Fiber optic sensors and detectors

Abstract

To provide a fiber sensor that can detect a detection object rapidly and easily.SOLUTION: The present invention relates to a fiber sensor FS for detecting an object by using an optical fiber 10 having a fiber body 11 and a covering unit 20 covering the fiber body. The fiber sensor includes: a gap formation unit 21 for forming a gap 23 in a top end of the fiber body, the gap including an axial line J of the fiber body and opening in the outside; and a light emission unit 30 in the gap, which can emit light. The light emission unit includes a light emission sensor having an intensity varied by a reaction with an object.SELECTED DRAWING: Figure 2

Description

[Technical field]

[0001] The present invention relates to fiber sensors and detection devices. [Background technology]

[0002] For example, in brain science research, technology is needed to selectively and quantitatively measure neurotransmitter molecules and ions to detect the activation of neurons and other subjects of research. In order to analyze behaviors controlled by specific brain regions, spatial resolution of several hundred microns and time resolution on the scale of seconds are required. It is also important to be able to analyze experimental animals while they are still alive.

[0003] For example, to detect neurotransmitter molecules, methods have been used that use microelectrodes to detect the electric current generated by the oxidation-reduction reaction of the molecules, or that insert a microdialysis probe with a semipermeable membrane at the tip to collect small molecules and then separate and analyze them using liquid chromatography.

[0004] The method of detecting the electric current generated by the redox reaction of molecules has the disadvantage that it is difficult to distinguish between molecules with similar redox potentials, and the method of separating and analyzing molecules using liquid chromatography has the problem that the time required for separation is extremely low, resulting in extremely low time resolution. Therefore, proteins that emit fluorescence in response to ions or neurotransmitter molecules have been developed, and techniques for detecting this fluorescence using optical fibers have been developed (see, for example, Non-Patent Document 1). [Prior art documents] [Non-patent literature]

[0005] [Non-Patent Document 1] A Genetically Encoded Fluorescent Sensor Enables Rapid and Specific Detection of Dopamine in Flies, Fish, and Mice. 1. Cell. 2018 July 12;174(2):481-496. Summary of the Invention [Problem to be solved by the invention]

[0006] The method described in the above-mentioned Non-Patent Document 1 requires transfection to express fluorescence in vivo, which poses the problem of being time-consuming and labor-intensive.

[0007] The present invention has been made in consideration of the above points, and an object of the present invention is to provide a fiber sensor and a detection device that can detect an object to be detected without requiring much time and effort. [Means for solving the problem]

[0008] According to a first aspect of the present invention, there is provided a fiber sensor for detecting an object using an optical fiber having a fiber body and a coating portion that coats the fiber body, the fiber sensor including a gap forming portion that forms a gap including an axis of the fiber body and opening to the outside, at a tip side of the fiber body, and a light emitting portion that is located in the gap and is capable of emitting light, the light emitting portion including a light emitting sensor whose light emission intensity changes when it reacts with the object.

[0009] According to a second aspect of the present invention, there is provided a detection device having the fiber sensor of the first aspect of the present invention, an imaging unit that captures an emission image from the light emitting unit via the optical fiber, and a detection unit that detects the object based on the captured emission image. Effect of the Invention

[0010] According to the present invention, it is possible to detect a detection target without spending much time and effort. [Brief description of the drawings]

[0011] [Figure 1] FIG. 1 is a diagram showing an embodiment of the present invention, and is a schematic configuration diagram of a detection device 1 having a fiber sensor FS of a first embodiment. [Diagram 2] 1 is a partial cross-sectional view showing the tip portion of an optical fiber 10. FIG. [Diagram 3] 1A to 1C are schematic perspective views showing a procedure for producing a fiber sensor FS. [Figure 4] 4 is a partial cross-sectional view showing the tip portion of an optical fiber 10 in second and third embodiments of the fluorescent sensor. FIG. [Diagram 5] 1 is a cross-sectional view showing an optical fiber 10 in a fiber sensor FS according to a second embodiment. [Figure 6] 10 is a perspective view showing a procedure for forming a gap 23 in the optical fiber 10 using a drill D. FIG. [Figure 7] 11 is an external perspective view showing an optical fiber 10 in a fiber sensor FS according to a third embodiment. FIG. [Figure 8] 13 is a diagram showing a fluorescent image of a light-emitting portion in a fiber sensor detected by a detection portion. FIG. [Figure 9] FIG. 1 is a graph showing the relationship between the molar concentration (μM) of cells and the change in fluorescence intensity. [Figure 10] FIG. 1 is a graph showing the relationship between response time and fluorescence intensity with respect to the time response of a specific cell to norepinephrine. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0012] DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS Hereinafter, an embodiment of a fiber sensor and a detection device according to the present invention will be described with reference to FIGS. The following embodiment shows one aspect of the present invention, does not limit the present invention, and can be modified as desired within the scope of the technical idea of ​​the present invention. In addition, in the following drawings, the scale and number of each structure are different from the actual structure in order to make each configuration easier to understand.

[0013] [First embodiment of fiber sensor] 1 is a schematic diagram of a detection device 1 having a fiber sensor FS according to a first embodiment of the present invention. The detection device 1 detects exogenous substances and endogenous substances (e.g., physiologically active substances) as targets using the fiber sensor FS. In the following, an example in which a physiologically active substance, which is an endogenous substance, is detected as a target will be described. Examples of physiologically active substances include neurotransmitter molecules, ions, and glucose.

[0014] The detection device 1 includes a fiber sensor FS, a light source unit 40, a dichroic mirror 50, a photodetector (imaging unit) 60, a preamplifier 70, a lock-in amplifier 80, and a detection unit 90. The light source unit 40 is a light source that emits excitation light. The light source unit 40 in this embodiment emits excitation light with a wavelength of 470 nm (blue), for example. The excitation light from the light source unit 40 is incident on the dichroic mirror 50 via a patch cable c1.

[0015] The dichroic mirror 50 transmits light of a predetermined wavelength and reflects light of other wavelengths. The dichroic mirror 50 of this embodiment reflects excitation light from the light source unit 40 and makes the light enter the optical fiber 10 via the patch cable c2. The patch cable c1 and the optical fiber 10 are optically connected. The dichroic mirror 50 transmits light (fluorescence; details will be described later) entering from the optical fiber 10 via the patch cable c2.

[0016] The photodetector 60 serves as an imaging section and captures a fluorescent image from the optical fiber 10 that has passed through the dichroic mirror 50. As an example, a photodiode is used as the photodetector 60. A signal captured by the photodetector 60 is sent to a preamplifier 70 and amplified. The signal amplified by the preamplifier 70 is sent to a lock-in amplifier 80.

[0017] The lock-in amplifier 80 receives the signal amplified by the preamplifier 70 and removes noise from the signal. The signal from which noise has been removed by the lock-in amplifier 80 is sent to a detection unit 90.

[0018] The detection unit 90 receives the signal from which noise has been removed by the lock-in amplifier 80, and detects the target object based on the signal in response to a change in the emission intensity of the fluorescent light from the optical fiber 10. As the detection unit 90, for example, a personal computer (PC) or a CPU in a PC can be used.

[0019] The fiber sensor FS has an optical fiber 10, an insertion tool 10A, and a light emitting unit 30. For example, at least the tip of the insertion tool 10A is inserted into a living body. The insertion tool 10A is inserted into a detection site in the living body that corresponds to the target. When the target is a neurotransmitter molecule, the insertion tool 10A is inserted into the brain. The insertion tool 10A is formed in a cylindrical shape. The material of the insertion tool 10A is not particularly limited as long as it does not have an adverse effect on the living body, and for example, a resin material is used.

[0020] As an example, the diameter of the insertion tool 10A is 0.1 to 10 mm, and the length is 1 to 10 mm. The diameter of the central hole of the insertion tool 10A is set to a size that allows the optical fiber 10 to be inserted therein.

[0021] The tip side of the optical fiber 10 is inserted into the center hole of the insertion tool 10 A. The base end side of the optical fiber 10 is connected to a patch cable c2. Fig. 2 is a partial cross-sectional view showing the tip portion of the optical fiber 10. As shown in Fig. 2, the optical fiber 10 has a fiber body 11, a coating portion 20, and a gap 23. The fiber body 11 extends along an axis J, and has a fiber core 11A located at the center and a fiber core 11B that coats the outer peripheral surface of the fiber core 11A. In the optical fiber 10 of this embodiment, the fiber core 11A is made of silica, and the fiber clad 11B is made of a synthetic resin.

[0022] As the fiber clad 11B, for example, methacrylic resin, polycarbonate resin (PC), polystyrene resin (PS), acrylic resin, fluorine-based resin, etc. can be used. As an example, the diameter of the fiber clad 11B is 230 μm. The refractive index of the fiber clad 11B is smaller than the refractive index of the fiber core 11A. Therefore, the excitation light incident on the fiber core 11A via the patch cable c2 is transmitted via the fiber core 11A.

[0023] The fiber core 11A has an emission part 12 that emits excitation light. The emission part 12 is a tip surface of the fiber core 11A.

[0024] The coating 20 coats the outer peripheral surface of the fiber clad 11B. The coating 20 has a diameter of 500 μm, for example. The coating 20 is made of a fluororesin, for example. Note that a clad that coats the outer peripheral surface of the fiber body 11 may be provided.

[0025] The coating portion 20 has a gap forming portion 21 and a dividing portion 22. The gap forming portion 21 forms a gap 23 that includes the axis J and opens to the outside, on the tip side in the axis J direction from the emission portion 12. The gap 23 in this embodiment faces the emission portion 12 and opens on the tip side in the axis J direction. Therefore, the gap 23 does not penetrate the optical fiber 10. The gap forming portion 21 has a cylindrical wall 24 that forms the gap 23. The cylindrical wall 24 is located radially outward from the fiber body 11 (fiber clad 11B) with the axis J as the center, and is formed in a cylindrical shape around the entire circumference. The cylindrical wall 24 is a region in the coating portion 20 that extends toward the tip side in the optical axis direction from the emission portion 12 of the fiber core 11A.

[0026] The dividing portion 22 is a region where the coating portion 20 is divided in the length direction (axis J direction) on the base end side of the emission portion 12 to expose the fiber body 11. In the axis J direction, the length L1 from the emission portion 12 to the tip of the cylindrical wall 24 is the same as the length L2 of the dividing portion 22. That is, the cylindrical wall 24 is formed by dividing the coating portion 20 in the axis J direction at a position on the base end side of the dividing portion 22 and moving the divided tip side coating portion 20 by the distance L1 toward the tip side relative to the fiber body 11.

[0027] The light-emitting unit 30 is located in the gap 23. The light-emitting unit 30 has a luminescence sensor whose luminescence intensity changes when it reacts with a detection target. The luminescence sensor is selected according to the target to be detected by the detection device 1. A number of embodiments of the luminescence sensor will be described below.

[0028] [First embodiment of the light emitting sensor] The light-emitting unit 30 has a cell 32 that is encapsulated in a hydrogel 31 and includes the light-emitting sensor of the first embodiment. The light-emitting sensor in this embodiment is a fluorescent sensor that emits fluorescence when irradiated with excitation light and changes the intensity of the fluorescence when it reacts with an object. The cell 32 has a protein sensor on its cell membrane, the fluorescence intensity of which increases in response to noradrenaline (norepinephrine). The protein sensor is produced by genetically modifying a noradrenaline receptor to insert a fluorescent protein (XFP) as a fluorescence emission site. The protein sensor may be located inside the cell.

[0029] Examples of the fluorescent moiety include green fluorescent proteins (GFP) such as TurboGFP, AcGFP, TagGFP, Azami-Green, ZsGreen, EmGFP, EGFP, GFP2, and HyPer; blue fluorescent proteins (BFP) such as EBFP; cyan fluorescent proteins (CFP) such as ECFP, TagCFP, AmCyan, and MidoriishiCyan; red fluorescent proteins (RFP) such as TurboRFP, DsRed-Express, DsRed2, TagRFP, DsRed-Monomer, AsRed2, and mStrawberry; and yellow fluorescent proteins (Yellow Proteins) such as TagYFP, EYFP, Venus, PhiYFP, PhiYFP-m, TurboYFP, ZsYellow, and mBanana. Examples of fluorescent proteins that emit infrared light include orange fluorescent proteins such as YFP, KusabiraOrange, and mOrange, and infrared fluorescent proteins such as TurboFP602, mRFP1, JRed, KillerRed, mCherry, HcRed, KeimaRed, mRasberry, and mPlum.

[0030] The hydrogel 31 is preferably made of a material that easily absorbs water, taking into consideration that the hydrogel 31 is retained in the voids 23 when it is wetted (swollen). As the components of the hydrogel 31, polysaccharides such as methylcellulose or dextran, acrylic hydrogels prepared by polymerizing monomers such as (meth)acrylamide, methylol acrylamide, or hydroxyethyl acrylate, or urethane hydrogels prepared from polyethylene glycol and diisocyanate can be used, and in addition to the above-mentioned examples, for example, polyvinyl alcohol, polyethylene glycol, polyhydroxyalkanoate, poly-(hydroxyethyl methacrylate), poly-(hydroxyethyl acrylate), polyvinylpyrrolidone, poly-(ethylene-co-vinyl alcohol), poly-(vinyl acetate-co-vinyl alcohol), poly(ethylene-co-vinyl acetate-co-vinyl alcohol), poly(ethylene glycol-co-propylene glycol), biopolymers such as collagen and gelatin can also be used.

[0031] FIG. 3 is a schematic perspective view showing a procedure for producing the above-mentioned fiber sensor FS. 3, when fabricating the fiber sensor FS, a portion of the tip side of the coating 20 is divided and moved toward the tip side relative to the fiber body 11 while the tip side of the optical fiber 10 is immersed in the hydrogel 31 containing the cells 32. When moving the portion of the tip side of the coating 20 toward the tip side, for example, a method can be used in which a slit is formed at the division position of the coating 20 and the portion of the tip side is pulled out relative to the fiber body 11. At this time, since the coating 20 is made of a fluorine-based resin and has low frictional resistance, the coating 20 can be easily pulled out.

[0032] By moving a part of the tip side of the covering part 20 toward the tip side, a gap 23 is formed on the tip side of the emission part 12, which is surrounded by a gap forming part 21 (cylindrical wall 24) and faces the emission part 12 and opens to the tip side in the direction of the axis J. The gap 23 is formed by moving the fiber body 11 relatively toward the base end side with respect to the covering part 20, and therefore has a cylindrical shape with the same diameter as the fiber body 11 (for example, 230 μm).

[0033] If the depth (distance L1) of the gap 23 is too large, the distance between the cell 32 in the gap 23 and the emission part 12 becomes large, which may reduce the accuracy of detecting the fluorescence from the cell 32. On the other hand, if the depth (distance L1) of the gap 23 is small, the cell 32 may not be sufficiently held in the gap 23. Therefore, the depth (distance L1) of the gap 23 is preferably 20 μm or more and 1000 μm or less.

[0034] Moreover, since the gap 23 formed by the relative movement of the fiber body 11 toward the base end with respect to the coating portion 20 becomes negative pressure, the hydrogel 31 containing the cells 32 is introduced through the opening and filled into the gap 23. Here, since the gap 23 is surrounded by the cylindrical wall 24 formed in a cylindrical shape all around, the cells 32 are uniformly diffused in the gap 23.

[0035] As a result, an optical fiber 10 is produced in which a gap 23 is formed at the tip and the hydrogel 31 containing cells 32 as light-emitting portions 30 that are uniformly dispersed in the gap 23 is filled. Then, the optical fiber 10 is inserted from the tip side into the central hole of the insertion tool 10A, thereby producing the fiber sensor FS.

[0036] In addition to the procedure of forming a gap 23 at the tip, providing a light-emitting unit 30 in the gap 23, and then inserting the optical fiber 10 into the central hole of the insertion tool 10A, it is also possible to select a procedure of forming a gap 23 in the optical fiber 10 inserted into the central hole of the insertion tool 10A by the procedure described above, and providing a light-emitting unit 30 in the gap 23.

[0037] When detecting a physiologically active substance in a living organism using the detection device 1 configured as above, first, the fiber sensor FS is inserted into the living organism. After the fiber sensor FS is inserted into the living organism, excitation light is emitted from the light source unit 40. Note that the emission of excitation light may be performed before the fiber sensor FS is inserted into the living organism.

[0038] The excitation light is incident on the dichroic mirror 50 via the patch cable c1, reflected therefrom, and then incident on the optical fiber 10 via the patch cable c2. The excitation light incident on the optical fiber 10 passes through the fiber body 11 and is emitted from the emission part 12 along the axis J. The excitation light emitted from the emission part 12 irradiates the cells 32 in the light-emitting part 30.

[0039] When the cell 32 is irradiated with the excitation light, the protein sensor emits fluorescence having a wavelength corresponding to the wavelength of the excitation light. A part of the fluorescence generated in the cell 32 enters the fiber body 11, then enters the dichroic mirror 50 via the patch cable c2 and is transmitted therethrough. An image of the fluorescence transmitted through the dichroic mirror 50 is captured by the photodetector 60. The signal of the fluorescence image captured by the photodetector 60 is amplified by the preamplifier 70, and after noise is removed by the lock-in amplifier 80, it is sent to the detection unit 90. Based on the input signal, the detection unit 90 monitors, for example, the fluorescence information from the cell 32 in a state where the cell 32 does not react with a physiologically active substance in the living body.

[0040] On the other hand, when the cell 32 reacts (comes into contact with) molecules of a physiologically active substance in a living body through the opening of the gap 23, the fluorescence emission characteristics of the protein sensor in the cell 32 change, causing a change in the fluorescence emission intensity. As a result, the fluorescence intensity from the cell 32 detected by the detection unit 90 changes. Here, the change in the fluorescence emission characteristics reaches a true measured value after the signal value starts to change (rise) a certain time after the cell 32 reacts with the physiologically active substance, but since the cell 32 is uniformly diffused in the gap 23 and the gap 23 is extremely small, the physiologically active substance immediately diffuses uniformly in the gap 23, allowing the change in the fluorescence intensity to be detected with a fast response. The detection unit 90 can detect a physiologically active substance in a living body by detecting a change in the fluorescence intensity to be monitored.

[0041] As described above, in the fiber sensor FS and detection device 1 of this embodiment, the gap 23 that includes the axis J of the excitation light and opens to the outside is provided on the tip side of the fiber body 11, and the light emitting section 30 including a fluorescent sensor that changes its fluorescence emission intensity when it reacts with an object is provided in the gap 23. Therefore, the fiber sensor FS and detection device 1 of this embodiment solve the problems of the difficulty in distinguishing between molecules with similar redox potentials that arise in a detection method using a molecular redox reaction, the problem of time-consuming separation that arises in a detection method using liquid chromatography, and the time and effort required in the method described in the above Non-Patent Document 1, and it becomes possible to detect an object to be detected without time and effort.

[0042] Furthermore, in the fiber sensor FS and detection device 1 of this embodiment, the gap 23 is formed by the cylindrical wall 24 that is positioned radially outward from the fiber body 11 and formed in a cylindrical shape around the entire circumference with the axis J as the center, so that the cells 32 can be uniformly diffused in the gap 23. Therefore, in the fiber sensor FS and detection device 1 of this embodiment, a slow response when detecting an object can be avoided by detecting the object based on a fluorescent signal from the cell 32 located away from the immediate vicinity of the emission part 12, as occurs when a part of the partition that forms the gap 23 is cut out and the diffusion of the cells 32 is not uniform.

[0043] Furthermore, in the fiber sensor FS and detection device 1 of this embodiment, the gap 23 has an opening but does not pass through the optical fiber 10, and negative pressure is generated even if the light-emitting section 30 held in the gap 23 is about to come off the opening, so that the light-emitting section 30 can be prevented from coming off the gap 23, as would occur if the gap 23 passed through the optical fiber 10.

[0044] In the above embodiment, a configuration in which a physiologically active substance is detected using a cell in which a protein sensor is expressed on the cell membrane as a fluorescent sensor is exemplified, but the present invention is not limited to this configuration. For example, instead of a cell, an artificially created liposome may contain a fluorescent sensor, and a physiologically active substance may be detected based on a change in the emission intensity of the fluorescence when the liposome reacts with the physiologically active substance.

[0045] [Second embodiment of the light emitting sensor] Next, a second embodiment of the luminescence sensor will be described with reference to FIG. In FIG. 4, the same elements as those in the first embodiment of the luminescence sensor shown in FIGS. 1 to 3 are denoted by the same reference numerals, and the description thereof will be omitted.

[0046] 4, the gap 23 is filled with a fluorescent sensor 32A constituted by a protein, a DNA probe, or beads encapsulated in a hydrogel 31 as a luminescence sensor in the light-emitting unit 30. As the protein, for example, the above-mentioned fluorescent protein (fluorescence-emitting site) can be used.

[0047] As the DNA probe, for example, a molecular beacon can be used, in which a single-stranded DNA recognizes a target substance (biologically active substance), partially hybridizes with the target substance, and emits light by fluorescence resonance energy transfer (FRET).

[0048] The beads are not particularly limited as long as the fluorescence intensity changes when they are converted into a physiologically active substance, and beads containing a fluorescent dye or fluorescently dyed beads can be used.

[0049] When the fluorescent sensor 32A having the above configuration is used, the fluorescent sensor 32A irradiated with excitation light emits fluorescence with a wavelength corresponding to the wavelength of the excitation light. On the other hand, when the fluorescent sensor 32A reacts with (comes into contact with) molecules of a physiologically active substance in a living body through the opening of the gap 23, the fluorescence emission characteristics of the fluorescent sensor 32A change, causing a change in the emission intensity of the fluorescence. Then, the detection unit 90 detects the change in the monitored fluorescence intensity, thereby making it possible to detect a physiologically active substance in a living body, as in the case of using the fluorescent sensor of the first embodiment described above.

[0050] [Third embodiment of the light emitting sensor] Next, a third embodiment of the luminescence sensor will be described. In the first and second embodiments of the luminescence sensor, a protein sensor whose fluorescence intensity increases in response to a physiologically active substance is used as a fluorescent sensor. 2+ The cell may have a protein on the cell membrane or inside the cell that causes the influx of ions such as ions, and may have a fluorescent indicator as the fluorescent sensor 32A whose fluorescent intensity changes in response to the influx of ions.

[0051] As the protein that induces the influx of ions, for example, a group of G protein-coupled receptors including acetylcholine receptors such as muscarinic receptors can be used. As the fluorescent indicator, for example, calcium ion indicators fura-2, indo-1, fluo-3, rhod-2 and members of the family thereof can be used.

[0052] When the fluorescent sensor 32A having the above configuration is used, the fluorescent sensor 32A irradiated with excitation light emits fluorescence with a wavelength corresponding to the wavelength of the excitation light. Meanwhile, in response to ions that flow in as a result of the protein reacting with a physiologically active substance in the living body through the opening of the gap 23, the fluorescent emission characteristics of the fluorescent indicator change, causing a change in the emission intensity of the fluorescence. Then, the detection unit 90 detects the change in the monitored fluorescence intensity, making it possible to detect a physiologically active substance in the living body.

[0053] The configuration using a protein that induces an influx of ions in response to the above-mentioned physiologically active substance and a fluorescent indicator that responds to the inflow of ions also makes it possible to detect physiologically active substances in the living body when liposomes are used instead of cells.

[0054] [Fourth embodiment of the light emitting sensor] Next, a fourth embodiment of the luminescence sensor will be described. In the first to third embodiments of the luminescence sensor, a configuration has been exemplified in which a fluorescent sensor emits fluorescence when irradiated with excitation light and the fluorescence intensity increases in response to a physiologically active substance, but for example, a configuration may be used in which a substance that does not emit light before reacting with a target and emits light when reacting with the target, thereby changing the luminescence intensity, is used as the luminescence sensor. In this case, the detection device 1 does not require a light source of excitation light.

[0055] As this type of luminescence sensor, for example, a luminescent enzyme (luciferase) derived from fireflies or luminescent bacteria can be used. In the detection device 1, a detection target in a living body can be detected by capturing a luminescence image of the luminescent enzyme emitting light in response to a substrate molecule that is a detection target.

[0056] The above-mentioned luminescent enzyme can be arranged in a configuration in which a protein sensor expressing the luminescent enzyme is placed on a cell membrane or inside a cell encapsulated in hydrogel 31 in void 23, on a cell membrane or inside a cell encapsulated in hydrogel 31 in void 23, or in a liposome instead of the above-mentioned cell.

[0057] [Second embodiment of fiber sensor] Next, a second embodiment of the fiber sensor FS will be described with reference to FIGS. In these figures, the same elements as those in the first embodiment shown in FIGS. 1 to 3 are designated by the same reference numerals, and the description thereof will be omitted. In addition, in the fiber sensor FS of the second embodiment, a configuration using the fluorescent sensor of the first embodiment is exemplified, but it is also applicable to configurations using the fluorescent sensors of the second and third embodiments.

[0058] FIG. 5 is a cross-sectional view showing an optical fiber 10 in a fiber sensor FS according to the second embodiment. 5, the void 23 of the optical fiber 10 according to this embodiment extends in a direction intersecting (orthogonal to) the axis J. The void 23 is located closer to the base end than the tip surface of the optical fiber 10. Therefore, the void forming portion 21 of this embodiment is composed of the fiber body 11 and the coating portion 20 that surround the periphery of the void 23. In addition, the emission portion 12 of the fiber body 11 is not the tip surface of the fiber body 11 but the surface facing the void 23.

[0059] The void 23 is cylindrical with a circular cross section. The void 23 passes through the fiber body 11, and is open to the coating 20 at one end (upper side in FIG. 5) and closed at the other end (lower side in FIG. 5). That is, the void 23 is provided so as not to pass through the optical fiber 10. The void 23 in the fiber body 11 and the void 23 in the coating 20 are coaxial, have the same diameter, and have the same cross-sectional shape. The other configurations are similar to those of the first embodiment.

[0060] The gap 23 can be formed by, for example, drilling. Fig. 6 is a perspective view showing a procedure for forming a gap 23 in an optical fiber 10 using a drill D. As shown in Fig. 6, a jig 100 having a groove 101 with a V-shaped cross section on its upper surface is used. Then, the optical fiber 10 placed in the groove 101 is drilled in the radial direction with the drill D, thereby forming a non-through gap 23 in the optical fiber 10. The above-mentioned light emitting unit 30 is injected into the gap 23 formed by drilling using a syringe or the like, thereby producing a fiber sensor FS.

[0061] In the fiber sensor FS of this embodiment, in addition to obtaining the same actions and effects as those of the first embodiment, the length of the gap 23 in the direction of the axis J can be shortened, and therefore the distance between the emission part 12 and the cell 32 can be shortened, making it possible to speed up the response when detecting an object.

[0062] [Third embodiment of fiber sensor] Next, a third embodiment of the fiber sensor FS will be described with reference to FIG. In this figure, the same elements as those in the first embodiment shown in FIGS. 1 to 3 are designated by the same reference numerals, and the description thereof will be omitted. In addition, in the fiber sensor FS of the third embodiment, a configuration using the fluorescent sensor of the first embodiment is exemplified, but it is also applicable to configurations using the fluorescent sensor of the second and third embodiments.

[0063] In the first and second embodiments described above, a portion of the optical fiber 10 constitutes the void forming portion 21 by processing the optical fiber 10. However, this embodiment differs from the first and second embodiments described above in that a void forming portion 21 is provided in the optical fiber 10.

[0064] FIG. 7 is an external perspective view showing the optical fiber 10 in the fiber sensor FS according to the third embodiment. 7, a gap forming portion 21 is provided at the tip of the optical fiber 10. The gap forming portion 21 has a disk portion 21A and a cylindrical portion (cylindrical wall) 21B.

[0065] The disk portion 21A covers the tip surface of the fiber body 11. The disk portion 21A is formed in a disk shape having substantially the same diameter (for example, about 125 μm) as the fiber body 11. The thickness of the disk portion 21A is, for example, about 200 μm.

[0066] The cylindrical portion 21B is formed in a cylindrical shape extending from the periphery of the disk portion 21A toward the tip side in the direction of the axis J. A gap 23 is formed between the surface on the tip side in the direction of the axis J of the disk portion 21A and the inner peripheral surface of the cylindrical portion 21B. The gap 23 is formed in a columnar shape that opens toward the tip side in the direction of the axis J.

[0067] As an example, the disk portion 21A and the cylindrical portion 21B are transparent or translucent to the above-mentioned excitation light and fluorescence, and are integrally formed from a photocurable resin that hardens when irradiated with energy light (e.g., laser light, ultraviolet light) (when energy is applied).

[0068] The void forming portion 21 having the above configuration can be formed by using a 3D printer that performs three-dimensional scanning relative to the photocurable resin attached to the tip surface of the fiber body 11 and repeats irradiation of the energy light E by the energy light irradiation portion 110 over multiple layers. In Fig. 7, the photocurable resin is configured to absorb the energy light E in two photons, but a single photon may also be used.

[0069] Specifically, the disk portion 21A and the cylindrical portion 21B are formed by, for example, focusing laser light on a photocurable resin attached to the tip surface of the fiber body 11. The light emitting portion 30 is injected into the gap 23 formed by the gap forming portion 21 using a syringe or the like. In this way, the fiber sensor FS is fabricated.

[0070] In the fiber sensor FS of this embodiment, in addition to obtaining the same actions and effects as those of the first embodiment described above, it becomes possible to easily form a void-forming portion 21 (i.e., a void 23) of any size and shape by controlling the focusing and scanning pattern of the energy light E such as laser light.

[0071] In the present embodiment, the configuration in which the void forming portion 21 has the disk portion 21A has been exemplified, but the present invention is not limited to this configuration, and the void forming portion 21 may have a configuration in which the void forming portion 21 is formed only by the cylindrical portion 21B without having the disk portion 21A. However, when the void forming portion 21 has the disk portion 21A, the contact area between the fiber body 11 and the void forming portion 21 increases, and the adhesive strength between the fiber body 11 and the void forming portion 21 can be increased.

[0072] [Example] The effects of the present invention will be more clearly understood by the following examples. Note that the present invention is not limited to the following examples, and can be modified as appropriate without departing from the spirit and scope of the present invention.

[0073] In this example, the detection target was noradrenaline (norepinephrine), a neurotransmitter, in a biochemical buffer (HEPES buffer). Collagen was used as the hydrogel described above, and a light-emitting unit was used that included human cultured cells (HEK293T) having GRAB-NE as a protein sensor that is encapsulated in the collagen and changes in the intensity of emitted fluorescence when reacting with noradrenaline. As the fiber sensor FS, a fiber sensor FS having the configuration of the first embodiment shown in Figures 2 and 3 was used.

[0074] Figure 8 shows a fluorescent image of the light-emitting part in the fiber sensor detected by the detection part. As shown in Figure 8, human cultured cells with GRAB-NE reacted with (contacted with) noradrenaline and were observed as bright spots of fluorescence (sensor cells expressing GRAB-NE).

[0075] Figure 9 is a graph showing the relationship between the molar concentration (μM) of cells and the change in fluorescence intensity. In Figure 9, a calibration curve is shown with the average value of 10 cells. The value of the change in fluorescence intensity (ΔF=F-F0) is a normalized value divided by the measured fluorescence intensity F.

[0076] As shown in Figure 9, a 10-20% increase in intensity was measured at a concentration of several μM. This confirmed that noradrenaline was detected by cells containing the protein sensor. Furthermore, when the change in fluorescence intensity for each cell was examined, the concentration of noradrenaline at which the fluorescence intensity was saturated varied from cell to cell, ranging from about 1 to 10 μM, although there were some cells for which the intensity hardly changed at all. This suggests that the number of cells in the hydrogel must be at least 10.

[0077] Fig. 10 is a graph showing the relationship between the response time and the fluorescence intensity with respect to the time response of a specific cell to noradrenaline. Fig. 10 shows the relationship between the response time and the fluorescence intensity for a cell immersed in a collagen solution with a molar concentration of 1 to 10 µM at a position 873 µm away from the emission part of the excitation light.

[0078] As shown in FIG. 10, by tracking the change in fluorescence intensity over time, it was confirmed that the fluorescence intensity was saturated in about 400 seconds. Using the diffusion equation x = √(2Dt), where D is the diffusion constant of noradrenaline in collagen, D = 0.95 × 10 -9 m 2 s -1 This value is similar to the diffusion constant of small molecules in water, confirming that for smaller hydrogels and devices, we can expect significantly faster time responses (e.g., 5 seconds for a 100 μm long hydrogel).

[0079] That is, it was suggested that the response speed (change) of the fluorescence intensity could be shortened to about 5 seconds by setting the distance L1 shown in Fig. 2 to 100 µm. In other words, it was suggested that noradrenaline could be detected by setting the monitoring of the fluorescence intensity by the detection unit 90 to every 5 seconds.

[0080] Although the preferred embodiment of the present invention has been described above with reference to the accompanying drawings, it goes without saying that the present invention is not limited to the examples. The shapes and combinations of the components shown in the above examples are merely examples, and various modifications can be made based on design requirements, etc., without departing from the spirit of the present invention.

[0081] For example, in the above embodiment, a configuration for detecting noradrenaline as a neurotransmitter molecule has been exemplified, but it is also possible to detect serotonin, dopamine, etc. in addition to noradrenaline. In this case, a fluorescent sensor whose fluorescence emission intensity changes upon reaction with each neurotransmitter molecule may be selected.

[0082] In addition, in the above embodiment, a configuration for detecting neurotransmitter molecules as physiologically active substances is exemplified, but the configuration may also be such that analytes in blood or body fluids (e.g., sugars such as glucose, pH, cholesterol, proteins, etc.) are detected in addition to neurotransmitter molecules. The above-mentioned hydrogel may include a fluorescent sensor as the light-emitting unit 30. The fluorescent sensor may be selected from those whose fluorescent emission intensity changes upon reaction with a physiologically active substance of the detection target.

[0083] For example, when sugars such as glucose are detected using a hydrogel containing a fluorescent sensor, the fluorescent sensor can be a substance that reversibly binds to glucose, such as a ruthenium organic complex, a fluorescent phenylboronic acid derivative, or fluorescein bound to a protein. In addition, instead of ruthenium in the ruthenium organic complex, an organic complex of osmium, iridium, rhodium, rhenium, chromium, or the like can be used. As the fluorescent phenylboronic acid derivative, a compound containing two phenylboronic acids and anthracene as a fluorescent residue can increase the detection sensitivity. Other suitable detection techniques include fluorescence resonance energy transfer (Förster resonance energy transfer, FRET), fluorescence energy transfer, fluorescence polarization, fluorescence quenching, phosphorescence, luminescence enhancement, and luminescence quenching. In particular, a luminescence method that applies FRET utilizing energy transfer between a donor (D) and an acceptor (A) is preferred, and when glucose is detected, a lectin (ConA) bound to a donor (D) and an analyte analog (dextran) bound to an acceptor (A) are used.

[0084] In addition, in the above embodiment, the configuration for detecting one type of physiologically active substance is exemplified, but the present invention is not limited to this configuration, and a configuration for detecting multiple types of physiologically active substances may be used. For example, when detecting noradrenaline and dopamine, a cell (or a liposome or a hydrogel) having a first fluorescent sensor whose fluorescence emission intensity changes upon reaction with noradrenaline and a second fluorescent sensor whose fluorescence emission intensity changes upon reaction with dopamine may be used as a light-emitting unit.

[0085] In this case, if the wavelength bands of the fluorescence of the first and second fluorescent sensors are the same (or overlap), there is a possibility that the physiologically active substances detected by the detection unit 90 cannot be distinguished. Therefore, the first and second fluorescent sensors may be configured to emit fluorescence of different wavelength bands when reacting with one of the physiologically active substances, and the wavelength band of the excitation light emitted by the light source unit may be configured to be switchable between a first excitation light corresponding to the wavelength band of the fluorescence emitted by the first fluorescent sensor and a second excitation light corresponding to the wavelength band of the fluorescence emitted by the second fluorescent sensor. In this case, if the above-mentioned monitoring of the fluorescence intensity by the detection unit 90 is performed while alternately switching between the first excitation light and the second excitation light every 2.5 seconds, it becomes possible to continuously monitor each physiologically active substance every 5 seconds.

[0086] In the above embodiment, a configuration for detecting an endogenous physiologically active substance is exemplified, but the present invention is not limited to this configuration, and it is also possible to detect, for example, an exogenous substance as a detection target. Examples of exogenous detection targets include odorous substances, taste substances, drugs, etc. In this case, a substance containing a receptor for the exogenous substance (for example, an olfactory receptor, a taste receptor, a drug receptor, etc.) may be used as the fluorescent sensor.

[0087] REFERENCE SIGNS LIST 1...detection device, 10...optical fiber, 11...fiber body, 11A...fiber core, 11B...fiber clad, 12...emission section, 20...covering section, 21...void forming section, 21B...cylindrical section (cylindrical wall), 22...severing section, 23...void, 24...cylindrical wall, 30...light emitting section, 31...hydrogel, 32...cell, 32A...fluorescence sensor, 60...photodetector (imaging section), 90...detection section, FS...fiber sensor, J...optical axis

Claims

1. A fiber sensor for detecting an object using an optical fiber having a fiber body provided with an emission part for emitting excitation light and a coating part for coating the fiber body, a gap forming portion that forms a gap including an axis of the fiber body and opening to the outside, on a tip side of the fiber body; a light-emitting portion located in the gap and capable of emitting light; Equipped with the light emitting unit includes a light emitting sensor whose light emission intensity changes when reacting with the object, the gap forming portion has a cylindrical wall that is positioned radially outward from the fiber body and is formed in a cylindrical shape around the entire circumference with the axis as the center, the optical fiber has a division portion where the coating portion is divided in a longitudinal direction to expose the fiber body, the cylindrical wall is formed by moving the divided coating portion toward the tip side of the fiber body, A fiber sensor, wherein the length of the divided portion in the longitudinal direction is the same as the length from the emission portion to the tip of the cylindrical wall.

2. A fiber sensor for detecting an object using an optical fiber having a fiber body provided with an emission part for emitting excitation light and a coating part for coating the fiber body, The fiber body has a fiber core located at the center and a fiber clad covering an outer circumferential surface of the fiber core, the coating portion coats an outer peripheral surface of the fiber clad, a gap forming portion that forms a gap including an axis of the fiber body and opening to the outside, on a tip side of the fiber body; a light-emitting portion located in the gap and capable of emitting light; Equipped with the light emitting unit includes a light emitting sensor whose light emission intensity changes when reacting with the object, The gap forming portion forms the gap extending in a direction intersecting the axis, The gap penetrates the fiber body at a position closer to the base end than the end face of the tip end of the optical fiber, and opens in the coating.

3. The coating portion is formed of a resin. The fiber sensor according to claim 2.

4. the light-emitting unit has cells encapsulated in a hydrogel; The cell has the luminescence sensor inside or on the cell membrane. The fiber sensor according to claim 1 .

5. The luminescence sensor is a protein contained in the cell. The fiber sensor according to claim 4.

6. The cell has a protein that causes an influx of a predetermined ion when reacting with the target substance, and a fluorescent indicator that changes in fluorescence emission intensity in response to the ion as the luminescence sensor. The fiber sensor according to claim 4.

7. the light-emitting unit has liposomes encapsulated in a hydrogel, The liposome contains the luminescence sensor. The fiber sensor according to claim 1 .

8. The light emitting unit has a hydrogel including the light emitting sensor. The fiber sensor according to claim 1 .

9. The light-emitting unit has a protein encapsulated in a hydrogel as the light-emitting sensor. The fiber sensor according to claim 1 .

10. The light emitting unit includes a DNA probe as the light emitting sensor contained in a hydrogel. The fiber sensor according to claim 1 .

11. The cylindrical wall is formed of a hardening material that is hardened by application of energy. The fiber sensor according to claim 1.

12. The void does not penetrate the optical fiber. The fiber sensor according to any one of claims 1 to 11.

13. The target substance is a physiologically active substance. A fiber sensor according to any one of claims 1 to 12.

14. A fiber sensor according to any one of claims 1 to 13; an imaging unit that captures an image of light emitted from the light emitting unit through the optical fiber; a detection unit that detects the object based on the captured luminescence image; A detection device having the following:

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