JP7341716B2 - Measurement cover member - Google Patents

Description

The present invention relates to a measuring cover member.

In recent years, attempts have been made to transplant various cells to repair damaged tissues and the like. For example, use of fetal cardiomyocytes, skeletal myoblasts, mesenchymal stem cells, cardiac stem cells, ES cells, iPS cells, etc. to repair myocardial tissue damaged by ischemic heart diseases such as angina pectoris and myocardial infarction. has been attempted (Non-Patent Document 1).

As part of such efforts, cell structures formed using scaffolds and sheet-like cell cultures formed by forming cells into sheets have been developed (see, for example, Patent Documents 1 to 3).

In order to transplant such a sheet-shaped cell culture to the target affected area (transplant site), for example, the end of the sheet-shaped cell culture is pinched with tweezers, etc., or scooped out with a spatula, etc. A series of operations are required, such as removing the sheet-shaped cell culture from the packaging container, transporting it to the affected area, and transplanting (affixing) it to the affected area, but the absolute physical strength of the sheet-shaped cell culture is low; Since wrinkles, tears, and damage are likely to occur, this series of operations requires a high degree of skill and great care.

Patent Document 1 describes a system that makes it possible to easily peel off a sheet-like cell culture in a culture container without causing contamination, wrinkles, tears, damage, etc. , by integrating it with a culture incubator, it is possible to automate the process from culture sheet creation to peeling.

Special Publication No. 2007-528755 Japanese Patent Application Publication No. 2010-81829 Japanese Patent Application Publication No. 2010-226991 Japanese Patent Application Publication No. 2011-115080

Haraguchi et al., Stem Cells Transl Med. 2012;1(2):136-41

The system described above performs a series of operations inside the storage compartment, including culturing cells while controlling the temperature, applying vibration to the culture container to peel off the sheet-shaped cell culture, and measuring its state with a sensor. However, when a temperature adjustment or vibration process is performed within the storage section, measurements by the sensor may be affected.

The present invention has been made in view of the above-mentioned problems, and it is an object of the present invention to provide a means for optimizing measurements in a culture device.

[1] A device for measuring a target object in a culture container, which includes a tray part and a holder part provided in the tray part, and the holder part positions and holds the culture container in the tray part. and the holder portion has an opening provided in the tray portion.
[2] The device according to [1], which includes a plurality of holder sections provided in the tray section and is capable of integrally positioning and holding a plurality of culture vessels.
[3] The device according to [1] or [2], further comprising a sample holder portion having smaller dimensions than the holder portion.

[4] The device according to any one of [1] to [3], further including a stopper portion disposed inside the opening.
[5] The device according to any one of [1] to [4], further including a handle portion disposed along the frame portion of the tray portion surrounding the holder portion.
[6] The device according to [5], further comprising a through hole passing through the tray part and the handle part in a direction perpendicular to the plane of the tray part.
[7] The device according to any one of [1] to [6], further including a fixing member provided with a notch portion into which the tray portion fits.
[8] The device according to any one of [1] to [7], wherein the object is a sheet-shaped cell culture.

[9] A cover member for measuring a target object in a culture container, wherein the culture container is housed in a culture device that includes a storage section, a substrate, and a sensor section, and the sensor section is attached to the culture container on the substrate. The cover member is a cylindrical member extending between the sensor section and the substrate.
[10] The cover member according to [9], wherein the cover member is configured to expand in diameter from the proximal end toward the distal end.
[11] The cover member according to [9] or [10], wherein the base end side of the cover member is configured to surround the sensor section.
[12] The cover member according to any one of [9] to [11], wherein the distal end side of the cover member has an outer shape that matches the arrangement of the culture container on the substrate.

[13] The cover member according to any one of [9] to [12], wherein the cover member is configured such that a lighting can be placed inside the cover member.
[14] The cover member according to any one of [9] to [13], wherein the cover member has an outer shape that matches the detection range of the sensor unit.
[15] A plurality of sensor sections are arranged in the housing section, and the outer shape of the cover member is configured to match the outer shape of the detection ranges of the plurality of sensor sections, [9] to [14] The cover member described in any one of.
[16] The cover member according to any one of [9] to [15], wherein the object is a sheet-shaped cell culture.

According to the present invention, measurements in a culture device can be optimized.

FIG. 1 is a conceptual diagram showing an example of use of a device according to a first embodiment of the present invention. FIG. 2 is a perspective view of the tray section according to the first embodiment of the invention. FIG. 3 is a top view of the fixing member according to the first embodiment of the present invention. FIG. 4 is a conceptual diagram of a cover member according to a second embodiment of the present invention. FIG. 5 is a conceptual diagram of the culture apparatus in the present invention.

Hereinafter, the present invention will be described in detail based on preferred embodiments with reference to the drawings. Note that the size of each member in the drawings is appropriately emphasized for the purpose of explanation, and does not indicate the actual proportion or size.

First Embodiment First, a first embodiment of the present invention will be described. FIG. 1 is a conceptual diagram showing an example of use of a device according to a first embodiment of the present invention. FIG. 2 is a perspective view of the tray section according to the first embodiment of the invention. FIG. 3 is a top view of the fixing member according to the first embodiment of the present invention.

FIG. 1 shows a conceptual diagram of a culture apparatus 3 using a device 1. The culture device 3 includes a storage section 31, a substrate 32, a sensor section 33, a temperature control section (not shown), a CO ₂ control section (not shown), and a shaking control section (not shown). The housing unit 31 can function as an incubator that houses the culture container D and maintains the internal temperature and CO ₂ concentration at values suitable for culture using a temperature control unit and a CO ₂ control unit. In this embodiment, the object to be measured is a sheet-like cell culture S, and the sensor unit 33 will be described as measuring the state of the sheet-like cell culture S cultured within the culture container D.

When using a temperature-responsive culture dish (for example, see JP-A-2-211865) as the culture container D, the sheet-like cell culture S can be cultured by lowering the temperature inside the storage part 31 using the temperature control part. It can be peeled off from the container D. The substrate 32 is a plate-shaped member that divides the inside of the storage section 31 into an upper/lower space, and the culture container D can be placed on it. In one aspect, the substrate 32 can be shaken (e.g., horizontal vibration, rocking, horizontal eccentric vibration, horizontal transverse vibration, etc.) by a shaking control unit, thereby peeling the sheet-like cell culture from the culture container D. You can also do that.

The state of the sheet-like cell culture S measured by the sensor section 33 includes, for example, the distance between the sensor section 33 and the sheet-like cell culture S, the refractive index, transmittance, reflectance, etc. of the sheet-like cell culture S. can be mentioned. From the viewpoint of data versatility and processing diversity, it is preferable that the adhesion state of the sheet-shaped cell culture S is acquired as image data. In this embodiment, the sensor section 33 is arranged in the lower space. The explanation will be given assuming that the image capturing unit is an image capturing unit that photographs (measures) the inside of the culture container D from below the substrate 32.

FIG. 1B is a diagram of the substrate 32 in FIG. 1A viewed from above the culture device 3. The substrate 32 is made of a light-transmitting material, and the sensor section 33 measures the state of the sheet-like cell culture S in the culture container D placed on the substrate 32 from below the substrate 32. be able to. In the device 1, a plurality of culture vessels D can be integrally arranged on the substrate 32. As shown in the figure, the device 1 arranges three culture vessels D in a substantially triangular shape according to the detection range of the sensor unit 33 (in this embodiment, the angle of view indicated by the dotted line in FIG. 1A). be able to.

As shown in FIG. 2, the device 1 includes a tray part 11 and a holder part 12. The tray section 11 functions as a tray on which a plurality of culture containers D can be placed and transported. The tray section 11 is provided with a plurality of holder sections 12. The holder part 12 has a function of positioning and holding the culture container D in the tray part 11. The holder part 12 positions the culture container D in the tray part 11 by suitable means such as steps, protrusions, restraints, adhesion, and magnetic force.

The holder part 12 can include an opening provided in the tray part 11. By designing the opening to have dimensions equal to or larger than the dimensions (for example, diameter) of the culture vessel D, the culture vessel D can be accommodated therein. By supporting the bottom edge of the culture container D by the stopper portion 13 disposed inside the opening, the culture container D can be positioned and held in the tray portion 11.

As illustrated, by providing a step between the stopper part 13 and the opening, the holder part 12 can be configured to position and hold the culture container D. The stopper part 13 only needs to be able to support at least the bottom edge of the culture container D, and a plurality of stopper parts 13 can be arranged at intervals in the circumferential direction of the holder part 12. Furthermore, by designing the stopper portion 13 to protrude by a length that does not block the sheet-like cell culture S in the culture container D, the sheet-like cell culture S can be suitably measured. . The stopper portion 13 can also be made of a light-transmitting material.

In another embodiment, the stopper portion 13 may be configured as a plate-shaped member made of a light-transmitting material. The bottom surface of the culture container D can also be stably supported by placing the stopper section 13 inside the opening, for example by pasting it on the bottom surface of the tray section 11. The opening may be designed to have a size that does not block the sheet-like cell culture S in the culture container D, that is, to be at least larger than the size of the sheet-like cell culture S. In this way, since the tray section 11 has an opening that facilitates observation of the inside of the culture container D, the object inside the culture container D can be accurately measured from above and below. This is particularly advantageous when photographing sheet-like cell cultures that are translucent and have low visibility.

As illustrated, the tray section 11 can be configured in a substantially triangular shape in which three holder sections 12 for holding three culture vessels D are provided, for example. Moreover, the tray part 11 can further be provided with a sample holder part 12' having a smaller dimension (for example, diameter) than the holder part 12. The sample holder part 12' can hold a sample culture container D' having smaller dimensions than the culture container D, for example. The sheet-like cell culture cultured in the culture container D' can also be used as a test sample for quality control. This allows the amount of cells used as a test sample to be minimized.

The device 1 may further include a handle portion 14 that functions as a handle when moving the tray portion 11. The handle part 14 can be connected to a frame part surrounding the holder parts 12, 12' of the tray part 11. As illustrated, the handle part 14 is connected to a plurality of positions of the tray part 11 and is provided so as to bridge between the plurality of connection parts along the frame part, so that observation of the culture container D is not blocked. It can be configured as follows. As illustrated, by making the center of gravity of the handle portion bridged between the plurality of connection portions correspond to the center of gravity of the tray portion 11, the plurality of culture vessels D can be stably lifted.

The device 1 may further include a through hole 15 passing through the tray part 11 and the handle part 14 in a direction perpendicular to the plane of the tray part 11, as shown. For example, by observing the two through-holes 15, it is possible to identify the center axis (in the case of this embodiment, the optical axis) of the sensor section 33 indicated by the arrow in the figure and the center passing through the center of the two through-holes 15. Misalignment with the axis can be detected. Thereby, it is possible to perform distortion correction (keystone correction, etc.) on the photographed image, and it is possible to accurately grasp the state of the sheet-shaped cell culture. Furthermore, the inclination and position of the tray portion 11 can also be accurately detected. Positioning can also be performed by providing a protrusion on the substrate 32 into which the through hole 15 fits.

As another aspect, the through hole 15 is made into a polygonal shape and designed so that the extension line of the center point of the through hole 15 and each vertex intersects with the center point of each holder part 12, so that the image of the through hole 15 is It is also possible to configure so that the position of each holder part 12 can be detected. With this, for example, when an image taken by the sensor unit 33 is analyzed by computer processing, the range to be analyzed on the image (i.e., the position of the sheet-shaped cell culture S) can be quickly specified. , the analysis process can be made more efficient.

As shown in FIG. 3, the device 1 may further include a fixing member 16 having a notch 17 cut out to match the outer shape of the tray portion 11. The fixing member 16 is a plate-shaped member adapted to the dimensions of the substrate 32 in FIG. can be fixed to the substrate 32. Thereby, the analysis process can be made more efficient, and furthermore, when the substrate 32 is shaken, it is possible to configure the culture container D to be appropriately applied with force.

Although the device 1 according to the first embodiment of the present invention has been described above, the present invention is not limited thereto. The external shape of the tray section 11 is determined according to the dimensions of the storage section 31, the number of sensor sections 33, the detection range of the sensor section 33, the dimensions of the culture container D, the number of culture containers D and D', the shape of the substrate 32, etc. Can be changed freely. For example, the tray part 11 can also be configured in a diamond shape in which all six culture vessels D shown in FIG. 1B can be arranged integrally.

Conventionally, when performing image analysis in randomly arranged culture vessels, especially in a plurality of culture vessels, it is first necessary to detect the position of each culture vessel. Since the culture vessels D can be positioned integrally, the analysis process can be made more efficient. In addition, conventionally, when multiple culture vessels are shaken, their positional relationships become inconsistent, but the device 1 of the present invention can fix these positional relationships, making analysis processing easier. It can be made more efficient.

In the present embodiment, two sensor sections 33 are arranged in the lower space of the storage section 31 to measure the inside of the culture container D from below the tray section 11, but the sensor sections 33 are arranged above the substrate 32. It is also possible to configure the culture container D so that it can be measured from above. The detection range of the sensor section 33 is determined by the distance between the sensor section 33 and the substrate 32, the detection angle, etc., so the number of sensor sections 33 (one, three or more may be used) depending on the area of the culture container to be measured. ), distance, detection angle, etc. can also be changed.
As described above, according to the device 1 of the present invention, measurements in the culture apparatus can be optimized.

Second Embodiment Next, a second embodiment of the present invention will be described.
FIG. 4 is a conceptual diagram showing an example of use of the cover member 2 according to the second embodiment of the present invention. In each figure in this application, the size of each member is appropriately emphasized for ease of explanation, and the illustrated member does not indicate its actual size.
In addition, in the drawings, the same components as those of the device 1 according to the first embodiment are given the same reference numerals, and the differences from the first embodiment will be explained in detail below, and similar matters will be explained in detail. , the explanation is omitted.

As shown in FIG. 4, the cover member 2 is a cylindrical member that surrounds the sensor section 33 and extends between the sensor section 33 and the substrate 32. As shown in FIG. The cover member 2 expands in diameter from the proximal end 21 side (sensor section 33 side) toward the distal end 22 side (substrate 32 side) so as not to block the detection range (angle of view, etc.) of the sensor section 33. It is configured as follows. That is, when the sensor section 33 is an imaging section, the cover member 2 has a shape like a lens hood and protects the lens from external light and dust. Conventional lens hoods extend from the periphery of the lens, but the cover member 2 of the present invention also has the function of protecting the sensor section 33 itself by surrounding the sensor section 33 on the base end 21 side. Fulfill.

As described above, the culture device 3 can function as an incubator that houses the culture container D and maintains the internal temperature and CO ₂ concentration at values suitable for culture using the temperature control section and the CO ₂ control section. . In the figure, for the sake of simplification, only the top surface of the storage section 31 is shown, but for example, the top surface may be configured to be openable and closable so that the culture container D can be taken in and taken out. Further, it is preferable that the upper end of the cover member 2 and the substrate 32 are not connected to each other and are provided with a slight distance so that the substrate 32 can be shaken for peeling off the sheet-like cell culture as described above.

When culturing cells in the housing section 31, the temperature inside the housing section 31 is normally maintained at 37°C. At this time, the liquid in the culture container D may evaporate and generate moisture. In addition, it is necessary to accommodate equipment such as the sensor section 33, temperature control section, CO ₂ control section, shaking control section, and wiring in the housing section 31, and if such wiring blocks the detection range of the sensor section 33, There is. On the other hand, the cover member 2 of the present invention can isolate the detection range of the sensor section 33 from moisture and shielding objects, so that measurements in the culture device 3 can be optimized. In addition, since the cover member 2 is tapered in diameter from the distal end 22 toward the proximal end 21, a space for arranging equipment and wiring can be secured within the accommodating portion 31.

Cell culture is usually carried out manually by workers with specialized knowledge in large-scale facilities called cell culture centers (CPCs) where cleanliness is maintained, and the manufacturing costs and expenses of such facilities are low. The effort is enormous. Therefore, in one embodiment, for example, an air filter unit (not shown) that combines a filter such as a HEPA filter and a fan is incorporated into the culture apparatus 3 to maintain the cleanliness inside the storage section 31 and to perform a series of operations. It can also be configured as a closed system so that everything is automated.

As another embodiment, the culture device 3 can be designed so that it can be used in relatively inexpensive, small-scale equipment, such as a clean bench where cleanliness is maintained. When the housing section 31, which is the casing of the culture device 3, is downsized, for example, the number of culture vessels D that can be installed on the substrate 32 is limited, the distance between the sensor section 33 and the substrate 32 is shortened, and the equipment and Various problems need to be taken into account, such as the difficulty of securing space to accommodate the wiring.

When the distance between the sensor section 33 and the substrate 32 is short, for example, as shown in FIG. 1, the number of culture vessels D that can be measured within the detection range of the sensor section 33 is three. However, four or more sheet-shaped cell cultures may be used in one treatment. Therefore, as shown in FIGS. 1 and 4, two sensor units 33 are arranged side by side in the storage unit 31, and a total of six (one is a spare) sheet-shaped cell culture S can be cultured. It can also be configured as follows.

When two sensor sections 33 are arranged side by side, the two tray sections 11 arranged on the substrate 32 can be measured separately, as shown in FIG. 1B. In this case, the outer shape of the tip 22 of the cover member 2 is set to be approximately elliptical (in this embodiment, approximately It is preferable to configure (into a shape). Similarly, as shown in FIG. 4, the base end 21 of the cover member 2 is preferably formed into a substantially elliptical shape to match the two sensor sections 33 arranged side by side.

In this way, the outer shape of the cover member 2 is preferably configured to match the outer shape of the detection ranges (angles of view, etc.) of the plurality of sensor sections 33 connected together. Moreover, when the substrate 32 is shaken in the left-right direction in FIG. 1B, when the shaking is stopped, the tray part 11 may stop moving to either the left or right. Therefore, it is preferable that the tip 22 of the cover member 2 has a shape that matches the shaking direction and the moving distance (amplitude) of the substrate 32 (in the case of this embodiment, the above-mentioned elliptical shape is further elongated laterally).

As described above, since the sheet-shaped cell culture is translucent and has low visibility, lighting for photographing can also be placed inside the housing section 31. When the substrate 32 is made of a light-transmitting material, the inside of the culture container D can be photographed from either above or below the substrate 32. Regarding the arrangement of illumination, those skilled in the art can select the optimal position for photographing the sheet-shaped cell culture according to the arrangement of the substrate 32, sensor section 33, tray section 11, culture container D, etc.

When photographing a target object, it is common to arrange the illumination and the imaging unit so that the target object is sandwiched between them. Therefore, the lighting and the sensor section 33 can be arranged so that the substrate 32 is sandwiched between them. Furthermore, the upper surface of the accommodating portion 31 may be made of a light-transmissive material so that images can be taken using external light instead of illumination. When culturing in the upper space of the culture container D, condensation may occur, so it is preferable to arrange lighting in the lower space. In this case, the cover member 2 can also be configured so that the illumination can be placed inside thereof so that the light from the illumination is not blocked by the cover member 2.
As described above, according to the cover member 2 of the present invention, measurements in the culture device can be optimized.

Although the cover member 2 according to the second embodiment of the present invention has been described above, the present invention is not limited thereto. The outer shape of the cover member 2 is based on the detection range of the sensor section 33, the number of sensor sections 33, the distance between the sensor section 33 and the substrate 32, the shaking direction and movement distance of the substrate 32, the area and arrangement of the culture container that is the measurement target, It can be changed as appropriate depending on the object to be measured, the intensity of illumination, etc. For example, as described above, the tray section 11 can be configured in a diamond shape, and the outer shape of the cover member 2 can be configured so that six culture vessels D can be measured with one sensor section 33.

The storage section 31 of the culture device 3 can be configured to have a substantially rectangular parallelepiped shape as shown in FIG. Then, an input/output unit (a touch panel may be used) such as a monitor or an operation unit is incorporated into the front side of the storage unit 31 to operate each device, monitor the status of each device, and perform measurements using the sensor unit 33 (for example, image ) can also be configured. Control of temperature, CO ₂ , shaking, etc. can also be performed manually while checking measurements (for example, images) by the sensor unit 33 on a monitor.

In the present invention, each configuration can be replaced with any one that can perform the same function, or any configuration can be added.

1 Device 11 Tray part 12 Holder part 13 Stopper part 14 Handle part 15 Through hole 16 Fixing member 17 Notch part 2 Cover member 21 Base end 22 Tip 3 Culture device 31 Storage part 32 Substrate 33 Sensor part D Culture container S Sheet cell culture

Claims (8)

A cover member for measuring a target object in a culture container,
The culture container is housed in a culture device including a storage part, a substrate, and a sensor part,
The sensor unit can measure the object in the culture container on the substrate,
The cover member is a cylindrical member extending between the sensor part and the substrate,
The cover member expands in diameter from the sensor side toward the board side,
The substrate can be shaken, and the outer shape of the cover member is configured to surround a moving distance of the culture container on the substrate in the shaking direction.
The cover member. The cover member according to claim 1 , wherein the cover member has an elliptical horizontal cross section . The cover member according to claim 1 or 2, wherein the sensor portion side of the cover member is configured to surround the sensor portion. The cover member according to any one of claims 1 to 3, wherein the substrate side of the cover member has an outer shape that matches the arrangement of the culture container on the substrate. A cover member according to any one of claims 1 to 4, wherein the cover member is configured such that a light can be arranged inside the cover member. The cover member according to any one of claims 1 to 5, wherein the cover member has an outer shape that matches the detection range of the sensor unit. Any one of claims 1 to 6, wherein a plurality of sensor sections are arranged in the housing section, and the outer shape of the cover member is configured to match the outer shape of connecting detection ranges of the plurality of sensor sections. The cover member described in. The cover member according to any one of claims 1 to 7, wherein the object is a sheet-shaped cell culture.