Patent Grant
11189476
The present disclosure relates to a sample support body, an ionization method, and a mass spectrometry method.
Conventionally, a laser desorption/ionization method is known as a method of ionizing a sample such as a biological sample to perform, for instance, mass spectrometry. As a sample support body used in the laser desorption/ionization method, one including a substrate in which a plurality of through-holes are formed and a conductive layer that is provided on at least one surface of the substrate is described in Patent Literature 1.
Patent Literature 1: Japanese Patent No. 6093492
Components of the ionized sample are detected in mass spectrometry, and mass spectrometry of the sample is performed on the basis of detection results. Therefore, an improvement in signal intensity (sensitivity) of the components of an ionized sample is desired in mass spectrometry.
The present disclosure is directed to provide a sample support body, an ionization method, and a mass spectrometry method capable of improving a signal intensity of components of an ionized sample in mass spectrometry.
According to an aspect of the present disclosure, there is provided a sample support body for ionization of a sample. The sample support body includes a substrate including a first surface and a second surface on sides opposite to each other, a first conductive layer provided on the first surface, and a second conductive layer provided on the second surface. A plurality of through-holes opening on the first surface and the second surface are formed in a predetermined region of the substrate, the predetermined region being for ionizing components of the sample on. A width of a first opening on the first surface side is larger than a width of a second opening on the second surface side in each of the plurality of through-holes.
In this sample support body, the width of the first opening on the first surface side is larger than the width of the second opening on the second surface side in each of the plurality of through-holes. For this reason, for example, when a solution including the sample is dropped to the plurality of through-holes from the first surface side, the solution moves to the second surface side through the plurality of through-holes, and components of the sample in the solution stay on the first surface side in an appropriate state. Therefore, when the first surface is irradiated with an energy beam while a voltage is applied to the first conductive layer, the components of the sample are reliably ionized. Further, for example, when the sample support body is disposed such that the first surface faces the sample, the components of the sample move smoothly to the second surface side through the plurality of through-holes and stay on the second surface side in an appropriate state. Therefore, when the second surface is irradiated with an energy beam while a voltage is applied to the second conductive layer, the components of the sample are reliably ionized. Thus, according to this sample support body, it is possible to improve the signal intensity of components of an ionized sample in mass spectrometry.
In the sample support body according to the aspect of the present disclosure, when viewed in a direction in which the first surface and the second surface are opposite to each other, an outer edge of the first opening may be located outside an outer edge of the second opening in each of the plurality of through-holes. Accordingly, for example, when a solution including the sample is dropped to the plurality of through-holes from the first surface side, components of the sample in the solution can stay on the first surface side in a more appropriate state. Further, for example, when the sample support body is disposed such that the first surface faces the sample, the components of the sample can move more smoothly to the second surface side through the plurality of through-holes, and the components of the sample can stay on the second surface side in a more appropriate state.
In the sample support body according to the aspect of the present disclosure, each of the plurality of through-holes may include a first portion on the first opening side and a second portion on the second opening side. The first portion may have a funnel shape expanding toward the first opening. Alternatively, in the sample support body according to the aspect of the present disclosure, each of the plurality of through-holes may have a frustum shape expanding toward the first opening. In both cases, for example, when a solution including the sample is dropped to the plurality of through-holes from the first surface side, components of the sample in the solution can stay on the first surface side in an appropriate state. Further, for example, when the sample support body is disposed such that the first surface faces the sample, the components of the sample can move smoothly to the second surface side through the plurality of through-holes, and the components of the sample can stay on the second surface side in an appropriate state.
In the sample support body according to the aspect of the present disclosure, a minimum value of the width may be 1 nm and a maximum value of the width may be 700 nm in each of the plurality of through-holes. Accordingly, for example, when a solution including the sample is dropped to the plurality of through-holes from the first surface side, components of the sample in the solution can stay on the first surface side in an appropriate state. Further, for example, when the sample support body is disposed such that the first surface faces the sample, the components of the sample can move smoothly to the second surface side through the plurality of through-holes, and the components of the sample can stay on the second surface side in an appropriate state.
According to another aspect of the present disclosure, there is provided a sample support body for ionization of a sample. The sample support body includes a conductive substrate including a first surface and a second surface on sides opposite to each other. A plurality of through-holes opening on the first surface and the second surface are formed in a predetermined region of the substrate, the predetermined region being for ionizing components of the sample. A width of a first opening on the first surface side is larger than a width of a second opening on the second surface side in each of the plurality of through-holes.
In this sample support body, the width of the first opening on the first surface side is larger than the width of the second opening on the second surface side in each of the plurality of through-holes. For this reason, for example, when a solution including the sample is dropped to the plurality of through-holes from the first surface side, the solution moves to the second surface side through the plurality of through-holes, and components of the sample in the solution stay on the first surface side in an appropriate state. Therefore, when the first surface is irradiated with an energy beam while a voltage is applied to the substrate, the components of the sample are reliably ionized. Further, for example, when the sample support body is disposed such that the first surface faces the sample, the components of the sample move smoothly to the second surface side through the plurality of through-holes, and stay on the second surface side in an appropriate state. Therefore, when the second surface is irradiated with an energy beam while a voltage is applied to the substrate, the components of the sample are reliably ionized. Thus, according to this sample support body, it is possible to improve the signal intensity of components of an ionized sample in mass spectrometry.
According to another aspect of the present disclosure, there is provided an ionization method including a first process of preparing the sample support body provided with the first conductive layer and the second conductive layer described above, a second process of mounting the sample support body on a mount surface of a mount portion such that the second surface faces the mount surface, and dropping a solution including the sample to the plurality of through-holes from the first surface side, and a third process of ionizing components of the sample staying on the first surface side by irradiating the first surface with an energy beam while applying a voltage to the first conductive layer.
According to another aspect of the present disclosure, there is provided an ionization method including a first process of preparing the sample support body provided with the first conductive layer and the second conductive layer described above, a second process of mounting the sample on a mount surface of a mount portion, and mounting the sample support body on the mount surface such that the first surface faces the sample, and a third process of ionizing components of the sample having moved to the second surface side through the plurality of through-holes by irradiating the second surface with an energy beam while applying a voltage to the second conductive layer.
According to these ionization methods, since the sample support body provided with the first conductive layer and the second conductive layer described above is used, it is possible to improve the signal intensity of components of an ionized sample in mass spectrometry.
According to another aspect of the present disclosure, there is provided a mass spectrometry method including the first process, the second process, and the third process of the ionization method described above; and a fourth process of detecting the components ionized in the third process.
According to this mass spectrometry method, since the sample support body provided with the first conductive layer and the second conductive layer described above is used, it is possible to improve the signal intensity of components of an ionized sample in mass spectrometry.
According to another aspect of the present disclosure, there is provided an ionization method including a first process of preparing the sample support body provided with the conductive substrate described above, a second process of mounting the sample support body on a mount surface of a mount portion such that the second surface faces the mount surface, and dropping a solution including the sample to the plurality of through-holes from the first surface side, and a third process of ionizing components of the sample staying on the first surface side by irradiating the first surface with an energy beam while applying a voltage to the substrate.
According to another aspect of the present disclosure, there is provided an ionization method including a first process of preparing the sample support body provided with the conductive substrate described above, a second process of mounting the sample on a mount surface of a mount portion, and mounting the sample support body on the mount surface such that the first surface faces the sample, and a third process of ionizing components of the sample having moved to the second surface side through the plurality of through-holes by irradiating the second surface with an energy beam while applying a voltage to the substrate.
According to these ionization methods, since the sample support body provided with the conductive substrate described above is used, it is possible to improve the signal intensity of components of an ionized sample in mass spectrometry.
According to another aspect of the present disclosure, there is provided a mass spectrometry method including the first process, the second process, and the third process of the ionization method described above; and a fourth process of detecting the components ionized in the third process.
According to this mass spectrometry method, since the sample support body provided with the conductive substrate described above is used, it is possible to improve the signal intensity of components of an ionized sample in mass spectrometry.
According to the present disclosure, a sample support body, an ionization method, and a mass spectrometry method capable of improving a signal intensity of components of an ionized sample in mass spectrometry can be provided.
Hereinafter, embodiments of the present disclosure will be described in detail with reference to the drawings. The same or equivalent portions are denoted by the same reference signs in each of the drawings, and duplicate description thereof will be omitted.
As illustrated in
The substrate 2 is formed of, for instance, an insulating material in a shape of a rectangular plate. When viewed in the thickness direction of the substrate 2, a length of one side of the substrate 2 is, for instance, approximately several centimeters, and a thickness of the substrate 2 is, for instance, within a range of approximately 1 μm to 50 μm. In each of the through-holes 20, a width of a first opening 20a on the first surface 2a side is larger than a width of a second opening 20b on the second surface 2b side. When viewed in the thickness direction of the substrate 2, an outer edge of the first opening 20a is located outside an outer edge of the second opening 20b in each of the through-holes 20. That is, when viewed in the thickness direction of the substrate 2, the outer edge of the first opening 20a encompasses the outer edge of the second opening 20b in each of the through-holes 20.
The width of the first opening 20a denotes the diameter of the first opening 20a in a case in which the first opening 20a has a nearly circular shape when viewed in the thickness direction of the substrate 2 and denotes the diameter (effective diameter) of a virtual largest circle fitted into the shape in a case in which it has a shape other than a nearly circular shape. Likewise, the width of the second opening 20b denotes the diameter of the second opening 20b in a case in which the second opening 20b has a nearly circular shape when viewed in the thickness direction of the substrate 2 and denotes the diameter (effective diameter) of a virtual largest circle fitted into the shape in a case in which it has a shape other than a nearly circular shape. In the present embodiment, the width of the first opening 20a is approximately twice the width of the second opening 20b.
Each of the through-holes 20 includes a first portion 21 on the first opening 20a side and a second portion 22 on the second opening 20b side. The first portion 21 has a funnel shape expanding toward the first opening 20a. The second portion 22 has a columnar shape. A center line of the first portion 21 and a center line of the second portion 22 coincide with each other. In each of the through-holes 20, a minimum value of the width is 1 nm and a maximum value of the width is 700 nm. Here, the width of the through-hole 20 denotes the diameter of the through-hole 20 in a case in which a sectional shape of the through-holes 20 perpendicular to the thickness direction of the substrate 2 is a nearly circular shape and denotes the diameter (effective diameter) of a virtual largest circle fitted into the sectional shape in a case in which it has a sectional shape other than a nearly circular shape. In the present embodiment, the minimum value of the width is the diameter of the second portion 22 and the maximum value of the width is the diameter of the first opening 20a.
The first conductive layer 41 is provided on the first surface 2a of the substrate 2. The first conductive layer 41 covers a part on the first surface 2a of the substrate 2 in which no through-hole 20 is formed. The second conductive layer 42 is provided on the second surface 2b of the substrate 2. The second conductive layer 42 covers a part on the second surface 2b of the substrate 2 in which no through-hole 20 is formed.
The first conductive layer 41 and the second conductive layer 42 are formed of a conductive material. In the present embodiment, the first conductive layer 41 and the second conductive layer 42 are formed of platinum (Pt) or gold (Au). In this way, for the reason described below, a metal having a low affinity (reactivity) with a sample and high conductivity is preferably used as a material for the first conductive layer 41 and the second conductive layer 42.
For example, if the first conductive layer 41 and the second conductive layer 42 are formed of a metal such as copper (Cu) that has a high affinity with a sample such as protein, a sample is ionized in a state in which Cu atoms adhere to sample molecules in a process (which will be described below) of ionizing the sample, and there is concern that detection results may deviate in mass spectrometry (which will be described below) according to the adhered amount of the Cu atoms. Therefore, a metal having a low affinity with a sample is preferably used as a material for the first conductive layer 41 and the second conductive layer 42.
Meanwhile, it becomes easier to apply a constant voltage to a metal having higher conductivity in an easy and stable way. For this reason, if the first conductive layer 41 and the second conductive layer 42 are formed of a highly conductive metal, a voltage can be uniformly applied to the first surface 2a and the second surface 2b of the substrate 2. Further, a metal having higher conductivity tends to have higher thermal conductivity. For this reason, if the first conductive layer 41 and the second conductive layer 42 are formed of a highly conductive metal, an energy of a laser beam (energy beam) with which the substrate 2 has been irradiated can be efficiently transmitted to a sample through the first conductive layer 41 and the second conductive layer 42. Therefore, a highly conductive metal is preferably used as a material for the first conductive layer 41 and the second conductive layer 42.
From the foregoing viewpoints, for example, Pt, Au, or the like is preferably used as a material for the first conductive layer 41 and the second conductive layer 42. For example, the first conductive layer 41 and the second conductive layer 42 are formed to have a thickness within a range of approximately 1 nm to 350 nm using a plating method, an atomic layer deposition (ALD) method, a vapor deposition method, a sputtering method, or the like. For example, chromium (Cr), nickel (Ni), titanium (Ti), or the like may be used as a material for the first conductive layer 41 and the second conductive layer 42.
The substrate 2 illustrated in
The substrate 2 may be formed by anodizing a valve metal other than Al, such as tantalum (Ta), niobium (Nb), titanium (Ti), hafnium (Hf), zirconium (Zr), zinc (Zn), tungsten (W), bismuth (Bi), or antimony (Sb). Alternatively, the substrate 2 may be formed by anodizing silicon (Si).
Next, an ionization method and a mass spectrometry method according to the embodiment using the sample support body 1 will be described. In
First, as illustrated in (a) of
Subsequently, as illustrated in (b) of
Accordingly, in each of the through-holes 20, the solution including the sample S enters the inside of the second portion 22 from the first portion 21, and a part of the solution including the sample S stays in the first portion 21 of each of the through-holes 20 due to surface tension (see
Subsequently, as illustrated in
In this way, when the first surface 2a of the substrate 2 is irradiated with the laser beam L while a voltage is applied to the first conductive layer 41, the components S1 of the sample S staying on the first surface 2a side of the substrate 2 are ionized, and sample ions S2 (ionized components S1) are discharged (third process). To be specific, an energy is transmitted from the first conductive layer 41 (see
The discharged sample ions S2 move while accelerating toward a ground electrode (not illustrated) provided between the sample support body 1 and an ion detection unit 15. That is, the sample ions S2 move while accelerating toward the ground electrode due to a potential difference between the first conductive layer 41 to which a voltage is applied and the ground electrode. The sample ions S2 are detected by the ion detection unit 15 of the mass spectrometer 10 (fourth process). In the present embodiment, the mass spectrometer 10 is a scanning mass spectrometer utilizing a time-of-flight mass spectrometry (TOF-MS) method. The foregoing first to fourth processes correspond to the mass spectrometry method according to the embodiment using the sample support body 1.
Next, an ionization method and a mass spectrometry method according to another embodiment using the sample support body 1 will be described. In
First, the aforementioned sample support body 1 is prepared (first process). The sample support body 1 may be prepared by being manufactured by a person who carries out the ionization method and the mass spectrometry method or may be prepared by being obtained from a manufacturer, a seller, or the like of the sample support body 1.
Subsequently, as illustrated in (a) of
Accordingly, as illustrated in (b) of
Subsequently, as illustrated in
In this way, when the second surface 2b of the substrate 2 is irradiated with the laser beam L while a voltage is applied to the second conductive layer 42, the components S1 of the sample S which have moved to the second surface 2b side of the substrate 2 through the plurality of through-holes 20 are ionized, and the sample ions S2 (ionized components S1) are discharged (third process). To be specific, an energy is transmitted from the second conductive layer 42 (see
The discharged sample ions S2 move while accelerating toward a ground electrode (not illustrated) provided between the sample support body 1 and the ion detection unit 15. That is, the sample ions S2 move while accelerating toward the ground electrode due to a potential difference between the second conductive layer 42 to which a voltage is applied and the ground electrode. The sample ions S2 are detected by the ion detection unit 15 of the mass spectrometer 10 (fourth process). In the present embodiment, the ion detection unit 15 detects the sample ions S2 in way of corresponding to a scanning position of the laser beam L. Accordingly, a two-dimensional distribution of molecules constituting the sample S can be imaged. Further, in the present embodiment, the mass spectrometer 10 is a scanning mass spectrometer utilizing a time-of-flight mass spectrometry method. The foregoing first to fourth processes correspond to the mass spectrometry method according to another embodiment using the sample support body 1.
As described above, in the sample support body 1, the width of the first opening 20a on the first surface 2a side is larger than the width of the second opening 20b on the second surface 2b side in each of the plurality of through-holes 20. For this reason, for example, if the solution including the sample S is dropped to the plurality of through-holes 20 from the first surface 2a side, the solution moves to the second surface 2b side through the plurality of through-holes 20, and the components S1 of the sample S in the solution stay on the first surface 2a side in an appropriate state. Therefore, when the first surface 2a is irradiated with the laser beam L while a voltage is applied to the first conductive layer 41, the components S1 of the sample S are reliably ionized. Further, for example, when the sample support body 1 is disposed such that the first surface 2a faces the sample S, the components S1 of the sample S move smoothly to the second surface 2b side through the plurality of through-holes 20 and stay on the second surface 2b side in an appropriate state. Therefore, when the second surface 2b is irradiated with the laser beam L while a voltage is applied to the second conductive layer 42, the components S1 of the sample S are reliably ionized. Therefore, according to the sample support body 1, a signal intensity of the ionized components S1 of the sample S can be improved in mass spectrometry.
Further, in the sample support body 1, when viewed in the thickness direction of the substrate 2, the outer edge of the first opening 20a is located outside the outer edge of the second opening 20b in each of the plurality of through-holes 20. Accordingly, for example, when the solution including the sample S is dropped to the plurality of through-holes 20 from the first surface 2a side, the components S1 of the sample S in the solution can stay on the first surface 2a side in a more appropriate state. Further, for example, when the sample support body 1 is disposed such that the first surface 2a faces the sample S, the components S1 of the sample S move more smoothly to the second surface 2b side through the plurality of through-holes 20, and the components S1 of the sample S can stay on the second surface 2b side in a more appropriate state.
Further, in the sample support body 1, each of the plurality of through-holes 20 includes the first portion 21 on the first opening 20a side and the second portion 22 on the second opening 20b side. The first portion 21 has a funnel shape expanding toward the first opening 20a. Accordingly, for example, when the solution including the sample S is dropped to the plurality of through-holes 20 from the first surface 2a side, the components S1 of the sample S in the solution can stay on the first surface 2a side in an appropriate state. Further, for example, when the sample support body 1 is disposed such that the first surface 2a faces the sample S, the components S1 of the sample S move smoothly to the second surface 2b side through the plurality of through-holes 20, and the components S1 of the sample S can stay on the second surface 2b side in an appropriate state.
Further, in the sample support body 1, in each of the plurality of through-holes 20, the minimum value of the width is 1 nm and the maximum value of the width is 700 nm. Accordingly, for example, when the solution including the sample S is dropped to the plurality of through-holes 20 from the first surface 2a side, the components S1 of the sample S in the solution can be made to stay on the first surface 2a side in an appropriate state. Further, for example, when the sample support body 1 is disposed such that the first surface 2a faces the sample S, the components S1 of the sample S can move smoothly to the second surface 2b side through the plurality of through-holes 20, and the components S1 of the sample S can stay on the second surface 2b side in an appropriate state.
According to the ionization method and the mass spectrometry method of the embodiment and another embodiment described above, since the sample support body 1 provided with the first conductive layer 41 and the second conductive layer 42 described above is used, it is possible to improve the signal intensity of the components S1 of the ionized sample S in mass spectrometry.
The present disclosure is not limited to the embodiments described above. For example, as illustrated in
Further, in the embodiments described above, one predetermined region (a predetermined region for ionizing components of the sample S) is provided in the substrate 2, but a plurality of predetermined regions may be provided in the substrate 2. Further, the plurality of through-holes 20 need not be formed in only the predetermined region. As in the embodiments described above, for example, the plurality of through-holes 20 may be formed in the entire substrate 2. That is, the plurality of through-holes 20 need only be formed in at least the predetermined region. Further, in the embodiments described above, the sample S is disposed such that one sample S corresponds to one predetermined region, but the sample S may be disposed such that a plurality of samples S correspond to one predetermined region. Further, a conductive layer may be provided on an inner surface of each of the through-holes 20. Further, a frame may be attached to the substrate 2. In such a case, handling of the sample support body 1 is facilitated, and deformation of the substrate 2 due to a temperature change or the like is curbed.
Further, the sample support body 1 may include a conductive substrate 2. In this case as well, for example, when a solution including the sample S is dropped to the plurality of through-holes 20 from the first surface 2a side, a solution moves to the second surface 2b side through the plurality of through-holes 20, and the components S1 of the sample S in the solution stay on the first surface 2a side in an appropriate state. Therefore, when the first surface 2a is irradiated with the laser beam (energy beam) L while a voltage is applied to the substrate 2, the components S1 of the sample S are reliably ionized. Further, for example, when the sample support body 1 is disposed such that the first surface 2a faces the sample S, the components S1 of the sample S move smoothly to the second surface 2b side through the plurality of through-holes 20 and stay on the second surface 2b side in an appropriate state. Therefore, when the second surface 2b is irradiated with the laser beam (energy beam) L while a voltage is applied to the substrate 2, the components S1 of the sample S are reliably ionized. Thus, even in a case in which the sample support body 1 is provided with the conductive substrate 2, the signal intensity of the components S1 of the ionized sample S in mass spectrometry can be improved. In this case, the first conductive layer 41 and the second conductive layer 42 can be omitted in the sample support body 1.
Further, in the second process, the sample support body 1 may be fixed to the slide glass 6 by means other than the tape 7 (for example, means using an adhesive, a fixing tool, or the like). Further, in the third process, a voltage may be applied to the first conductive layer 41 or the second conductive layer 42 without having the mount surface 6a of the slide glass 6 and the tape 7 therebetween. In such a case, the slide glass 6 and the tape 7 do not have to be conductive. Further, the tape 7 may be a part of the sample support body 1. In a case in which the tape 7 is a part of the sample support body 1 (that is, in a case in which the sample support body 1 includes the tape 7), for example, the tape 7 may be fixed to the first surface 2a side on a circumferential edge of the substrate 2 in advance.
Further, in the mass spectrometer 10, the laser beam irradiation part 13 may collectively irradiate the predetermined region on the first surface 2a or the predetermined region on the second surface 2b with the laser beam L, and the ion detection unit 15 may detect the sample ions S2 while maintaining two-dimensional information of the region. That is, the mass spectrometer 10 may be a projection mass spectrometer. Further, the ionization method described above can also be used for other measurements such as ion mobility measurements and other experiments.
Further, the purpose of the sample support body 1 is not limited to ionization of the sample S using irradiation with the laser beam L. The sample support body 1 can be used for ionization of the sample S using irradiation with an energy beam such as a laser beam, an ion beam, or an electron beam. In the ionization method and the mass spectrometry method described above, the sample S can be ionized using irradiation with an energy beam.
Further, in the ionization method and the mass spectrometry method according to another embodiment described above, the sample S is not limited to a water-containing sample and may be a dried sample. In such a case, in the second process, after the sample support body 1 is mounted on the mount surface 6a such that the first surface 2a faces the sample S, a predetermined solution is dropped to the plurality of through-holes 20 from the second surface 2b side, for example. Accordingly, the components S1 of the sample S can move smoothly to the second surface 2b side through the plurality of through-holes 20, and the components S1 of the sample S can stay on the second surface 2b side in an appropriate state. Further, in the second process, a solution including the sample S may be mounted on the mount surface 6a, and the sample support body 1 may be mounted on the mount surface 6a such that the first surface 2a faces the solution including the sample S. In such a case as well, the components S1 of the sample S can move smoothly to the second surface 2b side through the plurality of through-holes 20, and the components S1 of the sample S can stay on the second surface 2b side in an appropriate state.
1: sample support body, 2: substrate, 2a: first surface, 2b: second surface, 6: slide glass (mount portion), 6a: mount surface, 20: through-hole, 20a: first opening, 20b: second opening, 21: first portion, 22: second portion, 41: first conductive layer, 42: second conductive layer, L: laser beam (energy beam), S: sample, S1: component.
| Number | Date | Country | Kind |
|---|---|---|---|
| JP2018-021810 | Feb 2018 | JP | national |
| Filing Document | Filing Date | Country | Kind |
|---|---|---|---|
| PCT/JP2019/001113 | 1/16/2019 | WO | 00 |
| Publishing Document | Publishing Date | Country | Kind |
|---|---|---|---|
| WO2019/155834 | 8/15/2019 | WO | A |
| Number | Name | Date | Kind |
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| 20020094533 | Hess et al. | Jul 2002 | A1 |
| 20050130222 | Lee | Jun 2005 | A1 |
| 20170358436 | Naito et al. | Dec 2017 | A1 |
| 20200357621 | Kotani | Nov 2020 | A1 |
| 20210057198 | Kotani | Feb 2021 | A1 |
| Number | Date | Country |
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| 2004-354376 | Dec 2004 | JP |
| 3122331 | Jun 2006 | JP |
| 2007-247070 | Sep 2007 | JP |
| 2009-080106 | Apr 2009 | JP |
| 2009-120892 | Jun 2009 | JP |
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| WO-2014020939 | Feb 2014 | WO |
| WO-2017038709 | Mar 2017 | WO |
| Entry |
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| International Preliminary Report on Patentability dated Aug. 20, 2020 for PCT/JP2019/001113. |
| Office Action dated Jun. 30, 2021 in U.S. Appl. No. 16/965,460. |
| Number | Date | Country | |
|---|---|---|---|
| 20210043437 A1 | Feb 2021 | US |
