JP2005168384A - Culture vessel and automatic culture system - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To improve the productivity for cell culture by accommodating more culture vessels in a culture chamber, and to facilitate the unloading of closely arranged individual culture vessels from the culture chamber. <P>SOLUTION: The culture vessel 3 is constituted of a box-shaped vessel body 100, having the following construction: The outer side of one of two walls 101 and 102 ( e.g. 101 ) lying apart against each other is provided with projections 105a and 105b, and the outer side of the other wall ( e.g. 102 ) is provided with projections 105c and 105d at such positions as not to interfere with the projections 105a and 105b provided on an adjoining culture vessel 3 when the similar culture vessel 3 is arranged in the direction of the walls 101 and 102 lying apart from each other. <P>COPYRIGHT: (C)2005,JPO&NCIPI

Description

  The present invention relates to a culture container that performs cell culture in vitro, and an automatic culture apparatus that automatically performs cell culture using the culture container.

As a conventional automatic culture apparatus, an apparatus including a fixed storage shelf that can store a plurality of culture vessels and a conveying means that can move horizontally, vertically, and rotationally is known (see, for example, Patent Document 1). .)
This automatic culture apparatus includes a plurality of small rooms arranged in a vertical direction on a storage shelf arranged in a culture chamber, and accommodates one culture container in each small room for culturing, either in the middle of culture or in culture At the end, the conveying means is operated to take out the culture containers one by one from the small room, or to accommodate the culture containers in the small room.

In addition, this automatic culture apparatus is provided with a tip removal unit that automatically removes a used injection tip or a discharge tip into which a culture medium has been injected and discharged from a culture container from a medium injection needle or a medium discharge needle. In this way, a new sterilization chip with no medium attached can be automatically attached to the medium injection needle and the medium discharge needle, and the new sterilization chip is always in contact with the cells in the culture container. Therefore, the cells are not contaminated by dust or the like.
JP 2002-262856 A (FIG. 1 etc.) Japanese Patent Laid-Open No. 2002-077877

  In the automatic culture apparatus described in Patent Document 1, culture is performed by storing culture containers one by one in a small chamber in the culture chamber. However, if an attempt is made to improve productivity, large culture is performed. Needless to say, it is better to accommodate and culture as many culture vessels as possible in the room. However, if a plurality of culture vessels are densely arranged, it becomes difficult to take out the individual culture vessels from the culture chamber, and high work accuracy is required when transporting the culture vessels.

  The present invention has been made in view of the above circumstances, and it is possible to improve the productivity of cell culture by accommodating more culture vessels in the culture chamber, and to dispose the closely arranged individual culture vessels from the culture chamber. The purpose is to make it easier to remove.

As means for solving the above problems, a culture container and an automatic culture apparatus having the following configuration are employed.
That is, the culture container of the present invention is a box-shaped culture container having an open top surface, and a first protrusion is formed on the outer surface of one of the two wall parts facing and spaced apart from each other. When the same culture vessel is arranged on the outer surface of the other wall portion of the two wall portions in the direction in which the two wall portions are separated from each other, it does not interfere with the first protrusion provided on the adjacent culture vessel. A second protrusion is formed at the position.

In the present invention, when a plurality of culture vessels are arranged in a direction in which one wall portion and the other wall portion are separated from each other, adjacent culture vessels face one wall portion and the other wall portion. At this time, since the first protrusion and the second protrusion are formed at positions that do not interfere with each other, adjacent culture containers are arranged more closely, and more culture containers are provided in a limited space. Can be accommodated.
In addition, since the first protrusion and the second protrusion provided on the two closely contacting walls are formed at positions where they do not interfere with each other, even when one culture container is transported, The protrusion does not get in the way.

  The automatic culture apparatus of the present invention is an automatic culture apparatus provided with a transport apparatus that transports the culture container of the present invention, wherein the transport apparatus moves along the one wall portion and the first protrusions. And a second claw portion that moves along the other wall portion and engages with the second protrusion, the first claw portion and the second claw portion. The culture vessel is held by operating the claw portions of the same in synchronization.

  In the present invention, since both the first claw portion and the second claw portion move along two opposing wall portions of the container body and engage with the first and second protrusions, a plurality of cultures Even when the containers are closely arranged, it is possible to engage the first and second protrusions by inserting the first and second claw portions between adjacent culture containers.

In the present invention, the culture vessel is provided with a lid formed so as to cover the upper surface and cover the other wall, and the lid is provided with first and second protrusions provided on the culture vessel. It is preferable to form a notch for avoiding interference.
In the present invention, in order to prevent the first and second claws and the lid of the automatic culture apparatus from interfering with each other, the lower portions of the first and second protrusions are below the lower edge of the lid. It is preferable to be located at.
In the present invention, it is preferable that the dimension in the thickness direction of the first and second protrusions is substantially the same as the thickness of the other wall portion.
In the present invention, it is preferable that the cross-sectional shape along one or the other wall portion of the first and second protrusions is substantially the same as the shape of the notch portion.
In this invention, it is preferable to form the bulging part which contact | abuts the upper part of a 1st, 2nd protrusion in the cover body back surface near a notch part.

According to the present invention, even when a plurality of culture vessels are arranged closely, adjacent culture vessels are arranged more closely, and it is possible to accommodate more culture vessels in a limited space. Therefore, it is possible to improve the productivity of cell culture by accommodating more culture vessels in the culture chamber.
In addition, according to the present invention, even when a plurality of culture vessels are arranged closely, the first and second protrusions are engaged with the first and second claws between adjacent culture vessels. Can be combined. Thereby, since the workability | operativity by the machine at the time of conveying a culture container improves, it can make it easy to take out each culture container from a culture chamber.

One embodiment of the culture vessel of the present invention will be described with reference to FIGS.
As shown in FIG. 1, the culture container 3 of the present embodiment includes a container main body 100 and a lid 110 attached to the upper surface of the container main body 100. The container body 100 has a rectangular shape when viewed from above, and has a box shape with an open top surface and a uniform depth. The lid 110 has a rectangular shape like the container body 100, and an edge 111 is formed on the periphery so as to be folded downward. The lid 110 is attached to the container body 100 so that the upper edge of the container body 100 is stored inside the edge 111. In the present embodiment, the four wall portions 101 to 104 of the container body 100 are inclined so that the upper surface is wider than the bottom surface. However, the container body 100 may be configured without providing such an inclination. good.

  On the outer side surfaces of the two wall portions 101 and 102 that face and separate from each other in the width direction of the container main body 100, when the culture vessel 3 is transferred using the container transfer robot 200 described later, the claw portions 204 a to 204 a of the container transfer robot 200. Two protrusions 105a to 105d hooked on 204d are formed. Two protrusions (first protrusions) 105 a and 105 b formed on the outer surface of one wall portion 101 are provided apart from each other in a direction parallel to the bottom surface of the container body 100, and the outer surface of the other wall portion 102. The two protrusions (second protrusions) 105 c and 105 d formed in the above are spaced apart in a direction parallel to the bottom surface of the container main body 100. The dimension t in the thickness direction of each of the protrusions 105a to 105d is substantially the same as the thickness T of the wall portions 101 and 102, as shown in FIG.

  As shown in FIG. 3, a plurality of culture vessels 3 are arranged side by side in an automatic culture apparatus to be described later, but when two culture vessels 3 are arranged in the width direction of the vessel main body 100, The projections 105a and 105b are provided closer to the wall portion 103 so that the projections 105a and 105b provided on the portion 101 and the projections 105c and 105d provided on the wall portion 102 of the other container body 100 do not interfere with each other. The protrusions 105c and 10d are provided closer to the wall 104.

  The distance between the protrusions 105a and 105b along the longitudinal direction of the container body 100 and the distance between the protrusions 105c and 105d are both 2 × L1, and the distance between the protrusions 105b and 105c along the longitudinal direction of the container body 100 is L1. ing. This is equivalent to the fact that the distance between the protrusions 105a and 105c along the longitudinal direction of the container body 100, the distance between the protrusions 105c and 105b, and the distance between the protrusions 105b and 105d are all L1. Further, the distance from the wall portion 103 to the protrusion 105a along the longitudinal direction of the container body 100 and the distance from the wall portion 104 to the protrusion 105d are both L2. Further, the distances from the bottom surface of the container body 100 to the protrusions 105a to 105d are all equal. The container body 100 has a rotationally symmetric shape centered on the center of the bottom surface, and the protrusions 105a to 105d are always at the same position even when the walls 101 and 102 are interchanged.

  The height of the edge 111 of the lid 110 is substantially equal to the distance from the upper surface of the container main body 100 to the center of each of the protrusions 105a to 105d. Therefore, the edge 111 interferes with each of the protrusions 105a to 105d. In order to avoid this, a notch 112 is formed. In the state where the lid 110 is attached to the container main body 100, as shown in FIG. 4, the lowermost portions of the protrusions 105 a to 105 d protrude downward from the edge 111. This structure is employed in order to prevent interference between claws 204a to 204d of the container transport robot 200, which will be described later, and the lid 110.

The protrusions 105a to 105d and the notch 112 may have the following structure. For example, as shown in FIG. 5, each protrusion 105a-105d is expanded so that thickness t may be extended, and also the cross-sectional shape along the wall part 101 of the protrusion 105a and the shape of the notch part 112 are formed substantially identically. When such a structure is adopted, the protrusion 105a and the notch 112 are aligned with almost no gap when the lid 110 is attached to the container main body 100, so that foreign matter enters the inside of the container main body 100 from this position. There is nothing. Note that it is preferable that the protrusion 105a is not provided with a notch or the like but has a solid shape.
In addition, as shown in FIG. 6, a bulging portion 113 is formed on the back surface of the lid 110 so as to correspond to the position where the uppermost portions of the protrusions 105 a to 105 d abut. When such a structure is adopted, the upper edges of the wall portions 101 and 102 of the container main body 100 and the back surface of the lid 110 are in contact with each other without any gap, so that no foreign matter enters the inside of the container main body 100 from this location.

Next, an automatic culture apparatus 1 that performs automatic culture of cells using the culture container 3 having the above structure will be described with reference to FIG.
The automatic culture apparatus 1 of the present embodiment includes a first space S1 and a second space S2 that are sealed with a transparent wall material that can be observed from the outside and communicate with each other through an opening provided between the two. Is provided. A shutter 2 is provided in the opening.

  The first space S1 is divided into three sections. In the spaces S11 and S13 on both sides, two culture chambers 4 for storing the culture vessels 3 are provided, for a total of four. A tray transfer robot 5 that delivers the tray 7 between the culture chamber 4 and the conveyor 9 is provided. In the upper part of the central space S12, there is provided an air purifying unit 6 for sending a clean descending air flow in order to purify the air in the space S12.

  The culture chamber 4 has a box shape, an opening 4b is provided on a certain side surface, and an openable / closable door 4a is provided on the opening 4b. A plurality of rail-like tray holding members 4c are provided at corresponding height positions on the left and right side walls toward the opening 4b. In the culture chamber 4, each tray holding member 4c that forms a pair on the left and right sides is provided. The tray 7 can be accommodated in a plurality of stages in the vertical direction so as to be hung. A plurality of, for example, 10 culture vessels 3 can be placed side by side on the tray 7. The tray holding member 4c is not limited to a rail shape, and may be in any form as long as the tray 7 can be supported in a removable manner.

  Each culture chamber 4 is arranged with the door 4a facing the central space S12, and those facing each other across the central space S12 face the door 4a. All the culture chambers 4 are maintained at predetermined internal conditions such as a temperature of 37 ± 0.5 ° C., a humidity of 100%, and a CO2 concentration of 5%.

  Below each culture chamber 4 is provided a stocker 8 for accommodating a tray 7 on which a plurality of unused culture vessels 3 are placed. The stocker 8 has a box shape similar to that of the culture chamber 4, and an opening is provided on the entire side of the culture chamber 4 opposite to the door 4a, and an openable / closable door is provided in the opening. .

  The tray transfer robot 5 is disposed substantially in the center of the central space S12, and includes a first arm 5a that rotates horizontally, a second arm 5b that is connected to the tip of the first arm 5a and rotates about the vertical axis, A hand 5c that is connected to the tip of the second arm 5b and rotates about the vertical axis but does not itself have a mechanism for deteriorating the environment in the culture chamber 4, such as a drive unit and a transmission mechanism, and the first arm 5a, A lifting mechanism 5d for lifting and lowering the two arms 5b and the hand 5c is provided. By operating these mechanisms, the tray transfer robot 5 accesses all the trays 7 accommodated in each culture chamber 4 and conveys the conveyor 9 disposed between the first space S1 and the second space S2. It has an operation range in which the tray 7 can be handed over and an operation range in which at least the uppermost tray 7 in the stocker 8 can be accessed.

  The hand 5c is formed in a flat shape that extends in the horizontal direction so that the tray 7 can be placed thereon, and has a thickness that can be inserted into the gap between the upper and lower trays 7 accommodated in the culture chamber 4. . The hand 5c is lifted from a state where it is inserted into the gap between the upper and lower trays 7, so that the tray 7 is pushed up from below by the two arms and taken up from the tray holding member 4c, and the tray 7 is stably held. It is like that.

  The conveyor 9 includes two endless belts 9a arranged on the left and right sides with a larger distance than the width of the hand 5c of the tray transfer robot 5, and is laid between the two belts 9a. Thus, the tray 7 is placed. The belt 9a is not limited to endless.

  A culture treatment apparatus 30 is installed in the second space S2. The culture treatment apparatus 30 includes a supply / discharge robot 10 that supplies cells and supplies / recovers the cells to the culture container 3 on the tray 7 that has been conveyed from the first space S1 to the second space S2 by the conveyor 9, and a culture robot. A centrifuge 11 for separating cells from the medium in the container 3, and four dispensing robots 13 that can be horizontally rotated and moved up and down with an electric pipette 12 for dispensing various liquids such as serum and reagents, A plurality of disposable tips 14 attached to the tips of the electric pipettes 12 of the supply / discharge robot 10 and the dispensing robot 13 are accommodated, and three chip supply devices 15 that can be provided within the operation range of the supply / discharge robot 10 and the dispensing robot 13. And a chip collection unit 31 that discards and collects used chips 14, a reagent supply device 16 that stores various liquids such as serum and reagents in a plurality of containers, and the culture container 3. The culture vessel 3 is moved horizontally between the robots 10 and 13 and the microscope 17 capable of observing the state of the cells to be observed, the plurality of storage tanks 18 for storing the waste liquids discarded by replacing each reagent and medium, and the like. Between the horizontal movement mechanism 19 and the slider 20 of the horizontal movement mechanism 19, the culture vessel 3 received is placed and placed between the placement table 21 and the tray 7 and the placement table 21 that have been conveyed to the second space S2. And a container transfer robot (transfer apparatus) 200 for delivering the culture container 3. The second space S2 is also provided with an air purifier 32 that forms a clean downward airflow to purify the air in the second space S2.

  The second space S2 is arranged at an intermediate position in the height direction, and the first partition wall 33 that vertically divides the second space S2 into an upper space S21 and a lower space S22, and the first partition wall 33. Is divided into three spaces S21, S221, and S222 arranged in the vertical direction by a second partition wall 34 that further divides the inside of the lower space S22 vertically. The first partition wall 33 is disposed at the height of the conveyor 9, and in the space S21 above the first partition wall 33, an arm 13a of the dispensing robot 13, an arm 13a of the dispensing robot 13, and an XY table of the microscope 17 are provided. The mechanism part etc. above 17a are arrange | positioned.

  The first partition wall 33 has a long hole 35 for connecting the mounting table 21 to the horizontal movement mechanism 19 in the lower space S21 in order to move the mounting table 21 in the upper space S21. A through hole 36 for taking out the chip 14 from the chip supply device 15 arranged in the space S221 below the wall 33 and a disposal port 37 for discarding the used chip 14 are formed therethrough. Further, the first partition wall 33 is provided with vent holes 38 penetrating vertically along the side walls 30a and 30b (shaded portions).

  In the space S221 between the first partition wall 33 and the second partition wall 34, the body portion of the dispensing robot 13, the tip supply device 15, the reagent supply device 16, and the XY table 17a of the microscope 17 are below. The part, the horizontal movement mechanism 19, and a duct for connecting the disposal port 37 of the chip collection unit 31 and the disposal container 39 are provided. In the vicinity of the side walls 30a and 30b of the second partition wall 34, vent holes 43 penetrating vertically are provided along the side walls 30a and 30b (shaded portions). In the space S222 below the second partition wall 34, the centrifuge 11, the storage tank 18, the waste container 39, and the exhaust fan 41 are arranged. An HEPA filter 42 is provided at the outlet of the exhaust fan 41 in order to clean the exhausted air.

  The supply / discharge robot 10 sucks and injects the cells separated by the electric pipette 10a for supplying or sucking the medium from the upper opening of the culture vessel 3 with the lid 3b opened and the centrifuge 11 into the culture vessel 3. A head 10c having an electric pipette 10b, two arms 10d and 10e that can be turned horizontally, and a lifting mechanism 10f that is provided at the tip of the arm 10e and moves up and down the head 10c. Reference numeral 10 g denotes a duct that guides the culture medium from the storage tank 18 or discharges the culture medium into the other storage tank 18 as waste liquid.

  The supply / discharge robot 10 attaches the tip 14 supplied from the tip supply device 15 to the tip of the electric pipette 10b, aspirates the cells separated by the centrifuge 11, and mounts the culture vessel 3 mounted on the mounting table 21. It is designed to be supplied from the upper opening. Further, another tip 14 supplied from the tip supply device 15 is attached to the tip of the electric pipette 10a, and the medium is supplied into the culture vessel 3 mounted on the mounting table 21, or the medium is supplied from the inside of the culture vessel 3. Alternatively, the medium containing the cells is aspirated. The used chip 14 once used is removed and collected by the chip collection unit 31. Therefore, the supply / discharge robot 10 has various devices such as a mounting table 21, a chip supply device 15, a chip collection unit 31, and a cell supply device (not shown) from the centrifuge 11 disposed within its operating range. .

  The centrifuge 11 separates and sinks cells having a high specific gravity suspended in the medium by rotating the medium containing cells supplied from the supply / discharge robot 10 at a low speed.

  Each of the dispensing robots 13 includes a horizontally rotatable arm 13a provided with an electric pipette 12 to which a tip 14 is detachably attached to a tip, and an elevating mechanism 13b for raising and lowering the arm 13a. The dispensing robot 13 supplies a culture medium and various reagents into the culture vessel 3 that has been transported by the horizontal movement mechanism 19. Therefore, the dispensing robot 13 arranges various devices such as the mounting table 21, the chip supply device 15, the chip recovery unit 31, and the reagent supply device 16 on the horizontal movement mechanism 19 within the operation range.

  The tip supply device 15 accommodates a plurality of tips 14 in an arrayed state in a container 15a opened upward with the attachment ports to the electric pipettes 10a, 10b, 12 facing upward. When the robot 13 needs a new tip 14, the tip 14 is attached to the tip of the electric pipette 10a, 10b, 12 simply by inserting the electric pipette 10a, 10b, 12 from above. . The container 15a is attached to the moving mechanism 15b so as to be reciprocated in a direction intersecting the moving direction of the electric pipettes 10a, 10b, and 12 by the supply / discharge robot 10 and the dispensing robot 13. The tip supply device 15 that supplies the tip 14 to the dispensing robot 13 is provided with another moving mechanism 15c that moves the container 15a in a direction orthogonal to the moving direction of the moving mechanism 15b. Thus, the electric pipettes 10a, 10b, and 12 can access all the tips 14 in the container 15a.

The tip collection unit 31 includes a gripping device (not shown) that grips the tip 14 at the entrance of the disposal container 39, and the tip 14 used in the supply / discharge robot 10 and the dispensing robot 13 is inserted into the gripping device. Then, this is grasped. In this state, the supply / discharge robot 10 and the dispensing robot 13 move the electric pipettes 10a, 10b, 12 to remove the used tips 14 from the tips of the electric pipettes 10a, 10b, 12 and put them in the disposal container 39. It is collected via the duct 40. The disposal container 39 is detachably disposed in the space S222 and can be replaced as necessary.
When exchanging the duct 40 and the waste container 39, the door may be accessed from the outside of the culture treatment apparatus 30 by opening a door (not shown) provided on the side walls 30a and 30b of the culture treatment apparatus 30.

  The reagent supply device 16 has a plurality of reagent containers 16b mounted in a cylindrical casing kept at a constant temperature. Each reagent container 16b includes, for example, MEM (Minimal Essential Medium), DMEM (Dulbecco's Modified Eagle Medium), and FBS (Fetal Bovine Serum: fetal bovine serum) that constitute a medium necessary for culturing cells. ) And serum such as human serum, proteolytic enzymes such as trypsin that detach cells in the culture vessel 3, growth factors such as cytokines that grow cells during culture, differentiation induction such as dexamethasone that differentiates cells Factors, antibiotics such as penicillin antibiotics, hormones such as estrogen, and nutrients such as vitamins are stored.

  The microscope 17 is used in the case of observing the state of cells in the culture vessel 3 and the degree of proliferation or counting the number of cells in the middle of the culturing process or when exchanging the medium. ing. The XY stage 17a, working distance adjustment, magnification change, and the like of the microscope 17 can all be performed by remote control. It may be possible to observe the state of the cells in the culture vessel 3 from the outside of the automatic culture apparatus 1 by arranging an eyepiece lens toward the outside of the second space S2.

  The storage tank 18 stores, for example, MEM, PBS (phosphate buffered saline), etc. that can be used in common for all specimens, and in the reagent container 16b in the reagent supply device 16 as necessary. To supply. In addition, the storage tank 18 may be a waste liquid tank that stores waste medium and the like that are discharged during medium replacement.

  The horizontal movement mechanism 19 includes a slider 20 that can be moved in the horizontal direction by a linear movement mechanism. The mounting table 21 is mounted on the slider 20, and the culture vessel 3 mounted on the mounting table 21 can be moved from the conveyor 9 to the operating range of the dispensing robot 13.

  The mounting table 21 includes a holding mechanism (not shown) for mounting and holding the culture vessel 3 transferred from the tray 7 on the conveyor 9. Moreover, you may provide the vibration apparatus (illustration omitted) which gives this culture | cultivation container 3 a vibration. For example, in addition to a device that reciprocally swings the culture vessel 3 within a predetermined angle range, a device that applies ultrasonic vibration or a device that applies horizontal vibration may be employed as the vibration device.

  A container transport robot 200 that delivers the culture container 3 between the tray 7 transported to the second space S2 and the mounting table 21 is arranged in parallel with the transport direction of the tray 7 by the conveyor 9, as shown in FIG. The first bar 201 is provided, and the second bar 202 is disposed in a direction orthogonal to the first bar 201. The first bar 201 is fixed at a fixed position. The second bar 202 has a base end attached to the first bar 201 and is supported so as to be movable along the longitudinal direction of the first bar 201 (the direction of the arrow X in the figure).

  The second bar 202 is provided with a slider 203 supported so as to be movable in the longitudinal direction of the second bar 202 (the direction of the arrow Y in the figure). As shown in FIG. 9, the slider 203 is provided with claw portions 204 a to 204 d that engage with the protrusions 105 a to 105 d formed on the container body 100 of the culture container 3 to lift the culture container 3. The claw portions (first claw portions) 204a and 204b correspond to the two protrusions 105a and 105b provided on the wall portion 101 of the container body 100, respectively, and the claw portions (second claw portions) 204c and 204d are These correspond to the two protrusions 105c and 105d provided on the wall 102 of the container body 100, respectively.

  Each of the claw portions 204a to 204d has a shape obtained by punching a thin plate into an L shape, and is arranged following the arrangement of the protrusions 105a to 105d. Each of the claw portions 204a to 204d is supported by the slider 203 so as to be all up and down in a vertical direction, and descends to both sides of the container body 100 along the outer side surfaces of the wall portions 101 and 102. By horizontally moving in the longitudinal direction of the main body 100, it is inserted below the corresponding projections 105a to 105d, and when it is lifted, it engages with the projections 105a to 105d to lift the culture vessel 3.

Cell culture using the automatic culture apparatus 1 configured as described above will be described.
First, bone marrow fluid collected from a patient is placed in a centrifuge container (not shown), and centrifuged at a centrifuge 11 to separate bone marrow cells having a high specific gravity from the bone marrow fluid. The operation of putting the centrifuge container into the centrifuge 11 may be performed by an operator or may be performed by the supply / discharge robot 10.

  Next, the bone marrow cells separated from the bone marrow fluid are transferred from the centrifuge container to the culture container 3. First, the conveyor 9 is operated in advance, and the ten culture vessels 3 arranged in the vertical 5 rows and the horizontal 2 rows on the tray 7 are moved from the first space S1 to the second space S2. The transfer robot 200 is operated to transfer the culture vessels 3 one by one from the tray 7 onto the mounting table 21. The container transfer robot 200 first moves the second bar 202 along the first bar 201 and also moves the slider 203 along the second bar 202, above the certain culture container 3 on the tray 7. The slider 203 is disposed on the screen. Next, the claw portions 204a to 204d are lowered until the tips of the claw portions 204a to 204d are lower than the projections 105a to 105d, and then the claw portions 204a to 204d are moved laterally to move the projections 105a to 105d. Insert downward. Thereafter, when the claw portions 204a to 204d are raised, the claw portions 204a to 204d engage the corresponding projections 105a to 105d, and the culture vessel 3 is lifted.

The container transport robot 200 that lifts the culture vessel 3 moves the second bar 202 and moves the slider 203, and places the lifted culture vessel 3 above the mounting table 21. Next, the claw portions 204 a to 204 d are lowered and the culture vessel 3 is placed on the mounting table 21. Thereafter, when the claw portions 204a to 204d are moved laterally to be moved out from below the protrusions 105a to 105d and then lifted, the transfer of the culture vessel 3 onto the mounting table 21 is completed. Since the two culture vessels 3 are placed on the mounting table 21, the above operation is repeated to transfer the second culture vessel 3 onto the mounting table 32.
When the culture container 3 is transferred onto the mounting table 21, a culture container opening / closing device (not shown) is operated to release the lid 3 b from the culture container 3 mounted on the mounting table 21 to open the culture container 3.

  Next, the chip supply device 15 is operated to move the container 15 a into the operation range of the supply / discharge robot 10. When the container 15a is moved, the head 10c of the supply / discharge robot 10 is lowered to receive one unused tip 14 from the container 15a and is attached to the tip of the electric pipette 10a. When the chip 14 is delivered, the container 15a is immediately returned below the lid 15b.

  After the tip 14 is attached to the tip of the electric pipette 10a, the head 10c is turned to move the electric pipette 10a onto the centrifuge 11, and then the head 10c is lowered to bring the tip 14 into the bone marrow cells in the centrifuge 11. The bone marrow cells are aspirated into the chip 14. When the bone marrow cells are aspirated, the head 10c is raised, and then the head 10c is rotated to move the electric pipette 12 onto the mounting table 21, and the bone marrow cells sucked into the chip 14 are transferred to the culture container on the mounting table 21. Move in 3. When the bone marrow cells have been transferred, the used tip 14 is removed from the electric pipette 12 and put into a waste container (not shown) through the duct 40 and discarded.

  When the used tip 14 is discarded, one unused tip 14 is again received from the container 15 a and attached to the tip of the electric pipette 12. Thereafter, the supply / discharge robot 10 is operated in the same manner as described above, and DMEM and PBS (phosphate buffered saline) in the storage tank 18 are supplied into the culture vessel 3.

Next, the horizontal movement mechanism 19 is operated, and the culture vessel 3 containing the bone marrow cells is moved to the operation range of each dispensing robot 13 together with the mounting table 21. When the culture container 3 is moved, each dispensing robot 13 is operated to receive one unused tip 14 from the container 15a and is attached to the tip of the electric pipette 10a. When the chip 14 is delivered, the container 15a is immediately returned below the lid 15b. After the tip 14 is attached to the tip of the electric pipette 10a, the tip 14 is brought into contact with serum or various reagents in the container 16b of the reagent supply device 16 and sucked into the tip 14. When the serum and various reagents are aspirated, the arm 13 a is rotated to move the electric pipette 12 onto the mounting table 21, and the serum and various reagents sucked into the chip 14 are supplied into the culture container 3 on the mounting table 21. To do. The order of the bone marrow cell injection, the aspiration and supply of serum and various reagents may be changed as appropriate, but the chip 14 must be replaced once used.
By supplying the supply of serum and various reagents, the bone marrow cells are mixed in an appropriate medium in the culture vessel 3, but if necessary, in order to distribute the bone marrow cells uniformly in the medium. The culture vessel 3 may be vibrated together with the mounting table 21.

  When the inside of the culture vessel 3 is in a state where bone marrow cells are mixed in an appropriate medium, the horizontal movement mechanism 19 is operated to move the culture vessel 3 to the vicinity of the conveyor 9 and the culture vessel opening / closing device is operated to perform culture. The lid 3b is attached to the container 3 and the culture container 3 is closed. Subsequently, the container transport robot 200 is operated, and the same procedure as when the culture container 3 is transferred from the tray 7 onto the mounting table 21, The culture vessel 3 is transferred from the mounting table 21 onto the tray 7.

  When the above processing is performed on the ten culture containers 3 on the tray 7, the conveyor 9 is operated to move the ten culture containers 3 placed on the tray 7 from the second space S2 to the first space S1. . When the culture vessel 3 placed on the tray 7 is moved to the first space S1, the tray transfer robot 5 is operated to lift the tray 7 and transfer it to the front of the culture chamber 4 in which the tray 7 is to be stored. When the door 4a of the culture chamber 4 is opened, the tray 7 is inserted into the culture chamber 4 and placed on the empty tray holding member 4c. When the tray transfer robot 5 is retracted, the door 4a is closed, and the culture conditions in the culture chamber 4 are kept constant so that the cells are appropriately cultured.

  When exchanging the culture medium or the container, the tray transfer robot 5 is operated to take out the culture container 3 in the culture chamber 4 and the conveyor 9 is operated to operate the conveyor 9 in the same manner as described above to operate from the first space S1 to the second space S2. The culture vessel 3 placed on the tray 7 is moved. And trypsin is inject | poured into the culture container 3 moved to 2nd space S2, and the cell in the culture container 3 peels from the container main body 3a. Subsequently, the supply / discharge robot 10 is operated to put the detached cells into a centrifuge container, and the centrifuge 11 collects only necessary ones such as mesenchymal stem cells. Other processing steps are the same as described above. At this time, the culture container 3 of the same specimen may be placed on the tray 7, or the culture containers 3 of different specimens may be placed. Moreover, the culture container 3 of the same sample may be mounted on the mounting table 21, or the culture container 3 of a different sample may be mounted.

  By performing the culture process over a predetermined period by exchanging the medium and the container a plurality of times, a sufficient number of mesenchymal stem cells proliferate in the culture container 3. Whether the mesenchymal stem cells have grown to a sufficient number is determined by operating the supply / discharge robot 10 to transport the culture vessel 3 to the microscope 17 and observing the inside of the culture vessel 3 or measuring the number of cells. To judge.

  According to the automatic culture apparatus 1, mesenchymal stem cells having a sufficient number of cells can be automatically cultured from bone marrow fluid collected from a patient as described above. In addition, after sufficient mesenchymal stem cells are obtained, a biological tissue filling material such as calcium phosphate and a differentiation inducing factor such as dexamethasone are placed in the culture vessel 3 and the culture process is continued, so that a defective part of the living body is obtained. It is possible to manufacture a body tissue filling material that can be supplemented to the body.

  As described above, even when several culture containers 3 are closely arranged, the protrusions 105a and 105b provided on one culture container 3 and the other culture container 3 are adjacent to each other. Since the provided projections 105c and 105d do not interfere with each other, it is possible to arrange the two culture vessels 3 more closely, and accommodate many culture vessels in the very limited space of the culture chamber 4. Thus, the productivity of cell culture can be improved.

  Further, even when the plurality of culture containers 3 are closely arranged, the automatic culture apparatus 1 protrudes by inserting the claw portions 204a to 204d provided in the container transport robot 200 between the adjacent culture containers 3. 105a to 105d can be engaged with each other, and this improves the workability of the machine when transporting the culture container, so that the individual culture containers can be easily taken out from the culture chamber.

It is a perspective view which shows one Embodiment of the culture container of this invention. It is a perspective view which shows protrusion and the wall part of the periphery of it. It is the top view which looked at the container main body which comprises the culture container of FIG. 1 from upper direction. It is the side view which looked at the protrusion provided in the container main body of FIG. 2 from the side. It is a perspective view which shows the modification of the shape of protrusion. It is a perspective view which shows the modification of the shape of a cover body. It is a perspective view which shows one Embodiment of the automatic culture apparatus of this invention. It is the top view which looked at the container conveyance robot which comprises the automatic culture apparatus of FIG. 4 from upper direction. It is state explanatory drawing which shows the movement of a nail | claw part at the time of a container conveyance robot holding a culture container.

Explanation of symbols

3 Culture container 7 Tray 100 Container body 110 Lid 105a, 105b Protrusion (first protrusion)
105c, 105d Protrusion (second protrusion)
200 Container transfer robot (transfer device)
201 First bar 202 Second bar 203 Slider
204a, 204b Claw part (first claw part)
204c, 204d Claw part (second claw part)

Claims (7)

  A box-shaped culture container having an open upper surface, wherein a first protrusion is formed on an outer surface of one of the two wall portions facing and spaced apart from each other, and the other wall of the two wall portions When the same culture container is arranged in the direction in which the two wall portions are separated from each other on the outer surface of the part, the second protrusion is at a position that does not interfere with the first protrusion provided in the adjacent culture container. A culture vessel characterized by being formed.   The lid has a lid formed to cover the upper surface and to cover the one or the other wall, and the lid has a notch for avoiding interference with the first and second protrusions. The culture container according to claim 1, wherein the culture container is formed.   The culture container according to claim 2, wherein lower portions of the first and second protrusions are present below a lower edge of the lid.   The culture container according to any one of claims 1 to 3, wherein a dimension in a thickness direction of the first and second protrusions is substantially the same as a thickness of the one wall and the other wall. .   5. The cross-sectional shape of the first and second protrusions along the one and other wall portions is substantially the same as the shape of the cutout portion. 6. Culture container.   The culture according to any one of claims 2 to 5, wherein a bulging portion that abuts on top of the first and second protrusions is formed on the back surface of the lid in the vicinity of the notch. container. An automatic culture apparatus comprising a transport device for transporting the culture container according to claim 1,
The transport device moves along the one wall portion and engages with the first projection, and moves along the other wall portion and engages with the second projection. An automatic culture apparatus comprising: a second claw portion that is joined, and holding the culture vessel by operating the first claw portion and the second claw portion in synchronization.

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