JP2020184893A - Cover member for measurement - Google Patents

Abstract

To provide means for optimization of measurement in a culture device.SOLUTION: The present invention provides a cover member for measuring an object in a culture vessel, where the culture vessel is stored in a culture device that includes a storage part, a substrate and a sensor part, the sensor part can measure an object in the culture vessel on the substrate, and the cover member is a cylindrical member extending between the sensor part and the substrate.SELECTED DRAWING: None

Description

The present invention relates to a measurement cover member.

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

As a part of such an attempt, a cell structure formed by using a scaffold and a sheet-like cell culture in which cells are formed in a sheet shape 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 (transplantation site), for example, the end of the sheet-shaped cell culture is picked with a tweezers or scooped with a spatula or the like. A series of operations such as taking out the sheet-shaped cell culture from the packaging container, transferring it to the affected area, and transplanting (pasting) it to the affected area are required, but the sheet-shaped cell culture has low absolute physical strength. Since wrinkles, tears, and breaks are likely to occur, a high level of skill is required for this series of operations, and great care must be taken.

Patent Document 1 describes a system capable of easily peeling a sheet-shaped cell culture in a culture vessel without causing contamination, wrinkles, tearing, breakage, or the like. By integrating with the culture incubator, it is possible to automate the process from sheet preparation to peeling by culture.

Special Table 2007-528755 Japanese Unexamined Patent Publication No. 2010-81829 JP-A-2010-226991 Japanese Unexamined Patent Publication No. 2011-11580

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

In the above system, cells are cultured while controlling the temperature, the culture vessel is vibrated to peel off the sheet-shaped cell culture, and the state is measured by a sensor, and a series of operations are performed in the storage unit. However, when the temperature control or vibration process is performed in the storage unit, the measurement by the sensor may be affected.

The present invention has been made in view of the above problems, and an object of the present invention is to provide a means for optimizing measurement in a culture apparatus.

[1] A device for measuring an object in a culture vessel, which includes a tray portion and a holder portion provided on the tray portion, and the holder portion positions and holds the culture vessel in the tray portion. The device, wherein the holder portion has an opening provided in the tray portion.
[2] The device according to [1], which includes a plurality of holder portions provided on the tray portion and can integrally position and hold a plurality of culture vessels.
[3] The device according to [1] or [2], further including a holder portion for a sample having a size smaller than that of the holder portion.

[4] The device according to any one of [1] to [3], further including a stopper portion arranged inside the opening.
[5] The device according to any one of [1] to [4], further including a handle portion, which is arranged along a frame portion of a tray portion surrounding the holder portion.
[6] The device according to [5], further including a through hole, which penetrates the tray portion and the handle portion in a direction perpendicular to the surface of the tray portion.
[7] The device according to any one of [1] to [6], further comprising a fixing member, provided with a notch for fitting the tray portion.
[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 an object in a culture container, the culture container is housed in a culture apparatus including a storage part, a substrate and a sensor part, and the sensor part is a culture container on the substrate. The cover member is a tubular member that can measure an object inside and extends between the sensor portion and the substrate.
[10] The cover member according to [9], wherein the cover member is configured to increase in diameter from the proximal end side to the distal end side.
[11] The cover member according to [9] or [10], wherein the base end side of the cover member is configured to surround the periphery of the sensor portion.
[12] The cover member according to any one of [9] to [11], wherein the tip end side of the cover member has an outer shape suitable for the arrangement of the culture vessel on the substrate.

[13] The cover member according to any one of [9] to [12], wherein the cover member is configured so that lighting can be arranged 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 units are arranged in the accommodating portion, and the outer shape of the cover member is configured to match the outer shape connecting the detection ranges of the plurality of sensor units. [9] to [14] The cover member according to 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, the measurement in the incubator can be optimized.

FIG. 1 is a conceptual diagram showing a usage example of the device according to the first embodiment of the present invention. FIG. 2 is a perspective view of the tray portion according to the first embodiment of the present 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 according to the present invention.

Hereinafter, the present invention will be described in detail with reference to the drawings, based on preferred embodiments. The size of each member in the figure is emphasized as appropriate for explanation, and does not indicate the actual ratio or size.

First Embodiment First, the first embodiment of the present invention will be described. FIG. 1 is a conceptual diagram showing a usage example of the device according to the first embodiment of the present invention. FIG. 2 is a perspective view of the tray portion according to the first embodiment of the present 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 the device 1. The culture device 3 includes a housing unit 31, a substrate 32, a sensor unit 33, a temperature control unit (not shown), a CO ₂ control unit (not shown), and a shaking control unit (not shown). The storage unit 31 can function as an incubator that houses the culture vessel D and keeps the internal temperature and CO ₂ concentration at values suitable for culture by the temperature control unit and the CO ₂ control unit. In the present embodiment, the object to be measured is the sheet-shaped cell culture S, and the sensor unit 33 will be described as measuring the state of the sheet-shaped cell culture S cultured in the culture vessel D.

When a temperature-responsive culture dish (see, for example, Japanese Patent Application Laid-Open No. 2-211865) is used as the culture vessel D, the sheet-shaped cell culture S is cultured by lowering the temperature inside the storage unit 31 with the temperature control unit. It can be peeled off from the container D. The substrate 32 is a plate-shaped member that divides the inside of the accommodating portion 31 into an upper / lower space, and the culture container D can be placed on the plate-shaped member. In one embodiment, the substrate 32 can be shaken (eg, horizontal vibration, rocking, horizontal eccentric vibration, horizontal lateral shaking, etc.) by the shaking control unit, thereby peeling the sheet-shaped cell culture from the culture vessel D. You can also do it.

The state of the sheet-shaped cell culture S measured by the sensor unit 33 includes, for example, the distance between the sensor unit 33 and the sheet-shaped cell culture S, the refractive index, the transmittance, and the reflectance of the sheet-shaped cell culture S. Can be mentioned. From the viewpoint of versatility of data and versatility of processing, it is preferable that the adhesive state of the sheet-shaped cell culture S is acquired as image data, and in the present embodiment, the sensor unit 33 is arranged in the lower space. This is an imaging unit, and the inside of the culture vessel D will be photographed (measured) from below the substrate 32.

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

As shown in FIG. 2, the device 1 includes a tray portion 11 and a holder portion 12. The tray unit 11 functions as a tray on which a plurality of culture containers D can be placed and transported. The tray portion 11 is provided with a plurality of holder portions 12. The holder portion 12 has a function of positioning and holding the culture vessel D in the tray portion 11. The holder portion 12 positions the culture vessel D in the tray portion 11 by suitable means such as steps, protrusions, restraints, adhesions, and magnetic forces.

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

As shown in the drawing, by providing a step between the stopper portion 13 and the opening, the culture vessel D can be configured as a holder portion 12 for positioning and holding the culture vessel D. The stopper portions 13 need only be able to support at least the bottom edge portion of the culture vessel D, and a plurality of stopper portions 13 can be arranged at intervals in the circumferential direction of the holder portion 12. Further, the stopper portion 13 is designed so as to protrude by a length that does not shield the sheet-shaped cell culture S in the culture vessel D, so that the sheet-shaped cell culture S can be suitably measured. .. The stopper portion 13 can also be made of a light-transmitting material.

In another aspect, the stopper portion 13 can also be configured as a plate-shaped member made of a light-transmitting material. Then, by arranging the stopper portion 13 inside the opening by, for example, attaching it to the lower surface of the tray portion 11, the bottom surface of the culture vessel D can be stably supported. The opening may be designed to have a size that does not shield the sheet-shaped cell culture S in the culture vessel D, that is, at least larger than the size of the sheet-shaped cell culture S. As described above, since the tray portion 11 has an opening that facilitates observation inside the culture vessel D, the object in the culture vessel D can be accurately measured from above and below. This is particularly advantageous when imaging sheet-like cell cultures that are translucent and have low visibility.

As shown, the tray portion 11 can be configured, for example, in a substantially triangular shape provided with three holder portions 12 for holding the three culture vessels D. Further, the tray portion 11 may be further provided with a holder portion 12'for a sample having a size (for example, a diameter) smaller than that of the holder portion 12. The sample holder portion 12'can hold, for example, the culture vessel D'for samples having a size smaller than that of the culture vessel D. Then, the sheet-shaped cell culture cultured in the culture vessel D'can also be used as a test sample for quality control. As a result, the amount of cells used as a test sample can be minimized.

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

As shown in the figure, the device 1 may further include a through hole 15 that penetrates the tray portion 11 and the handle portion 14 in a direction perpendicular to the surface of the tray portion 11. Then, for example, by observing the two through holes 15, the central axis (in the case of the present embodiment, the optical axis) of the sensor unit 33 indicated by the arrow in the figure and the center passing through the centers of the two through holes 15 The deviation from the axis can be detected. As a result, distortion correction (trapezoidal correction, etc.) of the captured image can be performed, and the state of the sheet-shaped cell culture can be accurately grasped. In addition, this makes it possible to accurately detect the inclination and position of the tray portion 11. Positioning can also be performed by providing a protrusion on the substrate 32 for fitting the through hole 15.

As yet another embodiment, the through hole 15 has a polygonal shape, and the extension line between the center point of the through hole 15 and each vertex is designed so that the center point of each holder portion 12 intersects with the image of the through hole 15. Therefore, it can be configured so that the position of each holder portion 12 can be detected. Thereby, for example, when the image taken by the sensor unit 33 is analyzed by computer processing or the like, the range to be analyzed on the image (that is, 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 fit the outer shape of the tray 11. The fixing member 16 is a plate-shaped member conforming to the dimensions of the substrate 32 of FIG. 1B, and by fixing this to the substrate 32 and fitting the tray portion 11 into the notch portion 17, the culture container D Can be fixed to the substrate 32. As a result, the analysis process can be made more efficient, and further, when the substrate 32 is shaken, a force can be appropriately applied to the culture vessel D.

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 outer shape of the tray portion 11 is adjusted to the dimensions of the accommodating portion 31, the number of sensor portions 33, the detection range of the sensor portion 33, the dimensions of the culture vessel D, the number of culture vessels D and D', the shape of the substrate 32, and the like. You can change it freely. For example, the tray portion 11 can be configured in a rhombus shape in which all six culture vessels D shown in FIG. 1B can be integrally arranged.

Conventionally, when performing image analysis in randomly arranged culture vessels, particularly in a plurality of culture vessels, it is necessary to first detect the position of each culture vessel, but the device 1 of the present invention is a culture vessel D, particularly a plurality of culture vessels D. Since the culture vessel D of the above can be integrally positioned, the analysis process can be made more efficient. Further, conventionally, when a plurality of culture containers are shaken, their positional relationships become disjointed. However, since the device 1 of the present invention can fix these positional relationships, the analysis process is performed. It can be made more efficient.

In the present embodiment, two sensor units 33 are arranged in the lower space of the accommodating unit 31, and the inside of the culture vessel D is measured from below the tray unit 11, but the sensor unit 33 is arranged above the substrate 32. Therefore, the culture vessel D can be configured so that it can be measured from above. Since the detection range of the sensor unit 33 is determined by the distance between the sensor unit 33 and the substrate 32, the detection angle, etc., the number of sensor units 33 (one or three or more may be used) according to the area of the culture vessel to be measured. ), Distance, detection angle, etc. can be changed.
As described above, according to the device 1 of the present invention, the measurement in the culture apparatus can be optimized.

Second Embodiment Next, the second embodiment of the present invention will be described.
FIG. 4 is a conceptual diagram showing a usage example of the cover member 2 according to the second embodiment of the present invention. In each drawing of the present application, the size of each member is appropriately emphasized for ease of explanation, and each member in the drawing does not indicate an actual size.
Further, in the drawings, the same configurations as those of the device 1 according to the first embodiment are designated by the same reference numerals, and the differences from the first embodiment will be described in detail below. , The description is omitted.

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

As described above, the culture apparatus 3 can function as an incubator that houses the culture vessel D and keeps the internal temperature and CO ₂ concentration at values suitable for culture by the temperature control unit and the CO ₂ control unit. .. In the figure, for simplification, only the upper surface portion of the accommodating portion 31 is displayed, but for example, the upper surface portion may be opened and closed so that the culture container D can be taken in and out. Further, it is preferable that the upper end portion of the cover member 2 is not connected to the substrate 32 and a slight distance is provided so that the substrate 32 can be shaken for peeling of the sheet-shaped cell culture as described above.

When culturing cells in the containment section 31, the temperature inside the containment section 31 is usually maintained at 37 ° C. At this time, the liquid in the culture vessel D may evaporate to generate moisture. Further, when it is necessary to house equipment and wiring such as a sensor unit 33, a temperature control unit, a CO ₂ control unit, and a shaking control unit in the accommodating unit 31, such wiring shields the detection range of the sensor unit 33. There is. On the other hand, since the cover member 2 of the present invention can isolate the detection range of the sensor unit 33 from moisture and a shield, the measurement in the culture device 3 can be optimized. Further, since the cover member 2 has a reduced diameter from the tip end 22 toward the base end 21, it is possible to secure a space for arranging equipment and wiring in the accommodating portion 31.

Cell cultures are usually manufactured manually in large, clean facilities called cell culture centers (CPCs) by skilled workers, and the cost of manufacturing such facilities The effort is enormous. Therefore, in one embodiment, for example, an air filter unit (not shown), which is a combination of a filter such as a HEPA filter and a fan, is incorporated into the culture apparatus 3, and the cleanliness inside the accommodating portion 31 is maintained, and a series of operations is performed. Can also be configured as a closed system so that everything is automated.

In another aspect, the incubator 3 can be designed to be used in a relatively inexpensive, small-scale facility, such as a clean bench that maintains cleanliness. When the storage unit 31 which is the housing of the culture device 3 is miniaturized, for example, the number of culture containers D that can be installed on the substrate 32 is limited, the distance between the sensor unit 33 and the substrate 32 is shortened, the device or It is necessary to consider various problems such as difficulty in securing a space for accommodating wiring.

When the distance between the sensor unit 33 and the substrate 32 is short, for example, as shown in FIG. 1, the number of culture containers D that can be measured within the detection range of the sensor unit 33 is three. However, 4 or 5 or more sheet-shaped cell cultures may be used in one treatment. Therefore, as shown in FIGS. 1 and 4, two sensor units 33 can be arranged side by side in the accommodating 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 units 33 are arranged side by side, as shown in FIG. 1B, the tray units 11 arranged on the substrate 32 can be measured separately. In this case, the outer shape of the tip 22 of the cover member 2 is substantially elliptical in the case of the present embodiment so as to match the arrangement of the culture vessel D on the substrate 32 as shown by reference numeral 22 (white line) in FIG. 1B. It is preferable to configure it (in shape). Similarly, as shown in FIG. 4, the base end 21 of the cover member 2 is preferably formed in a substantially elliptical shape in accordance with the two sensor portions 33 arranged side by side.

As described above, it is preferable that the outer shape of the cover member 2 is configured to match the outer shape in which the detection ranges (angles of view, etc.) of the plurality of sensor units 33 are connected. Further, when the substrate 32 is shaken in the left-right direction in FIG. 1B, when the shaking is stopped, the tray portion 11 may be stopped in a state of being moved to the left or right. Therefore, it is preferable that the tip 22 of the cover member 2 is formed in a shape that matches the shaking direction of the substrate 32 and the moving distance (amplitude) (in the case of the present embodiment, the elliptical shape is further elongated).

As described above, since the sheet-shaped cell culture is translucent and has low visibility, lighting for photographing can be arranged in the accommodating portion 31. When the substrate 32 is made of a light-transmitting material, the inside of the culture vessel D can be photographed from either above or below the substrate 32. Those skilled in the art can select the optimum position for photographing the sheet-shaped cell culture according to the arrangement of the substrate 32, the sensor unit 33, the tray unit 11, the culture vessel D, and the like with respect to the arrangement of the illumination.

When photographing an object, it is common to arrange the illumination and the imaging unit so as to sandwich the object. Therefore, the substrate 32 can be arranged so as to sandwich the substrate 32 between the illumination and the sensor unit 33. Further, the upper surface portion of the accommodating portion 31 may be made of a light-transmitting material, and external light may be used instead of illumination for shooting. When culturing the culture vessel D in the upper space, dew condensation may occur, so it is preferable to arrange the lighting in the lower space. In this case, the cover member 2 may be configured so that the illumination can be arranged inside the cover member 2 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, the measurement in the culture apparatus 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 includes the detection range of the sensor unit 33, the number of sensor units 33, the distance between the sensor unit 33 and the substrate 32, the shaking direction and moving distance of the substrate 32, the area and arrangement of the culture vessel to be measured, and so on. It can be changed as appropriate according to the object to be measured, the intensity of illumination, and the like. For example, as described above, the tray portion 11 may be formed in a diamond shape, and the outer shape of the cover member 2 may be configured so that the six culture vessels D can be measured by one sensor portion 33.

The accommodating portion 31 of the incubator 3 can be configured to have a substantially rectangular parallelepiped shape as shown in FIG. Then, an input / output unit (which may be a touch panel) such as a monitor or an operation unit is incorporated in the front side surface of the accommodating unit 31, and operation of each device, monitoring of the state of each device, and measurement by the sensor unit 33 (for example, an image). ) Can be displayed. Control of temperature, CO ₂ , shaking, etc. can also be performed manually while checking the measurement (for example, image) by the sensor unit 33 on the monitor.

In the present invention, each configuration can be replaced with any configuration capable of exerting 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-like cell Culture

Claims (8)

A cover member for measuring an object in a culture vessel.
The culture vessel is housed in a culture device, including a storage unit, a substrate and a sensor unit.
The sensor unit can measure the object in the culture vessel on the substrate,
The cover member is a tubular member extending between the sensor portion and the substrate.
The cover member. The cover member according to claim 1, wherein the cover member is configured to increase in diameter from the proximal end side to the distal end side. The cover member according to claim 1 or 2, wherein the base end side of the cover member is configured to surround the periphery of the sensor portion. The cover member according to any one of claims 1 to 3, wherein the tip end side of the cover member has an outer shape suitable for the arrangement of the culture vessel on the substrate. The cover member according to any one of claims 1 to 4, wherein the cover member is configured so that lighting 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 portions are arranged in the accommodating portion, and the outer shape of the cover member is configured to conform to the outer shape connecting the detection ranges of the plurality of sensor parts. 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.