JP2020099211A - Cell culture container - Google Patents

Abstract

To provide a cell culture container which does not interfere with observation of cultured cells and accurately measures an electric resistance of the cells.SOLUTION: A cell culture container 1 comprises: a cell culture chamber 21; a cell culture film 20 partitioning the inside of the cell culture chamber 21; a channel connecting the outside and the cell culture chamber 21 in communication; a first electrode chamber 22 and a second electrode chamber 23 in direct communication with the cell culture chamber 21; a first action electrode 31 and a first reference electrode 32 arranged in the first electrode chamber 22; a second action electrode 41 and a second reference electrode 42 arranged in the second electrode chamber 23. The first electrode chamber 22 is arranged on an upper side of the cell culture film 20, and the second electrode chamber 23 is arranged on a lower side of the cell culture film 20. At least part of each of the first electrode chamber 22 and the second electrode chamber 23 does not overlap a cell culture region 200 which is a portion of the cell culture film 20 that lies within the cell culture chamber 21, in a vertical direction.SELECTED DRAWING: Figure 1

Description

The present invention relates to a cell culture container for culturing cells inside and measuring the electric resistance of the cultured cells.

Techniques for measuring the electrical resistance of cells in order to investigate the properties of the cultured cells and the state of culture are known. For example, in transepithelial electrical resistance (TEER) measurement, cells cultured in a membrane form are measured by disposing electrodes on one side and the other side of a cell culture membrane in a culture medium and measuring the electrical resistance value. Measure the electric resistance of itself. A technique for measuring the electric resistance value of a cell is described in Patent Document 1, for example.

On the other hand, the development of so-called micro flow channel devices is in progress. In the micro-channel device, the cell culture space is closed by removing the minute flow passages, so that external factors can be eliminated as much as possible for cell culture. A technique for measuring the electric resistance value of cells in such a microchannel device is described in, for example, Patent Document 2.

International Publication No. 2012/147463 Japanese Patent Publication No. 2017-513483

In the microchannel device described in Patent Document 2, since the cell culture region and the electrode are arranged at a position where they overlap, it is difficult to observe the entire cultured cells. On the other hand, when the electrodes are arranged at positions apart from the cell culture region, the electric resistance between the electrodes increases, and the accuracy of measuring the electric resistance of the cells themselves may decrease.

The present invention has been made in view of such circumstances, and an object of the present invention is to provide a cell culture container that does not hinder the observation of cultured cells and that can accurately measure the electrical resistance of the cells. ..

In order to solve the above problems, the first invention of the present application is a cell culture container for culturing cells inside and measuring the electrical resistance of the cultivated cells, which is a cell culture chamber capable of storing a culture solution. A cell culture membrane that divides the inside of the cell culture chamber into an upper space and a lower space between the bottom surface and the top surface of the cell culture chamber, and directly or indirectly between the outside and the cell culture chamber. A first electrode chamber that directly communicates with the cell culture chamber above the cell culture membrane, and a second electrode chamber that directly communicates with the cell culture chamber below the cell culture membrane. A first working electrode and a first reference electrode arranged in the first electrode chamber, and a second working electrode and a second reference electrode arranged in the second electrode chamber. At least a part of each of the one electrode chamber and the second electrode chamber does not vertically overlap with a cell culture region which is a part of the cell culture membrane inside the cell culture chamber.

2nd invention of this application is the cell culture container of 1st invention, Comprising: The minimum cross-sectional area of the space between said 1st working electrode and said cell culture area, and the space between said 2nd working electrode and said cell culture area. The minimum sectional area of is larger than the maximum channel area of the channel.

3rd invention of this application is a cell culture container of 1st invention or 2nd invention, Comprising: The housing|casing which comprises the said cell culture chamber is transparent in the position which overlaps with at least one part of the said cell culture area|region in the up-down direction. It has a portion formed of a material.

4th invention of this application is the cell culture container in any one of 1st invention thru|or 3rd invention, Comprising: The surface area in the said 1st electrode chamber of the said 1st working electrode is the said 1st electrode of the said 1st reference electrode. It is larger than the surface area in the chamber, and the surface area of the second working electrode in the second electrode chamber is larger than the surface area of the second reference electrode in the second electrode chamber.

5th invention of this application is a cell culture container in any one of 1st invention-4th invention, Comprising: The said 1st electrode chamber is a 1st direction orthogonal to an up-down direction with respect to the said cell culture chamber. And the second electrode chamber is disposed on the other side in the first direction with respect to the cell culture chamber.

According to the first to fifth inventions of the present application, it is possible to prevent the electrode from overlapping the cell culture region and obstructing the observation. In addition, it is possible to prevent the measurement accuracy of the electric resistance of the cell from decreasing due to the increase in the electric resistance between the electrodes.

Particularly, according to the second invention of the present application, the influence of the external environment in the cell culture chamber can be reduced by reducing the flow passage area of each flow passage. Further, it is easy to control the inflow/outflow amount of the culture solution or the reagent into/from the cell culture chamber. On the other hand, by increasing the minimum cross-sectional area of the space between each electrode and the cell culture region, the electrical resistance of the culture medium between each electrode and the cell culture region is reduced. Thereby, the electrical resistance of the cell can be measured more accurately.

Particularly, according to the third invention of the present application, it is easy to observe the state of the cells cultured in the cell culture region.

Particularly, according to the fifth invention of the present application, the electric resistance of the culture solution between the electrodes becomes smaller. Thereby, the electrical resistance of the cell can be measured with higher accuracy.

It is a longitudinal cross-sectional view of the cell culture container according to the first embodiment. It is a layout view of the cell culture container according to the first embodiment in a top view. It is an exploded perspective view of the cell culture container concerning a 1st embodiment. It is the schematic which showed the electrical connection of the measurement unit which concerns on 1st Embodiment. It is a longitudinal cross-sectional view of the cell culture container according to the second embodiment. It is a layout view of the cell culture container according to the second embodiment in a top view. It is an exploded perspective view of the cell culture container concerning a 2nd embodiment. It is an arrangement view in a top view of a cell culture container according to a modification. FIG. 8 is a layout view of a cell culture container according to another modification in a top view.

Embodiments of the present invention will be described below with reference to the drawings. In the following description, the direction parallel to the bottom surface of the cell culture container is referred to as “horizontal direction”, and the direction orthogonal to the horizontal direction is referred to as “vertical direction”. However, the posture of the cell culture container during use may not be such that the bottom surface of the cell culture container is horizontal.

<1. First Embodiment>
<1-1. Structure of cell culture vessel>
FIG. 1 is a vertical cross-sectional view of a cell culture container 1 according to the first embodiment of the present invention. FIG. 2 is a layout view showing the layout of each part in a top view of the cell culture container 1. FIG. 3 is an exploded perspective view of the cell culture container 1.

The cell culture container 1 is a container for culturing cells inside and measuring the electric resistance of the cultured cells. The cell culture container 1 is a so-called micro flow channel device in which the internal space becomes a closed space except for the fine flow channels 24, 25, 26, 27.

As shown in FIG. 1 and FIG. 2, the cell culture container 1 has a cell culture chamber 21, a first electrode chamber 22, a second electrode chamber 23, a first upper flow path 24, a second upper channel 24, and an inner space of the main body 10. It has an upper channel 25, a first lower channel 26, and a second lower channel 27. That is, the main body 10 is a housing that constitutes the cell culture chamber 21. Further, the cell culture container 1 has a first working electrode 31, a first reference electrode 32, a second working electrode 41, and a second reference electrode 42.

In addition, as shown in FIGS. 1 and 3, the main body 10 of the cell culture container 1 includes a resin-made first member 11, a second member 12, a third member 13, a fourth member 14, and a fourth member which are vertically overlapped with each other. The fifth member 15 and the sixth member 16 and the cell culture membrane 20. The main body part 10 is a first member 11, a second member 12, a third member 13, a cell culture membrane 20, a fourth member 14, a fifth member 15, and a sixth member 16 in this order from the bottom to the top. Composed of overlapping.

The cell culture chamber 21 is arranged in the center of the cell culture container 1. In the present embodiment, the cell culture chamber 21 is formed by the upper surface of the first member 1, the lower surface of the sixth member 16, and the inner surfaces of the second member 12 to the fifth member 15. A culture solution can be stored in the cell culture chamber 21. The cell culture membrane 20 is vertically spaced from each of the bottom surface and the top surface of the cell culture chamber 21. As a result, the cell culture membrane 20 partitions the inside of the cell culture chamber 21 into an upper space and a lower space between the top surface and the bottom surface of the cell culture chamber 21.

The cell culture film 20 is a film having cell adhesiveness. For the cell culture membrane 20 of the present embodiment, for example, a thin film called a membrane, which has many fine through holes, is used. As the material of the membrane, for example, PC (polycarbonate), PTFE (polytetrafluoroethylene), PET (polyethylene terephthalate), etc. are used. The surface of the membrane may be coated with collagen or the like to enhance cell adhesiveness. A portion of the cell culture membrane 20 exposed in the cell culture chamber 21 becomes a cell culture region 200. In the present embodiment, the portion other than the cell culture region 200 of the cell culture membrane 20 is covered with the third member 13 and the fourth member 14, and thus is not exposed. Instead of the cell culture membrane 20, a plate-shaped member having cell adhesiveness and provided with a large number of fine through holes may be used.

The first electrode chamber 22 directly communicates with the cell culture chamber 21 above the cell culture membrane 20. The first electrode chamber 22 is arranged on one side in the first direction with respect to the cell culture chamber 21. In the present embodiment, the upper surface of the fourth member 14, the lower surface of the sixth member 16, and the inner surface of the fifth member 15 form the first electrode chamber 22.

The second electrode chamber 23 directly communicates with the cell culture chamber 21 below the cell culture membrane 20. The second electrode chamber 23 is arranged on the other side in the first direction with respect to the cell culture chamber 21. In the present embodiment, the second electrode chamber 23 is formed by the upper surface of the first member 11, the lower surface of the third member 13, and the inner surface of the second member 12. Further, the first electrode chamber 22 and the second electrode chamber 23 do not vertically overlap with the cell culture region 200.

The “first direction” is a direction parallel to the bottom surface of the cell culture container 1 and orthogonal to the vertical direction. 1 and 2, the first direction is the left-right direction of the paper surface, one side of the first direction is the right side of the paper surface, and the other side of the first direction is the left side of the paper surface. The “second direction” is a direction parallel to the bottom surface of the cell culture container 1 and orthogonal to the first direction. In FIG. 2, the second direction is the vertical direction of the paper surface, one side of the second direction is the upper side of the paper surface, and the other side of the second direction is the lower side of the paper surface.

The first upper channel 24 and the second upper channel 25 are channels that communicate the outside of the cell culture container 1 and the cell culture chamber 21 above the cell culture membrane 20, respectively. One end of the first upper flow path 24 directly communicates with the cell culture chamber 21, and the other end opens to the end surface of the cell culture container 1 on the one side in the second direction. One end of the second upper flow path 25 directly communicates with the cell culture chamber 21, and the other end opens to the end surface of the cell culture container 1 on the other side in the second direction. In this embodiment, the upper surface of the fourth member 14, the lower surface of the sixth member 16, and the inner surface of the fifth member 15 form a first upper flow path 24 and a second upper flow path 25.

The first lower flow channel 26 and the second lower flow channel 27 are flow channels that connect the outside of the cell culture container 1 and the cell culture chamber 21 below the cell culture membrane 20, respectively. One end of the first lower channel 26 directly communicates with the cell culture chamber 21, and the other end opens to the end surface of the cell culture container 1 on the one side in the second direction. One end of the second lower flow path 27 is in direct communication with the cell culture chamber 21, and the other end is open to the end surface of the cell culture container 1 on the other side in the second direction. In the present embodiment, the first lower channel 26 and the second lower channel 27 are formed by the upper surface of the first member 11, the lower surface of the third member 13, and the inner surface of the second member. In the present embodiment, the first upper flow path 24 and the first lower flow path 26 are vertically overlapped with each other, and the second upper flow path 25 and the second lower flow path 27 are vertically overlapped with each other. Will be placed.

The first working electrode 31 and the first reference electrode 32 are arranged in the first electrode chamber 22. Specifically, the first working electrode 31 and the first reference electrode 32 are fixed to the top surface of the first electrode chamber 22. The first working electrode 31 and the first reference electrode 32 are formed, for example, by applying an electrode paste, which is a material of the electrode, to the lower surface of the sixth member 16 that constitutes the top surface of the first electrode chamber 22, and then drying it. It In addition, the first working electrode 31 and the first reference electrode 32 may be those in which a metal is vapor-deposited on the top surface of the first electrode chamber 22, or a plate-shaped electrode may be a top surface of the first electrode chamber 22. It may be adhered to.

The second working electrode 41 and the second reference electrode 42 are arranged in the second electrode chamber 23. Specifically, the second working electrode 41 and the second reference electrode 42 are fixed to the bottom surface of the second electrode chamber 23. The second working electrode 41 and the second reference electrode 42 are formed, for example, by applying an electrode paste, which is a material of the electrode, on the upper surface of the first member 11 that forms the bottom surface of the second electrode chamber 23, and then drying it. .. The second working electrode 41 and the second reference electrode 42 may be those in which a metal is vapor-deposited on the bottom surface of the second electrode chamber 23, or a plate-shaped electrode is bonded to the bottom surface of the second electrode chamber 23. It may be the one.

Thus, by arranging at least a part of the first electrode chamber 22 and the second electrode chamber 23 for arranging the electrodes in a position not vertically overlapping the cell culture region 200, the electrodes 31, 32, 41 are formed. , 42 can be prevented from overlapping the cell culture region 200 and obstructing the observation.

Further, by providing the first electrode chamber 22 and the second electrode chamber 23 at positions adjacent to the cell culture chamber 21, it is possible to prevent the distance between the cell culture region 200 and each electrode 31, 32, 41, 42 from increasing. it can. Therefore, it is possible to prevent the electric resistance between the working electrodes 31 and 41 and the reference electrodes 32 and 42 from increasing and the measurement accuracy of the electric resistance of the cell from decreasing.

In this embodiment, the minimum cross-sectional area of the space between the first working electrode 31 and the cell culture region 200 is larger than the maximum channel area of each channel 24-27. Further, the minimum cross-sectional area between the second working electrode 41 and the cell culture region 200 is larger than the maximum flow channel area of each of the flow channels 24 to 27. As described above, by reducing the flow passage area of each of the flow passages 24 to 27, the influence of the external environment in the cell culture chamber 21 can be reduced. Further, it is easy to control the inflow/outflow amount of the culture solution or the reagent into/from the cell culture chamber 21. On the other hand, the larger the minimum cross-sectional area of the space between each electrode 31, 32, 41, 42 and the cell culture region 200, the larger the culture medium between each electrode 31, 32, 41, 42 and the cell culture region 200. Has a smaller electric resistance. As a result, the electric resistance of the cells cultured on the cell culture region 200 can be measured more accurately.

In addition, in the present embodiment, the first electrode chamber 22 is arranged on one side in the first direction with respect to the cell culture chamber 21, and the second electrode chamber 23 is arranged on the other side in the first direction with respect to the cell culture chamber 21. It That is, the first electrode chamber 22 and the second electrode chamber 23 are arranged at positions facing each other across the cell culture chamber 21. As a result, the first working electrode 31, the cell culture region 200, and the second working electrode 41 are arranged on a substantially straight line. Further, the first reference electrode 32, the cell culture region 200, and the second reference electrode 42 are arranged in a substantially straight line. Therefore, the electrical resistance of the culture solution between the first working electrode 31 and the second working electrode 41 and the electrical resistance of the culture solution between the first reference electrode 32 and the second reference electrode 42 become smaller. Thereby, the electric resistance of the cells cultured on the cell culture region 200 can be measured with higher accuracy.

As shown in FIG. 2, in the present embodiment, the surface area of the first working electrode 31 inside the first electrode chamber 22 is larger than the surface area of the first reference electrode 32 inside the first electrode chamber 22. The surface area of the second working electrode 41 in the second electrode chamber 23 is larger than the surface area of the second reference electrode 42 in the second electrode chamber 23.

In order to increase the electric capacitance on the surfaces of the working electrodes 31, 41, it is preferable to design the surface areas of the working electrodes 31, 41 as large as possible. By arranging the electrodes 31, 32, 41, 42 in the electrode chambers 22, 23, the degree of freedom in designing the electrodes 31, 32, 41, 42 is increased. Therefore, as in the present embodiment, the surface areas of the working electrodes 31, 41 can be designed to be relatively large. Moreover, even when the surface area of the working electrodes 31, 41 is increased by disposing the electrodes 31, 32, 41, 42 in the electrode chambers 22, 23, the electrodes 31, 32, 41, 42 are It does not easily overlap the cell culture region 200 in the vertical direction.

In the present embodiment, each of the members 11 to 16 that form the main body 10 is made of PDMS (polydimethylsiloxane). PDMS has high transparency and low autofluorescence. Therefore, it is easy to observe the cells on the cell culture region 200 from the outside of the main body 10. In the present embodiment, all positions of the main body portion 10 that vertically overlap with the cell culture region 200 are formed of PDMS, which is a transparent material. However, it suffices that the main body portion 10 has a portion formed of a transparent material at a position that vertically overlaps at least a part of the cell culture region 200. For example, if the upper part of the cell culture region 200 is made of a transparent material, the cell culture region 200 can be observed.

<1-2. Measurement of electrical resistance of cultured cells>
Next, a method for measuring the electric resistance of cultured cells using the cell culture container 1 will be described with reference to FIG. FIG. 4 is a schematic diagram showing the electrical connection of the measurement unit 9. Note that FIG. 4 shows the electrical connection of the measurement unit 9 when the electrical resistance measurement is performed on one cell culture container 1. In the actual electrical resistance measurement, the measurement may be performed on a plurality of cell culture vessels 1 at the same time. In that case, for example, the connection destinations of the power supply device 91 and the voltmeter 92 of the measuring device 90 described later can be switched for each cell culture container 1.

As shown in FIG. 4, the measurement unit 9 includes the cell culture container 1 and the culture solution in the cell culture container 1 described above, and a measurement device 90 connected to the cell culture container 1.

The measuring device 90 includes a power supply device 91 and a voltmeter 92. The two output terminals of the power supply device 91 are connected to the first working electrode 31 and the second working electrode 41. The two input terminals of the voltmeter 92 are connected to the first reference electrode 32 and the second reference electrode 42. When measuring the electric resistance of cultured cells, the cell culture container 1 is filled with a culture solution.

In FIG. 4, the resistance Rm is an electrical resistance of the cell culture region 200 and cells cultured on the cell culture region 200 (hereinafter, referred to as “cell portion”). The resistance Rw1 is the electric resistance of the culture solution between the first working electrode 31 and the cell part. The resistance Rw2 is the electric resistance of the culture solution between the second working electrode 41 and the cell part. The resistance Rr1 is the electric resistance of the culture solution between the first reference electrode 32 and the cell part. The resistance Rr2 is the electric resistance of the culture solution between the second reference electrode 42 and the cell part.

The resistance values of the resistances Rw1, Rr1, Rw2, Rr2 between the electrodes 31, 32, 41, 42 and the cell portion and the resistance Rm of the cell culture region 200 in the state where the cells are not cultured are respectively set in advance. Measure as a control.

Then, the power supply device 91 is driven to flow a constant current between the first working electrode 31 and the second working electrode 41. At the same time, the voltmeter 92 measures the voltage value between the first reference electrode 32 and the second reference electrode 42. Since the working electrodes 31 and 41 and the reference electrodes 32 and 42 are arranged sufficiently close to each other, the current flowing between the reference electrodes 32 and 42 and the current flowing between the working electrodes 31 and 41 have the same current value. Can be approximated as Therefore, the electric resistance value between the reference electrodes 32 and 42 is calculated from the voltage value between the reference electrodes 32 and 42. Further, the resistance Rm of the cell part is calculated based on each value measured in advance. Thereby, the electrical characteristics of the cells cultured on the cell culture region 200 can be obtained.

When a potential is applied between the first working electrode 31 and the second working electrode 41 using the power supply device 91, an electric double layer is formed on the electrode surfaces of the first working electrode 31 and the second working electrode 41. It At this time, the output potential of the power supply device 91 is the potential applied to the entire circuit. Therefore, the output potential of the power supply device 91 does not have an accurate voltage value between the working electrodes 31 and 41. Therefore, the first reference electrode 32 and the second reference electrode 42 are arranged near the first working electrode 31 and the second working electrode 41, respectively, and the potential between the reference electrodes 32 and 42 is measured. The resistance Rm of the cell part can be more accurately measured by performing calculation using the measured potential.

<2. Second Embodiment>
Next, a cell culture container 1A according to the second embodiment of the present invention will be described with reference to FIGS. 5 and 6. FIG. 5 is a vertical sectional view of the cell culture container 1A. FIG. 6 is a layout diagram showing the layout of each part in a top view of the cell culture container 1A. FIG. 7 is an exploded perspective view of the main body 10A of the cell culture container 1A.

As shown in FIG. 5 and FIG. 6, the cell culture container 1A has a cell culture chamber 21A, a first electrode chamber 22A, a first electrode chamber 22A, and It has a two-electrode chamber 23A, a first upper channel 24A, a second upper channel 25A, a first lower channel 26A, and a second lower channel 27A. In addition, the cell culture container 1A has a first working electrode 31A, a first reference electrode 32A, a second working electrode 41A, and a second reference electrode 42A.

Further, as shown in FIGS. 5 and 7, the main body 10A of the cell culture container 1A includes a resin-made first member 11A, a second member 12A, a third member 13A, a fourth member 14A, and a fourth member 14A which are vertically overlapped with each other. The fifth member 15A and the sixth member 16A, and the cell culture membrane 20A. The main body 10A includes a first member 11A, a second member 12A, a third member 13A, a cell culture membrane 20A, a fourth member 14A, a fifth member 15A, and a sixth member 16A in this order from the bottom to the top. Composed of overlapping.

The cell culture chamber 21A, the first electrode chamber 22A, the second electrode chamber 23A, the flow paths 24A, 25A, 26A, 27A and the cell culture region 200A are arranged in the cell culture chamber of the cell culture container 1 according to the first embodiment. The arrangement of 21, the first electrode chamber 22, the second electrode chamber 23, the channels 24, 25, 26, 27 and the cell culture region 200 is the same.

Each of the first working electrode 31A, the first reference electrode 32A, the second working electrode 41A, and the second reference electrode 42A is a columnar electrode extending vertically. The main body portion 10A includes two first through holes 51A extending from the upper surface of the main body portion 10A to the top surface of the first electrode chamber 22A, and two second through holes extending from the top surface of the main body portion 10A to the top surface of the second electrode chamber 23A. It has a through hole 52A.

The first working electrode 31A and the first reference electrode 32A are respectively inserted in the first through hole 51A. The upper ends of the first working electrode 31A and the first reference electrode 32A are arranged above the upper surface of the main body 10A. Further, the lower ends of the first working electrode 31A and the first reference electrode 32A are arranged inside the first electrode chamber 22A.

The second working electrode 41A and the second reference electrode 42A are respectively inserted in the second through holes 52A. The upper ends of the second working electrode 41A and the second reference electrode 42A are arranged above the upper surface of the main body 10A. Further, the lower ends of the second working electrode 41A and the second reference electrode 42A are respectively arranged in the second electrode chamber 23A.

In this way, each of the electrodes 31A, 32A, 41A, 42A may be arranged so as to penetrate the main body 10A.

<3. Modification>
Although the main embodiments of the present invention have been described above, the present invention is not limited to the above embodiments.

FIG. 8 is a layout diagram showing the layout of each part in a top view of the cell culture container 1B according to a modification. As shown in FIG. 8, the cell culture container 1B has a cell culture chamber 21B, a first electrode chamber 22B, and a second electrode chamber as internal spaces of the main body 10B, as in the cell culture container 1 according to the first embodiment. 23B, the first upper channel 24B, the second upper channel 25B, the first lower channel 26B, and the second lower channel 27B. Further, the cell culture container 1B has a first working electrode 31B, a first reference electrode 32B, a second working electrode 41B, and a second reference electrode 42B.

The cell culture chamber 21B, the first electrode chamber 22B, the second electrode chamber 23B, the cell culture region 200B, and the electrodes 31B, 32B, 41B, 42B are arranged in the cell culture chamber 21 of the cell culture container 1 according to the first embodiment. The arrangement of the first electrode chamber 22, the second electrode chamber 23, the cell culture region 200, and the electrodes 31, 32, 41, 42 is the same.

In the above-described first embodiment and second embodiment, each flow path is arranged substantially at the center of the cell culture chamber in the first direction. That is, each flow path was in direct communication with the approximate center of the cell culture chamber in the first direction. However, the present invention is not limited to this.

In the example of FIG. 8, the first upper flow path 24B is connected to the cell culture chamber 21B at one side of the center in the first direction. The second upper flow path 25B is connected to the cell culture chamber 21B on the other side of the center in the first direction. The first lower channel 26B is connected to the cell culture chamber 21B on the other side of the center in the first direction. The second lower flow path 27B is connected to the cell culture chamber 21B at one side of the center in the first direction.

As a result, the inflow position and the outflow position of the culture solution are different in the top view between the upper side and the lower side of the cell culture region 200B. As in the example of FIG. 8, the positions of the flow paths 24B, 25B, 26B, 27B can be changed appropriately.

FIG. 9 is a layout diagram showing a layout of each part in a top view of a cell culture container 1C according to a modification. As shown in FIG. 9, the cell culture container 1C has a cell culture chamber 21C, a first electrode chamber 22C, and a second electrode chamber as the internal space of the main body 10C, as in the cell culture container 1 according to the first embodiment. 23C, the first upper channel 24C, the second upper channel 25C, the first lower channel 26C, and the second lower channel 27C. Further, the cell culture container 1C has a first working electrode 31C, a first reference electrode 32C, a second working electrode 41C, and a second reference electrode 42C.

The cell culture chamber 21C, the first electrode chamber 22C, the second electrode chamber 23C, the cell culture region 200C, and the electrodes 31C, 32C, 41C, 42C are arranged in the cell culture chamber 21 of the cell culture container 1 according to the first embodiment. The arrangement of the first electrode chamber 22, the second electrode chamber 23, the cell culture region 200, and the electrodes 31, 32, 41, 42 is the same.

In the above-described embodiments and modified examples, each flow path directly communicates with the cell culture chamber. However, the present invention is not limited to this.

In the example of FIG. 9, the first upper channel 24C is connected to the first electrode chamber 22C at one side of the cell culture chamber 21C in the first direction. As a result, the first upper flow path 24C indirectly communicates with the cell culture chamber 21C via the first electrode chamber 22C. The second upper flow path 25C directly communicates with the cell culture chamber 21C on the other side of the center in the first direction. The first lower flow path 26C is connected to the second electrode chamber 23C on the other side of the cell culture chamber 21C in the first direction. As a result, the first lower flow path 26C indirectly communicates with the cell culture chamber 21C via the second electrode chamber 23C. In addition, the second lower flow path 27C directly communicates with the cell culture chamber 21C on one side of the center in the first direction.

As a result, the inflow position and the outflow position of the culture solution are different in the top view between the upper side and the lower side of the cell culture region 200C. As in the example of FIG. 9, the respective channels 24C, 25C, 26C, 27C do not necessarily directly communicate with the cell culture chamber, but may communicate indirectly.

Further, in the above embodiment, the shape of the cell culture container is a rectangular parallelepiped, but the present invention is not limited to this. The shape of the cell culture container can be appropriately designed into a cylindrical shape, a polygonal prism shape, a hemispherical shape, and other shapes.

Further, in the above-described embodiment, the shape of the cell culture region is a substantially quadrangle in a top view, but the present invention is not limited to this. The shape of the cell culture region may be a circular shape, an elliptical shape, a polygonal shape in a top view, or any other complicated shape imitating a biological structure. Further, in the above-mentioned embodiment, the cell culture region is arranged horizontally, but the present invention is not limited to this. The cell culture region may have a gradient as a whole with respect to the horizontal direction, or may have a complicated shape in which the gradient changes partially.

Moreover, the detailed configuration of the cell culture container may be different from the drawings of the present application. Further, the respective elements appearing in the above-described embodiments and modified examples may be appropriately combined as long as no contradiction occurs.

1, 1A, 1B, 1C Cell culture container 10, 10A, 10B, 10C Body part 20, 20A Cell culture membrane 21, 21A, 21B, 21C Cell culture chamber 22, 22A, 22B, 22C First electrode chamber 23, 23A, 23B, 23C 2nd electrode chamber 24, 24A, 24B, 24C 1st upper side flow path 25, 25A, 25B, 25C 2nd upper side flow path 26, 26A, 26B, 26C 1st lower side flow path 27, 27A, 27B, 27C 2nd lower side flow path 31, 31A, 31B, 31C 1st working electrode 32, 32A, 32B, 32C 1st reference electrode 41, 41A, 41B, 41C 2nd working electrode 42, 42A, 42B, 42C 2nd reference Electrode 200, 200A, 200B, 200C Cell culture area

Claims (5)

A cell culture vessel for culturing cells inside and measuring the electrical resistance of the cultivated cells,
A cell culture room that can store culture fluid,
Between the bottom surface and the top surface of the cell culture chamber, a cell culture membrane that partitions the interior of the cell culture chamber into an upper space and a lower space,
A flow path that directly or indirectly connects the outside with the cell culture chamber,
A first electrode chamber that directly communicates with the cell culture chamber above the cell culture membrane,
A second electrode chamber that is in direct communication with the cell culture chamber below the cell culture membrane;
A first working electrode and a first reference electrode arranged in the first electrode chamber;
A second working electrode and a second reference electrode disposed in the second electrode chamber;
Have
A cell culture container in which at least a portion of each of the first electrode chamber and the second electrode chamber does not vertically overlap with a cell culture region that is a portion of the cell culture membrane inside the cell culture chamber. The cell culture container according to claim 1, wherein
The minimum cross-sectional area of the space between the first working electrode and the cell culture region and the minimum cross-sectional area of the space between the second working electrode and the cell culture region are larger than the maximum flow passage area of the flow passage. , Cell culture vessels. The cell culture container according to claim 1 or 2, wherein
The cell culture container, wherein a casing forming the cell culture chamber has a portion formed of a transparent material at a position that vertically overlaps at least a part of the cell culture region. The cell culture container according to any one of claims 1 to 3,
A surface area of the first working electrode in the first electrode chamber is larger than a surface area of the first reference electrode in the first electrode chamber,
The cell culture vessel, wherein the surface area of the second working electrode in the second electrode chamber is larger than the surface area of the second reference electrode in the second electrode chamber. The cell culture container according to any one of claims 1 to 4,
The first electrode chamber, with respect to the cell culture chamber, is disposed on one side of the first direction orthogonal to the vertical direction,
The second electrode chamber is a cell culture container arranged on the other side in the first direction with respect to the cell culture chamber.

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