JP6660544B2 - Cell clump sheet culture device - Google Patents

Description

  The present invention relates to a cell mass sheet culturing device, and more particularly, to a cell mass sheet culturing device in which cell masses are arranged in a plane on a flat culture tray, and the cell masses are cultured to form a cell mass sheet fused with each other. .

In regenerative medicine for restoring damaged biological functions using stem cells or the like, a cell sheet in which cells are cultured at a high density is used. Since this cell sheet is extremely thin, a method for producing a laminated cultured cell sheet in which several cell sheets are stacked is known (Patent Document 1).
On the other hand, a cell mass (spheroid) in which cells have been cultured in a substantially spherical shape in advance is arranged in a state of being in contact with each other and suspension culture is performed, and a cell mass having a thickness of 50 to 300 μm (cell A method for producing a lump sheet is known (Patent Document 2).
Furthermore, as a method for arranging a cell mass in an arbitrary space, a method for producing a three-dimensional structure of cells in which a cell mass is penetrated through a support made of a filament or a needle is known (Patent Document 3). ).

Japanese Patent No. 4921353 Japanese Patent No. 5523830 Japanese Patent No. 4517125

However, when the cell sheets are laminated as in Patent Literature 1, the cell sheets are extremely thin, so that they are difficult to handle and difficult to automate.
In addition, when a cell mass sheet is obtained by suspending and culturing a cell mass as in Patent Document 2, since the cell masses fuse irregularly, the shape of the obtained cell mass sheet is not determined, and the thickness of the obtained cell mass sheet is also irregular. There was a problem of uniformity.
Furthermore, when a cell mass is made to penetrate through a support made of a thread-like body or a needle-like body as in Patent Literature 3, there is a possibility that cells may be damaged.
In addition, when culturing a cell mass placed in a plane on the mounting surface, the cell mass located in the center is limited to contact with the culture solution only at the upper part, so that the cells at the peripheral part where the side part and the upper part come into contact with each other The growth of the lower part was slower than the mass, and in some cases it became necrotic.
In view of such a problem, the present invention provides a cell mass sheet culturing apparatus capable of bringing a culture solution into contact with a lower portion of a cell mass sheet.

That is, the cell mass sheet culturing apparatus according to the first aspect of the present invention prepares a cell mass sheet in which a plurality of cell masses are planarly arranged on a mounting surface in a culture vessel and the cell masses are cultured and fused with each other. A cell mass sheet culture device,
Pressing means for laterally moving the cell mass in culture on the mounting surface is provided.

According to the first aspect of the present invention, there is provided a pressing means for laterally moving the cell mass on the mounting surface , and the cell mass is laterally moved on the mounting surface during culture by the pressing means. The culture solution is caught between the surface and the cell mass sheet being cultured, and the culture solution sufficiently spreads to the lower portion of the cell mass located at the central portion, so that the portion can be cultured well. Further, activation of the cell mass is expected by the movement.

Configuration diagram of the cell mass sheet manufacturing apparatus according to the first embodiment Sectional view of the storage container and suction nozzle Perspective view of culture tray and culture vessel Top view of culture tray and culture vessel Side view of the culture vessel support Side view of inspection means Side view of culture solution supply means and cell mass sheet moving means Plan view of a culture tray usable in the first embodiment Top view of culture tray usable in the first embodiment and side view of pin Plan view of a culture tray usable in the first embodiment Side view of culture tray and culture vessel usable in the first embodiment Side view of culture solution supply means usable in the first embodiment Configuration diagram of the cell mass sheet manufacturing apparatus according to the second embodiment Top view of culture vessel Side view of culture vessel Plan view for explaining the holder Side view of cell mass sheet moving means Plan view of a culture vessel usable in the second embodiment Front view of isolator and incubator provided with cell mass sheet manufacturing equipment Configuration diagram of the cell mass sheet manufacturing apparatus according to the third embodiment

FIG. 1 to FIG. 7 illustrate a cell mass sheet manufacturing apparatus 1 according to a first embodiment, in which a plurality of cell masses 2 (spheroids: see FIG. 2) are cultured. A cell mass sheet 4 (see FIG. 6) is arranged in a plane on a flat mounting surface provided in 3, and the cell mass 2 is cultured and fused with each other.
The cell mass 2 can be produced, for example, by the method disclosed in Japanese Patent No. 4517125. In other words, when cells are seeded and cultured in a container having an inner surface that is non-adhesive, the cells adhere to each other in search of a scaffold to form small cell clumps, which are further fused to have an outer diameter of about 500 μm. A substantially spherical cell mass 2 is formed.
More efficiently, the cell mass 2 can be easily obtained by culturing the cells in the non-adhesive wells (substantially hemispherical storage portions) of the well plate. The method of preparing the cell mass 2 is not limited to this, and a swirling culture method in which the cell suspension is put in a swirling culture solution, a method in which the cell suspension is put in a test tube and precipitated by a centrifuge, Alternatively, it can be prepared by various known methods such as a method of culturing using alginate beads.
As described above, the cell mass 2 refers to a substantially spherical cell aggregate having an outer diameter of about 100 to 500 μm in which cells are aggregated and aggregated, and the cell mass sheet 4 in the present invention is referred to as such. Are formed into a sheet by fusing a plurality of cell clumps with a thickness of about 100 to 500 μm according to the size of the cell clump 2 to be used. It can be adjusted by the number of cell masses 2 to be made.
The cell mass sheet 4 thus produced is different from a cell sheet obtained by culturing a single cell into a sheet by culturing a cell suspension in a planar manner.

The cell mass sheet manufacturing apparatus 1 of the present embodiment is provided inside an isolator 61 in which the inside is maintained in an aseptic state and an incubator 62 which is connectable to the isolator 61 as shown in FIG. Further, the isolator 61 is provided with a pass box 63 for decontaminating the carried-in material.
FIG. 1 shows a configuration provided inside the isolator 61, and a storage container supporting portion 6 that supports a storage container 5 that stores a cell mass 2 and a culture that supports a culture container 3 in which a cell mass sheet 4 is cultured. A container support 7, a suction nozzle 8 as a plurality of holding means for holding the cell mass 2, and a nozzle as a moving means for moving the suction nozzle 8 relatively to the storage container 5 and the culture container 3 It comprises a moving means 9 and an inspection means 10 (see FIG. 6) for checking the culture state of the cell mass sheet 4 in the culture vessel 3.
On the other hand, FIG. 7 shows a configuration provided inside the incubator 62, in which a culture solution supply means 11 for supplying a culture solution to the cell mass 2 or the cell mass sheet 4 on the culture vessel 3, and a cell mass sheet in culture. 4 for moving the cell mass sheet. Since the cell mass 2 or the cell mass sheet 4 is cultured inside the incubator 62, the configuration inside these incubators 62 constitutes a cell mass culturing device or a cell mass sheet culturing device.
In the following description, the horizontal direction in FIG. 1 is the X direction, the vertical direction is the Y direction, and the vertical direction is the Z direction. In FIG. 4, the horizontal direction is the X direction and the vertical direction is the Y direction.

The inside of the isolator 61 is maintained in a sterile environment, and a one-way flow of sterile air from above to below is formed by sterile air supply means.
A glove 61a that can be worn by an operator is provided on the front of the isolator 61, so that various operations can be performed. In addition, a robot or a transfer means having a required configuration may be provided inside the isolator 61 to automatically perform these operations.
Next, the inside of the incubator 62 is maintained in a sterile environment and at a predetermined temperature and humidity suitable for culturing the cell mass sheet 4, and the cell mass 2 is cultured and fused to the cell mass sheet 4. During the formation, the incubator 62 can be separated from the isolator 61, and the cell mass 2 can be cultured at a location away from the isolator 61.
Therefore, the isolator 61 and the incubator 62 are connected by a connecting means 64, and a connecting means for connecting and disconnecting the incubator 62 and the isolator 61 while maintaining an aseptic state as described in, for example, Japanese Patent No. 4656485. Can be used.

FIG. 2 is a cross-sectional view of the storage container 5. The storage container 5 includes a plurality of storage recesses 5 a in a vertical and horizontal direction in a plan view, and each of the storage recesses 5 a has a substantially spherical cell mass 2 together with a culture solution. Is housed.
The housing recesses 5a of the housing container 5 are arranged by a predetermined number in the X direction and the Y direction, respectively, in plan view. In the present embodiment, the X of the cell mass 2 constituting the cell mass sheet 4 produced in the culture container 3 is set. The number is the same as the number in the direction and the Y direction, that is, seven in the X direction and seven in the Y direction.

The storage container supporting portion 6 that supports the storage container 5 positions the storage container 5 on an upper surface thereof with a positioning piece 6b, and includes a Y-direction table 6a that forms the nozzle moving unit 9.
As will be described later, since seven suction nozzles 8 of this embodiment are provided in the X direction and can be moved only in the X direction and the Z direction, the Y-direction table 6a moves the storage container 5 in the Y direction. , The suction nozzle 8 and the housing container 5 can be relatively moved in each of the XYZ directions.
More specifically, first, the Y-direction table 6a positions the first row of the storage recesses 5a of the storage container 5 below the suction nozzle 8, and the suction nozzle 8 is moved from the first row of the storage recesses 5a to the cell mass. When 2 is sucked and held, the Y-direction table 6a moves the storage container 5 by one row in the Y direction, and the second row of the storage recesses 5a is positioned below the suction nozzle 8.

As shown in FIG. 1, a culture tray 13 is accommodated in the culture vessel 3 so as to be able to move up and down, and the culture vessel 3 is supported by the culture vessel support 7.
As shown in FIG. 3, the culture tray 13 has a substantially square plate shape. As shown in FIG. 4, the upper surface thereof has a mounting surface 13a on which seven cell clusters 2 are arranged in the X and Y directions, respectively. The size is set so that they can be aligned in a state where they are in contact with each other.
The mounting surface 13a of the culture tray 13 on which the cell mass 2 is placed has a non-cell-adhesive property, so that the cell mass 2 and the cell mass sheet 4 are hardly adhered. Since the cell mass 2 is hard to adhere to the mounting surface 13a, the function of connecting the adjacent cell masses 2 to each other becomes stronger, and the formation of the cell mass sheet 4 is promoted.
The cell non-adhesiveness can be obtained by making the surface properties water-repellent or superhydrophilic, and can be realized by using a known material having these properties or by performing a surface treatment or a coating treatment. In addition, as a property of the surface of the mounting surface 13a, a DLC (diamond-like carbon) film can be formed.
Further, the culture tray 3 is made of a material that transmits light, and when a coating or the like is formed on the mounting surface 13a, the coating also transmits light.

The culture vessel 3 has a box shape with a bottom and contains therein a culture solution at a predetermined depth in advance, and a plurality of pins 14 as guide means G are regularly provided on the bottom surface.
The bottom surface of the culture container 3 is formed in substantially the same shape as the culture tray 13, whereby the cell mass 2 located on the outer peripheral side of the cell mass 2 aligned on the mounting surface 13 a of the culture tray 13 is , Thereby supporting the aligned cell mass 2 so as not to move from the outside.
Further, the culture vessel 3 is formed of a transparent resin or glass and can transmit light, so that the inspection means 10 can be inspected together with the culture vessel 3.

The culture tray 13 is located at a lowered position in contact with the bottom surface of the culture vessel 3 and at a raised position raised with respect to the lowered position.
Props 15 are provided at the four corners of the culture tray 13, and the adjacent supports 15 are mutually connected by a reinforcing member 16 provided thereabove. Each column 15 is allowed to move only in the vertical direction (Z direction) along the inner surface of the culture vessel 3, so that the culture tray 13 can be moved up and down inside the culture vessel 3 while maintaining a horizontal state. I have.
On the other hand, the culture vessel 3 is provided with a lock mechanism 17 which is provided at a position through which any of the columns 15 passes and which comes and goes with a spring or the like (not shown).
Of the supports 15, the support 15 that passes through the lock mechanism 17 is formed with a recess 15a that engages with the lock mechanism 17. When the culture tray 13 is at the raised position, the lock mechanism 17 projects and the recess is formed. 15a to hold the culture tray 13 at the raised position.
On the other hand, when the culture tray 13 is moved up and down from this elevated position, a predetermined force may be applied to the column 15 in the Z direction, whereby the lock mechanism 17 resists the urging force of the spring. And retreats from the recess 15a.

The pin 14 is fixed to the bottom surface of the culture vessel 3 so as to face upward. The culture tray 13 has a plurality of through holes 13b in accordance with the position of the pin 14.
Accordingly, when the culture tray 13 is located at the lowered position, the pins 14 pass through the through holes 13b and protrude from the upper surface of the culture tray 13, so that the guide means G is set on the culture tray 13.
On the other hand, when the culture tray 13 is located at the raised position, the culture tray 13 is located above the tip of the pin 14 and the pin 14 is detached from the through hole 13b. The guide means G is retracted from above.

Next, as shown in FIG. 4, in this embodiment, seven cell clusters 2 are arranged on the culture tray 13 in the X and Y directions vertically and horizontally. A space formed around the cell mass 2 and formed by the plurality of pins 14 and the pins 14 constitutes a housing portion Ga for housing each cell mass 2.
Specifically, one cell mass 2 is surrounded by four pins 14, and the distance between adjacent pins 14 is made smaller than the diameter of the cell mass 2.
As a result, by accommodating the cell mass 2 in the accommodation portion Ga, the cell mass 2 accommodated in the adjacent accommodation portion Ga can be brought into contact with each other. The distance between the cell clusters 2 and the degree of contact at this time can be adjusted by the distance between the pins 14 and the diameter of the pins 14.
When the cell mass 2 is aligned with the mounting surface 13a of the culture tray 13 as in the present embodiment and the cell mass 2 comes into contact with the wall surface of the culture vessel 3, The housing portion Ga is also formed by the pins and the pins 14.

As shown in FIG. 5, the culture vessel support unit 7 includes a guide moving unit 18 for moving the guide unit G, and a Y-direction table 7 a constituting the nozzle moving unit 9.
The guide moving means 18 holds the reinforcing member 16 of the column 15 provided on the culture tray 13 and holds the culture tray 13 at a predetermined height, and supports and raises and lowers the culture vessel 3. And lifting means 18b.
With the holding means 18a holding the culture tray 13, the elevating means 18b raises and lowers the culture container 3, thereby relatively raising and lowering the culture container 3 and the culture tray 13, and lowering the culture tray 3 to the lower position. And the raised position.
The Y-direction table 7a is, like the Y-direction table 6a constituting the storage container support portion 6, every time the cell mass 2 sucked and held by the suction nozzle 8 is stored in the storage portion Ga of the culture tray 13. The accommodating portions Ga of the culture tray 3 together with the culture container 3 are moved one row at a time in the Y direction.

Further, at a position adjacent to the culture vessel support part 7, a holding lid supply means (not shown) for supplying a holding lid 19 as a floating prevention means above the cell mass 2 aligned on the culture tray 13 is provided. I have.
It is possible to use a robot or the like as the presser lid supply means. When the cell mass 2 is aligned on the culture tray 13, the presser lid 19 is supplied above the aligned cell mass 2. I have.
The presser lid 19 is a member having a mesh shape or a large number of through holes, and the presser lid 19 does not hinder the supply of the culture solution to the cell mass 2 and the cell mass sheet 4 on the culture tray 13. It has become.
The holding lid 19 is made of a light-transmitting material like the culture tray 13, and the side of the holding lid 19 that contacts the cell mass 2 or the cell mass sheet 4 is similar to the mounting surface 13 a of the culture tray 13. , Cells are non-adherent.

Next, the suction nozzle 8 as the holding means will be described. As shown in FIG. 2, a main body 8a connected to a negative pressure generating means (not shown) and a tubular suction provided at a lower end of the main body 8a. 8b.
The inner diameter of the adsorbing portion 8b is smaller than that of the cell mass 2. When the cell mass 2 is adsorbed and held at the tip of the adsorbing portion 8b in the accommodation recess 5a of the accommodation container 5, the cell mass 2 is moved to the adsorbing portion. 8b is not sucked.
In addition, on the mounting surface 13a of the culture tray 13, the center position of the suction portion 8b is positioned substantially at the center of the accommodation portion Ga formed by the pins 14 and 14, and in this state, the negative pressure generated by the negative pressure generating means is set. Is solved, the cell mass 2 can be arranged in the accommodation part Ga.
The holding means may have a configuration such as holding the cell mass 2 by a gripper or the like in addition to the suction nozzle 8 for adsorbing and holding the cell mass 2.

Further, a nozzle moving means 9 as a moving means for moving the suction nozzle 8 includes an X-direction rail 21 provided in the X direction over the accommodation container support 6 and the culture container support 7, and along the X-direction rail 21. X-direction moving means 22 for moving the suction nozzle 8 in the X-direction, a Z-direction moving means 23 provided on the X-direction moving means 22 for raising and lowering the suction nozzle 8 in the Z-direction, It is constituted by the Y-direction tables 6a and 7a in the culture vessel supporting portion 7. The holding means such as the suction nozzle 8 and the moving means constitute a means for supplying the cell mass 2.
Further, seven suction nozzles 8 of this embodiment are arranged in the X direction, and the distance between these suction nozzles 8 in the X direction is changed by the space changing means 24.
Specifically, when the suction nozzle 8 is located above the storage container support 6, the spacing changing unit 24 sets the suction nozzle 8 at the same interval as the storage recesses 5 a of the storage container 5 aligned in the X direction. Change the interval.
On the other hand, when the suction nozzle 8 is located above the culture vessel supporting portion 7, the interval changing means 24 has the same interval as the interval in the X direction in the accommodating portion Ga formed by the pins 14 protruding on the mounting surface 13a. Then, the interval between the suction nozzles 8 is changed.
It is also possible to provide a plurality of rows of suction nozzles 8 in the Y direction, and when the interval between the accommodating recesses 5a of the accommodating container support 6 and the interval between the accommodating portions Ga formed on the culture tray 3 are the same. In this case, the interval changing means 24 can be omitted.
Further, for example, when the interval between the storage portions Ga is small, it is also possible to store the cell mass 2 in every other storage portion Ga by the suction nozzle 8, and in this case, first, the odd number on the mounting surface 13a is set. The cell mass 2 may be placed in the second accommodation portion Ga, and then the cell mass 2 may be placed in the even-numbered accommodation portion Ga.
It is sufficient that at least one suction nozzle 8 is provided. As a means for supplying the cell mass 2, in addition to the configuration for transferring the cell mass 2 as in this embodiment, a large number of cell masses 2 can be used. It may be configured to feed out one by one from the accommodated containers.

The X-direction rail 21 is provided with a camera 25 that moves in the X direction. When the cell mass 2 is placed on the culture tray 13 by the suction nozzle 8, the camera 25 moves the culture container support 7. It is configured to move upward to photograph the mounting surface 13a.
The photographed image confirms whether or not the cell clumps 2 are arranged in all the accommodating portions Ga on the mounting surface 13a. An instruction can be given to accommodate the cell mass 2.
When the nozzle moving means 9 positions the suction nozzle 8 above the culture vessel support 7, the camera 25 retreats from above the culture vessel support 7 to avoid contact with the suction nozzle 8. It has become.

The inspection means 10 shown in FIG. 6 includes an illumination means 26 provided below and a camera 27 provided above, and the culture vessel 3 is connected to the illumination means 26 and the camera 27 by a robot (not shown) or other means. Is to be held between.
As described above, since the culture tray 13, the bottom of the culture vessel 3, and the holding lid 19 are each made of a light-transmitting material, the light irradiated by the illumination unit 26 emits light from the culture tray 3 or the culture vessel 3. 3, the light passes through the cell mass 2 or the cell mass sheet 4 on the culture tray 13 and is received by the camera 27.
The camera 27 confirms these culture conditions from the photographed cell mass 2 or cell mass sheet 4, and can determine continuation or stop of the culture depending on the quality of the culture condition.
In the present embodiment, the illuminating means 26 and the camera 27 are provided inside the isolator 61, but a part of the isolator 61 is protruded to the outside, and the upper and lower parts of the protruding part are made of light transmitting material. As a member, the culture container 3 may be held between these members.

FIG. 7 shows a culture solution supply means 11 provided inside the incubator 62 for supplying a culture solution to the cell mass sheet 4 and a cell mass sheet movement for moving the cell mass sheet 4 being cultured on the culture tray 13. Means 12 are shown.
When a plurality of culture vessels 3 are accommodated in the incubator 62, the culture solution supply means 11 and the cell mass sheet moving means 12 can be provided for each of the culture vessels 3.
The culture container 3 is provided with a connection port 3a below it. Specifically, the connection port 3a is located below the culture tray 13 in a state where the culture tray 13 is located at the raised position. It is provided as follows.
The culture solution supply means 11 includes a pipe 31 having one end connected to the connection port 3a and the other end inserted into the liquid level of the culture solution from above the culture vessel 3, and a culture solution provided in the pipe 31. And a liquid feed pump 32 for feeding the liquid.
The liquid sending pump 32 sucks the culture solution from above the culture container 3 and flows the culture solution from below the culture tray container 3 through the connection port 3a. The culture solution is circulated.

On the other hand, the culture tray 13 is carried into the incubator 62 in a state where the culture tray 13 is located at the raised position in the culture container 3. Therefore, the pins 14 are located below the culture tray 13.
The culture tray 13 is provided with through-holes 13b vertically penetrating therethrough in accordance with the positions of the pins 14, and specifically, the cell mass 2 aligned on the mounting surface 13a of the culture tray 13. The cell tray 2 is provided at a position between the cell and the cell mass 2, and connects the mounting surface 13 a side and the lower surface side of the culture tray 13 to each other.
With such a configuration, the culture solution supplied by the culture solution supply means 11 passes through the through hole 13b, and is placed on the mounting surface 13a side from the lower surface side of the culture tray 13, that is, the mounting surface 13a and the cell mass sheet. 4, and the through holes 13b can be used as supply holes for the culture solution.
As a result, the culture solution can be supplied and brought into contact with the back side of the cell mass sheet 4 that comes into contact with the culture tray 13, and the supply of the culture solution tends to be insufficient due to the contact with the culture tray 13. Cultivation of the back side of the lump sheet 4 can be performed favorably.
When the liquid sending direction of the liquid sending pump 32 is reversed, the culture solution is sucked from the back side of the cell mass sheet 4 through the through-hole 13b, so that the culture from the periphery of the cell mass sheet 4 to the back side is performed. The liquid flows in, and the culture liquid is supplied. In this case, the through hole 13b functions as a discharge hole.

As the cell mass sheet moving means 12, a through-hole 13b formed in the culture tray 13 and a pin 14 provided in the culture vessel 3 and passing through the through-hole 13b can be used.
Further, in the incubator, there are provided a holding member 33 for holding the reinforcing member 16 of the column 15 provided on the culture tray 13, and elevating means 34 for elevating the holding member 33, and the holding member 33 is provided by the elevating means 34. The culture tray 13 moves up and down in the culture vessel 3 by raising and lowering 33.
As shown in FIG. 7B, when the elevating means 34 lowers the culture tray 13 and the pins 14 penetrate from below the culture tray 13, the pins 14 slightly protrude from the culture tray 13 and Is pushed up from below.
Here, since the mounting surface 13a of the culture tray 3 is formed to be non-adhesive, the cell mass sheet 4 can be easily separated from the mounting surface 13a by the pins 14.
The cell mass sheet moving means 12 operates at a predetermined interval, and by such an operation, the cell mass sheet 4 can be easily collected from the culture container 3 when the culture is completed. Inside, it is expected that the cell mass 2 will be stimulated to activate the cell mass 2.
The pin 14 does not need to be completely removed from the through hole 13b.

Hereinafter, a method for producing the cell mass sheet 4 using the cell mass sheet manufacturing apparatus 1 having the above configuration will be described.
First, the container 5 containing the cell mass 2 or the culture container 3 containing the culture tray 13 is loaded into the isolator 61 via the pass box 63 or a predetermined amount is stored in the culture container 3. A preliminary step of supplying the culture solution is performed.
At this time, if a plurality of the storage containers 5 and the culture containers 3 are carried into the isolator 61, a plurality of cell mass sheets 4 can be continuously formed using the plurality of culture containers 3.

Next, a guide setting step of setting the pins 14 as the guide means G on the mounting surface 13a of the culture tray 13 is performed. Specifically, in the culture vessel support portion 7, the guide moving means 18 positions the culture tray 13 at the lower position, whereby the pins 14 protrude from the upper surface of the culture tray 13 to form the accommodation portion Ga. The guide means G is set on the mounting surface 13a.
Note that this guide installation step can be performed simultaneously with the above-described preparation step by loading the culture vessel 3 in which the culture tray 13 has been previously positioned at the lowered position into the isolator.

After the guide means G is set on the culture tray 13 in this manner, the accommodation step of accommodating the cell mass 2 in the accommodation section Ga of the guide means G is then performed by moving the suction nozzle 8.
The nozzle moving means 9 moves the suction nozzle 8, and the interval changing means 24 changes the interval between the suction nozzles 8 while adsorbing and holding the cell mass 2 from the accommodation recess 5 a of the accommodation container 5. 2 is accommodated in the accommodation section Ga of the guide means G provided on the culture tray 13 of the culture vessel 3.
Since the accommodating portion Ga is constituted by four pins 14 surrounding the periphery of the cell mass 2, the cell mass 2 accommodated in the accommodating portion Ga is prevented from jumping out, and the alignment state of the cell mass 2 is reduced. Is to be maintained.
When the cell mass 2 is accommodated in all the accommodating portions Ga in this way, the camera 25 moves above the culture vessel supporting portion 7 to photograph the cell mass 2 on the mounting surface 13a, and It is confirmed whether or not is stored in all the storage portions Ga.

Subsequently, an anti-floating step is performed in which the holding lid 19 is disposed above the cell mass 2 arranged on the culture tray 13.
Specifically, a worker wearing a holding lid supply means or glove 61a (not shown) puts the holding lid 19 into the inside of the culture vessel 3, and the holding lid 19 raises the cell mass 2 on the mounting surface 13a. To prevent

Next, from the state where the cell mass 2 is accommodated in the accommodating portion Ga of the guide means G, a guide retreating step of retreating the pin 14 as the guide means G from the mounting surface 13a of the culture tray 13 is performed.
Specifically, the culture tray 13 is positioned at an elevated position with respect to the culture container 3 by the guide moving means 18 of the culture container support portion 7, whereby the pins 14 projecting from the upper surface of the mounting surface 13 a are mounted. The state is retracted from the placing surface 13a.
Further, since the adjacent cell clusters 2 are in contact with each other when housed in the housing section Ga, the cell clusters 2 arranged vertically and horizontally on the mounting surface 13a are kept in contact with each other. Will remain.

When the cell mass 2 is aligned on the culture tray 13 in this manner, the culture tray 13 is transferred to the incubator 62 together with the culture container 3 by a robot or an operator's manual operation, and aligned on the culture tray 13. A culture step of culturing the cell mass 2 is performed.
First, when the culture tray 13 is transferred to the incubator 62, the incubator 62 is separated from the isolator 61 and placed at a position separated from the isolator 61.
Then, the inside of the incubator 62 is maintained at a predetermined temperature and humidity and at a predetermined concentration of carbon dioxide and oxygen, and the cell mass 2 on the culture tray 13 is cultured, so that the adjacent cell masses 2 are fused with each other. As a result, one cell mass sheet 4 is finally formed.
At this time, since the cell mass 2 is aligned on the mounting surface 13a of the culture tray 13 and is kept in contact with the cell mass 2, the formed cell mass sheet 4 has the shape of the mounting surface 13a of the culture tray 13. Will have substantially the same shape, and the thickness can be made substantially uniform,

During the culture step, a culture solution supply step of supplying a culture solution to the cell mass 2 or the cell mass sheet 4 on the culture tray 13 at predetermined intervals in the incubator 62, and the cell mass sheet 4 on the culture tray 13 are moved. A cell mass sheet moving step is performed.
In the culture solution supply step, the culture solution supply means 11 operates at predetermined intervals, and generates a flow of the culture solution from below to above in the culture vessel 3, whereby the cell mass 2 or cell A new culture solution is supplied to the lump sheet 4.
Further, since the culture tray 13 is located at an elevated position in the culture vessel 3 and has a through-hole 13b as the supply hole, the cell mass sheet 4 and the mounting surface 13a are provided through the through-hole 13b. During this time, a culture solution can be supplied, and the back side of the cell mass sheet 4 can be efficiently cultured.
That is, when the cell mass 2 is aligned and the cell mass sheet 4 is prepared as in the present embodiment, there is a problem that the culture solution does not sufficiently spread on the back side of the cell mass 2 located at the center. By performing this culture solution supply step, it has become possible to satisfactorily culture cells located in the relevant portion.

When the adjacent cell clusters 2 are fused to form the cell cluster sheet 4, the culture tray 3 positioned at the ascending position is lowered by the elevating means 34 in the cell cluster sheet moving step.
Then, the pins 14 protrude from the upper surface of the culture tray 3, and the cell mass sheet 4 is pushed up from below by the pins 14, and the cell mass sheet 4 is separated from the culture tray 3. At this time, since the surface of the culture tray 3 is formed to be non-adhesive, the cell mass sheet 4 can be detached and moved upward with a small pressing force.
By performing the cell mass sheet moving step at predetermined intervals, it is possible to maintain a state where the cell mass sheet 4 does not adhere to the culture tray 3 or a state where the cell mass sheet 4 can be detached from the culture tray 3 with a smaller force. Can give a stimulus.

When the cell mass sheet 4 is formed to some extent by the culturing step, an inspection step of connecting the incubator 62 to the isolator 61 at regular intervals to check the culture state of the cell mass sheet 4 is performed.
In the test step, the culture vessel 3 is transferred from the connected incubator 62 to the isolator 61, and further transferred to the test means 10 shown in FIG.
In the inspection unit 10, the light irradiated from the illumination unit 26 from below passes through the culture container 3, the culture tray 13, the cell mass sheet 4, and the holding lid 19, and the culture of the cell mass sheet 4 is performed based on the image captured by the camera 27. Is checked.
Then, when it is determined in the inspection step that the cell mass sheet 4 is sufficiently cultured, a collection process of collecting the cell mass sheet 4 from the culture container 3 is performed.
By performing the cell mass sheet action step during the culture step, the cell mass sheet 4 can be separated from the mounting surface 13a and collected without using a chemical such as trypsin in the collection step, and the cell The damage to the can be suppressed as much as possible.

Although the guide retreating step in the above embodiment is performed before transferring the culture tray 3 to the incubator 62, the guide retreating step may be performed after transferring the incubator 62.
Specifically, the above-described culturing step is performed with the pins 14 as the guide means G projecting into the mounting surface 13a of the culture tray 13, and the adjacent cell clusters 2 are fused to form the cell cluster sheet 4. After that, the pins 14 can be removed from the formed cell mass sheet 4. Alternatively, the guide retreating step may be performed after the cell mass 2 has been cultured to some extent, and the culture may be continued.

  Further, the culture tray 13 is moved up and down during the culturing step with respect to the culture solution supply step in the above embodiment, and the pin 14 is inserted into and pulled out of the through hole 13b. Alternatively, the culture solution may be pushed into or drawn out of the cell mass and allowed to flow to supply the culture solution to the back surface of the cell mass sheet 4.

Then, for the cell mass sheet moving step in the above embodiment, the culture solution is supplied at a predetermined pressure by the culture solution supply means 11, and the culture solution is ejected upward from the through hole 13b of the culture tray 13. Alternatively, the cell mass sheet 4 may be pushed up from below.
In that case, the holding member 33 and the elevating / lowering means 34 as the cell mass sheet moving means 12 can be omitted, and the culture solution supply means 11 can be used as the cell mass sheet moving means 12.

FIG. 8 illustrates variations of the culture tray 13 and the pins 14 as the guide means G that can be used in the cell mass sheet manufacturing apparatus 1 of the first embodiment.
The cell mass 2 is placed on the culture tray 13 shown in FIG. 8 in a staggered manner. Specifically, the cell mass 2 is linearly arranged in the X direction, and the cell mass 2 is arranged in the Y direction. It is arranged so that it is shifted by half.
In order to align the cell clusters 2 in this manner, the through-holes 13b formed in the culture tray 13 and the pins 14 as the guide means G are accommodated by positioning six pins 14 around each cell cluster 2. It is arranged so as to form the portion Ga.
Also in this culture tray 13, the cell clusters 2 come into contact between the adjacent pins 14, and the cell clusters 2 are separated from each other even when the pins 14 are removed as in the above-described embodiment. The contact state can be maintained.
The cells are not limited to the arrangement of the through-holes 13b and the pins 14 in the culture tray 13 shown in FIGS. 4 and 8, but may be arranged in various forms, such as by arranging the cell mass 2 concentrically on the circular culture tray 13. The mass 2 can be aligned.

FIG. 9 also illustrates a variation of the culture tray 13 and the pins 14 as the guide means G that can be used in the cell mass sheet manufacturing apparatus 1 of the first embodiment.
As shown in FIG. 9A, four pins 14 are positioned around each cell mass 2 on the culture tray 13 of the present embodiment, as shown in FIG. To be aligned.
The pin 14 has a larger diameter than the pin 14 shown in FIG. 4, and the through hole 13b is formed to be larger than the diameter of the pin 14, so that a gap is provided between the pin 14 and the opening of the through hole 13b. Is formed. In addition, four arc-shaped relief portions are formed at the four corners of the through hole 13b in accordance with the shape of the cell mass 2, respectively.
By increasing the size of the through-hole 13b so that a gap is formed between the pin 14 and the opening of the through-hole 3a, more culture can be performed from the through-hole 3a as the supply hole in the culture solution supply step. The liquid can be supplied to the back surface of the cell mass 2 or the cell mass sheet 4.
On the other hand, as shown in FIG. 9B, a tapered portion 14a is formed at the tip of the pin 14, and the tapered portion 14a prevents contact between the cell masses 2 between the pins 14a and 14a. Not be able to.

FIG. 10 also illustrates a variation of the culture tray 13 that can be used in the cell mass sheet manufacturing apparatus 1 of the first embodiment. Here, the description will be made using the culture tray 13 used in FIG.
The culture tray 13 of the present embodiment is provided with a supply hole 13c at a position corresponding to the accommodation portion Ga, that is, a position where the cell mass 2 is placed, in addition to the through hole 13b through which the pin 14 passes. Has become.
That is, in the culture solution supply step, the culture solution can be supplied to the back side of the cell mass sheet 4 also from the through-hole 3a and the supply hole 3b, so that many culture solutions can be circulated. ing.
In any of the above embodiments, the pins 14 are not arranged on the outer peripheral side of the aligned cell mass 2, but the pins 14 are arranged on the outer peripheral side so that the cell mass 2 does not touch the inner wall of the culture vessel 3. It can also be configured as follows.
In this case, the outer peripheral pins 14 are configured to remain on the mounting surface 13a even if other pins 14 are removed from the mounting surface 13a in the guide unit retreating step.
More specifically, when the outer peripheral side pin 14 is made longer than the other pins 14 and the culture tray 13 is positioned at an elevated position with respect to the culture vessel 3 by the guide moving means 18, the outer peripheral side pin 14 becomes The pin 14 is kept in a state of protruding from the mounting surface 13a. When it is necessary to retract these outer pins 14 as well, the raising position may be set in two stages so that the culture tray 13 can be further raised from the above raising position.

FIG. 11 illustrates a variation of the culture tray 13 and the culture container 3 that can be used in the cell mass sheet manufacturing apparatus 1 of the first embodiment.
The culture tray 13 of this embodiment has a bottomed box shape, and has a mounting surface 13a on which the cell mass 2 is mounted and a side surface 13c provided adjacent thereto. Unlike the culture tray 13 and the culture container 3 in FIG. 3, the cell mass 2 arranged by the side surface 13c of the culture tray 13 is supported.
The bottom surface of the culture vessel 3 is formed slightly larger than the culture tray 13, so that a gap is formed between the side surface of the culture tray 13 and the inner wall of the culture vessel 3.
The gap is provided with a slide member 41 provided on the side surface of the culture tray 13 and a guide rail 42 provided in the culture vessel 3 in the vertical direction. Along the inside of the culture vessel 3.
The culture tray 13 is provided with a locking member 44 that is swingably provided via a hinge 43. When the culture tray 13 is at the raised position as shown in FIG. The locking member 44 is tilted outward by 43, and is locked to the upper end of the culture vessel 3.
As the guide means G, similarly to the culture tray 13 and the culture vessel 3 shown in FIG. 3, the pin 14 provided on the bottom surface of the culture vessel 3 is passed through a through hole 13b provided in the culture tray 13 so that the pin 14 is configured to protrude above the mounting surface 13 a of the culture tray 13.
As the guide moving means 18 for moving the culture tray 13 inside the culture vessel 3 and projecting the pins 14, a robot or the like can be used instead of the holding means 18a and the elevating means 18b in FIG. .

FIG. 12 illustrates a variation of the culture solution supply means 11 that can be used in the cell mass sheet manufacturing apparatus 1 of the first embodiment.
The culture solution supply means 11 shown in FIG. 7 is configured to supply the culture solution by circulating the culture solution in the culture vessel 3 using the pipe 31 and the liquid feed pump 32. The culture solution supply means 11 has a configuration capable of supplying a fresh culture solution into the culture container 3.
More specifically, the culture solution supply means 11 includes a culture solution tank 51 for storing fresh culture solution, and a supply pipe connected between the culture solution tank 51 and the connection port 3 a of the culture vessel 3. 52, a liquid feed pump 53 provided in the supply pipe 52, a discharge pipe 54 having one end inserted into the culture solution of the culture vessel 3 and the other end connected to a drain tank (not shown), and the discharge pipe. And a drainage pump 55 provided at 54.
According to the above configuration, the fresh culture solution in the culture solution tank 51 is supplied to the culture container 3 by the liquid supply pump 53, and the used culture solution in the culture container 3 is discharged by the drainage pump 55. Thus, a fresh culture solution can be circulated in the culture container 3.
In this case, the culture solution may be supplied below the culture tray 13 and discharged from above as shown in the drawing, or may be supplied upward and discharged from below.

Next, FIGS. 13 to 16 illustrate a cell mass sheet manufacturing apparatus 1 and a method of manufacturing the cell mass sheet 4 according to the second embodiment. The description of the configuration common to the first embodiment will be omitted, and the common configuration will be described with the same reference numerals.
As shown in FIGS. 13 and 14, the culture container 103 of this embodiment has a bottomed box shape and contains a culture solution in advance, and the cell mass 2 is placed on the mounting surface 103a, which is the bottom surface of the culture container 103. It is to be placed on.
The guide means G of this embodiment shown in FIG. 13A is mounted on the mounting surface 103a of the culture vessel 103, and is held and retracted from the mounting surface 103a. I have.
The guide means G includes a partition member 104 in which a plurality of storage portions Ga are partitioned, and is configured to align the cell mass 2 in the X and Y directions vertically and horizontally.
Here, the size of each accommodation part Ga is set slightly larger than the outer diameter of the cell mass 2.
After accommodating the cell mass 2 in the accommodating portion Ga, the guide means G is retracted from the culture tray 103, whereby the cell mass 2 can be arranged in an aligned state on the mounting surface 103a of the culture vessel 103. it can.

The culture vessel 103 of the present embodiment is provided with a pressing means 105 for pressing the cell masses 2 aligned on the mounting surface 103a from the lateral direction and bringing adjacent cell masses 2 into contact with each other.
The pressing unit 105 is configured to first approach the cell mass 2 arranged on the culture container 103 from a first opposing direction (Y direction) and press the cell mass 2 from the Y direction. A pressing member 111, a second pressing member 112 approaching each other from the second opposing direction (X direction) orthogonal to the Y direction and pressing the cell mass 2 in the X direction, and provided on the culture vessel support 7. And moving means 113 for moving the first and second pressing members 111 and 112 forward and backward.
Connection rods 111a and 112a penetrating from the outside to the inside are slidably provided on the four side surfaces 103b of the culture vessel 103, respectively. The first and second pressing members 111 and 112 are provided, respectively.
The connecting rods 111a and 112a are provided with two stopper members 111b and 112b with the side surface 103b of the culture vessel 103 interposed therebetween, and the stroke amount of the connecting rods 111a and 112a is determined by the interval between the stopper members 111b and 112b. Have been.
With such a configuration, the first and second pressing members 111 and 112 can reciprocate to the retracted position (FIG. 14A) or the advanced position (FIGS. 14C and 14D), respectively. As shown in FIG. 15, the upper part of the first and second pressing members 111 and 112 is covered with the pressing lid 19 to prevent the cell mass 2 from floating during the pressing operation.

The distance between the first pressing members 111 and the distance between the second pressing members 112 located at the retracted positions are set to be substantially the same as the lengths of the partition members 104 constituting the guide means G in the X and Y directions, respectively. .
When the first pressing member 111 is moved to the forward position as shown in FIG. 14C, the cell mass 2 is pressed in the Y direction by the first pressing member 111, and the gap in the Y direction in the aligned cell mass 2 is thereby adjusted. Is eliminated.
Similarly, when the second pressing member 112 is moved to the forward position as shown in FIG. 14D, the cell mass 2 is pressed in the X direction by the second pressing member 112, and thereby the aligned cell mass 2 The gap in the X direction is eliminated.
Here, when the second pressing member 112 is moved to the forward position, in order to avoid contact with the first pressing member 111, the first pressing member 111 is previously moved to the retracted position.
Then, by pressing the cell mass 2 from the Y direction and the X direction by the first and second pressing members 111 and 112, the cell mass 2 on the culture container 103 is brought into contact with the adjacent cell masses 2. They will be aligned in a state.

When the adjacent cell clusters 2 are aligned with each other by the pressing means 105 in this manner, a holder 114 as shown in FIG. 16 is attached to the outer edge of the aligned cell clusters 2. I have.
The holder 114 is a frame-shaped member that is set to have substantially the same width as the X direction and the Y direction of the cell mass 2 in contact with each other, and supports the cell mass 2 from the outside so that the alignment state of the cell mass 2 does not collapse. It has become something.
Further, the holder 114 is provided with a mesh on its upper surface, so that the lifting and movement of the cell mass 2 inside the holder 114 is restricted, and the flow of the culture solution is allowed so as not to hinder the culture of the cell mass 2 inside. It has become.

FIG. 17 shows the cell mass sheet moving means 121 provided in the incubator 62, the first and second pressing members 111 and 112 provided in the culture tray 103, and the cell mass moving means 121 provided in the incubator. A moving means 115 for reciprocating the first and second pressing members 111 and 112 is constituted.
The first and second pressing members 111 and 112 are reciprocated by the moving means 115 so as to press the cell mass 2 on which the holder 114 is mounted from the lateral direction together with the holder 114.
Since the mounting surface 103a of the culture container 103 is also non-cell-adhesive, the cell mass sheet 4 can be moved by pressing from the side.
Further, a connection port 103c may be provided in the culture vessel 103 to circulate the culture solution in the culture vessel 103, similarly to the culture solution supply means 11 described in the first embodiment.

A method for producing the cell mass sheet 4 using the cell mass sheet manufacturing apparatus 1 of the second embodiment having the above configuration will be described.
First, a loading step of loading the container 5 or the culture vessel 103 into the isolator and a preparation step of storing a culture solution in the culture vessel 103 are performed. When placing the culture vessel 103 on the culture vessel support part 7, the moving means 113 of the culture vessel support part 7 is connected to the connection rods 111a, 112a of the first and second pressing members 111, 112.
Next, as shown in FIG. 14A, a guide installation step of installing the guide means G in the culture vessel 103 is performed, and the accommodation section Ga of the guide means G installed in the culture vessel 103 by the suction nozzle 8 is performed. Then, a housing step of housing the cell mass 2 is performed.
When the cell mass 2 is accommodated in the accommodating portion Ga of the guide means G in the accommodating step, a guide retreating step of retreating the guide means G from the culture vessel 103 is performed as shown in FIG. A floating prevention step of arranging the holding lid 19 on the upper part of the cell mass 2 arranged in the step (1) is performed.
The guide installation step, guide removal step, and floating prevention step can be performed by an operator or a robot.

Subsequent to the guide retreating step and the floating prevention step, a pressing step of pressing the cell mass 2 on the culture vessel 103 from the lateral direction and bringing them into contact with each other is performed in the culture vessel support portion 7.
That is, in the state where the guide means G is retracted in the guide retracting step, although the cell clusters 2 are aligned, a gap may be formed between them, and even if culturing is performed in this state, In some cases, fusion of the cell masses 2 does not occur.
Therefore, first, as shown in FIG. 14C, the cell mass 2 is moved in the Y direction by the first pressing member 111, and then, as shown in FIG. 14D, the cell mass 2 is moved by the second pressing member 112. It is moved in the X direction, and the cell clusters 2 on the culture tray 103 are aligned while being in contact with each other.

When the cell masses 2 come into contact with each other in the pressing step, the holding lid 19 provided on the aligned cell masses 2 is removed, and the second pressing member 112 is located at the retracted position. Then, a holder mounting step of mounting the holder 114 on the outer periphery of 2 is performed.
By mounting the holder 114, the movement of the cell mass 2 inside the holder 114 is prevented, so that the cell mass 2 can maintain the aligned state in contact with each other.

Subsequently, the culture container 103 is transferred to the incubator 62, and a culture step is performed in the incubator 62.
When the culturing step is performed for a predetermined period and the cell clumps 2 are fused together inside the holder 114 to form the cell clump sheet 4, a cell clump sheet moving step of moving the cell clump sheet 4 is performed.
The cell clump sheet 4 on which the holder 114 is mounted is located in the culture vessel 103, and the moving means 115 provided in the incubator 62 moves the first and second pressing members 111 and 112 so that the holder 114 is moved. The cell sheet 4 is pressed from the side.
By moving the cell clump sheet 4 in the cell clump sheet moving step, the culture solution enters between the cell clump sheet 4 and the culture vessel 103. Can be supplied.
In the present embodiment, the cell mass sheet 4 in the holder 114 can be moved in the X direction and the Y direction by the first and second pressing members 111 and 112, but the first pressing member 111 or the second pressing member 112 can be moved. The moving means 115 may be connected to only one of them to move only one of the X direction and the Y direction.

FIG. 18 illustrates a culture vessel 103 that can be used in the cell mass sheet manufacturing apparatus 1 according to the second embodiment, and illustrates a variation of the culture vessel 103 according to the second embodiment.
Both ends of the first pressing member 111 are provided so as to be expandable and contractable by an urging force of a spring or the like. When the first pressing member 111 is extended, the guide unit G is retracted by the guide retreating step. In this state, it has the width of the cell mass 2 in which gaps are formed.
Therefore, by moving the first pressing member 111 to the forward position, the cell mass 2 is pressed in the Y direction, and the gap in the Y direction in the aligned cell mass 2 can be eliminated.
Since the first pressing member 111 is expandable and contractible, the second pressing member 112 can be positioned in an approaching state while the first pressing member 111 is positioned at the forward position.
Then, the second pressing member 112 also presses the first pressing member 111 while pressing the cell mass 2, whereby the first pressing member 111 can be contracted.
Thereby, the cell mass 2 can be aligned in a state where the first and second pressing members 111 and 112 are both located at the forward position, and when the second pressing member 112 is located at the advanced position, the cell mass 2 It is possible to prevent the displacement in the Y direction.

The contents of the first and second embodiments can be appropriately combined.
For example, the culture vessel 103 in the second embodiment is housed in the culture vessel 3 instead of the culture tray 3 in the first embodiment, and a through-hole is formed in the bottom of the culture vessel 103 and provided on the bottom of the culture vessel 3. If the pin 14 is penetrated, the pin 14 can be used as the guide means G.
In this case, since the culture vessel 103 is provided with the pressing means 105, it is not necessary to bring the cell mass 2 accommodated in the accommodation portion Ga of the guide means G into contact with each other. The cell mass 2 can be accommodated in the accommodating portion Ga formed by the above, and if the pressing unit 105 is then operated to perform a pressing step, the cell mass 2 can be aligned in a contact state.
According to such a configuration, the culture solution is supplied to the back side of the cell mass sheet 4 using the through holes provided in the culture vessel 103 as supply holes, so that the cells located on the back side can be cultured well. The cell clump sheet can be pushed up from the culture vessel 103 by the pins.
That is, in the cell mass sheet moving step, it is not necessary to move the cell mass sheet in the lateral direction as in the second embodiment, so that the first and second pressing members 111 and 112 are moved to the forward position in the pressing process. The holder 114 can be housed in the incubator while being kept, and in this case, the mounting of the holder 114 can be omitted.

FIG. 20 illustrates a culture vessel 3 used in the method for producing a cell mass sheet according to the third embodiment, and can be used, for example, in the cell mass sheet manufacturing apparatus 1 used in the first embodiment.
The culture vessel 3 of this embodiment has a box shape with a bottom, and its bottom surface constitutes a mounting surface 13a of the cell mass 2, and the mounting surface 13a has cell non-adhesiveness as in the above-described embodiment. .
The guide means G of the present embodiment is composed of a support frame 201 provided in a lattice shape and pins 202 provided downward from intersections of the lattice, and the intersections of the lattice and the positions of the pins 202 Are provided in the same arrangement as the pins 14 shown in FIG. 4 in the first embodiment, and the four pins 202 constitute a housing portion Ga for housing the cell mass 2.
By arranging the pins 202 in this manner, when the cell mass 2 is accommodated in the accommodation part Ga, the cell mass 2 can be aligned in a state where the adjacent cell masses 2 are in contact with each other.
Here, the size of each lattice constituting the support frame 201 is set slightly smaller than the diameter of the cell mass 2 because the pins 202 are provided in the above arrangement, but the size of the suction nozzle 8 This makes it possible to push the cell mass 2 from above.
Then, as in the above-described embodiment, when the cell mass 2 is accommodated in the accommodation portion Ga of the guide device G, the guide device G may be immediately taken out as a guide retreating step, and the culture of the cell mass 2 proceeds to some extent. May be evacuated.
Further, the cell container 3 of this embodiment is provided with a lid 203 on the upper part thereof, but the pipe 31 of the culture solution supply means 11 (not shown) is provided on the lid 203 as in the above-described embodiment. , A culture solution is supplied.

In each of the above embodiments, the cell mass 2 is arranged in a plane on the culture tray to culture the single-layer cell mass sheet 4, but the cell mass 2 is placed on the culture tray 3 in the accommodation step. If a plurality of cells are stacked, it is possible to culture a thicker cell mass sheet 4.
Further, in each of the above embodiments, one cell mass 2 is accommodated in the accommodating portion Ga of the guide means G, but a plurality of, for example, about 2 to 4 cells are accommodated in one accommodating portion Ga. A predetermined number of cell clusters 2 may be accommodated, and it is only necessary that each of the accommodation sections Ga individually accommodate a predetermined number of cell clusters 2.

DESCRIPTION OF SYMBOLS 1 Cell mass sheet manufacturing apparatus 2 Cell mass 3 Culture container 4 Cell mass sheet 5 Storage container 8 Suction nozzle 9 Nozzle moving means 10 Inspection means 11 Culture solution supply means 12 Cell mass sheet moving means 13 Culture tray 13a Mounting surface 13b Through hole 13c supply hole 14 pin 19 holding lid G guide means Ga accommodating part

Claims (2)

A cell mass sheet culturing apparatus for arranging a plurality of cell masses in a plane on a mounting surface in a culture vessel and producing a cell mass sheet in which the cell masses are cultured and fused with each other,
A cell mass sheet culturing device, comprising a pressing means for laterally moving the cell mass being cultured on a mounting surface . The cell clump sheet culture apparatus according to claim 1, wherein the culture vessel is housed in an incubator for culturing the cell clump.

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