JPWO2005090548A1 - Incubator holding device - Google Patents

Abstract

PROBLEM TO BE SOLVED: To facilitate handling of an incubator in a culture device which applies stress to cells in the incubator. The present invention relates to a first holding member and a second holding member on which an incubator is suspended, the first holding member and the second holding member being detachable from a main body of a culture device. There is provided a holding device for an incubator, comprising: a connecting tool that connects the first holding member and the second holding member.

Description

  TECHNICAL FIELD The present invention relates to an improvement of a holding device for holding an incubator in a culture device equipped with an incubator for culturing sample cells while generating stress.

It is known that when stress is applied to a sample cell and the cell is cultured, the stress stimulates the cell to undergo a specific change.
Conventionally, methods for applying stress to cultured sample cells have been proposed.
For example, Patent Documents 1 to 3 disclose techniques for deforming the bottom membrane of a container on which sample cells are placed by deforming the bottom membrane to generate stress on the sample cells.

The present inventors consider that it is difficult to apply uniform stress to the sample cells by the method of deforming the bottom membrane of the incubator with a suction device, and therefore the pulling force in the direction parallel to the bottom membrane of the incubator is applied to the incubator. The study has been advanced by paying attention to the addition type culture device (see Non-Patent Document 1).
In this culture device, as shown in FIG. 1, for example, a sample cell is placed on a transparent and deformable culture membrane 3 having a thickness of 200 μm coated with fibronectin, and a pair of opposite sides of the culture membrane 3 (thickness The side walls 5 and 5 are provided (see Non-Patent Document 1). The thick side wall 5 is used for uniformly extending the culture film 3 on which the sample cells are placed.

In FIG. 1, an engagement hole 7 is formed in the side wall 5.
A pin (not shown) of the extension device 10 is inserted into the engagement hole 7.
As shown in FIG. 2, the extension device 10 includes a fixed plate (first holding member) 11, a movable plate (second holding member) 13, a step motor 15, and a control device 17.
Pins are formed at predetermined positions on the fixed plate 11 and the movable plate 13, and the engagement holes 7 of the incubator 1 are fitted into the pins. As a result, the incubator 1 is suspended between the fixed plate 11 and the movable plate 13. The distance between the movable plate 13 and the fixed plate 11 changes according to the rotation of the step motor 15. As a result, the culture membrane 3 of the incubator 1 expands, and stress is applied to the sample cells. The rotation of the step motor 15 is controlled by the controller 17.

  See also Patent Documents 1 to 6 as related to the present invention.

US Pat. No. 4,789,601 US Pat. No. 4,822,741 US Pat. No. 4,839,280 JP, 2003-61642, A US Pat. No. 6,107,081 International publication WO02/46365 Involvement of SA channels in orienting response of cultured endothelial cells to cyclic stretch, the Journal of Physiology, 1998, H1532-1538, KEIJI NARUSE et al.

When the incubator 1 is set in the incubator shown in FIG. 1, the entire incubator including the step motor 15 and the controller 17 is put into a clean bench, and the fixed plate 11 and the movable plate 13 are incubated therein. I had attached vessel 1. After that, the culture device was moved to an environment suitable for cell culture and driven there, thereby stressing the cells in the incubator 1.
However, since the culture device including the drive unit such as the step motor 15 is large and heavy as a whole, it takes a lot of time to move it.
Further, since the fixed plate and the movable plate are connected only by the incubator 1, there is a risk that the positional relationship between the fixed plate and the movable plate may be irregularly displaced during the movement of the culture device. If such a deviation occurs, the incubator 1 is deformed and the cells in the incubator 1 are subjected to unexpected stress, which is not preferable.
That is, one object of the present invention is to make it easy to handle an incubator in an incubator that applies stress to cells in the incubator.

The present invention has been made to solve at least one of the above problems. That is,
A first holding member and a second holding member on which the incubator is suspended, the first holding member and the second holding member being detachable from the main body of the culture device;
A connector for connecting the first holding member and the second holding member,
An incubator holding device comprising:

According to the incubator holder thus configured, both the first holding member and the second holding member are attachable to and detachable from the incubator body. Therefore, the first holding member and the second holding member can be detached from the incubator main body and placed in the clean bench, and the incubator can be set therein. Then, both holding members with the incubator set are returned to the incubator body. As described above, according to the present invention, since the number of elements to be moved is reduced as much as possible, the handling thereof becomes easy. That is, it takes less time to move.
Further, since the first holding member and the second holding member are connected by the connecting tool, it is possible to prevent the occurrence of irregular deviation in the positional relationship between the two. Therefore, it is possible to prevent unexpected stress from being applied to the cells in the incubator.

In addition, in order to further stabilize the positional relationship between the first holding member and the second holding member, it is preferable to provide a fixture for fixing the two.
Further, a lid for covering at least the opening of the incubator is provided so that foreign matter such as dust does not enter the incubator during movement.

FIG. 1 shows a conventional incubator and its extension device. FIG. 2 is a perspective view of an incubator according to an embodiment of the present invention. FIG. 3 is a perspective view showing the configuration of the holding device of the embodiment. FIG. 4 is a plan view of the same. FIG. 5 is a sectional view showing the structure of a holding device of another embodiment. FIG. 6 is a plan view showing a schematic configuration of the culture device. FIG. 7 is a vertical cross-sectional view showing the schematic structure of the culture device. FIG. 8 is a front view of the culture device.

Embodiment of the invention

Hereinafter, each element of the present invention will be described in detail.
The culture device main body is provided with a first holding member seat and a second holding member seat for detachably fixing the first holding member and the second holding member. Further, a mechanism for changing the relative position of each holding member seat is provided. Although the first holding member seat is fixed and the second holding member seat is movable in the embodiment, both holding member seats may be movable.
The culture device main body can be made suitable for cell culture when the first holding member and the second holding member are attached and the atmosphere of both holding members is arbitrarily controlled to apply stress to them. .. In order to create the atmosphere, the culture device body is equipped with a temperature controller and a humidity controller.
Furthermore, cells can be incubated in the incubator by providing both holding members with an atmosphere filled with CO ₂ gas.

(First holding member and second holding member)
Engagement portions that engage with the incubator are formed on the first holding member and the second holding member. In the embodiment, a pin is erected from each holding member as this engaging portion. By fitting the engagement hole of the incubator to this pin, the incubator is suspended between the first holding member and the second holding member. The shape and structure of the engaging portion are not particularly limited as long as a sufficient force can be applied to stably deform the incubator.
The first holding member and the second holding member are detachably fixed to the holding member seat of the apparatus body. This facilitates handling of both holding members. In the embodiment, the holes of the respective holding members are fitted into the pins erected from the holding member seat, and further hooked so that the respective holding members and the holding member seat are detachably fixed. The mode of fixing the both is not limited to the structure of the embodiment, and a clip, screw, or other known structure can be adopted.
The shape, structure and material of the holding member are not particularly limited as long as they can stably hold the incubator. Although the plate-shaped one is adopted in the embodiment, it is also possible to use a rod-shaped one, a block-shaped one and the like. The number of incubators set on the holding member can be arbitrarily selected.

(Connector)
The first holding member and the second holding member are connected by a connecting tool. This connecting tool prevents at least both holding members from coming apart. This facilitates handling of the device. Furthermore, it is preferable that the connecting tool regulates the relative distance fluctuation between the two. As a result, it is possible to prevent the gap between the both holding members from being irregularly displaced during the movement and to give stress to the cells in the incubator.
As described above, in the embodiment as the connecting tool, the rod is expanded from the first holding member and the second holding member is slidably fitted to the rod. The structure of the connecting tool is not limited to this, and at least one of both holding members may be slidably attached to the rail as the connecting tool. Further, the connecting tool may be one in which both holding members are fixed and connected. For example, both holding members are clipped with a clip as a connecting tool. Further, it is possible to form the engaging holes in both the holding members, and to hang the connecting rod through the engaging holes so that the holding members can be fixed and connected. By fixing and connecting both holding members, deformation of the incubator can be reliably prevented.

(Lid)
A lid is used to cover the incubator set between the first holding member and the second holding member. Thus, it is possible to prevent dust and the like from entering the incubator when the device is moved (that is, when the incubator is moved).
The lid preferably has a length substantially equal to that of both holding members. As a result, it becomes possible to coat all the incubators set on both holding members.
The shape and material of the lid can be arbitrarily selected according to the target incubator. For example, a flat plate can be used as the lid.

(Incubator)
The incubator is capable of culturing cells and is capable of deforming itself and exerting a desired stress on the culturing cells. In addition to the known example shown in FIG. 1, the rectangular box type shown in FIG. 2 can be used.
Such an incubator is made of a deformable material. This is because the sample cells are indirectly stressed by extending the incubator.
As a material for forming the incubator, a material such as silicone elastomer that does not chemically interfere with sample cells is used.
It is preferable that the bottom film 23 has a rectangular shape in plan view and is formed to have the same film thickness as a whole so as to be uniformly extended. The bottom film 23 is preferably formed of a light-transmitting material so that the sample cells can be observed with an optical microscope.
It is preferable that the side wall is provided upright from the entire circumference of the bottom film. The side wall is made of a thick film to give it mechanical rigidity. This can prevent the bottom film from being deformed randomly.
It is preferable that the bottom film and the side wall are integrated from the viewpoint of reducing the number of components and, further, the manufacturing cost, but the bottom film and the side wall are not limited to be separate bodies.
Engagement holes 27 are provided in the side walls 25 provided at a pair of opposite edges of the rectangular bottom film. This engaging portion may engage with the first holding member and the second holding member. As a result, the position change of the two members whose relative positions can be controlled deforms the incubator.
Therefore, it is also possible to provide a protrusion from the side wall and engage it with each holding member.
When the bottom membrane is stretched, the peripheral edge 26 in the stretching direction is deformed so that the central portion is depressed. When such deformation occurs, the direction of the stress applied to the sample cell on the deformed portion differs from that of the other portions.
Therefore, the engagement hole 27 is formed on the extension line of the peripheral edge 26 in the extension direction of the rectangular bottom film. As a result, the force applied to the engaging portion is applied to the peripheral edge 26 in the extending direction of the bottom membrane, and this is reliably pulled. Therefore, the deformation of the peripheral edge 26 is prevented, so that the uniform stress can be applied to all the sample cells on the bottom membrane 23.
The type of sample cells used in the incubator 21 of this embodiment and the method of obtaining them are not particularly limited. For example, vascular endothelial cells (human, monkey, pig, cow, rat, mouse, rabbit, etc.), smooth muscle cells, cardiomyocytes, skeletal muscle cells, fibroblasts, osteoblasts, chondrocytes, osteoclasts, nerve cells Etc. can be used.

Hereinafter, the present invention will be described in more detail based on examples.
FIG. 3 is a perspective view showing the holding device 30 of this embodiment. FIG. 4 also shows a plan view. The holding device 30 of the embodiment is roughly composed of a fixed plate 31 as a first holding member, a movable plate 41 as a second holding member, a connector 50 and a lid 60.
A through hole 33 is formed in one long edge of the fixed plate 31. A pin 35 is provided upright on the other long edge. The engaging hole 27 of the incubator 21 shown in FIG. 2 is fitted into the pin 35.

A through hole 43 is also formed on one long edge of the movable plate 41, and a pin 45 is provided upright on the other long edge.
The incubator 21 is stably suspended between the fixed plate 31 and the movable plate 41 by fitting the engagement holes 27 of the incubator 27 into the pins 35 and 45 of the plates 31 and 41 in a forcefully fitted state. It As a result, the incubator 27 can be deformed in relation to the change in the distance between the fixed plate 31 and the movable plate 41. Therefore, it is possible to apply a desired stress to the cells in the incubator 27 by controlling the distance between the plates.

Two connecting rods 51 as connecting tools are fixed to the fixed plate 31. The movable plate 41 is slidably fitted to the connecting rod 51. By this connecting rod 51, the positional change of the movable plate 41 with respect to the fixed plate 31 is restricted to only one (the axial direction of the connecting rod 51). Therefore, it is possible to prevent irregular movement of the fixed plate 31 and the movable plate 41 in the state where the incubator 21 is set as shown in FIG.
Further, as shown in FIG. 5, holes 37 and 47 are formed on the side surfaces of the fixed plate 31 and the movable plate 41, and a fixing rod 55 as a fixing tool is fitted into the holes 37 and 47, thereby The plates 31 and 41 can be fixed. Thereby, the deformation of the incubator 21 can be completely prevented, and the cells in the incubator 21 can be prevented from being irregularly stressed.
Reference numeral 53 in the figure is a stopper that prevents the movable plate 41 from falling off.

  The lid 60 covers from above the four incubators 21 set on the plates 31 and 41. This prevents foreign matter such as dust from entering the incubator 21 even when the incubator 21 is taken out from the clean bench. The lid 60 is preferably made of a lightweight material such as resin for easy handling during transportation. Further, it is preferable that the incubator 21 is made of a transparent material so that the state of the incubator 21 can be observed. In this embodiment, the lid 61 is made of transparent acrylic resin.

The holding device 30 is set in the device body 71 of the culture device 70 as shown in FIGS. 6 and 7.
Here, the culture device 70 will be described. The culture device 70 is composed of the holding device 30 and the device body 71 described above. The apparatus main body 71 is provided with four fixed plate seats 75 for detachably fixing the fixed plate 31 and four movable plate seats 77 for detachably fixing the movable plate 41. In FIG. 6, the holding device 30 is set only on the upper two sets of the fixed plate seat 75 and the movable plate seat 77.
The movable plate seat 77 is moved by the step motor 79. Reference numeral 81 is a rail for moving the movable plate 77, and reference numeral 83 is a control device for controlling the rotation of the step motor 79. The stepper motor 79 converts the rotational movement of the shaft 85 into the axial movement of the rod 89 by the converter 87. A movable plate seat 77 is connected to the rod 89. Therefore, the movable plate seat 77 moves in the left-right direction in FIGS. 6 and 7 as the step motor 79 rotates.
This controls the distance between the fixed plate seat 75 and the movable plate seat 77, and thus controls the distance between the fixed plate 31 and the movable plate 41. That is, the control device 83 and the step motor 79 constitute a gap control means between the fixed plate 31 and the movable plate 41.

Positioning pins 91 and 92 are erected on the fixed plate seat 75 and the movable plate seat 77, and are fitted into the through holes 33 of the fixed plate 31 and the through hole 43 of the movable plate 41, respectively. Hooks 93 and 94 are rotatably attached to the fixed plate seat 75 and the movable plate seat 77, respectively. The hook 93 rotates so as to press the upper surface of the fixed plate 31 placed on the fixed plate seat 75. Similarly, the hook 94 rotates so as to press the upper surface of the movable plate 41 placed on the movable plate seat 77. The plates 31, 41 are stably fixed to the plate seats 75, 77 by the positioning pins 91, 92 and the hooks 93, 94, respectively. The plates 31 and 41 can be detached from the plate seats 75 and 77 by rotating the hooks 93 and 94 by 90 degrees. That is, the hooks 93 and 94 constitute a detachable means, whereby the fixed plate 31 and the movable plate 41 are detachably attached to the culture device body 71.
The method of detachably fixing the fixed plate and the movable plate to each plate seat is not limited to the above. For example, a clip separate from both may be prepared and the plate and the plate seat may be fixed by the clip.

In the device body 71 of the culture device 70, the chamber portion 100 in which the holding device 30 is set becomes airtight by closing the lid 101. A temperature controller 104 and a humidity controller 105 are provided in the chamber portion 100. Furthermore, it is possible to control to 5% carbon dioxide and 100% humidity.
FIG. 8 shows a front view of the culture device 70. The operator inputs the extension condition of the incubator 21 (that is, the stress condition applied to the cell) via the input device 110. Also, the environmental conditions (temperature, etc.) of the chamber portion 100 are input. Based on the input, the control device 83 controls the rotation of the step motor 79 and also controls the temperature controller 104 and the humidity controller 105. Reference numeral 111 in the figure is a display.

According to the culture device 70 of the embodiment configured as described above, by opening and closing the hooks 93 and 94, the fixed plate 31 and the movable plate 41 can be attached to and detached from the fixed plate seat 75 and the movable plate seat 77, respectively. Fixed. That is, the holding device 31 can be attached to and detached from the device body 71. This allows the incubator 21 to be moved independently of the device main body 71 with the incubator 21 set in the holding device 31. Therefore, handling of the incubator 21 after being set on the fixed plate 31 and the movable plate 41 becomes easy. Since the positional change between the fixed plate 31 and the movable plate 41 is regulated by the connecting rod 51, irregular deformation of the incubator 21 is prevented. As a result, unnecessary stress is not applied to the cells in the incubator 21 during movement. Further, by using the lid body 61, it is possible to prevent foreign matter from mixing into the cells in the incubator 21 during movement.
As described above, the culture device 70 of this embodiment has excellent operability.

Claims (4)

A first holding member and a second holding member on which the incubator is suspended, the first holding member and the second holding member being detachable from the culture device body;
A connector for connecting the first holding member and the second holding member,
An incubator holding device comprising: The holding device according to claim 1, further comprising a lid that covers the first holding member and the second holding member from above. The holding device according to claim 1 or 2, further comprising a fixture that fixes the positions of the first holding member and the second holding member. A culture device main body having means for controlling a distance between the first holding member and the second holding member on which the incubator is suspended;
Means for detachably attaching the first holding member and the second holding member to the culture device body,
A connector for connecting the first holding member and the second holding member,
A culture device comprising:

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