JP7307477B2 - Auxiliary device for extracellular potential measurement - Google Patents

Description

TECHNICAL FIELD The present invention relates to an extracellular potential measurement auxiliary device used for measuring extracellular potential based on cell activity of a cell to be measured, and more particularly to an extracellular potential measurement auxiliary device capable of measuring extracellular potential with high sensitivity.

Conventional extracellular potential measurement will be described using the MEGA system 10 shown in FIG. The example MEGA system 10 comprises a Grid 1 shaped bottom surface. This grid 1 comprises at least an electrical and/or optical multi-electrode array. The multi-electrode array comprises an electrode array, represented by black dots 8 in the illustrated example. The MEGA system 10 further comprises a second grid 4 shaped top surface. This grid 4 also comprises at least an electrical and/or optical multi-electrode array. The multi-electrode array comprises an array of electrodes, represented by black dots 9 in the illustrated example. The grids 1 , 4 are provided with electronic components, for example chips with said electronic components and electrically connected to the electrodes 8 , 9 . Furthermore, a foundation 2 is provided under said grid 1 . Base 2 provides mechanical support and strength to system 10 . The base 2 is further provided with a microfluidic system 3 . The microfluidic system 3 may be composed of biocompatible polymers. The grids 1, 4 may be aligned bottom to top with each other and held in place using means 5 for anchoring tissue sections 6 or cell cultures between both substrates. Such means 5 may be, for example, microdrives for adapting in a direction perpendicular to either grid 1, 4 (see above).

JP 2011-239779 A

The MEGA system 10 described above has the following points to be improved. Means 5 in the MEGA system 10 are means for aligning the grids 1, 4, bottom to top with each other and anchoring tissue sections 6 or cell cultures between both substrates. On the other hand, pressing the cells against the electrodes 8, 9 neither fixes the object against the electrodes nor improves the intensity of the signal. Therefore, there is a point to be improved in that if the cells to be detected have low signals, measurement data that can be analyzed cannot be obtained.

SUMMARY OF THE INVENTION Accordingly, an object of the present invention is to provide an extracellular potential measurement auxiliary device capable of measuring an extracellular potential with high sensitivity.
That is, the present invention is as follows.
[1]
An extracellular potential measurement auxiliary device for use in measuring extracellular potential based on cell activity of cultured cells,
a first layer located on the cell side;
a second hierarchical section positioned above the first hierarchical section;
Extracellular potential measurement auxiliary device having.
[2]
In the extracellular potential measurement auxiliary device according to [1],
The first layer section further includes:
a through hole connecting the bottom surface of the first layer portion and the top surface of the first layer portion and/or connecting the bottom surface of the first layer portion and the side surface of the first layer portion; to have
An extracellular potential measurement auxiliary device characterized by:
[3]
In the extracellular potential measurement auxiliary device according to [1] or [2],
The second layer section further includes:
a through hole connecting the bottom surface of the second layer portion and the top surface of the second layer portion and/or connecting the bottom surface of the second layer portion and the side surface of the second layer portion; to have
An extracellular potential measurement auxiliary device characterized by:
[Claim 4]
In any one of the extracellular potential measurement auxiliary devices according to [1] to [3],
The first layer section includes
having insulating properties;
An extracellular potential measurement auxiliary device characterized by:
[5]
In the extracellular potential measurement auxiliary device according to [4],
The first layer section includes
having flexibility;
An extracellular potential measurement auxiliary device characterized by:
[6]
In any one of the extracellular potential measurement auxiliary devices according to [1] to [5],
The second layer section includes
being made of metal;
An extracellular potential measurement auxiliary device characterized by:
[7]
An extracellular potential measurement auxiliary device for use in measuring extracellular potential based on cell activity of cultured cells,
a first layer located on the cell side;
a second hierarchical section positioned above the first hierarchical section;
An extracellular potential measurement method for measuring an extracellular potential using an extracellular potential measurement auxiliary device having

Means for solving the problems in the present invention and effects of the invention are shown below.

An extracellular potential measurement auxiliary device according to the present invention is an extracellular potential measurement auxiliary device used when measuring an extracellular potential based on cell activity of cultured cells, and is located on the cell side. , and a second layer section positioned above the first layer section.

As a result, it is possible to adjust two factors such as the weight of each layer and the buoyancy of each layer. Optimal pressure can be determined. Therefore, the extracellular potential can be measured with high sensitivity.

Moreover, when measuring the extracellular potential, a predetermined pressure can be applied to the cell to be measured, so the position of the cell can be fixed.

Furthermore, since the cell to be measured can be pressed toward a measurement electrode (described later), the intensity of the signal generated by cell activity can be increased compared to the conventional technique. That is, the signal intensity can be improved, the S/N ratio can be improved when measuring the extracellular potential, and as a result, the extracellular potential can be measured with high sensitivity. As a result, the extracellular potential can be measured even in cells with weak signal intensity generated by cell activity.

In the extracellular potential measurement auxiliary device according to the present invention, the first layer further connects the bottom surface of the first layer and the top surface of the first layer, and/or connects the first layer to the top surface of the first layer. It is characterized by having a through hole connecting the lower surface of the layered portion and the side surface of the first layered portion.

This allows medium to be provided from the upper surface to the cells located below the lower surface of the first layer, thereby maintaining the cells in an appropriate state.

In the extracellular potential measurement auxiliary device according to the present invention, the second layer further connects the bottom surface of the second layer with the top surface of the second layer, and/or connects the second layer with the top surface of the second layer. It is characterized by having a through hole connecting the lower surface of the layered portion and the side surface of the second layered portion.

Thereby, the culture medium can be provided from the upper surface to the lower surface of the second layer.

Therefore, even if the first layer and the second layer are arranged so as to overlap each other, it is possible to form a through hole communicating from the upper surface of the second layer to the lower surface of the first layer. That is, fresh medium outside the extracellular potential measurement auxiliary device can be provided to the cells located under the bottom surface of the first layer, and thus the cells can be maintained in an appropriate state.

The extracellular potential measurement auxiliary device according to the present invention is characterized in that the first layer section has flexibility.

As a result, it is possible to reduce the influence of the weight of the auxiliary device for extracellular potential measurement 100 on the cells located under the first layer.

The extracellular potential measurement auxiliary device according to the present invention is characterized in that the first layer section has insulating properties.

This makes it possible to accurately measure the extracellular potential without affecting the potential to be measured when measuring the extracellular potential.

The extracellular potential measurement auxiliary device according to the present invention is characterized in that the second layer section is made of a heavy material.

As a result, the size of the second layer can be reduced, and the size of the extracellular potential measurement auxiliary device can be reduced.

An extracellular potential measurement method according to the present invention is an extracellular potential measurement auxiliary device used for measuring extracellular potential based on cell activity of cultured cells, comprising: a first layer located on the cell side;
An extracellular potential is measured using an extracellular potential measurement auxiliary device having a second layer section positioned above the first layer section.

As a result, it is possible to adjust two factors such as the weight of each layer and the buoyancy of each layer. Optimal pressure can be determined. Therefore, the extracellular potential can be measured with high sensitivity.

BRIEF DESCRIPTION OF THE DRAWINGS FIG. 1 is a diagram showing an extracellular potential measurement auxiliary device 100 which is an embodiment of an extracellular potential measurement auxiliary device according to the present invention; FIG. 2 is a diagram showing a first layer section 101 of the auxiliary device for extracellular potential measurement 100; FIG. 3 is a diagram showing a second layer section 103 of the auxiliary device for extracellular potential measurement 100; It is a figure which shows the cell culture apparatus C. FIG. FIG. 2 shows an MEA probe P; FIG. 4 is a diagram showing a state of measuring an extracellular potential using the MEA probe P, the cell culture device C, and the extracellular potential measurement auxiliary device 100. FIG. FIG. 4 is a diagram showing the results of measuring the extracellular potential of myocardial cells using the extracellular potential measurement auxiliary device 100. FIG. FIG. 4 is a diagram showing the results of measuring the extracellular potential of myocardial cells using the extracellular potential measurement auxiliary device 100. FIG. FIG. 3 is a diagram showing extracellular potentials of myocardial cells measured using the extracellular potential measurement auxiliary device 100. FIG. FIG. 10 is a diagram showing another embodiment of the extracellular potential measurement auxiliary device according to the present invention; FIG. 10 is a diagram showing another embodiment of the extracellular potential measurement auxiliary device according to the present invention; FIG. 10 is a diagram showing another embodiment of the extracellular potential measurement auxiliary device according to the present invention; FIG. 10 is a diagram showing another embodiment of the extracellular potential measurement auxiliary device according to the present invention; It is a figure which shows the conventional extracellular potential measurement auxiliary device.

BEST MODE FOR CARRYING OUT THE INVENTION Hereinafter, embodiments of the present invention will be described in detail with reference to the drawings.

An auxiliary device for extracellular potential measurement according to the present invention will be described using an auxiliary device for extracellular potential measurement 100 as an example. The extracellular potential measurement auxiliary device 100 is an auxiliary device used when measuring an extracellular potential based on the activity of a cell to be measured.

First Configuration The configuration of the extracellular potential measurement auxiliary device 100 will be described with reference to FIG. Here, FIG. 1A shows a plan view of the auxiliary device for extracellular potential measurement 100, and FIG. 1B shows a front view of the auxiliary device for extracellular potential measurement 100, respectively. As shown in FIGS. 1A and 1B, the extracellular potential measurement auxiliary device 100 has a first layer section 101 and a second layer section 103 .

The first layer section 101 and the second layer section 103 are arranged in layers. The first layer portion 101 is arranged at a position closer to the cell than the second layer portion 103 is. The second layer section 103 is arranged above the first layer section 101 so as to overlap the first layer section 101 .

A configuration of the first layer section 101 will be described with reference to FIG. Here, FIG. 2A shows a plan view of the first layer portion 101, and FIG. 2B shows a cross-sectional view along the line X1-X1 of the first layer portion 101 shown in FIG. 2A. As shown in FIG. 2A, the first layer portion 101 has a rectangular plate shape. As shown in FIG. 2B, the first layer portion 101 has a lower surface 101a, an upper surface 101b, and a through hole 101c. The lower surface 101a is a surface located on the side closer to the cells. On the other hand, the upper surface 101b is a surface located opposite to the lower surface 101a and located farther from the cells. The through hole 101c is a hole that connects the lower surface 101a and the upper surface 101b. The first layer portion 101 has a plurality of through holes 101c.

The first layer 101 is made of an insulating and flexible material such as PDMS (PolyDiMethylSiloxane). By forming the first layer portion 101 from an insulating material, the extracellular potential can be accurately measured without affecting the potential to be measured. In addition, by forming the first layer section 101 from a flexible material, it is possible to reduce weight damage to cells from the extracellular potential measurement auxiliary device 100 .

The configuration of the second layer section 103 will be described with reference to FIG. Here, FIG. 3A shows a plan view of the second layer portion 103, and FIG. 3B shows a cross-sectional view along the line X2-X2 of the second layer portion 103 shown in FIG. 3A. As shown in FIG. 3A, the second layer portion 103 has a torus shape. As shown in FIG. 3B, the second layer portion 103 has a lower surface 103a, an upper surface 103b, and a through hole 103c. The lower surface 103 a is a surface that contacts the first layer section 101 . On the other hand, the upper surface 103b is a surface positioned facing the lower surface 103a. Through hole 103c is a hole that connects lower surface 101a and upper surface 101b. The second layer portion 103 has one through hole 103c in the center.

The second layer portion 103 is made of a heavy material such as metal. By forming the second layer part 101 with an insulating material, the extracellular potential can be accurately measured without affecting the potential to be measured.

In addition, by forming the first layer section 101 from a flexible material, it is possible to reduce weight damage to cells from the extracellular potential measurement auxiliary device 100 .

Further, by forming the through hole 101c in the first layer portion 101 and the through hole 103c in the second layer portion 103, even if the first layer portion 101 and the second layer portion 103 are arranged to overlap each other, the second layer portion A through-hole 101c and a through-hole 103c communicating from the upper surface 103b of the portion 103 to the lower surface 101a of the first layer portion 101 can be formed. As a result, the fresh medium outside the extracellular potential measurement auxiliary device 100 can be provided to the cells located under the lower surface 101a of the first layer section 101, thereby maintaining the cells in an appropriate state. can be done.

Thus, by forming the extracellular potential measurement auxiliary device 100 with a plurality of layers, it is possible to adjust two factors such as the weight of each layer and the buoyancy of each layer. This makes it possible to determine the optimum pressure for measuring the extracellular potential according to the type and state of the cell whose extracellular potential is to be measured.

By using the extracellular potential measurement auxiliary device 100, it is possible to apply a predetermined pressure to the cell to be measured when measuring the extracellular potential, so that the position of the cell can be fixed. In addition, since the cell to be measured can be pressed toward the measurement electrode (described later), the intensity of the signal generated by cell activity can be increased compared to the conventional technique. That is, the signal intensity can be improved, the S/N ratio can be improved when measuring the extracellular potential, and as a result, the extracellular potential can be measured with high sensitivity. As a result, the extracellular potential can be measured even in cells with weak signal intensity generated by cell activity.

In this way, by using the extracellular potential measurement auxiliary device 100, it is possible to provide the cell to be measured with a sufficiently fresh medium necessary for cell activity when measuring the extracellular potential, and a light load can be applied. extracellular potential can be measured without inhibiting cell activity. Furthermore, even when measuring the extracellular potential with the drug added, the added drug can be supplied to the cells along with the medium, so the extracellular potential can be measured without inhibiting drug responsiveness.

Second Method of Using the Auxiliary Device for Measuring Extracellular Potential 100 Regarding the method of using the auxiliary device for extracellular potential measurement 100, the extracellular potential generated by the cell activity of cardiomyocytes formed in the cell culture device C is measured by using an MEA (Multi-Electrode Array) probe. An example of measurement using P will be described with reference to FIGS. 4 to 6. FIG.

Here, the cell culture apparatus C will be briefly described with reference to FIG. Here, FIG. 4A shows a plan view of the cell culture apparatus C, and FIG. 4B shows a cross-sectional view of the cell culture apparatus C along the line X3-X3 shown in FIG. 4A. As shown in FIGS. 4A and 4B, the cell culture device C has a cell culture sheet CS and a frame CF. The cell culture sheet CS functions as a scaffold for cell culture. A fiber sheet made of a polymer material is used as the cell culture sheet CS. The cell culture sheet CS has an oriented structure, a non-oriented structure, or a mixed structure of oriented and non-oriented structures by polymers.

The frame CF functions as a holding member that flatly holds the cell culture sheet CS. The frame CF has a prismatic shape with a cell culture sheet arrangement space F1 in the center. Note that the frame CF has a predetermined thickness TS and a predetermined height HS. A cell culture sheet CS is attached to one end face of the frame CF. That is, the cell culture sheet CS is held in the cell culture sheet arrangement space CA.

The cell culture device C is arranged in a predetermined well filled with a predetermined medium, and cardiomyocytes are placed on the cell culture sheet CS held in the cell culture sheet arrangement space CA of the cell culture device C. The cardiomyocytes are seeded at a high density and cultured on the surface of the cell culture sheet CS. Cardiomyocytes are generally cultured on both surfaces of the cell culture sheet CS.

An MEA probe P is used for extracellular potential measurement. The MEA probe P is a device for measuring extracellular potentials generated by cell activity. MEA probe P will be briefly described with reference to FIG. Here, FIG. 5A shows a plan view of the MEA probe P, and FIG. 5B shows a cross-sectional view of the MEA probe P taken along line X4-X4 shown in FIG. 5A. As shown in FIGS. 5A and 5B, the MEA probe P has a substrate portion PB and a wall portion PW. The substrate part PB has a thin plate shape. The weld wall portion PW is formed on the substrate portion PB. The melted wall portion PW has a cylindrical shape with one end open, stores a predetermined culture medium inside, and forms a cell arrangement space PS for arranging the cells to be measured.

The MEA probe P also has a measurement electrode E1 and a reference electrode E2. The measurement electrode E1 is formed substantially in the center of the molten wall portion PW along the substrate portion PB. The measurement electrodes E1 are arranged in a 4×4 matrix. A predetermined number of reference electrodes E2 are arranged around the measurement electrode E1. The MEA probe P measures an extracellular potential due to cell activity based on the potential difference between the measurement electrode E1 and the reference electrode E2. A lead line (dotted line in the figure) is arranged from each of the measurement electrode E1 and the reference electrode E2.

After pouring a predetermined amount of culture medium into the cell placement space PS of the MEA probe P, the cell culture device C in which cardiomyocytes are cultured is placed in the cell placement space PS. At this time, the cell culture device C is placed on the measurement electrode E1 of the MEA probe P so that the cell culture sheet placement space F1 is positioned.

Further, the extracellular potential measurement auxiliary device 100 is arranged on the cell culture sheet CS in which the cardiomyocytes of the cell culture device C are cultured. FIG. 6A is a plan view of the extracellular potential measurement auxiliary device 100 arranged on the cell culture sheet CS in which the cardiomyocytes are cultured in the cell culture device C, and FIG. 6B is the X5-X5 section of FIG. 6A. show. As shown in FIGS. 6A and 6B, the extracellular potential measurement auxiliary device 100 is arranged in the cell culture sheet arrangement space F1 of the cell culture apparatus C. As shown in FIGS. At this time, the first layer part 101 of the extracellular potential measurement auxiliary device 100 is located on the cell culture sheet CS side, that is, the lower surface 101a of the first layer part 101 is positioned in the cell culture sheet arrangement space F1. It arrange|positions so that the cardiomyocytes cultured on the culture sheet CS may be touched. After placing the first layer part 101 , the second layer part 103 is placed on the first layer part 101 . As a result, the through-hole 101c of the first layer 101 and the through-hole 103c of the second layer 103 are communicated with each other, and fresh culture medium is constantly provided to the cells located under the auxiliary device for extracellular potential measurement 100. .

In addition, if necessary, the amount of culture medium is adjusted so that both the first layer section 101 and the second layer section 103 are located in the culture medium.

After that, using the MEA probe P, the extracellular potential due to the cell activity of the myocardial cells cultured in the cell culture apparatus C is measured. By arranging the extracellular potential measurement auxiliary device 100 in this way, the movement of cells can be measured without interfering with the activities of the cells.

Third Experimental Example As the first layer part 101 in the extracellular potential measurement auxiliary device 100, 23 cylindrical through-holes 101c with a diameter of 1 mm were formed. was used.

As the second layer part 103 in the auxiliary device for extracellular potential measurement 100, there are 7 kinds of metal toric disks with an outer diameter of 8 mm and an inner diameter of 5 mm, and weights of 100 mg, 200 mg, 300 mg, 400 mg, 500 mg, 600 mg, and 700 mg. was used.

A cell culture device C in which predetermined myocardial cells were cultured was placed in the cell placement space PS of the MEA probe P filled with a predetermined medium. When the auxiliary device for extracellular potential measurement 100 is not used, and when the auxiliary device for extracellular potential measurement (total weight 40 mg) having only the first layer portion 101 is used, the extracellular potential measurement aid device having the second layer portion 103 weighing 100 mg is used. When the auxiliary device for potential measurement 100 (total weight of 140 mg) is used, the second layer portion 103 to weigh 300 mg when the extracellular potential measurement auxiliary device 100 (total weight of 240 mg) having the second layer portion 103 to weigh 200 mg is used. When using the auxiliary extracellular potential measurement device 100 (total weight of 340 mg) having the When using the auxiliary extracellular potential measurement device 100 (total weight 540 mg) having the second layer portion 103, when using the extracellular potential measurement auxiliary device 100 (total weight 640 mg) having the second layer portion 103 weighing 600 mg, When using the extracellular potential measurement auxiliary device 100 (total weight: 740 mg) having the second layer portion 103 weighing 700 mg, the extracellular potential was measured in each case. The results are shown in FIG.

From the results of FIG. 7, when the extracellular potential measurement aid device 100 having the second layer portion 103 weighing 500 mg is used, that is, when the total weight of the extracellular potential measurement aid device 100 is 540 mg or more, the S/N ratio It can be seen that the degree of improvement in

FIG. 8A shows the result of extracellular potential measurement when the extracellular potential measurement auxiliary device 100 is not used. FIG. 8B shows the results of extracellular potential measurement using an extracellular potential measurement auxiliary device (total weight: 40 mg) having only the first layer portion 101 . Furthermore, FIG. 8C shows the result of extracellular potential measurement using the extracellular potential measurement auxiliary device 100 (total weight: 140 mg) having the second layer portion 103 weighing 100 mg. Furthermore, FIG. 8D shows the result of extracellular potential measurement using the extracellular potential measurement auxiliary device 100 (total weight: 740 mg) having the second layer portion 103 weighing 700 mg. The measurement time was 5 minutes for each measurement.

From the experimental results of FIGS. 8A, 8B, 8C, and 8D, the S/N ratios in each measurement were 2.5, 2.0, 71, and 420, respectively. For this reason, using the second layer section 103, a predetermined pressure is applied to the myocardial cells in the direction of the measurement electrode E1 of the MEA probe P to fix the positions of the myocardial cells to be measured, and to fix the positions of the myocardial cells to be measured. By pressing the cell toward the measurement electrode E1, the intensity of the signal generated by the cell activity is increased and the signal intensity is improved compared to the conventional method, and as a result, the S/N ratio during extracellular potential measurement is improved. can be improved. In other words, it can be seen that by using the extracellular potential measurement auxiliary device 100, the extracellular potential can be measured even in a cell with a weak signal intensity generated by cell activity. Also, it can be seen that the S/N ratio can be dramatically improved by increasing the weight of the second layer portion 103 .

As shown in FIG. 9, the extracellular potential measured in the myocardial cell uses the potential difference V1 obtained at the first peak of one signal, which is the potential change obtained by one heartbeat of the myocardial cell.

[Other embodiments]
(1) Through-holes 101a of first layer 101: In the first embodiment described above, 23 cylindrical through-holes 101c with a diameter of 1 mm were formed in the first layer 101. It is not limited to the example as long as it connects the lower surface 101a and the upper surface 101b of the . For example, as shown in FIG. 10 (A is a plan view and B is a cross-sectional view of X6-X6 in FIG. 10A), a square plate of 8 mm×8 mm and a diameter of 1.5 mm Eighteen cylindrical through-holes 201c may be formed.

Here, FIG. 11 shows an extracellular potential measurement auxiliary device 200 formed using the first layer section 201 and the second layer section 103 . In FIG. 1, A is a plan view, and B is a sectional view taken along line X7-X7 of FIG. 11A.

12A to E (in FIG. 12, A is a plan view, B is a front view, C is a right side view, D is a cross-sectional view of X8-X8 in FIG. 12A, and E is a cross-sectional view of X9-X9 in FIG. 12A. ), through grooves 301c1 and 301c2 may be formed to penetrate the side surface 301d and connect the lower surface 301a and the side surface 301d. The first layer portion 301 has a through groove 301c1 communicating with the upper surface 301b and a through groove 301c2 communicating with the lower surface 301a. The through grooves 301c1 and the through grooves 301c2 are formed hierarchically vertically and perpendicular to each other.

This allows the through groove 301c1 and the through groove 301c2 to communicate with each other. In addition, by using the first layer portion 301, the culture medium can be provided from the upper surface 301b of the first layer portion 301 to the lower surface 301a, and the culture medium can be provided from the side surface 301d to the lower surface 301a. Note that the first layer portion 301 can be formed of PDMS.

(2) Shape of the first layer portion 101: In the first embodiment described above, the first layer portion 101 has a rectangular plate shape, but other shapes such as a disk shape (see FIG. 11), an elliptical plate shape, It may have a polygonal board shape.

(3) Material of the first layer portion 101: In the first embodiment described above, the first layer portion 101 is made of PDMS. are not limited to those of For example, silicon, polycarbonate, and other plastic materials other than PDMS may be used.

In addition, due to the conditions such as the potential obtained by the cell activity of the cell to be measured, the size, thickness, type of the cell, and the strength against weight, a material with low flexibility and insulation, such as polycarbonate, is used. , may form the first layer.

(4) Position of the first layer 101: In the first embodiment described above, the lower surface 101a of the first layer 101 is in contact with the cell to be detected. are not limited to those of For example, a buffer such as a sheet may be placed on the cells to indirectly press the cells.

(5) Through hole 103c of second layer portion 103: In Example 1 described above, the second layer portion 103 had one through hole 103c in the center, but the second layer portion shown in FIG. Like 303, a plurality of through holes 303c may be formed. Moreover, the plurality of through holes 303c formed in the second layer portion 303 may have different shapes instead of the same shape.

Note that FIG. 12 shows the extracellular potential measurement auxiliary device 300 formed using the second layer section 303 and the first layer section 301 . 13, A is a plan view, B is a front view, C is a right side view, D is a cross-sectional view along X10-X10 in FIG. 12A, and E is a cross-sectional view along X11-X11 in FIG. 13A.

Also, if the medium can be supplied to the cells from the side surface 301d like the first layer portion 301 shown in FIG. 11, it is not necessary to form the through holes in the second layer portion.

(6) Shape of the second layer portion 103: In the first embodiment, the second layer portion 103 has a toroidal disk shape, but other shapes such as a disk shape, an elliptical plate shape, and a polygonal disk shape are used. It may be in shape.

(7) Cells to be measured: In Example 1 described above, cardiomyocytes were used as the cells to be detected. For example, nerve cells may be used.

Also, the cells to be measured may be single cells cultured on a planar sheet. Furthermore, cells cultured in a general cell sheet may be used instead of the cells cultured in the cell culture sheet CS. Furthermore, the cells may be three-dimensionally cultured instead of planarly cultured on the cell culture sheet CS. Furthermore, tissue sections taken from organisms such as humans may be used instead of cultured cells.

(8) Use of cell culture apparatus C: In the above-described Example 1, the cells to be measured were cultured using the cell culture apparatus C, but if the cells to be measured can be cultured, It is not limited to the illustrated one. For example, cells may be directly cultured in a predetermined container such as a cell culture dish.

(9) Use of MEA probe P: In Example 1 described above, the MEA probe P was used to measure the extracellular potential.

(10) Extracellular potential: In the above-described Example 1, the potential difference V1 obtained at the first peak of one signal obtained by one heartbeat was used as the extracellular potential. As long as it is an external potential, it is not limited to the example.

The extracellular potential measurement auxiliary device according to the present invention can be used, for example, to measure extracellular potential based on cell activity of cultured myocardial cells.

100 Extracellular Potential Measurement Auxiliary Device 101 First Layer Part 101a Lower Surface 101b Upper Surface 101c Through Hole 103 Second Layer Part 103a Lower Surface 103b Upper Surface 103c Through Hole C Cell Culture Device P MEA Probe 200 Extracellular Potential Measurement Auxiliary Device 201 First Layer Part 203a Lower surface 303b Upper surface 403c Through hole 300 Extracellular potential measurement auxiliary device 301 First layer part 301a Lower surface 301b Upper surface 301c Through hole 301d Side surface 303 Second layer part 303a Lower surface 303b Upper surface 303c Through hole

Claims (7)

An extracellular potential measurement auxiliary device for use in measuring extracellular potential based on cell activity of cultured cells,
a first layer located on the cell side;
a second hierarchical section positioned above the first hierarchical section;
wherein the first layer is made of an insulating or flexible material, the second layer is made of a heavy material made of metal , and the first layer is The extracellular potential measurement auxiliary device, wherein the part is arranged at a position closer to the cell than the second layer part, and can press the cell to be measured toward the measurement electrode. In the extracellular potential measurement auxiliary device according to claim 1,
The first layer section further includes:
a through hole connecting the bottom surface of the first layer portion and the top surface of the first layer portion and/or connecting the bottom surface of the first layer portion and the side surface of the first layer portion; to have
An extracellular potential measurement auxiliary device characterized by: In the extracellular potential measurement auxiliary device according to claim 1 or claim 2,
The second layer section further includes:
a through hole connecting the bottom surface of the second layer portion and the top surface of the second layer portion and/or connecting the bottom surface of the second layer portion and the side surface of the second layer portion; to have
An extracellular potential measurement auxiliary device characterized by: In the extracellular potential measurement auxiliary device according to any one of claims 1 to 3,
The first layer section includes
having insulating properties;
An extracellular potential measurement auxiliary device characterized by: In the extracellular potential measurement auxiliary device according to claim 4,
The first layer section includes
having flexibility;
An extracellular potential measurement auxiliary device characterized by: In the extracellular potential measurement auxiliary device according to any one of claims 1 to 5,
The second layer section includes
being made of metal;
An extracellular potential measurement auxiliary device characterized by: An extracellular potential measurement method for measuring an extracellular potential using the extracellular potential measurement auxiliary device according to any one of claims 1 to 6 .

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