JP2019205400A - Cell-treating method, device and system - Google Patents

Abstract

To realize a rational cell-treating method, device and system having a simple constitution.SOLUTION: A device that is used by being installed to a liquid injection container includes: a lid body which can be installed to the liquid injection container freely detachably; a liquid injection tube which can be fitted into a first through hole provided for the lid body; and an air intake tube which can be fitted into a second through hole provided for the lid body. The device is constituted such that, in the state in which the device is installed to the liquid injection container, a bottom end of the air intake tube which protrudes from the lid body into the liquid injection container is arranged adjacent to a bottom surface of the liquid injection container.SELECTED DRAWING: None

Description

  The present invention relates to a cell processing method, a device, and a system.

  In recent years, attempts have been made to transplant various cells in order to repair damaged tissues and the like. For example, use of fetal cardiomyocytes, skeletal myoblasts, mesenchymal stem cells, cardiac stem cells, ES cells, cardiomyocytes, etc. to repair myocardial tissue damaged by ischemic heart disease such as angina pectoris and myocardial infarction Has been tried. As part of such attempts, cell structures formed using scaffolds and sheet-shaped cell cultures in which cells are formed into sheets have been developed.

  Conventionally, these cell cultures are manufactured manually by workers having specialized knowledge in a clean room called a cell processing center (CPC: Cell Processing Center), and the cost and cost of manufacturing such cell cultures. The labor required for this is large, and its efficiency is desired. In view of this, an automatic cell culture apparatus has been proposed in which work related to the culture of these cells is performed by an articulated robot (Patent Document 1). However, there is a problem that it is difficult to automate advanced tasks that depend on the operator's technique in cell culture.

  In cell culture, the medium exchange process including waste liquid work and liquid injection work is a process that depends on the operator's technique.For example, in the liquid injection work, the culture solution is aspirated and poured by a pipette. It includes advanced operations such as wiping off the cultured medium and pipette replacement. In particular, when sucking the culture solution in the injection bottle with a pipette and injecting it into a culture flask, etc., there are problems that it takes time for complicated pipette operation and the risk of dripping from the tip of the pipette is high. .

International Publication 2016/104666

  Under such circumstances, the present inventors, in developing a means for efficiently performing cell treatment, are difficult to carry out the liquid injection work depending on the operator's advanced technique efficiently and promptly. And faced problems. Accordingly, an object of the present invention is to solve such problems and to realize a reasonable cell processing method, device and system with a simple configuration.

  In the course of diligent research to solve the above-mentioned problems, the inventors have focused on the occurrence of dripping, variation in the amount of liquid to be injected, etc., when the liquid injection operation is performed quickly. As a result of further research, it has been found that the above problem can be solved by using a means for limiting injection, and the present invention has been completed.

That is, the present invention relates to the following.
[1] A device that is used by being attached to a liquid injection container, and includes a lid that can be detachably attached to the liquid injection container, a liquid injection tube that can be fitted into a first through hole provided in the lid, and a lid. And an intake tube that can be fitted into a second through hole provided in the body.
[2] The configuration according to [1], wherein the lower end of the intake tube that protrudes from the lid into the liquid injection container in a state where the device is attached to the liquid injection container is disposed close to the lid. Devices.
[3] The device according to [1] or [2], wherein a check valve is provided in the intake tube.
[4] The device according to any one of [1] to [3], wherein the first through hole is provided in the vicinity of the peripheral edge of the lid.

[5] A method for injecting liquid, wherein (a) a step of preparing an injection container containing the liquid, (b) a lid body detachably attached to the injection container, and the lid body Attaching a device including a liquid injection tube fitted in a first through hole provided in the liquid crystal and an intake tube fitted in a second through hole provided in the lid to the liquid injection container; (c) injection The method, comprising: rotating a liquid container to inject a liquid through an injection tube; and (d) ending the injection by rotating the liquid injection container in a reverse direction.
[6] The method according to [5], further including step (e) in which step (c) and step (d) are repeated to continuously perform a plurality of injection operations.
[7] The method according to [5] or [6], further including a step (f) of determining an injection amount from an injection time after step (c) and before step (d).

[8] A device that is used by being attached to a liquid injection container, and includes a lid that can be detachably attached to the liquid injection container, a liquid injection tube that can be fitted into a first through hole provided in the lid, and a lid. An intake tube that can be fitted into a second through-hole provided in the body, and the lower end of the intake tube that protrudes from the lid into the injection container when the device is mounted on the injection container The device configured to be disposed proximate to a bottom surface.
[9] The device according to [8], wherein the intake tube is provided with a check valve.
[10] The device according to [8] or [9], wherein the first through hole is provided in the vicinity of the peripheral edge of the lid.
[11] The liquid injection container is a container for containing and injecting a culture medium, and the liquid injection is performed by rotating the liquid injection container. device.

[12] A method for injecting a culture medium, wherein (a) a step of preparing an injection container containing the culture medium, (b) a lid body detachably attached to the injection container, and a lid body Attaching a device including a liquid injection tube fitted in a first through hole provided in the liquid crystal and an intake tube fitted in a second through hole provided in the lid to the liquid injection container; (c) injection The method, comprising: rotating a liquid container to inject a medium through an injection tube; and (d) ending the injection by rotating the liquid injection container in the reverse direction.
[13] The method according to [12], further including step (e) in which step (c) and step (d) are repeated to continuously perform a plurality of injection operations.
[14] The method according to [12] or [13], further including the step of (f) determining an injection amount after the step (c) and before the step (d).

[15] A robot system for injecting a liquid by rotating an injection container containing a liquid around an axis perpendicular to the major axis of the injection container. Liquid injection start control to rotate the liquid, liquid injection control to stop the rotation for a predetermined time, liquid injection, and liquid injection end control to reversely rotate the liquid injection container around a predetermined axis, The container can be fitted into a lid that can be detachably attached to the liquid filling container, a liquid filling tube that can be fitted into a first through hole provided in the lid, and a second through hole provided in the lid. The robot system is equipped with a device including a suction tube, and a predetermined axis is set at a distal end portion of the liquid injection tube.
[16] The robot system according to [15], wherein a coordinate system (TCP) for position / attitude control of the robot system is set on a predetermined axis.
[17] The robot system according to [15] or [16], wherein the injection container is rotated through a lid gripped by an end effector of the robot system.
[18] The device is configured such that the lower end of the intake tube that protrudes from the lid into the liquid injection container in the state where the device is mounted on the liquid injection container is disposed close to the lid. 17]. The robot system according to any one of [17].
[19] The injection is performed on the cell culture flask, the predetermined axis and the culture surface of the cell culture flask are parallel, and the cell culture flask is disposed with the culture surface facing upward. The robot system according to any one of [15] to [18].

[20] A method of injecting a liquid by rotating an injection container containing a liquid around an axis perpendicular to the long axis of the injection container, the injection container being rotated about a predetermined axis The liquid injection container includes a liquid injection start step, a liquid injection step in which the rotation is stopped for a predetermined time and liquid is injected, and a liquid injection end step in which the liquid injection container is reversely rotated around a predetermined axis. A lid that can be detachably attached to the liquid container, a liquid injection tube that can be fitted into a first through-hole provided in the lid, and an intake tube that can be fitted into a second through-hole provided in the lid. The method is characterized in that the device is mounted and the predetermined axis is set at the tip of the injection tube.
[21] A program for controlling a robot for injecting a liquid by rotating a liquid injection container containing a liquid around an axis perpendicular to the long axis of the liquid injection container, the program around a predetermined axis Liquid injection start control for rotating the liquid injection container, liquid injection control for stopping the rotation for a predetermined time and liquid injection, and liquid injection end control for reversely rotating the liquid injection container around a predetermined axis, The liquid injection container, which is executed by the computer, includes a lid body that can be detachably attached to the liquid injection container, a liquid injection tube that can be fitted into a first through hole provided in the lid body, and a first liquid pipe provided in the lid body. 2. The program according to claim 1, wherein a device including an intake tube that can be fitted into the two through-holes is mounted, and a predetermined axis is set at a distal end portion of the liquid injection tube.

[22] A robot system for injecting a liquid by rotating an injection container having a constant amount of liquid containing a liquid around an axis perpendicular to the long axis of the injection container. Injection start control for rotating the injection container around the axis, injection control for stopping the rotation for a predetermined time and injecting the liquid, and injection end control for rotating the injection container in the reverse direction around the predetermined axis , And the predetermined time is calculated based on the injection flow rate Q [ml / s] measured in real time.
[23] The system according to [22], wherein the predetermined time is further calculated based on a time ΔT until the liquid is not injected in reverse rotation.
[24] The predetermined time is calculated based on the final liquid injection ratio X% when the liquid injection end step is performed with respect to the liquid injection amount when the liquid injection end step is not performed. 22] or the system according to [23].
[25] A lid body in which the liquid injection container can be detachably attached to the liquid injection container, a liquid injection tube that can be fitted into a first through hole provided in the lid body, and a second through hole provided in the lid body A device including an intake tube that can be fitted into the hole is attached, and a predetermined axis is set at a distal end portion of the liquid injection tube, according to any one of [22] to [24] System.
[26] The configuration according to [25], wherein the lower end of the intake tube protruding from the lid into the liquid injection container in a state where the device is mounted on the liquid injection container is arranged close to the lid. System.

[27] A method for injecting a liquid by rotating an injection container containing a liquid in a constant amount around an axis perpendicular to the major axis of the injection container, the rotation being about a predetermined axis A liquid injection start step for rotating the liquid injection container, a liquid injection step for stopping the rotation for a predetermined time to inject the liquid, and a liquid injection end step for rotating the liquid injection container in the reverse direction around a predetermined axis. The method is characterized in that the predetermined time is calculated based on the injection flow rate Q [ml / s] measured in real time.
[28] A program for controlling a robot for injecting liquid by rotating a liquid injection container having a constant liquid injection volume around an axis perpendicular to the long axis of the liquid injection container. Injecting start control for rotating the injection container around a predetermined axis, injection control for stopping the rotation for a predetermined time and injecting liquid, and reverse rotation of the injection container around a predetermined axis The program is executed by causing the computer to execute injection completion control, and the predetermined time is calculated based on an injection flow rate Q [ml / s] measured in real time.

  According to the present invention, by using a means for limiting the injection, it is possible to perform an efficient and highly accurate injection operation, and it is possible to prevent the occurrence of dripping, so that cell culture in a clean room or the like is possible. Suitable for manufacturing goods.

FIG. 1 shows a schematic view of a device 1 according to a first embodiment of the present invention. FIG. 2 is a schematic diagram for explaining a liquid injection operation using the device 1 of FIG. FIG. 3 is a conceptual diagram illustrating the arrangement of the storage container 10. FIG. 4 is a conceptual diagram of a robot system according to the second embodiment of the present invention. FIG. 5 is a schematic diagram for explaining the operation of the robot system of FIG. FIG. 6 is a conceptual diagram of the robot system according to the third embodiment. FIG. 7 shows a flow diagram of the medium exchange process. FIG. 8 shows a graph of the injection amount and time during the injection operation. FIG. 9 shows a graph of the injection speed and time during the injection operation. FIG. 10 shows a method for predicting the injection amount after the start of the injection end operation. FIG. 11 shows a flow diagram of parameter derivation and validity testing. FIG. 12 shows a prediction flow chart of the injection end operation start time. FIG. 13 shows a graph of the amount of injection and time during the injection operation. FIG. 14 shows the validity test result of the liquid injection work.

  The components constituting the liquid in the present invention include, for example, water, physiological saline, physiological buffer (eg, HBSS, PBS, EBSS, Hepes, sodium bicarbonate, etc.), medium (eg, DMEM, MEM, F12, DMEM / F12, DME, RPMI 1640, MCDB, L15, SkBM, RITC80-7, IMDM, etc., sugar solution (sucrose solution, Ficoll-paque (registered trademark) PLUS, etc.), seawater, serum-containing solution, Renographin (registered trademark) solution , Metrizamide solution, meglumine solution, glycerin, ethylene glycol, ammonia, benzene, toluene, acetone, ethyl alcohol, benzol, oil, mineral oil, animal fat, vegetable oil, olive oil, colloidal solution, liquid paraffin, turpentine oil, linseed oil, castor Oil etc. It is below.

The container in the present invention is not particularly limited, and examples thereof include a cell culture container, a cell culture flask for adherent cells, and a cell culture flask for floating cells. A cell culture flask has a substantially rectangular main body part, and at least one flat surface of the main body part is subjected to a surface treatment necessary for cell culture, and a plurality of cell culture flasks are stacked with the cell culture surface facing down. Thus, it refers to a container capable of multistage culture.
The liquid injection container in the present invention is not particularly limited as long as it can accommodate the medium to be injected into the storage container, for example, shaker flask, Erlenmeyer flask, roller bottle, liquid injection bottle, beaker, medium bottle, corner Examples include mold medium bottles, sterile bottles, and sterile bottles.

  The robot in the present invention is not particularly limited, and examples thereof include a linear motion / rotation device, a manipulator, and an articulated robot. Examples of the multi-joint robot include a 2-axis multi-joint robot, a 3-axis multi-joint robot, a 4-axis multi-joint robot, a 5-axis multi-joint robot, a 6-axis multi-joint robot, and a 7-axis multi-joint robot.

In the present invention, the “predetermined axis” refers to an axis that serves as a rotation center when the injection container is rotated. When the injection container is a general vertically long container, the predetermined axis is the long axis of the container. Set as vertical axis.
In the present invention, “TCP” refers to a tool center point, which is a coordinate system for expressing the position and orientation of a control object such as a tool, gripper, and work object located at the tip of a robot. Say. TCP, for example, can be set to any position and orientation (position and orientation convenient for operation and control) such as end effectors (grippers, tools, etc.) and work objects (flasks, bottles, etc.). In the case of a robot, the definition is usually made with respect to the coordinate system of the sixth axis of the robot.

  In the present invention, “rotate around a predetermined axis” means to rotate an object around a predetermined axis. For example, when a predetermined axis is set at one end of the opening of the liquid injection container, the liquid in the liquid injection container can be discharged only by a rotation operation around the one end. Further, for example, even when a predetermined axis is set at the center (center of gravity) of the liquid injection container, the rotational operation around the central axis of the liquid injection container and the translation operation of the circular arc trajectory are combined as described above. In addition, the liquid injection container can be rotated around one end of the opening of the liquid injection container.

  For example, when the robot is an articulated robot, the robot control can be made more efficient by associating a predetermined axis with TCP. If the robot is, for example, a 6-axis articulated robot, the rotation axis of the sixth axis and the rotation axis of the TCP are made parallel with the rotation axis of the TCP, and a little of the first to fifth axes. In operation, the liquid injection container can be rotated as described above. Further, when the rotation axis of the sixth axis and the rotation axis of the TCP coincide with each other, only the rotation operation of the sixth axis as described above. The injection container can be rotated.

  Examples of cells in the present invention include, but are not limited to, adherent cells (adherent cells). Adherent cells include, for example, adherent somatic cells (eg, cardiomyocytes, fibroblasts, epithelial cells, endothelial cells, hepatocytes, pancreatic cells, kidney cells, adrenal cells, periodontal cells, gingival cells, periosteum cells, skin Cells, synovial cells, chondrocytes, etc.) and stem cells (eg tissue stem cells such as myoblasts, cardiac stem cells, embryonic stem cells, pluripotent stem cells such as induced pluripotent stem (iPS) cells, mesenchymal stem cells, etc.) Etc. Somatic cells may be differentiated from stem cells, particularly iPS cells. Non-limiting examples of cells that can form a sheet-like cell culture include, for example, myoblasts (eg, skeletal myoblasts), mesenchymal stem cells (eg, bone marrow, adipose tissue, peripheral blood, skin, hair roots) , Muscle tissue, endometrium, placenta, umbilical cord blood, etc.), cardiomyocytes, fibroblasts, cardiac stem cells, embryonic stem cells, iPS cells, synovial cells, chondrocytes, epithelial cells (eg, oral mucosal epithelium) Cells, retinal pigment epithelial cells, nasal mucosal epithelial cells, etc.), endothelial cells (eg, vascular endothelial cells), hepatocytes (eg, liver parenchymal cells), pancreatic cells (eg, islet cells), kidney cells, adrenal glands Examples include cells, periodontal ligament cells, gingival cells, periosteal cells, skin cells and the like. In the present invention, those forming a monolayer cell culture, such as myoblasts or cardiomyocytes, are preferred, and skeletal myoblasts or iPS cell-derived cardiomyocytes are particularly preferred.

Hereinafter, preferred embodiments of the present invention will be described in detail with reference to the drawings.
[First Embodiment]
One aspect of the present invention is a device that is used by being attached to a liquid injection container, and includes a lid that can be detachably attached to the liquid injection container, and a liquid injection that can be fitted into a first through hole provided in the lid. It is related with the said device containing a tube and the intake tube which can be inserted in the 2nd through-hole provided in the cover body.

First, the first embodiment of the present invention will be described.
FIG. 1 is a schematic view of a device 1 according to the first embodiment of the present invention, FIG. 2 is a schematic view for explaining an injection operation using the device 1 of FIG. 1, and FIG. A conceptual diagram is shown. In this embodiment, the liquid is a culture medium, the injection container 30 is an injection bottle that stores the culture medium, the storage container 10 is a cell culture flask, and the injection operation is performed in a clean room. Will be described.
In each drawing in the present application, for ease of explanation, the size of each member is emphasized as appropriate, and the illustrated members do not indicate actual sizes.

  As shown in FIG. 1, the device 1 according to the first embodiment of the present invention includes a lid 5 that can be detachably attached to a mouth 33 of an injection container 30 having an opening 32 containing a culture medium, and a lid. The liquid injection tube 6 that can be fitted into the first through hole 51 provided in the top plate portion 50 of the body 5 and the intake tube 7 that can be fitted into the second through hole 52 provided in the top plate portion 50 of the lid 5. Including. The top plate portion 50 has a disk shape that can cover the opening 32 of the liquid injection container 30 and has a cylindrical skirt wall 53 that hangs down from the peripheral edge portion. An inner screw (not shown) is provided on the inner peripheral surface of the cylindrical skirt wall 53, and is screwed into an outer screw (not shown) provided on the outer peripheral surface of the mouth portion 33 of the liquid injection container 30. be able to.

  The first through-hole 51 is arranged near the peripheral edge of the top plate portion 50, and the liquid injection tube 6 fitted into the first through-hole 51 in the state where the lid body 5 is attached to the liquid injection container 30 is poured. It is arrange | positioned near the inner peripheral surface of the opening | mouth part 33 of the liquid container 30, and when the liquid injection container 30 is inclined and a culture medium is discharged | emitted, it is comprised so that a liquid may be discharged | emitted without remaining in the liquid injection container 30. . The length of the injection tube 6 is not particularly limited, but is preferably 0 to 100 mm, more preferably 10 to 60 mm, and the inner diameter is preferably 1 to 10 mm, more preferably 3 to 5 mm. The length of the liquid injection tube 6 that protrudes from the top plate portion 50 when fitted into the first through hole 51 is not particularly limited, but can preferably be set to 0 to 100 mm, more preferably 10 to 40 mm. .

  The length of the intake tube 7 is not particularly limited, but is preferably 0 to 100 mm, more preferably 10 to 60 mm, and the inner diameter is preferably 0.5 to 10 mm, more preferably 2 to 4 mm. Moreover, it is preferable to make it a small diameter compared with an injection tube. The length of the intake tube 7 that protrudes from the top plate portion 50 when fitted into the second through hole 52 is such that the lower end 71 of the intake tube 7 is disposed close to the top plate portion 50 of the lid 5. If it is, it will not specifically limit, Preferably it is 0-100 mm, More preferably, it can be set as 0-40 mm. The intake tube 7 is provided with a check valve 71 so that air from the outside of the liquid injection container 30 is allowed to pass therethrough but a medium from the inside of the liquid injection container 30 is not allowed to pass therethrough. The check valve may be disposed inside the liquid injection container 30. The liquid injection tube 6 and the intake tube 7 may be formed integrally with the lid 5.

  As shown in FIG. 2, when using the device 1, the device 1 is attached to the injection container 30 containing the culture medium and tilted, and the tip 61 of the injection tube 6 is injected into the opening 12 of the storage container 10. Position on the liquid container 30 side. At this time, the liquid injection tube 6 (first through hole) is rotated and positioned so as to be lower than the intake tube 7 (second through hole). It is preferable that the liquid move to the liquid injection tube 6 side instead of the intake tube 7 side. Then, the liquid injection container 30 is rotated in the direction of the arrow with the tip 61 of the liquid injection tube 6 as the rotation axis, and the medium in the liquid injection container 30 is moved to the device 1 side (liquid injection start operation). When the rotation is stopped for a predetermined time so that the opening 32 of the liquid injection container 30 comes to the lower side, the culture medium is discharged from the liquid injection tube 6 and injected into the storage container 10 (liquid injection operation). When the amount of liquid injection in the storage container 10 reaches a predetermined amount, the liquid injection container 30 is rotated in the direction opposite to the arrow direction to stop the liquid injection, and the medium is moved to the opposite side of the device 1 to complete the liquid injection. (Injection end operation).

  In the liquid injection operation, the liquid injection amount from which the culture medium is discharged (liquid injection) from the liquid injection tube 6 and the air intake amount of the air flowing from the air intake tube 7 are the same. Since the liquid injection amount and the air intake amount are limited by the dimensions of the liquid injection tube 6 and the air intake tube 7, the liquid injection amount is limited as compared with the case where the tube is not used (the liquid injection is limited). means). Further, when the air flowing into the liquid injection container 30 becomes continuous bubbles, and the amount of intake per unit time becomes constant, the amount of liquid injected per unit time also becomes constant. In this case, since the injection time and the injection volume in the injection operation are proportional, the relationship between the injection volume and the time required for injection is measured in advance, and the injection is started from the injection time based on the measurement result. By determining the amount of liquid, a highly accurate liquid injection operation can be performed.

  Conventionally, for example, when injecting 75 ml of medium into a culture flask, a pipette is inserted into the liquid injection bottle, the medium is aspirated, the amount of 75 ml is visually confirmed, the pipette is moved and poured into the culture flask. It was necessary to repeat the troublesome work of liquefying. On the other hand, by using the device 1 of the present invention, for example, the device 1 is attached to the injection container 30 containing about 480 ml of the medium, and the injection operation is performed for the time required for the 75 ml injection measured in advance. This can be carried out continuously (six times) while exchanging the six containers 10. As a result, the conventional troublesome work can be replaced with a simple work of repeating the rotation of the liquid injection container 30, pipetting operation, movement, visual confirmation of the amount, etc. are not required, and the time required for the liquid injection work Can be greatly shortened.

  Conventionally, in the case of using a pipette, since the medium is sucked and discharged through one flow path, dripping has occurred due to an erroneous operation or the like. On the other hand, the use of the device of the present invention eliminates the need for the medium suction operation, and discharge of the medium and air suction are performed in separate flow paths. it can. Furthermore, since the medium is injected through the tube by using the device of the present invention, the injection direction and the injection range of the medium are limited by the protruding direction and the diameter of the tube (the injection is limited). Means). Thereby, the accuracy of aiming of the liquid injection is improved, for example, it is possible to perform the liquid injection work by accurately aiming the storage container having a small opening, such as the opening of the cell culture flask, The occurrence of dripping can be minimized.

  And arrangement | positioning of the storage container 10 which inject | pours a culture medium can take various forms. For example, when the container 10 is a cell culture flask, it is preferable to arrange the main surfaces 14 and 15 so as to be parallel to the rotation axis as shown in FIG. 3A. Further, the storage container 10 may be installed on the base S or the like, and may be arranged to be inclined with the main surface 14 (culture surface) of the storage container 10 facing upward. Thereby, since the culture medium discharged | emitted from the injection tube 6 flows along an inclined surface, foaming does not occur easily. Similarly, as shown in FIG. 3B, the storage container 10 is placed in an upright posture so that the main surface 14 (culture surface) faces the proximal side of the injection container 30 and the main surface 15 side faces the distal side. You may arrange in. Thereby, not only does the culture medium flow along a small step between the one side of the mouth portion 13 and the main surface 15 and foaming hardly occurs, but the culture medium does not directly contact the main surface 14 (culture surface). .

  As described above, when the lower end 71 of the intake tube 7 is disposed close to the top plate portion 50 of the lid body 5, continuous bubbles are generated in the culture medium, but the length of protrusion of the intake tube 7 is increased. By doing so, it is also possible to configure so that bubbles do not occur in the liquid injection container 30 and it is difficult for foaming of the medium to occur. That is, when the injection container 30 is tilted and the culture medium is moved to the device 1 side, an air layer is formed on the bottom surface side of the injection container 30 and the lower end 71 of the intake tube 7 is positioned in the air layer. Configure. In this case, the length protruding from the top plate portion 50 of the intake tube 7 is not particularly limited as long as the lower end 71 of the intake tube 7 is disposed close to the bottom surface of the liquid injection container 30. The length from the bottom surface of the liquid container 30 is preferably 0 to 100 mm, more preferably 0 to 50 mm. And, as described above, the relationship between the injection volume and the time required for injection is measured in advance, and the injection volume is determined from the injection time based on the measurement result, so that the highly accurate injection work Can also be done.

  As described above, according to the liquid injection device 1 according to the first embodiment of the present invention, by using the means for limiting liquid injection, an efficient and quick liquid injection operation is possible, and the occurrence of liquid dripping occurs. Therefore, it is suitable for manufacturing a cell culture in a clean room or the like.

[Second Embodiment]
Another aspect of the present invention is a robot system for injecting a liquid by rotating an injection container containing a liquid around an axis perpendicular to the long axis of the injection container, the robot system having a predetermined axis Liquid injection start control for rotating the liquid injection container, liquid injection control for stopping the rotation for a predetermined time and liquid injection, and liquid injection end control for reversely rotating the liquid injection container around a predetermined axis, Executed, the liquid injection container has a lid that can be detachably attached to the liquid injection container, a liquid injection tube that can be fitted into a first through hole provided in the lid, and a second that is provided in the lid. The robot system is characterized in that a device including an intake tube that can be fitted into a through hole is mounted, and a predetermined axis is set at a distal end portion of the liquid injection tube.

Next, a second embodiment of the present invention will be described.
FIG. 4 is a conceptual diagram of the robot system according to the second embodiment of the present invention, and FIG. 5 is a schematic diagram for explaining the operation of the robot system of FIG. In each drawing in the present application, for ease of explanation, the size of each member is emphasized as appropriate, and the illustrated members do not indicate actual sizes.
In the figure, the same components as those of the device 1 according to the first embodiment are denoted by the same reference numerals, and differences from the first embodiment are described in detail below. The description is omitted.

  As shown in FIG. 4, the robot 20 is a 6-axis vertical articulated robot disposed on a base. The robot 20 includes a base 21 that can turn with respect to a base, a first arm 22 that is connected to the base 21 and can be tilted with respect to a vertical axis in the turning direction of the base 21, and a tip side of the first arm 22. The second arm base end 23 that is connected and tiltable with respect to the first arm, and is connected to the front end side of the second arm base end 23 and is rotatable with respect to the axial direction of the second arm base end 23. A second arm tip 24, a hand portion 25 connected to the tip side of the second arm tip 24 and tiltable with respect to the axial direction of the second arm tip 24, and a gripper connected to the hand portion 25. 26 (end effector). The hand unit 25 is configured to be rotatable along the axial direction thereof.

  Further, the robot 20 can communicate with a control device 40 including a storage unit 41 that stores a program describing a control procedure of the robot 20 and a processing unit 42 that processes the program and controls the robot 20. The robot 20 can automatically operate according to a control signal from the control device 40. With such a configuration, the robot 20 can automatically determine the position and orientation of the gripper 26 and rotate, open and close the gripper 26. The gripper 26 can automatically transfer, tilt, and rotate the liquid injection container 30. It can be carried out.

  Hereinafter, the liquid injection system according to this embodiment using the above-described robot system will be described. In this embodiment, the robot system uses the liquid injection container 30 as shown in the first embodiment to inject liquid into the storage container 10 via the gripper 26 (end effector) of the robot 20. The liquid injection container 30 is rotated to discharge the liquid and used to inject the liquid into the storage container 10. In this embodiment, the liquid is a culture medium, the injection container 30 is an injection bottle that stores the culture medium, the storage container 10 is a cell culture flask, and the injection operation is performed in a clean room. Will be described. Further, the cell culture flask has two main surfaces 14 and 15 as side surfaces, and one main surface 14 is subjected to surface processing so that cells can be cultured with the main surface 14 as the lower surface. It will be explained as being.

  Referring to the schematic diagram of FIG. 5, first, when using the robot system, the control device 40 is activated to cause the processing unit 42 to read the program stored in the storage unit 41. The processing unit 42 controls the robot 20 based on the program, and grips the lid 5 of the liquid injection container 30 to which the device 1 is attached by the gripper 26 (not shown). At this time, the first through hole 51 side of the device 1 is set to face the container 10. Next, the robot 20 is controlled to perform rotation and translation, and the distal end portion 61 of the liquid injection tube 6 fitted in the first through hole 51 is moved to the liquid injection container 30 side of the opening 12 of the storage container 10. Positioning is performed, and an axis perpendicular to the long axis of the liquid injection container 30 that intersects the distal end portion 61 is set as the rotation axis A (positioning control).

  At this time, a camera (not shown) for monitoring the positions and angles of the storage container 10 and the liquid injection container 30 and the robot 20 may be linked to perform accurate and automatic alignment. Therefore, the robot system may include a monitoring camera that can communicate with the control device 40, and the program may be a program that controls the robot 20 based on the position and angle monitored by the camera. The rotation axis A is set so as to be perpendicular to the long axis of the liquid injection container 30, the tip 61 is disposed above the opening 12 of the container 10, and the distance from the opening 12 is 0 to 3 cm. It is preferable to set in the range.

  Next, the robot 20 is controlled to rotate the liquid injection container 30 attached with the lid 5 around the rotation axis A in the direction of the arrow (liquid injection start control). As shown in FIG. 5, in this embodiment, the start angle of the liquid injection container 30 in the liquid injection start control is 30 deg from the horizontal with the liquid injection tube 6 upward, and the liquid injection tube 6 is downward with the stop angle. However, the present invention is not limited to this. For example, the stop angle may be set in a range of 5 to 85 deg. Accordingly, the medium discharged from the tip 61 of the liquid injection tube 6 is injected obliquely with respect to the inner wall of the mouth portion 13 of the container 10 and the inner wall of the container main body 11 so that the foam of the medium is increased. You may comprise so that it may become difficult to occur.

  Next, when the robot 20 is controlled to stop the rotation for a predetermined time so that the mouth 33 of the liquid injection container 30 is on the lower side, the medium in the liquid injection container 30 moves to the lid 5 side. The liquid is discharged from the liquid injection tube 6 and injected into the container 10 (liquid injection control). Next, when a predetermined time elapses, the liquid injection container 30 is rotated in the direction opposite to the arrow direction to stop at the start angle, and the medium is moved in the direction opposite to the mouth 33 side to end the liquid injection. (Injection end control). Then, the liquid injection operation to the plurality of storage containers 10 can be efficiently and quickly performed by causing the robot 20 to repeatedly execute the injection start control, the injection control, and the injection end control while exchanging the storage containers 10. it can. The predetermined time may be determined by measuring the relationship between the liquid injection amount and the liquid injection time in advance, and setting the time (predetermined time) for stopping the rotation based on the measured value and the target liquid injection amount. Good.

  On the other hand, when the remaining amount of the medium in the liquid injection container 30 decreases, only the liquid injection container 30 is removed from the lid body 5 held by the gripper 26, and a new liquid injection container 30 is attached to the lid body 5. The robot 20 may repeatedly execute the liquid injection start control, the liquid injection control, and the liquid injection end control. In this way, by repeating the replacement of the container 10 and the replacement of the liquid injection container 30, the setting (positioning control) of the rotation axis A of the liquid injection container 30 is omitted, and the interruption time of the liquid injection operation is minimized. Can do. The robot system of the present invention only needs to be able to execute at least the rotation operation related to the liquid injection start control, liquid injection control, and liquid injection end control. For example, the robot system is rotated using a simple device such as a linear motion / rotation device as a robot. Alternatively, the stop and reverse rotation steps may be repeated. The rotary shaft may be set to the lid 5 or the liquid injection container 30 so that the distal end portion 61 of the liquid injection tube 6 and the opening 12 are close to each other at least during liquid injection control.

  As described above, the storage container 10 is preferably disposed so that the main surfaces 14 and 15 of the storage container 10 (cell culture flask) are parallel to the rotation axis A as described above. You may comprise as follows. That is, for example, the placement of the container 10 is confirmed with a camera that can communicate with the robot system, the robot 20 is controlled, and the positioning control of the rotation axis A is performed in accordance with the orientation of the main surfaces 14 and 15 of the container 10. As described above, the program and the processing unit 42 may be configured. In addition, it may be configured to measure the amount of liquid injection in the container 10 in real time with such a camera and shift to liquid injection end control when the liquid injection amount reaches a predetermined amount, or can communicate with the robot system. The liquid injection amount in the storage container 10 may be measured in real time by the weight using an electronic balance, and the liquid injection end control may be shifted to when the liquid injection amount reaches a predetermined amount.

  The present invention can also be realized by executing the following processing. That is, a process in which software (program) that realizes the functions of the above embodiments is read by a robot via a network or various storage media, and a processing unit (CPU, MPU, etc.) built in the robot executes the program. It is. The present invention can also be realized by transmitting a control signal to the robot and operating it from a control device including the storage unit that stores the program and the processing unit that processes the program.

  As described above, according to the robot system according to the second embodiment of the present invention, it is possible to efficiently and quickly perform the liquid injection work by using the means for limiting the liquid injection and to prevent the occurrence of liquid dripping. Therefore, it is suitable for manufacturing a cell culture in a clean room or the like.

[Third embodiment]
Another aspect of the present invention is a robot system for injecting liquid by rotating an injection container having a constant amount of liquid containing the liquid around an axis perpendicular to the long axis of the injection container. The injection start control for rotating the injection container around a predetermined axis, the injection control for stopping the rotation for a predetermined time and injecting the liquid, and the injection container reversed around the predetermined axis The liquid injection end control to be rotated is executed, and the predetermined time is calculated based on the liquid injection flow rate Q [ml / s] measured in real time.

Next, a third embodiment of the present invention will be described. FIG. 6 is a conceptual diagram of the robot system according to the third embodiment.
As shown in FIG. 6, the robot system in the present embodiment can communicate with the electronic balance 80, and the electronic balance 80 measures the weight of the container 10 in real time to determine the injection amount and the injection flow rate. It can be calculated and fed back to the control device 40.

  In this embodiment, the robot system sets a predetermined time based on the injection flow rate Q [ml / s] measured in real time by the electronic balance 80 when the rotation is stopped for a predetermined time after shifting to the injection control. can do. The predetermined time is set, for example, by calculating the time required to reach the target injection amount from the inclination of the injection flow rate Q [ml / s]. When the predetermined time is reached, the liquid injection container is rotated in the reverse direction around the predetermined axis, and the flow shifts to liquid injection end control. In this way, by configuring so as to set the predetermined time based on the liquid injection flow rate Q [ml / s], the accuracy of the liquid injection amount injected into the storage container 10 is increased.

  On the other hand, due to the start angle, stop angle, rotation speed (angular speed) in the liquid injection start control, the reverse rotation speed in the liquid injection end control, the remaining amount of liquid in the liquid injection container 30, and the like during the liquid injection end control ( The liquid in the liquid injection tube 6 may be discharged (liquid injection) during reverse rotation). Therefore, the robot system according to the present embodiment is configured such that the predetermined time in the liquid injection control is set in consideration of the liquid injection amount that may be injected during the liquid injection end control. You can also

  For example, a series of liquid injection start control, liquid injection control, and liquid injection end control are performed in advance until all liquid in the liquid injection tube 6 is discharged (liquid injection) in the liquid injection end control (reverse rotation). The time ΔT is measured. The time ΔT is stored in the storage unit 41, and in actual liquid injection control, the time ΔT is subtracted from the predetermined time calculated based on the liquid injection flow rate Q [ml / s] as described above. The predetermined time can also be set as the predetermined time. Thereby, the transition from the liquid injection control to the liquid injection end control can be performed earlier by the time ΔT, and even when the liquid in the liquid injection tube 6 is discharged during the liquid injection end control, the container The accuracy of the amount of liquid injected into the liquid 10 can be increased.

  Further, when the reverse rotation speed (angular velocity) in the liquid injection end control is low, the liquid injection flow rate is gradually reduced, that is, the liquid injection flow rate at time ΔT during the liquid injection end control is the liquid injection flow rate Q [ml / s. Draw a gentle curve instead of a proportional relationship like Therefore, at time ΔT, there may be a difference between the injection amount when the injection completion control is not performed and the injection amount when the injection completion control is performed. The robot system in the present embodiment can be configured such that a predetermined time in the liquid injection control is set in consideration of such a difference.

  For example, the ratio of the liquid injection amount (final liquid injection ratio X%) when the liquid injection end control is performed with respect to the liquid injection amount when the liquid injection end control is not performed is measured in advance and stored in the storage unit 41. In the actual liquid injection control, the liquid injection volume Vx [ml] after the time ΔT is calculated from the liquid injection flow rate Q [ml / s], the time ΔT, and the final liquid injection ratio X%. And even if it is a case where a difference arises in the injection volume by setting the time to reach the injection volume Ve [ml] obtained by subtracting the injection volume Vx [ml] from the target injection volume as a predetermined time The accuracy of the amount of liquid injected into the storage container 10 can be increased.

  Hereinafter, the robot system according to the third embodiment will be described in more detail with reference to examples. However, this shows a specific example of the present invention, and the present invention is not limited thereto. Absent.

Example 1: Manual pouring operation FIG. 7 shows a flow chart of a general medium exchange process. The medium exchange process is a series of operations to drain the culture medium in the flask and inject a new culture medium into the flask, and it must be performed quickly without dripping. Relying on work. The medium exchange process is composed of a waste liquid process and a liquid injection process, and the liquid injection process is composed of operations such as removing a cap, injecting a new culture solution with a pipetter, and attaching a cap. The liquid injection process includes not only aspirating and injecting the culture solution with a pipette, but also removing and attaching the cap of the flask, wiping away the dipped culture solution, and replacing the pipette.

  As the container 10, a cell culture flask for adherent cells (manufactured by Thermo Fisher, T500 flask) was used. The outer diameter of the discharge port (opening 12) of the T500 flask was 28.2 [mm], the inner diameter was 25.8 [mm], and the thickness was 1.2 [mm]. In an actual medium replacement operation, a cell processing worker performed an operation of injecting the culture solution in the injection bottle into the flask using an electric pipette. 75 ml of the culture solution was poured into one set of 8 flasks. A work analysis of two sets. A work time of 550 s to 560 s was required per set, and a result that the liquid injection work time per flask was about 70 s was obtained.

Example 2: As a liquid injection check valve 71 using a device, 105-15001 manufactured by Kijima Corporation, as a robot, a 6-axis vertical articulated industrial robot: MOTOMAN-MH3F manufactured by Yaskawa Electric (first arm: 260 mm, The second arm: 270 mm) was used as a controller, and a system based on the RTLinux (registered trademark) environment was used as an electronic balance. The device is mounted on the injection bottle, the lid of the device is fixed to the gripper of the robot, the tip of the injection tube is TCP (Tool Center Point), and the injection into the T500 flask is performed only by changing the posture around the TCP. Worked. Further, by aligning the rotation axis of the robot's sixth axis with the TCP, the liquid injection operation can be performed only by the operation of the sixth axis.

  As shown in FIG. 5, the flask is rotated from an initial posture tilted upward by 30 deg from the horizontal to a posture tilted downward by 45 deg from the horizontal, stopped, and cultured in an injection bottle into a flask placed on an electronic balance. The liquid was poured. FIG. 8 and FIG. 9 show the injection volume, time, and injection velocity when all 480 ml of culture solution is injected. Since the injection speed is almost constant in the section excluding the start and end of injection (injection time 3 to 53 s, injection volume 10 to 412 ml), it should be a method suitable for controlling the injection volume. Was confirmed.

Example 3: Infusion work by robot The robot was controlled to inject 480 ml of the culture solution into 6 portions and inject 75 ml each. In consideration of errors, the volume of the culture solution was more than 75 ml × 6 servings. The target accuracy for one injection was ± 2%. The liquid injection operation by the robot is as follows: (1) The liquid is started from the initial position where the flask is tilted upward by 30 deg from the horizontal to the position where the flask is tilted downward by 45 deg from the horizontal, and liquid injection is started (control) 2) Liquid injection operation (control) in which the flask is poured in a state where the flask is tilted 45 degrees downward from the horizontal position, and (3) The liquid injection end operation is the initial position from the position where the flask is inclined 45 degrees downward from the horizontal position. Returning to step 3, the injection was completed, and the injection was completed in three stages.

  In order to realize smooth operation without dripping, the liquid injection start operation was 32 deg / s, and the liquid injection end operation was 108 deg / s (angular velocity: axis velocity). In order to reduce the amount of one injection to 75 ml, it is necessary to predict the injection amount after the start of the injection end operation and shift from the injection operation to the injection end operation. FIG. 10 shows a method for predicting the injection amount after the start of the injection end operation. The time (ΔT) from the start of the injection end operation until the injection amount stops increasing (ΔT), and the final injection ratio when the injection end operation is performed with respect to the injection amount when the injection end operation is not performed (X %) Was set as a parameter, and the amount of liquid injected when the liquid injection end operation was not performed was calculated from the slope of the measured value obtained during the liquid injection operation, and the amount of liquid injected after the start of the liquid injection end operation was calculated therefrom . Note that the angular velocity pattern around the predetermined axis (the angular velocity pattern of the sixth axis) is a square in the figure, but may be an acceleration pattern such as a trapezoid or an S-shape, and is not limited.

In the procedure of FIG. 11, parameter derivation and operation confirmation using parameters were performed. Initially, the parameters were determined by analyzing the data when the injection operation was repeated. Three sets of experiments were conducted in which the liquid injection end operation start liquid injection volume was assumed to be 70 ml, and the 480 ml culture liquid was injected in five portions. FIG. 12 shows a prediction flowchart of the liquid injection end operation start time, FIG. 13 shows the liquid injection amount at that time, and Table 1 shows the analysis result.

  As shown in FIG. 13, the acquired data from the electronic balance reaches the final value after reaching the maximum value once, but the time until it first reaches the stabilized value is ΔT. From the experiment of Example 2, it was found that the injection speed was stabilized when the injection volume exceeded about 10 ml. However, it is considered that the value close to the injection end operation can be predicted more accurately. A 60 ml slope was used. The average value in Table 1 was adopted as a parameter, and ΔT was determined to be 1.07 s, and the liquid injection rate during the liquid injection completion operation was 80.88%.

  Next, operation confirmation was performed using the determined parameters. FIG. 14 and Table 2 show the results of carrying out three sets of experiments in which 480 ml of the culture solution was divided into 6 portions and poured into 75 ml portions. The liquid injection operation could be carried out within the target accuracy range of ± 2% (required accuracy) without causing dripping out of the flask or at the tube tip. According to the analysis result of Example 1, the liquid injection work time per time in the case of manual work was about 70 s. On the other hand, from the result of FIG. 13, the time for one injection when the robot was used was about 15 s. In the method implemented this time, the suction work was unnecessary because it was injected directly from the injection bottle. It was confirmed that when the robot work and manual work were combined, the work time could be reduced to less than half even if the cap was removed and attached manually.

Claims (7)

A device that is used by being attached to a liquid injection container, and is provided on the lid, a lid that can be detachably attached to the liquid injection container, a liquid injection tube that can be fitted in a first through hole provided in the lid, and An intake tube that can be fitted into the second through hole formed,
The device, wherein the lower end of the intake tube that protrudes from the lid into the liquid injection container with the device mounted on the liquid injection container is arranged close to the bottom surface of the liquid injection container.   The device of claim 1, wherein the intake tube is provided with a check valve.   The device according to claim 1 or 2, wherein the first through hole is provided in the vicinity of the peripheral edge of the lid.   The device according to any one of claims 1 to 3, wherein the liquid injection container is a container for containing and injecting a culture medium, and the liquid injection is performed by rotating the liquid injection container. A method for injecting a medium,
(A) preparing an injection container containing a medium;
(B) A lid body that can be detachably attached to the liquid injection container, a liquid injection tube fitted into a first through hole provided in the lid body, and a second through hole provided in the lid body. Attaching a device including an intake tube to an injection container;
(C) rotating the liquid injection container to inject the medium through the liquid injection tube, and (d) reversing the liquid injection container to terminate the liquid injection.   The method according to claim 5, further comprising a step (e) of repeating the step (c) and the step (d) to continuously perform a plurality of liquid injection operations.   The method according to claim 5 or 6, further comprising the step of (f) determining an injection volume after step (c) and before step (d).