JP7362119B2 - measuring device - Google Patents

Description

The present invention relates to a measuring device.

2. Description of the Related Art Devices that measure the electrochemical properties of a sample using a biosensor are known as devices for measuring the concentration of a substance to be measured in a biological sample.

Patent Document 1 describes a measuring device in which a biosensor is removably installed in the main body of the device, and the biosensor includes a suction path for sucking a biological sample, a measurement device for measuring the concentration of a substance to be measured in the biological sample. comprising an electrode and a detection electrode for detecting completion of aspiration of a biological sample, the device body comprising a power source, a color liquid crystal display, an insertion port into which the biosensor is inserted, and a light source for illuminating the vicinity of the insertion port; The light source lights up when the power source is turned on, and turns off after the biosensor is attached to the device main body and the detection electrode of the biosensor detects a biological sample. ” is written.

International Publication 2014/181753

When trying to use a biosensor to measure electrochemical signals derived from activities such as metabolism of microorganisms in a sample containing microorganisms, the measurement sensitivity is insufficient with the device described in Patent Document 1. I discovered something.

Therefore, an object of the present invention is to provide a measuring device that can detect electrochemical signals derived from microbial metabolism and the like with excellent sensitivity.

As a result of intensive studies to achieve the above-mentioned problems, the present inventors have found that the above-mentioned problems can be achieved by the following configuration.

[1] A lower base material, a working electrode disposed on surface A of the lower base material and extending from one end of the lower base material in a direction along the surface A to approximately the center of the lower base material. at least two electrodes comprising: an electrolyte gel disposed on the electrode so as to be in direct contact with at least a portion of the electrode; a cover member covering the electrolyte gel in contact with a surface of the electrolyte gel opposite to the electrode side; and an electrochemical property of a microorganism connected to the electrode and sandwiched between the working electrode and the electrolyte gel of a biosensor having an upper base material supporting the cover member and disposed on the lower base material. A measuring device comprising: an electrochemical measuring section for measuring; and a pressure sealing section for pressing and pressurizing the electrolyte gel against the working electrode while sandwiching the lower base material and the upper base material from both sides.
[2] The measuring device according to [1], wherein the electrolyte gel contains a hydrophilic polymer, an electrolyte, and an oxygen adsorbent.
[3] A spacer layer is further provided between the upper base material and the lower base material, and the upper base material has a through hole along the thickness direction from one surface B to the other surface C. The spacer layer is recessed in a shape that fits around the outer periphery of the electrolyte gel, and the spacer layer extends from the side surface of the recessed portion toward the opening of the through hole disposed on the surface B. A cavity defined by the surface A of the upper base material, the surface B, and the side surface of the first notch is formed through the opening. The measuring device according to [1] or [2], wherein a flow path communicating with the through hole and extending from the surface C of the upper base material to the electrolyte gel is formed.
[4] The spacer layer and the upper base material further have a second notch extending in a direction parallel to the surface B from the side surface thereof toward the center thereof, and the second notch and the above The measuring device according to [3], wherein a part of the electrode is exposed in the recess defined by the surface A.
[5] The pressurized sealing part has a pair of plate-like members whose distance from each other can be adjusted, and the clearance distance when the plate-like members are brought closest is equal to or less than the maximum thickness of the biosensor, [ 4].
[6] The pressurized sealing part further includes a spacer disposed on at least one surface of the pair of plate-shaped members, and when the pressurization is applied, the spacer closes the opening of the through hole on the surface C side. The measuring device according to [5], which is occluded.

According to the present invention, a measuring device having excellent sensitivity can be provided.

A sample solution containing Shewanella oneidensis MR-1 (Optical density: 1) and lactic acid (1 mM) was inoculated onto carbon electrodes (working electrode and counter electrode), and agar containing indium tin oxide was inoculated therein. It is a graph showing the relationship between culture time and current value when carrying out aerobic culture by applying +0.2V to the working electrode with the silver electrode as the reference electrode. FIG. 2 is a perspective view of a biosensor that can be used in the measuring device according to the present invention. It is an exploded perspective view of the above-mentioned biosensor. It is a perspective view of a lower base material. FIG. 3 is a cross-sectional view taken along line PP' of the biosensor. FIG. 1 is a schematic perspective view of an embodiment of a measuring device according to the present invention. FIG. 3 is a perspective view of the measuring device in a state where the upper teeth of the clip are open. FIG. 3 is a perspective view of the measuring device with the biosensor inserted into the insertion port. FIG. 3 is a partially enlarged view of the clip in a state where a biosensor is inserted. It is a QQ' sectional view of the above-mentioned clip. FIG. 2 is a hardware configuration diagram of a measuring device. FIG. 2 is a functional block diagram of the measuring device. It is an operation flow of a control unit that operates according to a program.

The present invention will be explained in detail below.
Although the description of the constituent elements described below may be made based on typical embodiments of the present invention, the present invention is not limited to such embodiments.
In this specification, a numerical range expressed using "~" means a range that includes the numerical values written before and after "~" as the lower limit and upper limit.

The present inventors have diligently continued various studies to measure electrochemical signals derived from the metabolic activities of microorganisms with higher sensitivity. Among these, we found that it is effective to press the microorganisms to be measured against the working electrode, that is, to perform measurements under pressurized conditions.

Figure 1 shows that a sample solution containing Shewanella oneidensis MR-1 (optical density: 1) and lactic acid (1mM) is inoculated onto carbon electrodes (working electrode and counter electrode), and indium tin oxide is added thereto. It is a graph showing the relationship between culture time and current value when carrying out aerobic culture by applying agar containing agar and applying +0.2V to the working electrode using a silver electrode as a reference electrode.

In Figure 1, "1.5% Agar-ITO" indicates that the sample solution was not inoculated, and "1.5% Agar-ITO + MR1" indicates that the sample solution was inoculated but was not pressurized. The one that was not found, "1.5% Agar-ITO + MR1 + Weight", indicates that the sample solution was inoculated and then a weight was placed on the agar and pressurized.

From the results in Figure 1, it can be seen that "1.5% Agar-ITO + MR1 + Weight" in which the sample solution was pressurized from above the agar to the electrode side had a larger current than "1.5% Agar-ITO + MR1" in which no pressure was applied. It was found that a value (electrochemical signal) could be obtained.
In the above, it can be seen that the current value obtained by pressurization becomes large, but in our study, we investigated not only the magnitude of the current value but also the rise of the current (the difficulty in obtaining an electrochemical signal). It has also been found that the time it takes to The present invention has been completed based on the above novel findings.

[Biosensor]
FIG. 2 shows a perspective view of a biosensor 100 as an example of a biosensor that can be used in the measuring device according to the present invention. Further, FIG. 3 shows an exploded perspective view of the biosensor 100.
The biosensor 100 includes a lower base material 101, a spacer layer 106, an upper base material 108, and a cover member 109 in this order. Furthermore, an opening 113 for introducing a liquid sample is provided on the surface C of the upper base material 108.

FIG. 4 is a perspective view of the lower base material 101. On the surface A of the lower base material 101, there are electrodes 102 (working electrodes) each extending from one end of the lower base material 101 in a direction along the surface A of the lower base material 101 to approximately the center of the lower base material 101. 103, a counter electrode 104, and a working electrode 105) are arranged.
When the biosensor 100 is inserted into a measuring device to be described later, each of the above electrodes is connected to a connecting terminal provided in the measuring device.
The material for the lower base material 101 is not particularly limited, but resin, glass, and the like are preferable because they have superior insulation properties.
Further, the material of the electrode is not particularly limited, and materials known as electrodes for biosensors can be used without particular limitation.

Returning to FIG. 3, the biosensor 100 has a spacer layer 106 on the lower base material 101. The spacer layer 106 has a function of bonding the upper base material 108 and the lower base material 101, and is typically formed of an adhesive.
Further, the spacer layer 106 has a recessed part, and an electrolyte gel 107 having an outer periphery that fits with the inner periphery of the recessed part 110 is disposed so as to be fitted therein.

The electrolyte gel 107 has a substantially rectangular parallelepiped shape, is placed on the lower base material 101, and is in contact with the electrode 102. Since the recessed part 110 of the spacer layer 106 has a shape that fits around the outer periphery of the electrolyte gel 107, the electrolyte gel 107 can directly contact the electrode 102. Therefore, although the upper base material 108 and the lower base material 101 are bonded together by the spacer layer 106, one surface of the electrolyte gel 107 directly contacts the surface A of the lower base material 101 without intervening the spacer layer 106. In this part (namely, the recessed part 110), the surface of the electrolyte gel 107 and the surface A of the lower base material 101 are not bonded to each other.

The upper base material 108 has a through hole 111 along the thickness direction from the surface B on one side to the surface C on the other side. The through hole 111 has an opening 112 and an opening 113 on the surface B side and the surface C side of the upper base material 108, respectively.

The spacer layer 106 further includes a first notch 115 extending from the side surface of the recess 110 toward the opening 112 in a direction along the surface of the spacer layer 106 .
A cavity CAV is defined by a side surface of the first notch 115, a surface A, and a surface B.
The cavity CAV communicates with the through hole 111 via the opening 112, and as a result functions as a flow path from the surface C of the upper base material 108 to the electrolyte gel 107. Therefore, the liquid sample introduced from the opening 113 is guided to the electrolyte gel 107 via the flow path.

As already explained, since the electrolyte gel 107 and the electrode 102 are not bonded together, the liquid sample provided through the flow path infiltrates between the electrolyte gel 107 and the electrode 102.

Preferably, the liquid sample contains microorganisms. When the liquid sample enters between the electrolyte gel 107 and the electrode 102, microorganisms contained in the liquid sample are sandwiched between the electrolyte gel 107 and the electrode 102. Thereby, electron transfer between the microorganism and the electrode 102 can be measured as an electrochemical property.
In this measuring device, the electrolyte gel 107 is pressed toward the electrode 102 by the clip 505, so that excellent sensitivity can be obtained.
It is already known that electron transfer between microorganisms and electrodes reflects the metabolic activity of microorganisms, and this measuring device allows the metabolic activity of microorganisms to be measured with higher sensitivity.

The liquid sample is not particularly limited as long as it contains microorganisms or has the possibility of containing microorganisms. For example, saliva, sputum, etc. derived from humans can be used, and blood, urine, synovial fluid, etc. can also be used when some kind of infectious disease is suspected. Among these, saliva is preferred. Moreover, agricultural water, water and sewage water, etc. may be used.
Furthermore, the target microorganisms are not particularly limited. Among these, pathogenic microorganisms are preferred, and periodontal disease bacteria (Aggregatibacter actinomycetemcomitans, Prophyromonas gingivalis, etc.) are particularly preferred.

The spacer layer 106 and the upper base material 108 have a second notch 114 extending from the side surface (one end) in a direction along the surface B of the upper base material 108 (along the longitudinal direction of the upper base material 108). . The electrode 102 disposed on the surface A of the lower base material 101 is exposed from the recess defined by the second notch 114 and the surface A of the lower base material 101.
Thereby, when the biosensor 100 is inserted into a measuring device to be described later, the measuring device and the biosensor 100 can be electrically connected more easily.

FIG. 5 shows a PP' cross-sectional view of the biosensor 100. The cover member 109 is arranged to directly cover the electrolyte gel 107, and its end portion is supported by the upper base member 108. The cover member 109 is made of an elastic material, and when the cover member 109 is pressurized from the outside, the electrolyte gel 107 and the electrode 102 disposed on the lower base material 101 come into stronger contact.

According to the present biosensor 100, since the electrolyte gel 107 and the electrode 102 are not bonded together, the liquid sample passes through the electrolyte from the opening 113 through the flow path constituted by the through hole 111, the opening 112, and the cavity CAV. It penetrates between the gel 107 and the electrode 102. In this state, by further pressing the cover member 109 made of an elastic material, the electrolyte gel 107 and the electrode 102 can be brought into contact in a pressurized state, so that better sensitivity can be obtained.

[measuring device]
Next, one embodiment of the measuring device according to the present invention will be described using the drawings.
FIG. 6 is a schematic perspective view of an embodiment of the measuring device according to the present invention. The measuring device 500 is an electronic device in which electronic components are housed in a main body 501. The front side of the main body 501 is provided with a touch panel 502, a clip 505 for arranging and holding the biosensor, and an insertion port 506 for inserting the biosensor. 505 so that it can be held between the two.

The clip 505 is composed of a pair of plate-like members (upper teeth 503, lower teeth 504), and the upper teeth 503 are movable by a hinge 507, so that the clearance distance between the pair of plate-like members can be adjusted.
The clearance distance is the distance between the surfaces of the pair of plate-like members (indicated by "L" in FIG. 2) when the clip 505 is closed.
The clearance distance L of the clip 505 of the measuring device 500 is less than or equal to the maximum thickness of the biosensor 100, and is configured so that the cover member 109 can be pressed when the biosensor 100 is held.

FIG. 7 is a perspective view of the measuring device 500 in a state where the upper teeth 503 of the clip 505 are open. Opening the clip 505 increases the clearance distance, making it easier to insert the biosensor 100 into the insertion port 506.
FIG. 8 is a perspective view of the measuring device 500 with the biosensor 100 inserted into the insertion port 506. One end of the biosensor 100 is inserted into the insertion port 506 and electrically connected to an electrode terminal (not shown) of the measurement device 500. When the biosensor 100 is inserted, the clip 505 (upper teeth 503) is closed by the user. Note that, although the upper teeth 503 of the measuring device 500 are closed by the user (indicated as "F" in FIG. 8), the measuring device according to the present invention is not limited to the above, and the measuring device can automatically open and close the clip. It may have a mechanism.

FIG. 9 is a partially enlarged view of the clip 505 in a state where the biosensor 100 is inserted, and FIG. 10 is a sectional view taken along the line QQ'.
The plate member includes a base material and a spacer extending from an end of the base material along the thickness direction of the biosensor 100. As shown in FIG. 10, the upper tooth 503 of the clip 505 includes a base material 901 and a spacer 902 extending from the base material 901 in the thickness direction of the biosensor 100, and has a hook-shaped cross section.

The opening 113 is closed by the spacer 902 of the upper tooth 503. This simultaneously presses the electrolyte gel and closes the flow path, resulting in better sensitivity.

The electrolyte gel has the function of supplying insufficient water and electrolyte to a liquid sample and absorbing excess water and electrolyte. Electrolyte gel generally contains a hydrophilic polymer and an electrolyte, but if it also contains an oxygen adsorbent, the flow path, electrolyte gel, and electrode surroundings can be maintained anaerobically, making it more accurate. And quick measurements can be made.

FIG. 11 is an explanatory diagram of the hardware configuration of the measuring device 500.
The measuring device 500 includes a processor 1001, a storage device 1002, an input device 1003, an output device 1004, a measuring device 1005, a clip 505, and a connection terminal 1006.
The connection terminal 1006 of the measurement device 500 is electrically connected to the electrode 102 of the biosensor 100.
The processor 1001, storage device 1002, input device 1003, output device 1004, and measurement device 1005 are configured to be able to exchange data with each other via a bus 1007.

Processor 1001 controls measurement device 500. The storage device 1002 becomes a work area for the processor 1001. Furthermore, the storage device 1002 is a non-temporary or temporary recording medium that stores various programs and data.

Examples of the processor 1001 include a processor (CPU), microprocessor, processor core, multiprocessor, ASIC (application-specific integrated circuit), FPGA (field programmable gate array), and GPGPU (gener). al-purpose computing on graphics processing units ) etc.

Examples of the storage device 1002 include ROM (Read Only Memory), RAM (Random Access Memory), HDD (Hard Disk Drive), flash memory, and SSD (Solid State Drive).

Input device 1003 inputs data.
The input device 1003 includes a touch panel 502. In addition to the touch panel, the input device 1003 includes, for example, a keyboard, buttons, a mouse, a numeric keypad, a scanner, and the like.
Output device 1004 outputs data.
The output device 1004 includes a touch panel 502. In addition to the touch panel, the output device 1004 includes, for example, a display and a printer.

The measurement device 1005 controls the electrodes 102 (working electrode, counter electrode, and reference electrode) of the biosensor 100 via electrically connected connection terminals to measure electrochemical measurements of the sample. Typically a potentio/galvanostat can be used.

FIG. 12 is a functional block diagram of the measuring device 500.
The measuring device 500 includes a control section 1101, a storage section 1102, an input section 1103, an output section 1104, a pressure sealing section 1105, and an electrochemical measurement section 1106.

The control unit 1101 includes a processor 1001, and controls the following units to realize each function of the measuring device 500.

The pressurized sealing part 1105 is a function realized by the clip 505. By holding the biosensor 100 between the clips 505, the electrolyte gel 107 is pressed toward the electrode 102 through the cover member 109, and the electrode 102 and the electrolyte gel 107 are brought into close contact with each other, so that measurement can be performed under pressure. . Furthermore, the opening 113 of the biosensor 100 is closed by the spacer 902, and the inside of the cell, which is made up of the through hole 111, the cavity CAV, and the space between the electrolyte gel 107 and the lower base material 101, is isolated from the outside air. Furthermore, when the electrolyte gel 107 contains an oxygen adsorbent, it becomes easier to maintain the inside of the cell as an anaerobic environment, and better sensitivity can be obtained.

The electrochemical measurement unit 1106 is realized by a program stored in the storage unit 1102 executed by the control unit 1101, and a measurement device 1005 controlled thereby, a connection terminal 1006 connected to the electrode 102 of the biosensor 100, etc. This is a function that is
Specifically, by measuring the electrode potential of the working electrode 103 or measuring the magnitude of the flowing current, data for evaluating the metabolic activity, etc. of the microorganism to be measured can be obtained.

The input unit 1103 is a function realized by the input device 1003 controlled by the control unit 1101 executing a program stored in the storage unit 1102. The input unit 1103 receives a user's instruction to start measurement, input of various information, and the like.
The output unit 1104 is a function that is realized by a program stored in the storage unit 1102 executed by the control unit 1101 and an output device 1004 controlled thereby.
The output unit 1104 displays the status of the measuring device 500, the progress of measurement, the measurement results, etc. on the output device 1004.

Next, the operation of measuring device 500 will be explained.
The measuring device 500 operates as follows according to the program. FIG. 13 is an operation flow of the control unit 1101 of the measuring device 500 that operates according to the above program.

Typically, the above operation is started by receiving an instruction from the user to start measurement through the input unit. The biosensor 100 is typically prepared by a user and inserted into the insertion port 506 of the measurement device 500 prior to operation of the measurement device 500.

First, the biosensor 100 is inserted into the insertion port 506 of the measurement device 500, the clip 505 is closed, the electrolyte gel 107 is pressurized via the cover member 109, and the opening 113 is closed.
Note that when the measuring device 500 has a clip opening/closing mechanism, the control unit 1101 executes the program stored in the storage unit 1102, controls the clip 505 to close it, and adds the electrolyte gel 107. You can also press it.

When the input unit 1103 receives an instruction to start measurement, the electrochemical measurement unit 1106 controls the electrode 102 of the biosensor 100, and the electrochemical properties of the sample are measured (step S1201).
Next, the measurement results are displayed on the output device 1004 (step S1202).

This measuring device has a pressurized sealing section that sandwiches the lower and upper substrates from both sides and presses the electrolyte gel against the working electrode, allowing it to detect signals originating from microbial metabolism with higher sensitivity. can be detected. This measuring device can more sensitively and quickly obtain information on metabolism derived from microorganisms in samples of saliva, sputum, and other human fluids, making it possible to diagnose various diseases. , can be used for progress management, etc.

100: Biosensor 101: Lower base material 102: Electrode 103: Working electrode 104: Counter electrode 105: Working electrode 106: Spacer layer 107: Electrolyte gel 108: Upper base material 109: Cover member 110: Recessed part 111: Through hole 112: Opening 113: Opening 114: Second notch 115: First notch 207: Electrolyte gel 500: Measuring device 501: Main body 502: Touch panel 503: Upper tooth 504: Lower tooth 505: Clip 506: Insertion port 507: Hinge 901: Base material 902: Spacer 1001: Processor 1002: Storage device 1003: Input device 1004: Output device 1005: Measurement device 1006: Connection terminal 1007: Bus 1101: Control section 1102: Storage section 1103: Input section 1104: Output section 1105 : Pressure sealed part 1106 : Electrochemical measurement part

Claims (6)

a lower base material, at least two working electrodes disposed on a surface A of the lower base material and extending from one end of the lower base material in a direction along the surface A to approximately the center of the lower base material; an electrolyte gel disposed on the electrode so as to be in direct contact with at least a portion of the electrode, a cover member that covers the electrolyte gel in contact with a surface of the electrolyte gel opposite to the electrode side, and the A biosensor having an upper substrate supporting a cover member and disposed on the lower substrate, an electric current connected to the electrode and measuring electrochemical properties of microorganisms sandwiched between the working electrode and the electrolyte gel. Chemical measurement department,
A measuring device comprising: a pressurizing and sealing part that presses and presses the electrolyte gel against the working electrode while sandwiching the lower base material and the upper base material from both sides. The measuring device according to claim 1, wherein the electrolyte gel contains a hydrophilic polymer, an electrolyte, and an oxygen adsorbent. Further comprising a spacer layer between the upper base material and the lower base material,
The upper base material has a through hole along the thickness direction from one surface B to the other surface C,
The spacer layer is recessed in a shape that fits around the outer periphery of the electrolyte gel, and the surface of the spacer layer is formed from the side surface of the recessed portion toward the opening of the through hole disposed on the surface B. a first notch extending in the direction along the
A cavity defined by the surface A of the upper base material, the surface B, and the side surface of the first notch communicates with the through hole through the opening, and The measuring device according to claim 1 or 2, wherein a flow path leading to the electrolyte gel is formed. The spacer layer and the upper base material are
The electrode further includes a second notch extending in a direction parallel to the surface B from the side surface to the center thereof, and a part of the electrode is placed in a recess defined by the second notch and the surface A. 4. The measuring device according to claim 3, which is exposed. According to claim 4, the pressurized sealing part has a pair of plate-like members whose distance from each other can be adjusted, and the clearance distance when the plate-like members are brought closest is equal to or less than the maximum thickness of the biosensor. Measuring device as described. The pressure sealing part further includes a spacer disposed on the surface of at least one of the pair of plate-like members, and when the pressure is applied, the spacer closes the opening of the through hole on the surface C side. The measuring device according to claim 5.