region 522.

[0066] At least a portion of the periphery of the cell colony 530 is in contact with the frame member 520. Chemical bonding between the periphery of the cell colony 530 and the frame member 520 is not necessarily required. Figure 6 As shown, in a given cross section, the entire periphery of the frame member 520 may be covered by a portion of the cell population 530 (see Figure 6 ), a portion of the cell population 530 may cover a portion of the periphery of the frame member 520 (see Figure 6 From the viewpoint of improving the holding force, it is preferred that the entire periphery of the frame member 520 is covered by a portion of the cell population 530 in a given cross section.

[0067] like Figure 6 As shown, the portion of the cell group 530 occupying the predetermined holding area 522 is in a state of floating in the air. "Floating in the air" means that one face and / or the other face of the portion of the cell group 530 occupying the predetermined holding area 522 is exposed, and a space without interference exists on one face side and the other face side of the cell group 530. Thus, for example, when observing cells using a camera 310 or the like, there is no obstacle such as a support, and therefore, the support that can provide cell adhesion does not become an obstacle to cell observation and can hold the cell holding member and the cell holding method thereof.

[0068] use Figure 7 Modifications of the holding member will be described. Figure 7 1 is a schematic plan view showing a modified example of the holding member. The basic structure of the holding member 710 according to the modified example is the same as that of the present embodiment. The structure different from the present embodiment will be described below.

[0069] The frame member 720 involved in the modification is a member having a shape in which a part of the frame member 520 is cut away. The frame member 720 surrounds a part of the predetermined holding area 722. Thus, when the frame member 720 is an open structure (a structure with a cutout), the predetermined holding area 722 is defined by including a line connecting the end points of the open structure part at the shortest distance as the inner part of the frame member. The rod member 724 is the same as the rod member 524.

[0070] According to the modified example of the retaining member 710, in the cutout portion 726 of the frame member 720, it becomes easy for the camera 310 to photograph the cell group 530 and for the camera 310 to photograph from a direction parallel to the predetermined retaining area 722, thereby being able to provide a retaining member and a cell retaining method that can retain cells without providing a support for cell adhesion that becomes an obstacle to cell observation.

[0071] The sensor cell may be fixed at a fixed position by a light-transmitting member such as a test slide. By fixing the sensor cell at a fixed position, the fixed part of the sensor cell can be stably photographed by the camera 310 .

[0072] A method for holding cells on the holding member 510 (holding member 710) is described. A culture medium, sensor cells, and a frame member 520 (frame member 720) are prepared. The frame member 520 (frame member 720) is immersed in the culture medium containing the sensor cells. The immersion time of the frame member 520 (frame member 720) can be set to, for example, 1 hour or more, 2 hours or more, 5 hours or more, 10 hours or more, or 24 hours or more.

[0073] When the frame member 520 (frame member 720) is immersed, the holding member 510 (holding member 710) having the cell group 530 (cell group 730) formed in a membrane in the holding area is obtained. According to such a cell holding method, the holding member can easily hold the cell group 530 (cell group 730).

[0074] By using the holding member produced by the above method, the scaffold for cell adhesion does not become an obstacle to cell observation and the cells can be held, so that the manipulation of the cell group in micrometer units becomes easy. The holding member can be used in the produced culture container or taken out for use.

[0075] A certain amount of culture medium is accumulated in the simulated culture tank 110. In the simulated culture tank 110, a culture environment for sensor cells is provided by the culture medium. In the simulated culture tank 110, the culture medium can flow. At least a portion of the culture medium flowing into the simulated culture tank 110 is the culture medium flowing out of the cell culture tank 100. The culture medium in the simulated culture tank 110 is shared with the culture medium in the cell culture tank 100. By sharing the culture medium in the cell culture tank 100 and the simulated culture tank 110, a culture environment common to the culture environment of the product cells is provided to the sensor cells.

[0076] <Pipe 200C and Pipe 200D> The pipe 200C and the pipe 200D are connected to the simulated culture tank 110 .

[0077] The simulated culture tank 110 has an opening (not shown) for the pipe 200C. The simulated culture tank 110 is connected to the pipe 200C. The simulated culture tank 110 is connected to the organ cell tank 120 through the pipe 200C. The culture medium flows from the organ cell tank 120. The culture medium flows through the pump 210 described later. The position and size of the opening of the pipe 200C can be appropriately determined.

[0078] The simulated culture tank 110 has an opening (not shown) for the pipe 200D. The simulated culture tank 110 is connected to the pipe 200D. The simulated culture tank 110 is connected to the pump 210 described later through the pipe 200D. The culture medium flows out to the pump 210. The position and size of the opening of the pipe 200D may be appropriately determined.

[0079] <Flow of Culture Medium in Simulated Culture Tank 110 > The culture medium flows in from the organ cell tank 120, flows in the simulated culture tank 110, and flows out to the pump 210. The culture medium flows in through the pipe 200C and flows out through the pipe 200D. The culture medium always flows in the simulated culture tank 110. The culture medium is smoothly supplied to the simulated culture tank 110, providing a culture environment suitable for the cultivation and maintenance of sensor cells.

[0080] <<Cultivation environment in simulated culture tank 110>> The culture medium is caused to flow by the pump 210. Specifically, the culture medium circulates between the cell culture tank 100, the simulated culture tank 110, the organ cell tank 120, and the buffer tank 130. Figure 1 In the example shown, the culture medium refluxes in the order of the cell culture tank 100 → the organ cell tank 120 → the simulated culture tank 110 → the pump 210 → the buffer tank 130. By the reflux of the culture medium, the culture medium is shared by both the cell culture tank 100 and the simulated culture tank 110. By sharing the culture medium, the sensor cells in the simulated culture tank 110 are provided with a culture environment that is the same as the culture environment of the product cells in the cell culture tank 100.

[0081] <<Organ cell tank 120>> The organ cell tank 120 is a tank for accommodating feeder cells. A culture medium is stored in the organ cell tank 120. The feeder cells are kept immersed in the culture medium in the same manner as the product cells and the sensor cells.

[0082] The culture medium can flow in the organ cell tank 120. In the organ cell tank 120, the feeder cells are entirely immersed in the culture medium. In the organ cell tank 120, the surface of the feeder cells is maintained in contact with the culture medium.

[0083] The organ cell tank 120 has a holding member (not shown) and a locking member (not shown) for holding feeder cells. The feeder cells are held or locked without contacting the side walls (not shown) of the organ cell tank 120. In the organ cell tank 120, feeder cells can be housed in a desired environment.

[0084] <Pipe 200B and Pipe 200C> The organ cell tank 120 is connected to the pipe 200B and the pipe 200C. The organ cell tank 120 is connected to the cell culture tank 100 via the pipe 200B. The organ cell tank 120 is connected to the simulated culture tank 110 via the pipe 200C. The culture medium flows into the cell culture tank 100, flows in the organ cell tank 120, and flows out to the simulated culture tank 110.

[0085] <<Buffer tank 130>> The culture medium is stored in the buffer tank 130. The pipe 200E and the pipe 200A are connected to the buffer tank 130. The buffer tank 130 is connected to the pump 210 through the pipe 200E. The buffer tank 130 is connected to the cell culture tank 100 through the pipe 200A. The culture medium is refluxed through the buffer tank 130.

[0086] The buffer tank 130 has a culture medium supply port 140. The culture medium supply tank 144 is connected to the culture medium supply port 140 via a valve 142. The valve 142 is opened and closed by the control device 330. A new culture medium can be supplied from the culture medium supply tank 144 to the buffer tank 130. It should be noted that it is preferable to supply the new culture medium via a filter (not shown) that does not allow fungi to pass. The buffer tank 130 can be kept clean.

[0087] The buffer tank 130 has a nutrient supply port 150. A nutrient supply tank 154 is connected to the nutrient supply port 150 via a valve 152. The valve 152 is opened and closed by the control device 330. Necessary nutrients can be supplied from the nutrient supply tank 154 to the buffer tank 130. It should be noted that it is preferable to supply necessary nutrients via a filter (not shown) that does not allow fungi to pass through. The buffer tank 130 can be kept clean.

[0088] The buffer tank 130 has a culture medium discharge port 160. A part of the culture medium can be discharged from the culture medium discharge port 160 via a valve 162. The valve 162 is controlled to open and close by the control device 330.

[0089] For example, a given amount of medium (e.g., half the amount of the buffer tank 130) is discharged from the medium discharge port 160, and at the same time, an equal amount of new medium is introduced from the medium supply tank 144 through the medium supply port 140. In this way, a part of the medium can be replaced. Specifically, the medium is replaced under the condition that the pH value of the sampled medium and the mass of the protein contained in the sampled medium are lower than a preset threshold value.

[0090] <<Pump 210>> The pump 210 has an electric motor (not shown). The culture medium is caused to flow by the electric motor. The flow rate of the culture medium per unit time is controlled by the rotation speed of the electric motor. The pump 210 is driven by a control signal from the control device 330. The control signal includes information indicating the rotation speed of the electric motor.

[0091] The pump 210 connects the pipe 200D and the pipe 200E. The pump 210 is connected to the simulated culture tank 110 through the pipe 200D. The pump 210 is connected to the buffer tank 130 through the pipe 200C. The culture medium flows from the simulated culture tank 110 into the pump 210, and flows out from the pump 210 to the buffer tank 130 by the driving force of the pump 210.

[0092] By driving the pump 210, the culture medium can be refluxed. Figure 1 In the illustrated example, the culture medium is refluxed in the order of the cell culture tank 100 →the organ cell tank 120 →the simulated culture tank 110 →the pump 210 →the buffer tank 130 .

[0093] <<Estimation device 300>> The estimation device 300 includes a camera 310, an analysis and measurement device 320, and a control device 330. As described above, the sensor cells in the simulated culture tank 110 are provided with a culture environment that is the same as the culture environment of the product cells in the cell culture tank 100. Therefore, the state of the sensor cells is made the same as the state of the product cells, and the state of the product cells can be estimated. The estimation device 300 estimates the state of the product cells, controls the opening and closing of the valves 142, 152, and 162 of the buffer tank 130, or controls the flow rate of the culture medium using the pump 210.

[0094] <Camera 310> The camera 310 photographs the sensor cells to obtain appearance information of the sensor cells. The health status of the sensor cells can be determined based on the appearance information of the sensor cells. The appearance information of the sensor cells can be information of a narrow area of ​​the sensor cells or information of a wide area of ​​the sensor cells. The appearance information can be used as long as it can determine the health status of the sensor cells.

[0095] The camera 310 has an imaging element such as a CCD element or a CMOS element. The imaging element may be any imaging element suitable for imaging the appearance of the surface of the sensor cell, etc. The imaging element has a given resolution. The resolution may be any resolution required for estimating the health status of the sensor cell.

[0096] The camera 310 may also include an objective lens (not shown). The objective lens can be used to capture the appearance of the sensor cells at a desired magnification. The magnification may be set to a magnification required for determining the health status of the sensor cells.

[0097] Furthermore, an illumination device (not shown) may be provided. The brightness may be set to a required brightness for determining the health status of the sensor cells. The illumination device illuminates the surface of the sensor cells. The illumination device preferably does not change the cultivation environment such as the temperature of the sensor cells and the culture medium. The illumination device emits light of a wavelength required for photographing the appearance, such as white light. The brightness and wavelength preferably do not affect the cultivation of the sensor cells.

[0098] The shooting of the camera 310 is controlled by a control signal from the control device 330. The shooting conditions are determined according to the control signal, the shooting is started according to the control signal, and the shooting is ended according to the control signal.

[0099] The image captured by the camera 310 may be a single static image or a dynamic image. The image captured by the camera 310 may also be a plurality of static images captured at each given time. The image captured by the camera 310 may be any image as long as it can determine the health status of the sensor cells.

[0100] <Analysis and measurement device 320> The analysis and measurement device 320 is connected to the simulated culture tank 110 via a sampling tube 322. The analysis and measurement device 320 analyzes / measures the culture medium sampled from the simulated culture tank 110. The analysis and measurement device 320 mainly analyzes / measures the culture medium that provides the culture environment for the sensor cells. With such a structure, the analysis and measurement device 320 can analyze / measure the culture medium without easily affecting the culture environment of the product cells.

[0101] The analysis and measurement device 320 includes an optical sensor (not shown), a potential sensor (not shown), and the like.

[0102] <Optical sensor (spectroscopic analysis)> The optical sensor emits light from a light source and irradiates the sampled culture medium. The analytical measuring device 320 quantifies the absorbance of the sampled culture medium. The analytical measuring device 320 can obtain the mass of the protein contained in the sampled culture medium. The analytical measuring device 320 obtains the mass of the protein contained in the culture medium that provides the cultivation environment for the sensor cells. It should be noted that the analytical measuring device 320 can also measure the concentration of glucose.

[0103] <Potential sensor> The potential sensor quantifies the potential difference of the sampled culture medium. The analysis and measurement device 320 can obtain the pH value of the sampled culture medium. The analysis and measurement device 320 obtains the pH value of the culture medium that provides the culture environment for the sensor cells.

[0104] <Other> The analysis and measurement device 320 may also measure the temperature of the culture medium. The analysis and measurement device 320 only needs to obtain parameters that change as the product cells and sensor cells grow. The obtained parameters only need to be parameters that can be used to estimate the state of the product cells based on the values ​​of the parameters.

[0105] For example, the analysis and measurement device 320 can also detect the fouling of the culture medium (accumulation of metabolites), obtain data for instructions for replacement of the culture medium, instructions for operation of the dialysis device, or predict the replacement time based on the obtained data. Specifically, it is possible to predict changes caused by nutritional starvation of cells, excess metabolites, etc.

[0106] In addition, the analysis and measurement device 320 can also detect the components of metabolites and determine the deterioration state and health state of the culture medium.

[0107] <Control device 330> like Figure 2 As shown, the control device 330 mainly includes a CPU (central processing unit), a ROM (read only memory), a RAM (random access memory), a HDD (hard disk drive), an input / output interface, etc. (none of which are shown in the figure). The CPU, ROM, RAM, HDD, input / output interface, etc. constitute a captured image acquisition unit 340, an analysis and measurement result acquisition unit 350, a state estimation unit 360, a valve control unit 370, and a flow control unit 380.

[0108] <Photographed Image Acquisition Unit 340> The captured image acquisition unit 340 is electrically connected to the camera 310 via the input / output interface. The captured image acquisition unit 340 outputs a control signal indicating a shooting condition, a control signal for starting shooting, a control signal for ending shooting, etc. to the camera 310 at a given timing.

[0109] The captured image acquisition unit 340 acquires appearance information of the sensor cell from the captured image signal outputted from the camera 310. The captured image acquisition unit 340 receives the captured image signal outputted from the camera 310 and forms the captured image as appearance information. The appearance information is sequentially stored in a HDD or the like.

[0110] The appearance information of the sensor cells is stored in a HDD etc. A change in the health state of the sensor cells can be determined based on the appearance information of the sensor cells.

[0111] <Analysis and Measurement Result Acquisition Unit 350> The analysis and measurement result acquisition unit 350 is electrically connected to the above-mentioned analysis and measurement device 320 (optical sensor, potential sensor) via the input and output interface. The analysis and measurement result acquisition unit 350 acquires the protein mass (glucose concentration) of the culture medium from the output signal of the optical sensor, or acquires the pH value of the culture medium from the output signal of the potential sensor.

[0112] <State Estimation Unit 360> The state estimating unit 360 estimates the health state of the product cells based on the appearance information of the sensor cells, the quality of the protein in the culture medium (glucose concentration), and the pH value of the culture medium.

[0113] <Acquiring cell appearance information, obtaining protein quantity (obtaining glucose concentration), and obtaining pH value> For the sake of convenience, the names are as follows. The appearance information of the sensor cell obtained by the captured image acquisition unit 340 is referred to as the acquired cell appearance information. The mass of the protein (glucose concentration) of the culture medium obtained by the analysis and measurement result acquisition unit 350 is referred to as the acquired protein mass (acquired glucose concentration). The pH value of the culture medium obtained by the analysis and measurement result acquisition unit 350 is referred to as the acquired pH value.

[0114] <State space table for estimation> The state estimation unit 360 has an estimation state space table. The estimation state space table is a table to be referred to in order to estimate a state of a product cell based on one acquisition of cell appearance information, one acquisition of protein amount (one acquisition of glucose concentration), and one acquisition of pH value.

[0115] The estimated state space table stores a plurality of product cell states (health state, growth state) in correspondence with a plurality of estimated cell appearance information of sensor cells, a plurality of estimated protein amounts (a plurality of estimated glucose concentrations) of the culture medium, and a plurality of estimated culture medium pH values ​​of the culture medium. Specifically, in the estimated state space table, one piece of estimated cell appearance information, one piece of estimated protein amount, and one culture medium pH value are stored in correspondence with one state of the product cell.

[0116] <An example of a state space table for estimation> Figure 4 is an example of a state space table for estimation. Figure 4 In the estimated state space table shown, "estimated cell appearance information" is represented as "cell appearance state pattern", "estimated protein amount" is represented as "spectroscopic analysis result", "estimated culture medium pH value" is represented as "hydrogen ion concentration (pH)", and "a state of product cells" is represented as "estimated state".

[0117] In addition, Figure 4 The estimated state space table shown also includes the number of days (elapsed days) from the start of cultivation and the temperature of the culture medium. By also using these elapsed days, temperature, etc., the state of the product cells can be determined more accurately.

[0118] <Estimated cell appearance information, estimated protein quality (estimated glucose concentration), estimated culture medium pH value> The plurality of estimated cell appearance information is information of appearance information of a plurality of sensor cells respectively corresponding to a plurality of states of product cells obtained in advance through preliminary experiments. The plurality of estimated protein amounts (a plurality of estimated glucose concentrations) is information of a plurality of estimated glucose concentrations of a plurality of culture medium protein amounts respectively corresponding to a plurality of states of growth of product cells obtained in advance through preliminary experiments. The plurality of estimated culture medium pH values ​​is information of a plurality of culture medium pH values ​​respectively corresponding to a plurality of states of product cells obtained in advance through preliminary experiments.

[0119] The state estimating unit 360 refers to the estimated state space table, and selects one piece of estimated cell appearance information that is closest to one piece of acquired cell appearance information among the plurality of pieces of estimated cell appearance information.

[0120] The state estimating unit 360 refers to the estimated state space table and selects one estimated protein amount (one estimated glucose concentration) among the plurality of estimated protein amounts (one estimated glucose concentration) that is closest to one acquired protein amount (one acquired glucose concentration).

[0121] The state estimating unit 360 refers to the estimation state space table, and selects one estimation medium pH value that is closest to one acquired pH value among the plurality of estimation medium pH values.

[0122] The state estimation unit 360 determines a state of the product cell by referring to the estimated state space table based on the selected one piece of estimated cell appearance information, one piece of estimated protein amount (plurality of estimated glucose concentrations), and one piece of estimated culture medium pH value. The one state determined by referring to the estimated state space table is determined as the state of the product cell.

[0123] In this way, the state estimating unit 360 can determine a state of the product cells based on one acquisition of cell appearance information, one acquisition of protein amount (one acquisition of glucose concentration), and one acquisition of pH value.

[0124] <Specific example 1 of status estimation> For example, when an estimated culture medium has a high protein quality (high protein concentration); and the appearance information of an estimated cell is unhealthy; and the pH value of an estimated culture medium is 6 (weakly acidic), the state estimation unit 360 determines that the product cell at that moment is in an unhealthy state caused by excessive metabolites (lactic acid).

[0125] <Specific example 2 of status estimation> In addition, when an estimated culture medium has a low protein quality (low protein concentration); and an estimated cell appearance information is unhealthy; and an estimated culture medium pH value is 7 (neutral), the state estimation unit 360 determines that the product cells at that time point are in an unhealthy state caused by protein deficiency.

[0126] <Prediction of the future status of product cells> By determining the state of the product cells in sequence at each given time, the temporal change of the state of the product cells can be obtained. The state of the product cells in the future can be estimated based on the temporal change of the state of the product cells. For example, the time when the growth of the product cells is completed, the possibility that the product cells will become unhealthy in the future, etc. can be estimated.

[0127] <Differences between the state of product cells and the state of sensor cells> In the above example, the state of the sensor cell is assumed to be the same as the state of the product cell to determine or estimate the state of the product cell. However, the culture environment of the product cell is different from the culture environment of the sensor cell. Sometimes, there is a difference between the state of the product cell and the state of the sensor cell. Therefore, by storing information related to the difference between the state of the product cell and the state of the sensor cell in the estimation state space table, even when there is a difference, one state of the product cell can be determined or estimated.

[0128] <Valve control unit 370> The valve control unit 370 controls the opening and closing of the valves 142, 152, and 162 according to the determined health status of the product cells. These valves are opened or closed by sending an opening control signal or a closing control signal.

[0129] It should be noted that the valves may be opened or closed and their opening degrees may be controlled to prevent the culture environment of the cell culture tank 100 and the simulated culture tank 110 from being suddenly changed due to a sudden change in the buffer tank 130 .

[0130] <Flow rate control unit 380> The flow control unit 380 controls the flow rate of the pump 210 according to the determined health status of the product cells. The flow control unit 380 outputs a signal indicating the flow rate of the culture medium. The electric motor (not shown) of the pump 210 rotates at a rotation speed according to the flow rate.

[0131] Feedback control can be performed using the valve control unit 370 and the flow rate control unit 380 .

[0132] <<Product cell state determination process performed by the state estimation unit 360>> use Figure 3 The product cell state determination process will be described below. First, the processor of the state estimation unit 360 determines whether it is time to obtain cell appearance information (step S311).

[0133] When the processor of the state estimating unit 360 determines that it is time to obtain the cell appearance information (Yes), it obtains the appearance information of the sensor cell through the camera 310 (step S313 ).

[0134] Next, the processor of the state estimating unit 360 specifies and stores the estimated cell appearance information based on the cell appearance information acquired in the process of step S313 (step S315 ).

[0135] When the processor of the state estimating unit 360 determines in step S311 that it is not the time to acquire cell appearance information (No) or executes the process of step S315 , it determines whether it is the time to perform analysis / measurement (step S317 ).

[0136] When the processor of the state estimating unit 360 determines that it is time to perform analysis / measurement (Yes), the analyzing / measuring device 320 selects and stores the estimated analysis / measurement result of the liquid culture medium (step S319 ).

[0137] When the processor of the state estimating unit 360 determines in step S317 that it is not the time to perform analysis / measurement (No) or executes the process of step S319, it determines whether the estimated cell appearance information and the estimated analysis / measurement result are stored (step S321).

[0138] When the processor of the state estimating unit 360 determines that the estimated cell appearance information and the estimated analysis / measurement results are stored (Yes), it determines a state of the product cell based on the estimated cell appearance information and the estimated analysis / measurement results (step S323), and ends this subroutine.

[0139] When the processor of the state estimating unit 360 determines that the estimated cell appearance information and the estimated analysis / measurement results have not been stored (No), it returns the process to step S311.

[0140] <<<Sequence, configuration (parallel, series), quantity, etc. of each slot>>> In the above example, the structure is shown in which the cells are arranged in the order of cell culture tank 100 → organ cell tank 120 → simulated culture tank 110 → pump 210 → buffer tank 130, but the order is not limited to this. For example, the pump 210 may be arranged before the cell culture tank 100. The culture medium in the buffer tank 130 can be quickly supplied to the cell culture tank 100.

[0141] In the above example, a structure is shown in which each of the cell culture tank 100, the simulated culture tank 110, the organ cell tank 120, the buffer tank 130, and the pump 210 is provided with one, but the number is not limited thereto. By providing a plurality of cell culture tanks 100, the number of product cells to be cultured can be increased. By providing a plurality of simulated culture tanks 110, the state of the product cells can be determined more accurately without relying on individual differences of the sensor cells. By providing a plurality of buffer tanks 130, the buffer tanks 130 can be separated into those for culture medium and those for nutrition, thereby accurately controlling the ratio of culture medium to nutrition.

[0142] In the above example, a structure in which the cell culture tank 100, the simulated culture tank 110, the organ cell tank 120, the buffer tank 130 and the pump 210 are arranged in series is shown, but the arrangement is not limited to this. A plurality of cell culture tanks 100 may also be arranged in parallel. This makes it difficult for the health status of the product cells to be affected by each other. In addition, a plurality of columns of cell culture tanks 100, simulated culture tanks 110, organ cell tanks 120 and pumps 210 arranged in series may be prepared, and only the buffer tank 130 may be shared and arranged in parallel. This makes it difficult for the status of the product cells, sensor cells and feeder cells to be affected by other columns. In this way, a mixed arrangement of series and arrangement is possible.

[0143] In addition, it is also possible to branch or detour in the middle. For example, the cell culture tank 100 may be connected to both the organ cell tank 120 and the buffer tank 130. A part of the culture medium flowing out of the cell culture tank 100 can be supplied to the organ cell tank 120, and the remaining part of the culture medium flowing out of the cell culture tank 100 can be returned to the buffer tank 130.

[0144] <<<<Scope of implementation>>>> As described above, the present embodiment is described, but the description and drawings constituting a part of the present disclosure should not be construed as limiting, and various embodiments not described here are included.

[0145] Explanation of symbols 10 Cell Culture Presumptive System 100 Cell Culture Tanks 110 Simulation culture tank 120 organ cell tank 300 Prediction Device 310 Camera 310 320 Analysis and measurement device 340 Photographed image acquisition unit 350 Analysis and measurement result acquisition unit 360 Status Estimation Department 510 Retaining member 520 frame components 522 Maintaining the Reserved Area 524 Rod components 525 First end 526 Second End 530 Cell population 710 Retaining member 722 Maintaining the Reserved Area 724 Rod components 726 Incision 730 Cell population.

Claims

1. A cell state estimation system comprising: an appearance information acquisition unit that acquires appearance information of a second cell different from a first cell to be cultured, wherein at least a portion of a culture medium that provides a culture environment for the first cell provides a culture environment for the second cell; a physical property measuring unit configured to measure at least one physical property of at least one of a culture medium providing a culture environment for the second cells and the second cells; and An estimation reference information unit stores estimation reference information for reference in estimating a state of at least one of the first cell and the culture medium, wherein the estimation of the state of at least one of the first cell and the culture medium is based on at least one of a result obtained by the appearance information acquisition unit and a result measured by the physical property measurement unit.

2. The cell state estimation system according to claim 1, wherein: The cell state estimation system further comprises: at least one first tank for immersing the first cells in a culture medium for accommodation; at least one second tank for immersing the second cells in a culture medium for accommodation; as well as A connecting portion connects the first groove and the second groove to allow the culture medium to flow between the first groove and the second groove.

3. The cell state estimation system according to claim 2, wherein: The cell state estimating system further includes at least one third tank that accommodates third cells for culturing the first cells and in which the culture medium can flow.

4. The cell state estimation system according to claim 1, wherein: The cell state estimation system further includes a control unit configured to determine a flow rate of the culture medium based on a reference result of the estimation reference information.

5. The cell state estimation system according to claim 1, wherein: The cell state estimation system further includes a control unit that determines a timing for replacing the culture medium based on a reference result of the estimation reference information.

6. The cell state estimation system according to claim 1, wherein: The cell state estimation system further includes a control unit configured to determine a timing for adding nutrients to the culture medium based on a reference result of the estimation reference information.

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