JP2019070557A - Calorimetric sensor, manufacturing method therefor, and inspection device using the same - Google Patents

Abstract

The present invention provides a colorimetric biosensor with improved detection sensitivity.
A reference electrode (2) is formed on a thermoelectric conversion thin film (10). The reaction electrode 4 is formed on the thermoelectric conversion thin film 10, and the enzyme that reacts with the sample is fixed. The calorimetric sensor 1 is configured to be able to measure the potential difference between the reference electrode 2 and the reaction electrode 4. The reference electrode 2 and the reaction electrode 4 are separated in a direction inclined with respect to one side of the chip on which the calorimetric sensor 1 is formed.
[Selected figure] Figure 2

Description

  The present invention relates to a calorimetric sensor.

  Biosensors are used to detect specific components in liquid samples. A calorimetric method has been proposed as one of the biosensor methods. A calorimetric biosensor (hereinafter referred to as a calorimetric biosensor or simply a calorimetric sensor) includes a pair of electrodes forming a thermocouple. An enzyme that reacts with the target substance in the solution is applied to one of the reaction electrodes, and heat is generated by the reaction between the enzyme and the target substance. The other reference electrode is thermally isolated from the reaction electrode so that the temperature does not change. When a temperature rise corresponding to the concentration of the target material occurs in the reaction electrode, a potential difference corresponding to the temperature rise is observed (see Patent Document 1 and Patent Document 2).

Unexamined-Japanese-Patent No. 2015-200611 gazette JP, 2016-173273, A Unexamined-Japanese-Patent No. 2015-21779 JP 2012-242335 A JP-A-61-212750

  The present invention has been made in such a situation, and one of the exemplary objects of one embodiment of the present invention is to provide a calorimetric biosensor with improved detection sensitivity.

  One aspect of the present invention relates to a colorimetric sensor. The calorimetric sensor comprises a thermoelectric conversion thin film, a reference electrode formed on the thermoelectric conversion thin film, and a reaction electrode formed on the thermoelectric conversion thin film and on which an enzyme that reacts with an analyte is fixed. The potential difference between the electrode and the reaction electrode is configured to be measurable. The reference electrode and the reaction electrode are separated in a direction inclined with respect to one side of the chip on which the colorimetric sensor is formed.

  Another aspect of the present invention relates to an inspection apparatus. The inspection apparatus includes the above-described colorimetric sensor, a sense amplifier that generates a detection signal according to the potential difference between the reference electrode of the calorimetric sensor and the reaction electrode, and a processing unit that processes the detection signal. The processing unit offsets the detection signal such that (i) the value after the lapse of the first predetermined time from the start of the reaction becomes the first predetermined value, and (ii) the waveform of the detection signal after the offset has a second predetermined value. Integration is performed over a second predetermined time from the time that is a value.

  Another aspect of the present invention relates to a method of manufacturing a colorimetric sensor. The manufacturing method includes the steps of ion implantation into an SOI (Silicon On Insulator) substrate, forming an ohmic electrode, forming a SiO2 layer and etching a part, forming a wiring, and forming a SiO2 layer. The method includes the steps of forming and partially etching, forming an electrode pad, removing a device layer and a BOX layer of an SOI substrate, and removing a support layer immediately below a sensing region.

  It is to be noted that any combination of the above-mentioned constituent elements, or one in which the constituent elements or expressions of the present invention are mutually substituted among methods, apparatuses, etc. is also effective as an aspect of the present invention.

  According to an aspect of the present invention, a colorimetric biosensor with improved detection sensitivity can be provided.

It is a perspective view of a calorimetric sensor concerning an embodiment. FIGS. 2A and 2B are a plan view and a cross-sectional view of a calorimetric sensor according to an embodiment. It is a perspective view of a thermoelectric conversion thin film. It is a figure which shows an example of wiring structure. FIGS. 5A and 5B illustrate the use of a calorimetric sensor. FIGS. 6 (a) and 6 (b) illustrate the advantages of the calorimetric sensor. FIGS. 7 (a) to 7 (d) are cross-sectional views showing a method of manufacturing a calorimetric sensor. 8 (a) to 8 (c) are cross-sectional views showing a method of manufacturing a calorimetric sensor. FIG. 2 shows an inspection device with a calorimetric sensor. FIGS. 10 (a) and 10 (b) are diagrams showing measurement results by the calorimetric sensor. It is a figure which shows the output of the sensor which gave arithmetic processing. It is a figure which shows the value of integrating the waveform of FIG. 11 over the predetermined time from time 0, and the relationship of density.

(Overview)
A calorimetric sensor is disclosed herein as an embodiment of the present invention. The calorimetric sensor comprises a thermoelectric conversion thin film, a reference electrode formed on the thermoelectric conversion thin film, and a reaction electrode formed on the thermoelectric conversion thin film and on which an enzyme that reacts with an analyte is fixed. The potential difference between the electrode and the reaction electrode can be measured, and the reference electrode and the reaction electrode are spaced apart in a direction inclined with respect to one side of the chip on which the colorimetric sensor is formed.

  According to this embodiment, the distance between the reference electrode and the reaction electrode can be increased compared to when the reference electrode and the reaction electrode are separated in the direction parallel to one side of the chip, so that their thermal isolation can be enhanced. , Detection sensitivity can be improved.

  In one embodiment, the calorimetric sensor may further comprise a substrate supporting the thermoelectric conversion thin film. In the rectangular sensing area including the reaction electrode and the reference electrode, the lower portion of the thermoelectric conversion thin film may be a cavity. Since the heat capacity of the sensing area can be reduced by this, the detection sensitivity can be further improved.

  In one embodiment, the thermoelectric conversion thin film and the substrate may be an outermost device layer and a lowermost support layer of an SOI (Silicon On Insulator) substrate, respectively. Thereby, the thermoelectric conversion thin film and the base can be easily formed.

  The reaction electrode may be located substantially at the center of the sensing area, and the reference electrode may be located at a corner of the sensing area. Thereby, the reaction electrode can be kept away from the surrounding pads and other circuit elements, and the detection sensitivity can be enhanced.

In one embodiment, the thermoelectric conversion thin film has a first portion on which a reaction electrode is formed, a second portion formed in an I / O region surrounding the sensing region, and a first corner of the sensing region at one end. And the other end is connected to the second portion at the second corner diagonal to the first corner of the sensing area, and the other end is connected to the second portion at the other end. And a second bridge portion connected to the first portion.
Thereby, the reaction electrode can be supported by the beam structure of the first bridge portion and the second bridge portion.

  In one embodiment, the calorimetric sensor connects the first pad and the reference electrode via the first bridge portion or the second bridge with the first pad and the second pad formed in the I / O region. The semiconductor device may further include a first wire and a second wire connecting the second pad and the reaction electrode via the first bridge portion or the second bridge.

  In one embodiment, the reaction electrode may be a rectangle inclined 45 degrees to the tip. Thereby, the thermal isolation between the reaction electrode and the reference electrode can be enhanced as compared with the case where the reaction electrodes of the same size are rectangular parallel to the chip.

  In one embodiment, the calorimetric sensor may further include a first sealing tape provided on the surface of the calorimetric sensor and having an opening overlapping the reaction electrode.

  In one embodiment, the calorimetric sensor may further include a second sealing tape provided on the back surface of the calorimetric sensor. This can prevent the sample from infiltrating in the space below the sensing area.

Embodiment
Hereinafter, the present invention will be described based on preferred embodiments with reference to the drawings. The same or equivalent components, members, and processes shown in the drawings are denoted by the same reference numerals, and duplicating descriptions will be omitted as appropriate. In addition, the embodiments do not limit the invention and are merely examples, and all the features and combinations thereof described in the embodiments are not necessarily essential to the invention.

  The dimensions (thickness, length, width, etc.) of each member described in the drawings may be scaled as appropriate for ease of understanding. Furthermore, the dimensions of a plurality of members do not necessarily represent the magnitude relationship between them, and even if one member A is drawn thicker than another member B in the drawing, the member A is a member B It may be thinner than that.

  FIG. 1 is a perspective view of a colorimetric sensor 1 according to the embodiment. FIGS. 2A and 2B are a plan view and a cross-sectional view of the colorimetric sensor 1 according to the embodiment.

  The colorimetric sensor 1 is, for example, a rectangular chip 40 of several millimeters square. A rectangular area near the center of the chip 40 is referred to as a sensing area 50, and a rectangular area surrounding it is referred to as an I / O area 52.

  The colorimetric sensor 1 includes a reference electrode 2 and a reaction electrode 4 included in a sensing area 50. On the surface of the reaction electrode 4, an enzyme that reacts with a specific component (substrate of the sample) in the liquid sample is fixed. The method for fixing the enzyme is not particularly limited. For example, the enzyme may be mixed with a water-soluble photosensitive resin (for example, Biosurfine manufactured by Toyo Gosei Co., Ltd.), applied to the reaction electrode 4 and irradiated with ultraviolet light to be cured. Alternatively, an enzyme adhesive layer may be formed on the surface of the reaction electrode 4, and the enzyme may be adsorbed to the adhesive layer.

  The calorimetric sensor 1 is configured to be able to measure the potential difference ΔV between the reference electrode 2 and the reaction electrode 4. In the I / O area 52, a first pad PAD1 and a second pad PAD2 are provided. The first pad PAD1 is electrically connected to the reference electrode 2 via a first wiring (not shown), and the second pad PAD2 is electrically connected to the reaction electrode 4 via a second wiring (not shown). For example, the potentials can be measured by bringing the probes 60 and 62 into contact with the first pad PAD1 and the second pad PAD2.

  As shown in FIG. 1 or 2 (a), the reference electrode 2 and the reaction electrode 4 are separated in a direction inclined with respect to one side of the chip 40 on which the colorimetric sensor 1 is formed.

  Please refer to FIG. 2 (b). The reference electrode 2 and the reaction electrode 4 are formed on the thermoelectric conversion thin film 10. The material of the thermoelectric conversion thin film 10 is not particularly limited, but it is preferable to use Si (silicon). In addition to having a large Seebeck coefficient, Si has the advantage of being highly compatible with general semiconductor manufacturing processes.

  As shown in FIG. 2 (b), the thermoelectric conversion thin film 10 is supported by the base 20 in the I / O region 52. The base 20 is provided with a space 22 in a rectangular sensing area 50 including the reaction electrode 4 and the reference electrode 2. For example, the base 20 is in the shape of a rectangle surrounding the sensing area 50.

In the present embodiment, the colorimetric sensor 1 is formed on an SOI (Silicon On Insulator) substrate 42. The SOI substrate 42 comprises a Si support layer 44, a Si device layer 48, and an SiO ₂ insulating layer (BOX layer: Buried Oxide) 46 therebetween. The thermoelectric conversion thin film 10 is a Si device layer 48, and the substrate 20 is a Si support layer 44. For example, the thickness of the Si support layer 44 is 350 μm, and the Si device layer 48 is 5 μm.

  The reaction electrode 4 is a rectangle inclined 45 degrees with respect to the chip 40, and is located substantially at the center of the sensing area 50. On the other hand, the reference electrode 2 is located at the corner of the sensing area 50.

  As shown in FIG. 2 (b), the first sealing tape 6 is provided on the surface of the calorimetric sensor 1 and has an opening 7 which overlaps the reaction electrode 4. The back surface of the colorimetric sensor 1 is sealed by a second sealing tape 8. The first sealing tape 6 and the second sealing tape 8 may be heat-resistant adhesive tapes (polyimide films).

  As shown in FIG. 1, the thermoelectric conversion thin film 10 has a crosslinked structure. FIG. 3 is a perspective view of the thermoelectric conversion thin film 10. The thermoelectric conversion thin film 10 includes a first portion 12, a second portion 14, a first bridge portion 16, and a second bridge portion 18. The first portion 12 is a portion on which the reaction electrode 4 is formed. The second portion 14 is a portion corresponding to the I / O region 52. The first portion 12 is supported above the space 22 by the first bridge portion 16 and the second bridge portion 18. One end of the first bridge portion 16 is connected to the second portion 14 at the first corner C1 of the sensing region 50, and the other end is connected to the first portion 12. One end of the second bridge portion 18 is connected to the second portion 14 at a second corner C 2 diagonal to the first corner C 1 of the sensing region 50, and the other end is connected to the first portion 12.

  The width of the first bridge portion 16 and the second bridge portion 18 is preferably as narrow as possible. As described later, in the case of forming the wiring in the first bridge portion 16 (the second bridge portion 18), it is preferable to make the wiring as narrow as possible within the range in which the desired wiring can be formed. Thereby, the heat capacity of the portion contributing to the sensing of the thermoelectric conversion thin film 10 can be reduced, and the detection sensitivity can be enhanced. For example, the width of the first bridge portion 16 and the second bridge portion 18 may be about 50 μm to 200 μm.

  FIG. 4 is a view showing an example of the wiring structure. In the first bridge portion 16 of the thermoelectric conversion thin film 10, a first wire W1 and a second wire W2 are formed. The first wiring W1 connects the first pad PAD1 and the reference electrode 2 via the first bridge portion 16. The second wiring W2 connects the second pad PAD2 and the reaction electrode 4 via the first bridge portion 16. The first wire W1 is in contact with the reference electrode 2 through the via hole VIA1, and the second wire W2 is in contact with the reaction electrode 4 through the via hole VIA2. An interlayer insulating film (not shown) is formed between the wiring layer in which the first wiring W1 and the second wiring W2 are formed and the wiring layer in which the reference electrode 2 and the reaction electrode 4 are formed.

The above is the structure of the calorimetric sensor 1. Subsequently, its use will be described.
FIGS. 5A and 5B illustrate the use of the calorimetric sensor 1. As shown in FIG. 5A, a droplet 80 of a liquid sample may be dropped and used in the opening 7 of the first sealing tape 6. Alternatively, as shown in FIG. 5 (b), the colorimetric sensor 1 may be immersed in a container 84 filled with the liquid sample 82.

(First advantage)
FIG. 6A is a view for explaining the first advantage of the colorimetric sensor 1. The reference electrode 2 and the reaction electrode 4 are separated in a direction inclined with respect to one side of the chip. The distance between the two electrodes at this time is l ₁ . In FIG. 5, the reference electrode 3 separated in a direction parallel to one side of the chip is shown by a broken line with respect to the reaction electrode 4 for comparison, and the distance between the reference electrode 3 and the reaction electrode 4 is l ₂ is there. As is clear from FIG. 6A, l ₁ > l ₂ holds, and the thermal isolation of the two electrodes can be enhanced by laying out the two electrodes in a diagonal direction. Since the temperature of the reference electrode 2 is less susceptible to the thermal reaction in the reaction electrode 4, the detection sensitivity can be enhanced.

(Second advantage)
FIG. 6 (b) is a diagram for explaining the second advantage of the colorimetric sensor 1. The reaction electrode 4 is a rectangle inclined 45 degrees with respect to the chip. In FIG. 6 (b), for comparison, is shown the reaction electrode 5 are not inclined with respect to the chip, the distance of the reference electrode 2 and the reaction electrode 5 is l _3.
As apparent from FIG. 6 (b), l ₁ > l ₃ holds, and the thermal isolation between the reaction electrode 4 and the reference electrode 2 can be enhanced by inclining the reaction electrode 4. Since the temperature of the reference electrode 2 is less susceptible to the thermal reaction in the reaction electrode 4, the detection sensitivity can be enhanced.

(Third advantage)
As shown in FIGS. 1 and 2B, in the sensing region 50, a space 22 is provided immediately below the reaction electrode 4. Thereby, the heat capacity of the portion (mainly the thermoelectric conversion thin film 10) contributing to the sensing of the calorimetric sensor 1 can be reduced, and the detection sensitivity can be enhanced.

  Subsequently, an example of a method of manufacturing the colorimetric sensor 1 will be described. FIGS. 7A to 7D and FIGS. 8A to 8C are cross-sectional views showing a method of manufacturing the colorimetric sensor 1.

  As shown in FIG. 7A, in the device layer 48 of the SOI (Silicon On Insulator) substrate 42, ions are implanted into the regions 102 and 104 where the reference electrode 2 and the reaction electrode 4 are to be formed. Subsequently, as shown in FIG. 7B, the ohmic electrodes 112 and 114 functioning as the reference electrode 2 and the reaction electrode 4 are formed in the regions 102 and 104 in which the ion implantation has been performed. As a material of the electrode, Al (aluminum) can be used, and after sputtering, it is sintered.

Subsequently, as shown in FIG. 7C, a SiO ₂ layer 120 is formed, and partial regions 122 and 124 are etched. The region 122 is a region for forming a via hole for taking a contact between the reference electrode 2 and the first wiring W1 to be formed later, and the region 124 is a reaction electrode 4 and the second wiring W2 to be formed later Area for forming a via hole for making a contact.

  Subsequently, as shown in FIG. 7D, the wiring layer 130 is formed. The wiring layer 130 includes a first wiring W1 and a second wiring W2. Further, the wiring layer includes the land area 132 of the first pad PAD1 (second pad PAD2).

Subsequently, as shown in FIG. 8A, a SiO ₂ layer 140 is formed, and then a partial region thereof is etched to provide openings 142 and 144. The opening 144 is formed in a portion corresponding to the reaction electrode 4. The opening 142 is formed in a region where the first pad PAD1 and the second pad PAD2 are to be formed.

  Subsequently, as shown in FIG. 8B, the electrode pads 152 and 154 are formed on the openings 142 and 144, respectively. The electrode pad 152 corresponds to the first pad PAD1. The electrode pad 154 is a reaction electrode 4.

  Subsequently, portions of the Si device layer 48 and the BOX layer 46 of the SOI substrate 42 are removed from the upper side. This situation is not shown in FIG. 8 (c). Furthermore, the Si support layer 44 immediately below the sensing region 50 of the SOI substrate 42 is removed.

  Thereafter, the first sealing tape 6 is attached to the surface of the colorimetric sensor 1 and the second sealing tape 8 is attached to the back surface.

  FIG. 9 is a view showing an inspection apparatus 200 provided with the calorimetric sensor 1. The inspection apparatus 200 includes a sense amplifier 202 and a processing unit 210 in addition to the colorimetric sensor 1. The sense amplifier 202 amplifies the potential difference between the first pad PAD1 and the second pad PAD2, in other words, between the reference electrode 2 and the reaction electrode 4. The output voltage of the sense amplifier 202 is converted by an A / D converter (digitizer) 204 into a digital detection signal. The processing unit 210 acquires the concentration of the specific component in the liquid sample based on the detection signal. The sense amplifier 202 and the A / D converter 204 may be integrated on the same chip as the colorimetric sensor 1.

  FIGS. 10A and 10B are diagrams showing the measurement results by the calorimetric sensor 1. The chip size of the colorimetric sensor 1 used for measurement is 5 mm square, and the reaction electrode 4 is 1 mm square. Creatinase was used as an enzyme, and the colorimetric sensor 1 was immersed in creatinine solutions of various concentrations. As a preparation, the container was filled with saline, and after drawing out the saline, the creatinine solution was immediately injected. FIG. 10 (a) is a solution with a concentration of 20 mg / dl, and FIG. 10 (b) is a solution with a concentration of 50 mg / dl.

  From the waveforms of FIGS. 10A and 10B, it may be difficult to determine the concentration. Therefore, the processing unit 210 may perform waveform shift and the like.

FIG. 11 is a diagram showing an output of a sensor subjected to arithmetic processing. Here, (i) the detection signal is offset so that the value after the elapse of the first predetermined time T ₁ (five minutes) from the start of the reaction becomes the first predetermined value (here, 0). Further, (ii) in the waveform of the detection signal after offset, the time at which the value is the second predetermined value (−300 μV) is taken as time 0.

  As shown in FIG. 11, it can be seen that the time constant of the waveform after arithmetic processing depends on the concentration. FIG. 12 is a diagram showing the relationship between the value obtained by integrating the waveform of FIG. 11 over a predetermined time from time 0 and the concentration. As can be seen from FIG. 12, it can be seen that there is a correlation between the integrated value and the concentration of the waveform after arithmetic processing.

  From these results, according to the colorimetric sensor 1 according to the embodiment, it has been demonstrated that the concentration of the sample in the sample is detected.

  Although the present invention has been described based on the embodiments, the embodiments only show the principles and applications of the present invention, and the embodiments deviate from the concept of the present invention defined in the claims. However, many variations and modifications of the arrangement are permitted.

<Modification>
In the embodiment, the concentration is measured by immersing the calorimetric sensor 1 in a liquid. In this method, the reaction time is extended to several minutes because of the large volume of liquid. Therefore, a minute flow path (reaction chamber) may be formed on the upper side of the calorimetric sensor 1, and the sample solution confined in the reaction chamber may be measured.

Reference Signs List 1 calorimetric sensor 2 reference electrode 4 reaction electrode 10 thermoelectric conversion thin film 12 first portion 14 second portion 16 first bridge portion 18 second bridge portion 20 base 6 first sealing tape 8 second sealing tape W1 first wiring W2 Second Wiring 40 Chip 42 SOI Substrate 44 Si Support Layer 46 SiO2 Insulating Layer 48 Si Device Layer PAD1 First Pad PAD2 Second Pad 50 Sensing Area 52 I / O Area 60, 62 Probe 200 Inspection Device

Claims (12)

Thermoelectric conversion thin film,
A reference electrode formed on the thermoelectric conversion thin film;
A reaction electrode which is formed on the thermoelectric conversion thin film and on which an enzyme which reacts with the sample is fixed;
And the potential difference between the reference electrode and the reaction electrode can be measured,
A colorimetric sensor characterized in that the reference electrode and the reaction electrode are separated in a direction inclined with respect to one side of a chip on which the calorimetric sensor is formed. The substrate further comprises a substrate supporting the thermoelectric conversion thin film,
The calorimetric sensor according to claim 1, wherein the substrate is provided with a space in a rectangular sensing area including the reaction electrode and the reference electrode.   The calorimetric sensor according to claim 2, wherein the thermoelectric conversion thin film and the substrate are respectively a Si device layer as an outermost layer of an SOI (Silicon On Insulator) substrate and a Si support layer as a lowermost layer. The reaction electrode is located approximately at the center of the sensing area,
The calorimetric sensor according to claim 2, wherein the reference electrode is located at a corner of the sensing area. The thermoelectric conversion thin film is
A first portion on which the reaction electrode is formed;
A second portion formed in an I / O region surrounding the sensing region;
A first bridge portion having one end connected to the second portion at a first corner of the sensing area and the other end connected to the first portion;
A second bridge portion having one end connected to the second portion at a second corner diagonal to the first corner of the sensing area and the other end connected to the first portion;
The calorimetric sensor according to any one of claims 2 to 4, comprising First and second pads formed in the I / O region;
A first wire which is in contact with the reference electrode and connected to the first pad via the first bridge portion or the second bridge portion;
A second wire which is in contact with the reaction electrode and is connected to the second pad via the first bridge portion or the second bridge portion;
The calorimetric sensor according to claim 5, further comprising:   The colorimetric sensor according to any one of claims 1 to 6, wherein the reaction electrode is a rectangle inclined 45 degrees with respect to the chip.   The calorimetric sensor according to any one of claims 1 to 7, further comprising a first sealing tape provided on the surface of the calorimetric sensor and having an opening overlapping the reaction electrode.   The colorimetric sensor according to any one of claims 1 to 8, further comprising a second sealing tape provided on the back surface of the colorimetric sensor. A calorimetric sensor according to any one of claims 1 to 9,
A sense amplifier that generates a detection signal according to the potential difference between the reference electrode and the reaction electrode;
A processing unit that processes the detection signal;
An inspection apparatus comprising:   The processing unit offsets the detection signal such that (i) the value after a first predetermined time has elapsed from the start of the reaction becomes a first predetermined value, and (ii) the waveform of the detection signal after the offset has a value 11. The inspection apparatus according to claim 10, wherein integration is performed over a second predetermined time from a time that is a second predetermined value. A method of manufacturing a colorimetric sensor, comprising
Implanting ions into an SOI (Silicon On Insulator) substrate;
Forming an ohmic electrode;
Forming a SiO ₂ layer and partially etching it;
Forming a wire;
Forming a SiO ₂ layer and partially etching it;
Forming an electrode pad;
Removing a portion of the device layer and the BOX layer of the SOI substrate;
Removing the support layer directly below the sensing area;
A manufacturing method comprising: