JP5814806B2 - Optical waveguide type measuring system, measuring method, and optical waveguide type sensor chip - Google Patents

Description

  Embodiments described below generally relate to an optical waveguide measurement system, a measurement method, and an optical waveguide sensor chip.

  Conventionally, a measurement target is formed on the surface of an optical waveguide by an antigen-antibody reaction using a fine particle on which an antibody that specifically binds to the measurement target substance is immobilized and an optical waveguide on which an antibody that specifically binds to the measurement target substance is immobilized. An optical waveguide sensor chip that binds fine particles via a substance is disclosed. Absorbance caused only by fine particles bound to the surface of the optical waveguide due to the antigen-antibody reaction is detected by evanescent light near the surface of the optical waveguide, so the measurement target substance can be quantified without including the procedure for washing excess specimens and secondary antibodies. Is possible. However, microparticles may be adsorbed on the surface of the optical waveguide regardless of the antigen-antibody reaction, and light is absorbed and scattered by the microparticles adsorbed regardless of the antigen-antibody reaction. In some cases, measurement errors may occur.

  Thus, assuming an inspection item that requires detection with higher sensitivity, development of a technique for improving the detection sensitivity of the measurement target substance with higher accuracy is desired.

JP 2009-133842 A

  The problem to be solved by the present invention is to provide an optical waveguide measurement system, a measurement method, and an optical waveguide sensor chip capable of improving the detection sensitivity of a measurement target substance with higher accuracy.

The optical waveguide type measurement system according to the embodiment includes an optical waveguide having a sensing area in which a first substance that specifically binds to a measurement target substance is fixed, and a second substance that specifically binds to the measurement target substance. It is fixed and has a particle size of 0.2 μm or more and 20 μm or less, is provided so as to cover the first core and the first core, includes magnetic nanoparticles, and has a refractive index of the first core A magnetic fine particle having a high refractive index, a magnetic field applying unit for generating a magnetic field for moving the magnetic fine particle, a light source for causing light to enter the optical waveguide, and light emitted from the optical waveguide. A light receiving element for receiving light.

It is a figure which shows the structure of the optical waveguide type measuring system which concerns on 1st Embodiment. It is a schematic diagram which shows the form of a magnetic microparticle. (A)-(c) is process drawing which shows the method of measuring the measuring object substance in a sample solution. It is a figure which shows the example of a time-dependent change of detection signal intensity ratio. It is a figure which shows the calibration curve of a measurement result. It is a figure which shows the structure of the optical waveguide type measuring system which concerns on 2nd Embodiment. (A)-(c) is process drawing which shows the method of measuring the measuring object substance in a sample solution. It is a figure which shows the structure of the optical waveguide type measuring system which concerns on 3rd Embodiment. (A)-(c) is process drawing which shows the method of measuring the measuring object substance in a sample solution. (A) is a schematic diagram for illustrating the structure of a magnetic field application part, (b) is an AA arrow directional view in (a), (c) is a schematic perspective view. (A) is a schematic diagram for illustrating the structure of a magnetic field application part, (b) is a BB arrow line view in (a), (c) is a schematic perspective view. It is a graph for demonstrating the effect | action and effect of a magnetic field application part. (A) has a step of removing magnetic fine particles that may be noise by the magnetic field application unit, and (b) is a case of further having a step of stirring the magnetic fine particle dispersion and the sample solution by the magnetic field application unit. . It is a graph for demonstrating the effect | action and effect of a magnetic field application part. (A) performs only application of the upper magnetic field by the magnetic field application unit, and (b) and (c) perform combination of application of the lower magnetic field by the magnetic field application unit and application of the upper magnetic field by the magnetic field application unit. Is the case. (A)-(d) is process drawing which shows the method of measuring the measuring object substance in a sample solution.

Hereinafter, embodiments will be illustrated with reference to the drawings. In addition, in each drawing, the same code | symbol is attached | subjected to the same component and detailed description is abbreviate | omitted suitably.
In this specification, “upper”, “upward”, and “upward” are “upper”, “upward”, and “upward” in the direction of gravity, and “lower”, “lower”, and “downward”. And “lower”, “lower” and “down” in the direction of gravity.

[First embodiment]
FIG. 1 is a diagram showing a configuration of an optical waveguide measurement system according to the present embodiment. The optical waveguide measurement system 30 according to the present embodiment includes an optical waveguide sensor chip 100, a light source 7, a light receiving element 8, and a magnetic field application unit 10 (corresponding to an example of a first magnetic field application unit). .

  Further, the optical waveguide sensor chip 100 according to the present embodiment includes the substrate 1, the grating 2, the optical waveguide 3 on which the first substance 6 that specifically reacts with the measurement target substance 14 is immobilized, and the protection. A film 4, a frame 5, and magnetic fine particles 9 on which a second substance 13 that specifically reacts with the measurement target substance 14 is immobilized.

  As the optical waveguide 3, for example, a planar optical waveguide can be used. The optical waveguide 3 can be formed of, for example, a thermosetting resin or photocurable resin such as phenol resin, epoxy resin, or acrylic resin, or non-alkali glass. Specifically, the material used here is a material having transparency to predetermined light, and is preferably a resin having a higher refractive index than that of the substrate 1, for example. The immobilization of the first substance 6 that specifically reacts with the measurement target substance 14 in the sample solution in the sensing area 101 that is the detection surface is, for example, a hydrophobic interaction with the sensing area 101 that is the surface of the optical waveguide 3. This can be done by chemical bonding.

  As the first substance 6, for example, when the measurement target substance 14 in the sample solution is an antigen, an antibody (primary antibody) can be used.

  The magnetic fine particles 9 are held in a dispersed state on the sensing area 101 or are held in another space or a container (not shown). Here, “the magnetic fine particles are held in a dispersed state on the sensing area” means that the magnetic fine particles 9 are held in a dispersed state directly or indirectly above the sensing area 101. Examples of the form “the magnetic fine particles are indirectly dispersed above the sensing area 101” include a form in which the magnetic fine particles 9 are dispersed on the surface of the sensing area 101 via a blocking layer. The blocking layer includes a water-soluble substance such as polyvinyl alcohol, bovine serum albumin (BSA), polyethylene glycol, phospholipid polymer, gelatin, casein, and sugars (for example, sucrose and trehalose). As another example, there is a form in which the magnetic fine particles 9 are arranged above the sensing area 101 with a space therebetween. For example, a support plate (not shown) facing the sensing area 101 may be arranged, and the magnetic fine particles 9 may be held in a dispersed state on the surface of the support plate facing the sensing area 101. In this case, it is desirable that the magnetic fine particles 9 be held in a dry or semi-dry state. Although it is desirable to easily re-disperse when it comes into contact with a dispersion medium such as a sample solution, the form held in a dry or semi-dry state does not necessarily need to be in a completely dispersed state. When it is held in another space or container, it may be in a state of being dispersed in a dispersion, a state of being settled in a dispersion medium, etc. in addition to a dry or semi-dry state.

FIG. 2 is a schematic diagram showing the form of the magnetic fine particles 9.
FIG. 2A is a schematic diagram for illustrating the appearance of magnetic fine particles, and FIGS. 2B and 2C are schematic diagrams for illustrating a cross section of the fine particles.
As shown in FIG. 2A, the magnetic fine particles 9 are obtained by fixing the second substance 13 on the surface of the fine particles 12. As the second substance 13, for example, when the measurement target substance 14 in the sample solution is an antigen, an antibody (secondary antibody) can be used.
In this case, as shown in FIG. 2B, fine particles 12b in which the magnetic nanoparticles 12a are wrapped with a polymer material, or as shown in FIG. 2C, a core 12c and a shell provided so as to cover the core 12c. Thus, the fine particles 12d can be obtained.
The core 12c can be formed from a polymer material. The shell is formed of a polymer material and can include the magnetic nanoparticle 12a.
Alternatively, it may be a fine particle itself made of a magnetic material, and in this case, it is desirable to have a functional group that binds the substance to be measured to the fine particle surface. Examples of the magnetic material used for the fine particles 12 include various ferrites such as γ-Fe 2 O 3. In this case, it is preferable to use a material having superparamagnetism that quickly loses magnetism when the application of the magnetic field is stopped.
In general, superparamagnetism is a phenomenon that occurs in nanoparticles of several tens of nm or less. On the other hand, in order for light scattering to occur, the size of the fine particles needs to be several hundred nm or more. Therefore, as the magnetic fine particles 9 in the present embodiment, those in which magnetic nanoparticles 12a are wrapped with a polymer material or the like as shown in FIG. 2B or FIG. 2C are suitable.
In general, the refractive index is often about 1.5 to 1.6 for polymer materials, and about 3.0 for ferrites. Further, when the magnetic fine particles 9 are near the surface of the optical waveguide 3, the higher the refractive index, the easier it is to scatter light. For this reason, it is considered that the magnetic nanoparticles 12a distributed near the surface of the fine particles 12 to which the evanescent light is applied can be detected with higher sensitivity.
Here, as shown in FIG. 2B, in the fine particles 12b in which the magnetic nanoparticles 12a are simply wrapped with a polymer material, the magnetic nanoparticles 12a are distributed throughout the fine particles. Therefore, from the viewpoint of detection sensitivity, as shown in FIG. 2C, a structure in which the core-shell type fine particles 12d are included and the magnetic nanoparticles 12a are included in the shell at a high density is suitable.
The particle size of the fine particles 12 is preferably 0.05 μm or more and 200 μm or less, more preferably 0.2 μm or more and 20 μm or less. Since the light scattering efficiency is increased by using this particle size, detection sensitivity can be improved in the optical waveguide measurement system 30 that detects the measurement target substance 14 using light.

  The combination of the measurement target substance 14 and the first substance or the second substance that specifically binds to the measurement target substance 14 is not limited to a combination of an antigen and an antibody. Other examples include sugars and lectins, nucleotide chains and complementary nucleotide chains, ligands and receptors, and the like.

  At both ends of the main surface of the substrate 1, an incident side grating 2a and an emission side grating 2b are provided. The substrate 1 is, for example, alkali-free glass. The gratings 2a and 2b are formed of a material having a higher refractive index than the substrate. The optical waveguide 3 having a flat surface is formed on the main surface of the substrate 1 including the gratings 2a and 2b. The protective film 4 is coated on the optical waveguide 3. The protective film 4 is a resin film having a low refractive index, for example. The protective film 4 is provided with an opening through which a part of the surface of the optical waveguide 3 located between the gratings 2a and 2b is exposed. The opening can be, for example, rectangular, and the surface of the optical waveguide 3 exposed in the opening serves as the sensing area 101. The frame 5 is formed on the protective film 4 so as to surround the sensing area 101.

  The first substance 6 that specifically reacts with the measurement target substance 14 in the sample solution is immobilized in the sensing area 101 by, for example, a hydrophobization process using a silane coupling agent. Alternatively, a functional group may be formed in the sensing area 101, and an appropriate linker molecule may act to immobilize the sensing area 101 by chemical bonding. The second substance 13 that specifically reacts with the measurement target substance 14 in the sample solution is immobilized on the surface of the fine particle 12 by, for example, physical adsorption or chemical bonding via a carboxyl group, an amino group, or the like. The magnetic fine particles 9 on which the second substance 13 is immobilized are dispersed and held in the sensing area 101 on which the first substance 6 is immobilized. The dispersion and holding of the magnetic fine particles 9 are formed by, for example, applying and drying a slurry containing the magnetic fine particles 9 and a water-soluble substance on the sensing area 101 or a surface (not shown) facing the sensing area 101 and the like. The Alternatively, the magnetic fine particles 9 may be dispersed in a liquid and held in a space different from the reaction space 102 or a container (not shown).

  The light source 7 irradiates the above-described optical waveguide sensor chip 100 with light. The light source 7 is, for example, a red laser diode. The light incident from the light source 7 is diffracted by the incident side grating 2 a and propagates through the optical waveguide 3. Thereafter, the light is diffracted and emitted by the emission side grating 2b. The light emitted from the emission side grating 2b is received by the light receiving element 8, and the light intensity is measured. The light receiving element 8 is, for example, a photodiode. The amount of the magnetic fine particles 9 is measured by comparing the intensity of the incident light and the emitted light and measuring the light absorption rate. Then, the antigen concentration in the sample solution is obtained based on the measured amount of the magnetic fine particles 9. Details regarding the determination of the antigen concentration in the sample solution based on the measured amount of the magnetic fine particles 9 will be described later.

The magnetic field application unit 10 applies a magnetic field to the optical waveguide sensor chip 100. The magnetic field application unit 10 generates a magnetic field and applies the generated magnetic field to the optical waveguide sensor chip 100 to move the magnetic fine particles 9 according to the magnetic field. The magnetic field application unit 10 is disposed in a direction opposite to the direction in which the optical waveguide 3 exists when viewed from the magnetic fine particles 9. In the present embodiment, the magnetic field application unit 10 is installed in the upward direction in FIG. The magnetic field application unit 10 is, for example, a magnet or an electromagnet. In order to dynamically adjust the magnetic field strength, a method of adjusting with an electric current using an electromagnet is desirable, but using a ferrite magnet or the like, the magnetic field strength is adjusted according to the strength of the magnet itself or the distance from the optical waveguide sensor chip 100. May be.
For example, the magnetic field strength can be adjusted by disposing a ferrite magnet above the optical waveguide sensor chip 100 and changing the thickness of the ferrite magnet through a spacer between the magnet and the optical waveguide sensor chip 100. In addition, the magnetic field strength can be adjusted by changing the relative position of the ferrite magnet and the optical waveguide sensor chip 100 using an actuator such as a linear motor.
When an electromagnet is used, the coil is disposed on the side opposite to the settling direction (the direction of the optical waveguide 3) when viewed from the magnetic fine particles 9, and a current is applied to the coil. Can be adjusted.

  In this embodiment, by applying a magnetic field to the magnetic fine particles 9 by the magnetic field application unit 10, the magnetic fine particles 9 adsorbed on the sensing area 101 can be peeled off from the sensing area 101 without depending on the antigen-antibody reaction. it can. As a result, it is possible to measure the absorbance due to only the magnetic fine particles 9 bound to the sensing area 101 via the measurement target substance 14 by the antigen-antibody reaction, and reduce measurement errors.

  At this time, as the fine particles 12 of the magnetic fine particles 9, it is preferable to use those having superparamagnetism that quickly loses magnetization when the application of the magnetic field is stopped. Thereby, even if the magnetic fine particles 9 aggregate due to magnetization when a magnetic field is applied, they can be redispersed by stopping the application of the magnetic field. For example, even if a magnetic field is applied when the measurement target substance 14 is not present in the sample solution, an aggregate of the magnetic fine particles 9 may be generated and hardly peeled off from the sensing area 101 in some cases. Such agglomerates of magnetic fine particles 9 cause measurement errors. In this case, if the fine particles 12 have superparamagnetism, aggregation of the magnetic fine particles 9 can be suppressed, so that generation of measurement errors can be suppressed.

  Further, in order to further improve the redispersibility when the application of the magnetic field is stopped, the surface of the fine particles 12 may have a positive or negative charge. Alternatively, a dispersant such as a surfactant may be added to the dispersion medium of the magnetic fine particles 9.

  Furthermore, in the present embodiment, the naturally settled magnetic fine particles 9 can be pulled back upward by the magnetic field application unit 10. By repeating the natural sedimentation of the magnetic fine particles 9 and the upward pulling back by the magnetic field applying unit 10, the sample solution and the magnetic fine particles 9 can be stirred. Thereby, the coupling | bonding by the antigen antibody reaction of the magnetic microparticle 9 and the sensing area 101 via the antigen (measuring target substance 14) contained in the sample solution is promoted, and high detection sensitivity can be obtained in a shorter time. Therefore, when the measurement target substance 14 has a low concentration, the detection sensitivity can be increased.

  If the surface of the fine particles 12 is positively or negatively charged, or a dispersant such as a surfactant is added, the magnetic fine particles 9 can be easily redispersed when the application of the magnetic field is stopped, and stirring is further performed. Can be promoted. Thereby, it is possible to further improve the detection sensitivity.

FIG. 3 is a process diagram showing a method for measuring a substance to be measured in a sample solution.
Here, a method for measuring the amount of the measurement target substance 14 using the optical waveguide type measurement system 30 described above will be described with reference to FIGS.
The state in the reaction space 102 will be described.

First, an optical waveguide type measurement system 30 shown in FIG. 1 is prepared. Next, as shown in FIG. 3A, the sample solution is introduced onto the optical waveguide 3 in which the magnetic fine particles 9 are dispersed and held, and the magnetic fine particles 9 are redispersed. When the magnetic fine particles 9 are held in a space other than the optical waveguide 3 or in a separate container, a mixed dispersion of the sample solution and the magnetic fine particles 9 is introduced. Alternatively, the dispersion of the magnetic fine particles 9 and the sample solution may be introduced separately, such as first introducing the dispersion of the magnetic fine particles 9 and then introducing and mixing the sample solution.
That is, the sample solution containing the measurement target substance 14 and the magnetic fine particles 9 having magnetism in which the second substance 13 that specifically binds to the measurement target substance 14 is immobilized are provided in the optical waveguide sensor chip 30 and the measurement target. The first substance 6 that specifically binds to the substance 14 may be in contact with the immobilized sensing area 101.
As an introduction method, for example, dripping or inflow can be considered.

  Next, as shown in FIG. 3B, the magnetic fine particles 9 settle toward the sensing area 101 by their own weight. At this time, the first substance 6 (for example, a primary antibody) immobilized on the sensing area 101 and the second substance 13 (for example, a secondary antibody) immobilized on the surface of the fine particles 12 are measured substances 14 ( For example, it binds by antigen-antibody reaction via antigen). Thereby, the magnetic fine particles 9 are coupled to the sensing area 101.

  Next, as shown in FIG. 3C, a magnetic field is applied from a direction different from the settling direction as viewed from the magnetic fine particles 9 (for example, upward), thereby adsorbing to the sensing area 101 without passing through the measurement target substance 14. The magnetic fine particles 9 are removed from the sensing area 101 by moving in a direction (for example, upward) different from the settling direction.

  At this time, by setting the magnetic field strength to an appropriate value, the magnetic fine particles 9 bound to the sensing area 101 through the measurement target substance 14 by the antigen-antibody reaction are not peeled off, and the sensing area is not passed through the measurement target substance 14. Only the magnetic fine particles 9 adsorbed on 101 can be removed.

In the present embodiment, it is conceivable to obtain an appropriate magnetic field strength as follows. The states of the magnetic fine particles 9 that can be detected by near-field light such as evanescent light can be classified into the following (State 1) to (State 3) depending on the strength of interaction with the sensing area 101. Stated in the order of strong interaction, (State 1) is the state of the magnetic fine particles 9 bound to the sensing area 101 by binding of the measurement target substance 14 and the molecule that specifically binds to it, such as antigen-antibody binding, (State 2) is a state of the magnetic fine particles 9 non-specifically adsorbed on the sensing area 101 due to intermolecular force or hydrophobic interaction, and (state 3) is a state of the magnetic fine particles 9 floating in the vicinity of the sensing area 101, It is. The magnetic fine particles 9 in (State 1) are magnetic fine particles 9 that should contribute to the concentration detection of the measurement target substance 14, and the magnetic fine particles 9 in (State 2) or (State 3) become a measurement error factor (noise). Magnetic fine particles 9 that can be obtained. The magnetic fine particles 9 in (State 1) will be appropriately referred to as magnetic fine particles 9 combined with the sensing area 101.
Further, the magnetic fine particles 9 in (state 2) will be appropriately referred to as magnetic fine particles 9 adsorbed on the sensing area 101.

  Here, “the vicinity of the surface” of the optical waveguide 3 that can be detected by near-field light is, for example, evanescent light that permeates the surface of the propagation body when light propagates by total reflection, and the permeation distance d is expressed by It is calculated | required by (1). From formula (1), it can be seen that the bleed distance d is approximately a fraction of the wavelength of the light used for the measurement.

d = λ / {2π (n ₁ ² sin ² θ−n ₂ ² ) ^1/2 } (1)
Where d is the evanescent light penetration distance, λ is the wavelength of light used for measurement, n1 is the refractive index of the optical waveguide 3, n2 is the refractive index of the dispersion medium in which the magnetic fine particles 9 are dispersed, and θ is the total reflection angle. is there.
Therefore, the magnetic field application unit 10 applies a magnetic field having such a magnetic field strength that the magnetic fine particles 9 are separated from the sensing area 101 by a distance L that satisfies the following expression (2).
L> λ / {2π (n ₁ ² sin ² θ−n ₂ ² ) ^1/2 } (2)
Here, L is the distance that the magnetic fine particles 9 are separated from the sensing area 101, λ is the wavelength of light used for measurement, n1 is the refractive index of the optical waveguide 3, n2 is the refractive index of the dispersion medium that disperses the magnetic fine particles 9, and θ is The total reflection angle.
For example, if λ = 635 nm, n1 = 1.58, n2 = 1.33 (when the dispersion medium is water), and θ = 78 °, L> 130 nm. Therefore, the measurement error can be sufficiently reduced by applying the magnetic field and moving the magnetic fine particles 9 in (State 2) and (State 3) from the sensing area 101 by only a few hundred nm. Therefore, the time required to move the magnetic fine particles 9 in (State 2) or (State 3) away from the sensing area 101 to a distance that does not cause an error in detection sensitivity is very short. If the time is within an allowable range, the magnetic fine particles 9 in (State 2) and (State 3) are moved to a distance that does not cause measurement error even with a weak magnetic field strength, even if some time is required. It becomes possible. Thereby, the possibility that the magnetic fine particles 9 required for the measurement of (state 1) are excessively peeled off can be reduced. That is, the magnetic fine particles 9 (state 2) and (state 3) that can contribute to measurement without causing the magnetic fine particles 9 in (state 1) to be removed from the sensing area 101 to be measured from the sensing area 101 are measured from the sensing area 101. Since it can be peeled off to a distance that does not affect, the S / N ratio can be improved.

  Thus, the appropriate magnetic field strength means that the magnetic fine particles 9 in (state 1) that should contribute to the measurement do not peel off from the sensing area 101, and can cause noise in the measurement (state 2) or (state 3). The magnetic field intensity is appropriate for peeling the fine particles 9 from the sensing area 101 to a distance that does not affect the measurement. As described above, a method of optimally adjusting the magnetic field intensity with an electric current using an electromagnet is desirable. However, using a ferrite magnet or the like, the strength of the magnet itself, or the relative position between the optical waveguide sensor chip 100 and the magnet. May be changed to adjust the magnetic field strength. When an electromagnet is used, the coil is disposed on the side opposite to the settling direction (the direction of the optical waveguide 3) when viewed from the magnetic fine particles 9, and a current is applied to the coil. Can be adjusted.

In order to optimally adjust the magnetic field strength, the optical waveguide measurement system 30 according to the present embodiment includes a control unit 20 (a second control unit) that controls the magnetic field strength of the magnetic field applied by the magnetic field application unit 10. (Corresponding to an example). By performing the control as described above by the control unit 20, the magnetic field strength can be adjusted to an appropriate strength. For example, the magnetic particles 9 (state 2) and (state 3) that can contribute to the measurement without causing the magnetic particles 9 in (state 1) to be removed from the sensing area 101 can be measured from the sensing area 101. The magnetic field strength can be adjusted so that the magnetic field can be peeled to a distance that does not affect the magnetic field.
Further, when the magnetic field intensity is adjusted as needed, it can be dynamically controlled by adjusting the control unit 20.
For example, the control unit 20 (corresponding to an example of the first control unit) that controls at least one of the timing and the length of time of applying the magnetic field by the magnetic field application unit 10 may be used.

  Then, by measuring the difference in the detection signal intensity ratio in the light receiving element 8, the amount (for example, antigen concentration) of the measurement target substance 14 in the sample solution can be measured. Specifically, in FIG. 1, laser light is incident from the light source 7 to the optical waveguide 3 from the incident side grating 2a, propagates through the optical waveguide 3, and evanescent light or the like near the surface (exposed surface in the sensing area 101). Of near-field light. In this state, when a mixed dispersion of the sample solution and the magnetic fine particles 9 is introduced onto the sensing area 101, the magnetic fine particles 9 settle immediately after (FIG. 3 (a)) and near the sensing area 101, for example, evanescent light. The region is reached (FIG. 3 (b)). Since the magnetic fine particles 9 are involved in the absorption and scattering of the evanescent light, the intensity of the reflected light is attenuated. As a result, when the laser beam emitted from the emission side grating 2b is received by the light receiving element 8, the intensity of the emitted laser beam decreases with time due to the influence of the combined magnetic fine particles 9. Thereafter, when the upper magnetic field is applied by the magnetic field application unit 10, the magnetic fine particles 9 in (state 2) and (state 3) move out of the evanescent light region (FIG. 3 (c)). Recovers to the value of. The received light intensity at this time is compared with the received light intensity in the state shown in FIG. In addition, the light intensity of light emitted from the optical waveguide sensor chip 100 (corresponding to an example of the first light intensity) after the sample solution or the like is introduced into the optical waveguide sensor chip 100 and before the magnetic field is applied. ). Further, after application of the magnetic field, the light intensity of light emitted from the optical waveguide sensor chip 100 (corresponding to an example of the second light intensity) is measured. Then, the measurement target substance 14 can be quantified based on the difference between these light intensities.

  The decreasing rate of the intensity of the laser beam received by the light receiving element 8 depends on the amount of the magnetic fine particles 9 bound to the sensing area 101 mainly by an antigen-antibody reaction. That is, it is proportional to the antigen concentration in the sample solution involved in the antigen-antibody reaction. Therefore, obtain a fluctuation curve of laser light intensity over time in a sample solution with a known antigen concentration, obtain the rate of decrease in laser light intensity at a predetermined time after application of the upper magnetic field of this fluctuation curve, and determine the antigen concentration And a calibration curve showing the relationship between the laser beam intensity reduction rate and the laser beam intensity reduction rate. Next, a decrease rate of the laser light intensity at a predetermined time is obtained from the time measured by the above method in a sample solution with an unknown antigen concentration and a fluctuation curve of the laser light intensity, and the decrease rate of the laser light intensity is calculated using the calibration curve. Can be used to measure the antigen concentration in the sample solution.

  Next, an example in which the measurement of the present embodiment is performed by experiment will be described. The following specific numerical values and materials are examples, and are not limited to these numerical values and materials.

  In the experiment, a titanium oxide film having a refractive index of 2.2 to 2.4 is formed on a light-transmitting substrate 1 such as glass to a thickness of 50 nm by a sputtering method, and a lithography method and a dry etching method are performed. Thus, gratings 2a and 2b were formed. An ultraviolet curable acrylic resin film having a film thickness of about 10 μm was formed on the substrate 1 on which the gratings 2 a and 2 b were formed by a spin coating method and ultraviolet irradiation to obtain an optical waveguide 3. The refractive index after curing is 1.58.

  The protective film 4, which is a low refractive index resin film, includes a region corresponding to the upper side of the gratings 2 a and 2 b on the surface of the optical waveguide 3, and screen printing is performed so as to surround the antibody immobilization region that is the sensing area 101. Formed using. The refractive index after drying of the protective film 4 is 1.34. In order to form a liquid reservoir for holding the sample solution and the like, the resin frame 5 was fixed with double-sided tape. The first substance 6 for the measurement target substance 14 was immobilized on the surface of a region where no protective film was formed between the gratings by a covalent bond method.

  In the present embodiment, rat insulin is used as the measurement target substance 14, and an anti-rat insulin antibody is used as the first substance 6 immobilized on the sensing area 101. In addition, the magnetic fine particles 9 have core-shell type fine particles 12d containing a high density of magnetic nanoparticles 12a in the shell. The average particle diameter of the fine particles 12d was 1.1 μm. On the surface of the fine particles 12d, an anti-rat insulin antibody was immobilized as the second substance 13 by a covalent bond method. A dispersion containing such magnetic fine particles 9 was separately prepared.

  Next, light having a central wavelength of 635 nm from the light emitting diode 7 is incident from the grating 2a on the incident side, and the light intensity of the light emitted from the grating 2b on the emitting side is measured by the photodiode 8, while the sample solution and the magnetic fine particles are measured. After mixing with 9 dispersion liquid, it was introduced into the sensing area 101 (inside the frame 5). Thereafter, measurement was performed according to the measurement procedure described above.

  In this embodiment, the ferrite magnet is disposed above the optical waveguide sensor chip 100, a spacer is provided between the ferrite magnet and the optical waveguide sensor chip 100, and the magnetic field strength is changed by changing the thickness of the spacer. I let you.

FIG. 4 is a diagram illustrating an example of a change over time in the detection signal intensity ratio.
FIG. 4 shows an example of the change over time of the detection signal intensity ratio in the measurement method according to the present embodiment.
FIG. 5 is a diagram showing a calibration curve of measurement results.
As shown in FIG. 4, when the dispersion of magnetic fine particles 9 and the sample solution are first mixed and introduced into the sensing area 101, the detection signal intensity ratio increases according to the increase in fine particle density in the vicinity of the sensing area 101 due to sedimentation of the magnetic fine particles 9. descend. After that, when the upper magnetic field is applied by the magnetic field application unit 10, the magnetic fine particles 9 that can become noise adsorbed to the sensing area 101 are removed, so that the detection signal intensity ratio rises again and is lower than the initial detection signal intensity ratio. Saturates. The difference between the detection signal intensity ratio and the initial detection signal intensity ratio at this stage is defined as the ratio to the initial detection signal intensity ratio, that is, the signal decrease rate. A calibration curve shown in FIG. 5 represents the relationship between the signal decrease rate and the rat insulin concentration.

  4 and 5, when the measurement method of the present embodiment is used, by removing the magnetic fine particles 9 due to non-specific adsorption that cause measurement errors as noise from the evanescent wave region with an appropriate magnetic field, an extremely low concentration can be obtained. It was confirmed that rat insulin could be detected.

  According to the present embodiment, by applying a magnetic field to the magnetic fine particles 9 in a direction different from the settling direction, the magnetic fine particles 9 that may become noise adsorbed on the sensing area 101 without depending on the antigen-antibody reaction or the like are sensed. It can be peeled off from the area 101. Thereby, since the light absorbency resulting from the magnetic fine particle 9 couple | bonded with the sensing area 101 via the measurement object substance 14 by antigen antibody reaction etc. can be measured, a measurement error can be reduced.

  In addition, since the magnetic fine particles 9 that can become noise by applying a magnetic field can be removed, the work of removing such magnetic fine particles 9 by washing becomes unnecessary.

  According to the present embodiment, since the optical waveguide sensor chip 100 is used and measurement is performed by near-field light such as evanescent light, the distance for peeling the magnetic fine particles 9 from the sensing area 101 to a range that does not affect the measurement is short. Tesumu. Thereby, the time required for peeling off the magnetic fine particles 9 from the sensing area 101 by applying the upper magnetic field can be shortened. Alternatively, the magnetic fine particles 9 can be peeled from the sensing area 101 to a range that does not affect the measurement by a weaker magnetic field.

  In addition, according to the present embodiment, since the magnetic field strength can be controlled, the magnetic fine particles 9 that can be measurement noise can be removed from the sensing area 101 without peeling the magnetic fine particles 9 that should contribute to the measurement from the sensing area 101. It can be peeled to a distance that does not affect the measurement. As a result, the S / N ratio can be improved.

  Moreover, according to the present embodiment, the control accuracy can be kept high by dynamically controlling the magnetic field intensity by the control unit 20.

Further, if the magnetic fine particles 9 have superparamagnetism that quickly loses magnetization when the application of the magnetic field is stopped, the magnetic fine particles 9 can be easily redispersed when the application of the magnetic field is stopped. be able to. For this reason, even when the measurement target substance 14 does not exist in the sample solution, the generation of aggregates of the magnetic fine particles 9 is suppressed, so that generation of measurement errors can be suppressed.
Further, by using the magnetic fine particles 9 having the core-shell type fine particles 12d including the magnetic nano fine particles 12a in the shell, the scattering intensity of the evanescent light can be increased. As a result, highly sensitive detection can be performed.

  Further, by adding a positive or negative charge to the surface of the fine particles 12 in the magnetic fine particles 9 or adding a dispersant such as a surfactant, the magnetic fine particles 9 are redispersed when the application of the magnetic field is stopped. It is also possible to reduce the measurement error.

  Further, according to the present embodiment, the naturally settled magnetic fine particles 9 can be pulled back by applying a magnetic field in a direction different from the settling direction. By repeating the natural sedimentation of the magnetic fine particles 9 and the upward pulling back by the magnetic field applying unit 10, the sample solution and the magnetic fine particles 9 are agitated, so that the measurement target substance 14 (for example, an antigen) contained in the sample solution and The antigen-antibody reaction with the magnetic fine particles 9 is promoted, and high detection sensitivity can be obtained in a shorter time. Therefore, when the measurement target substance 14 has a low concentration, the detection sensitivity can be increased.

  At this time, further, the surface of the fine particles 12 in the magnetic fine particles 9 is given a positive or negative charge, or a dispersing agent such as a surfactant is added, so that the magnetic fine particles 9 are removed when the application of the magnetic field is stopped. It is possible to facilitate redispersion, promote stirring, and improve detection sensitivity.

  In addition, according to the present embodiment, the optical waveguide sensor chip 100 is used to measure the amount and concentration of the measurement target substance 14 using near-field light such as evanescent light. In this case, if the magnetic fine particles 9 having a particle diameter of 0.05 μm or more and 200 μm or less, preferably 0.2 μm or more and 20 μm or less are used, the light scattering efficiency can be increased. Detection sensitivity can be improved.

[Second Embodiment]
In the first embodiment, the case where a magnetic field is applied in a direction opposite to the direction of the optical waveguide 3 as viewed from the magnetic fine particles 9 has been described. However, in the second embodiment, the optical waveguide 3 as viewed from the magnetic fine particles 9 is described. A case where a magnetic field is applied in both the direction and the opposite direction will be described.

  FIG. 6 is a diagram illustrating a configuration of an optical waveguide measurement system according to the second embodiment. The optical waveguide measurement system 30a according to the present embodiment further includes a magnetic field application unit 11 (corresponding to an example of a second magnetic field application unit) in addition to the optical waveguide measurement system 30 of the first embodiment shown in FIG. It is added. Since the other configuration is the same as that of the first embodiment, the description thereof is omitted.

  The magnetic field application unit 11 applies a magnetic field to the optical waveguide sensor chip 100 in the direction of the optical waveguide 3 when viewed from the magnetic fine particles 9. Thereby, the magnetic fine particles 9 can be moved in the direction of the optical waveguide 3.

  The magnetic field application unit 11 is provided in the direction in which the optical waveguide 3 exists as viewed from the magnetic fine particles 9. In the present embodiment, the magnetic field application unit 11 is provided in the downward direction of the sensor chip 100.

  Similar to the magnetic field application unit 10, the magnetic field application unit 11 is a magnet or an electromagnet. In order to dynamically adjust the magnetic field strength, a method of adjusting the current by using an electromagnet is desirable, but using a ferrite magnet or the like, the strength of the magnet itself or the relative position of the optical waveguide sensor chip 100 and the magnet is changed. To adjust the magnetic field strength. For example, the magnetic field strength can be adjusted by disposing a ferrite magnet below the optical waveguide sensor chip 100 and changing the thickness of the ferrite magnet through a spacer between the magnet and the optical waveguide sensor chip 100. In the case of using an electromagnet, the coil is disposed in the direction of the optical waveguide 3 when viewed from the magnetic fine particles, and a current is applied to the coil. The magnetic field strength can be adjusted by changing the current value.

  Here, the optical waveguide type measurement system 30a of the present embodiment may further include a control unit 20a. The control unit 20a controls the strength of the magnetic field applied by the magnetic field application unit 10, the magnetic field application unit 11, or one of them. In this case, for example, as shown in FIG. 6, a common control unit 20 a and a changeover switch 20 a 1 can be provided for the magnetic field application unit 10 and the magnetic field application unit 11. In addition, independent control units may be provided for the magnetic field application unit 10 and the magnetic field application unit 11, respectively. In addition, a control unit that controls the magnetic field strength at the same time for the magnetic field application unit 10 and the magnetic field application unit 11 may be provided. Moreover, it is good also as the control part 20a which controls so that it may become an appropriate magnetic field strength dynamically by controlling a magnetic field strength as needed.

  Further, the control unit 20a may control the timing of applying the magnetic field in each of the magnetic field applying unit 10 and the magnetic field applying unit 11. Thereby, the magnetic field application unit 10 and the magnetic field application unit 11 can alternately apply a magnetic field according to a predetermined condition (for example, a predetermined time or a time during which a predetermined magnetic field is continuously applied).

FIG. 7 is a process diagram showing a method for measuring a substance to be measured in a sample solution.
Here, a method of measuring the amount of the measurement target substance 14 using the optical waveguide type measurement system 30a described above will be described with reference to FIGS.
The state in the sensing area 101 will be described.
7A and 7C are the same as those in the first embodiment shown in FIG.
Further, the measurement of the antigen concentration in the sample solution by measuring the difference in the detection signal intensity ratio in the light receiving element 8 is the same as in the first embodiment, and thus the description thereof is omitted.

  FIG. 7B is different from the first embodiment shown in FIG. 3 and will be described below. In FIG. 7B, the magnetic field application unit 11 applies a lower magnetic field in the settling direction (the direction of the optical waveguide 3, for example, the lower direction in FIG. 6) as viewed from the magnetic fine particles 9. Thereby, the magnetic fine particles 9 are attracted to the sensing area 101. At this time, the first substance 6 (for example, primary antibody) immobilized on the sensing area 101 and the second substance 13 (for example, secondary antibody) immobilized on the fine particles 12 of the magnetic fine particles 9 are measured substances. It binds by antigen-antibody reaction via 14 (for example, antigen). Thereby, the magnetic fine particles 9 are coupled to the sensing area 101.

  In the present embodiment, the downward magnetic field application shown in FIG. 7B and the upward magnetic field application shown in FIG. 7C may be alternately repeated.

  When the magnetic fine particles 9 are attracted to the optical waveguide 3 by applying a magnetic field in the downward direction shown in FIG. 7B, the substance 14 to be measured is in the first substance 6 and the second substance 13 in the sample solution. It remains in a state where it is not bonded or in a state where it is bonded to the second substance 13 immobilized on the surface of the fine particle 12 but not bonded to the first substance 6 immobilized in the sensing area 101. In addition, the magnetic fine particles 9 adsorbed nonspecifically exist in the sensing area 101.

  Therefore, in FIG. 7C, a magnetic field having a strength that does not peel off the magnetic fine particles 9 bound by the antigen-antibody reaction or the like is applied, and the magnetic fine particles 9 not bound by the antigen-antibody reaction or the like are moved in a direction different from the optical waveguide 3. Move.

  After that, as shown in FIG. 7B again, a magnetic field is applied in the direction of the optical waveguide 3 to attract the magnetic fine particles 9 that are not bound by antigen-antibody reaction or the like. Then, the measurement target substance 14 and the measurement target substance 14 bound to the second substance 13 immobilized on the surface of the fine particle 12 are newly bound to the first substance 6 immobilized in the sensing area 101.

  By repeating this, the number of magnetic fine particles 9 that are not bonded to the sensing area 101 due to an antigen-antibody reaction or the like can be reduced, and the number of magnetic fine particles 9 that are bonded to the sensing area 101 due to an antigen-antibody reaction or the like can be increased. As a result, the S / N ratio can be improved.

  According to the present embodiment, the magnetic fine particles 9 can be attracted to the sensing area 101 by applying a magnetic field to the magnetic fine particles 9 by the magnetic field applying unit 11. Thereby, since it becomes easier to couple the magnetic fine particles 9 to the sensing area 101, the detection sensitivity of the measurement target substance 14 can be improved.

  In addition, since the magnetic fine particles 9 and the sample solution are introduced into the reaction space 102 and the magnetic fine particles 9 are quickly drawn in the direction of the sensing area 101, the time for waiting for the natural precipitation of the magnetic fine particles 9 can be shortened. Measurement can be performed in a short time. In addition, the coupling between the magnetic fine particles 9 and the sensing area 101 can be promoted before the reaction or aggregation of the magnetic fine particles 9 proceeds. Thereby, since the utilization factor of the measurement target substance 14 for the binding between the magnetic fine particles 9 and the sensing area 101 can be further increased, higher detection sensitivity can be obtained.

  Furthermore, the specimen solution and the magnetic fine particles 9 can be stirred by moving the magnetic fine particles 9 by both or one of the magnetic field applying unit 10 and the magnetic field applying unit 11. As a result, the antigen-antibody reaction between the measurement target substance 14 (for example, antigen) contained in the sample solution and the magnetic fine particles 9 is promoted, and high detection sensitivity can be measured in a shorter time. Further, the magnetic fine particles 9 can be further stirred by repeatedly applying the upper magnetic field by the magnetic field applying unit 10 and applying the lower magnetic field by the magnetic field applying unit 11. As a result, the opportunity for the magnetic fine particles 9 to bind to the sensing area 101 via the measurement target substance 14 increases, so that the measurement target substance 14 can be detected in a shorter time. In addition, the probability that the magnetic fine particles 9 are bonded to the sensing area 101 can be improved, and the detection sensitivity and measurement accuracy of the measurement target substance 14 can be improved. For example, it is effective when the measurement target substance 14 has a low concentration.

  In the present embodiment, since the magnetic fine particles 9 are stirred using a magnetic field, a manual stirring operation or a stirring mechanism having a pump or the like is not necessary, and a simple measurement and a small measurement system can be realized. For example, if the application of the magnetic field by the control unit 20a is automated, measurement can be performed by only one operation in which the measurer introduces the sample solution into the sensor chip 100.

  Furthermore, if the magnetic fine particles 9 have superparamagnetism that quickly loses magnetization when the application of the magnetic field is stopped, the magnetic fine particles 9 may aggregate due to magnetization when the magnetic field is applied. The dispersion can be re-dispersed by stopping the application of the magnetic field. Even if the magnetic fine particles 9 aggregate when the magnetic field is applied, by stopping the application of the magnetic field before the aggregate of the magnetic fine particles 9 reaches the vicinity of the sensing area 101, the aggregate of the magnetic fine particles 9 is regenerated. Can be dispersed. Therefore, the magnetic fine particles 9 can reach the sensing area 101 in a dispersed state. Accordingly, it is possible to prevent an increase in measurement noise due to aggregation of the magnetic fine particles 9.

  Further, in order to further improve the redispersibility when the application of the magnetic field is stopped, the surface of the fine particles 12 in the magnetic fine particles 9 may be given a positive or negative charge. Alternatively, a dispersant such as a surfactant may be added as a dispersion medium to the surface of the fine particles 12 in the magnetic fine particles 9.

  In addition, according to the present embodiment, the detection sensitivity and measurement accuracy of the measurement target substance 14 can be improved by appropriately controlling the magnetic field strengths of the magnetic field application unit 10 and the magnetic field application unit 11 by the control unit 20a.

  In this embodiment, the same effect as that of the first embodiment can be obtained.

[Third embodiment]
In the first and second embodiments, the case where the optical waveguide is arranged in the natural sedimentation direction as viewed from the magnetic fine particles has been described. However, in the third embodiment, in the direction opposite to the natural sedimentation direction as viewed from the magnetic fine particles. A case in which an optical waveguide is present will be described.

FIG. 8 is a diagram showing a configuration of an optical waveguide type measurement system according to the third embodiment. The optical waveguide measurement system 30b according to the present embodiment uses a cap 15 having an enclosure shape in which liquid does not fall, instead of the frame 5 in the optical waveguide measurement system 30a of the second embodiment shown in FIG. The entire optical waveguide type measurement system 30a of the second embodiment shown in FIG. 6 is turned upside down. That is, in the present embodiment, the magnetic field application unit 10 is disposed below the optical waveguide sensor chip 100 and the magnetic field application unit 11 is disposed above the optical waveguide sensor chip 100. Therefore, in this embodiment, the magnetic field application unit 10 applies the lower magnetic field, and the magnetic field application unit 11 applies the upper magnetic field.
The magnetic field application unit 10 is not always necessary. Since the other configuration is the same as that of the second embodiment, description thereof is omitted.

  The measurement system shown in FIG. 8 includes a cap 15 having a concave cross section instead of the frame 5 in order to hold a mixed dispersion of the sample solution and the magnetic fine particles 9. The cap 15 and the sensing area 101 form a reaction space 102a that becomes a semi-enclosed space except for an opening for introducing liquid and an air vent hole (both not shown).

  Here, the optical waveguide measurement system 30b of the present embodiment may further include a control unit 20b. The control unit 20b controls the strength of the magnetic field applied by the magnetic field application unit 10 and the magnetic field application unit 11, or one of them. In this case, for example, as shown in FIG. 8, independent control units 20 b 1 and 20 b 2 can be provided for the magnetic field application unit 10 and the magnetic field application unit 11, respectively. Further, a common control unit for the magnetic field application unit 10 and the magnetic field application unit 11 and a changeover switch (not shown) may be provided. In addition, a control unit that controls the magnetic field strength at the same time for the magnetic field application unit 10 and the magnetic field application unit 11 may be provided. Alternatively, the control unit 20b may be configured to control the magnetic field strength as needed to dynamically adjust the magnetic field strength to an appropriate value.

  Further, the control unit 20b may control the timing of applying the magnetic field in each of the magnetic field application unit 10 and the magnetic field application unit 11. Thereby, the magnetic field application unit 10 and the magnetic field application unit 11 can alternately apply a magnetic field according to a predetermined condition (for example, a predetermined time or a time during which a predetermined magnetic field is continuously applied).

FIG. 9 is a process diagram showing a method of measuring a substance to be measured in a sample solution.
Here, a method of measuring the amount of the measurement target substance 14 using the optical waveguide type measurement system 30b described above will be described with reference to FIGS.
The state in the reaction space 102a will be described.
Further, the amount and concentration (for example, antigen concentration) of the measurement target substance 14 in the sample solution by measuring the difference in the detection signal intensity ratio in the light receiving element 8 is the same as in the case of the first embodiment. The description is omitted for the same reason.

  First, a measurement system shown in FIG. 8 is prepared. Next, as shown in FIG. 9A, a reaction space 102 a formed by the frame 5 and the sensing area 101 is filled with a mixed dispersion of the sample solution and the magnetic fine particles 9. The method for this is the same as the method described in the first embodiment. In addition, it is desirable that the sample solution or the like is introduced by inflow through an opening (not shown) for introducing the liquid. Here, the sample solution may contain a contaminant 17 that settles under its own weight. Examples of the contaminant 17 include blood cell components in blood. If such a contaminant 17 exists in the vicinity of the sensing area 101, it becomes a scatterer itself and causes measurement noise, or the reaction of the magnetic fine particles 9 binding to the sensing area 101 is hindered. Measurement accuracy may be reduced.

  Next, as shown in FIG. 9B, the magnetic field application unit 11 applies a magnetic field in the direction of the sensing area 101 when viewed from the magnetic fine particles 9. Thereby, the magnetic fine particles 9 are attracted to the sensing area 101. At this time, the first substance 6 (for example, a primary antibody) immobilized on the sensing area 101 and the second substance 13 (for example, a secondary antibody) immobilized on the surface of the fine particles 12 are measured substances 14 ( For example, it binds by antigen-antibody reaction via antigen). Thereby, the magnetic fine particles 9 are coupled to the sensing area 101. At the same time, the sedimentary contaminant 17 moves in the downward direction in FIG. 9B (the direction opposite to the sensing area 101) by its own weight.

  Next, as shown in FIG. 9C, the magnetic field applying unit 10 applies a downward magnetic field shown in FIG. 9C. Then, regardless of the antigen-antibody reaction, the magnetic fine particles 9 adsorbed on the sensing area 101 without passing through the measurement target substance 14 move in the settling direction and are removed from the sensing area 101. Here, even if the application of the magnetic field in the upward direction shown in FIG. 9 (b) is simply stopped using a measurement system that does not have the magnetic field application unit 10, the measurement target substance 14 is not involved regardless of the antigen-antibody reaction or the like. The magnetic fine particles 9 adsorbed on the sensing area 101 can be moved downward by their own weight. However, with this method, it is difficult to remove the magnetic fine particles 9 adsorbed on the sensing area 101 when the attractive force of the magnetic fine particles 9 on the sensing area 101 is superior to the downward force corresponding to its own weight. It becomes. In the step shown in FIG. 9C, the sedimentary contaminant 17 continues to move downward (in the direction opposite to the optical waveguide 3) in FIG. 9C due to its own weight.

  Also in the present embodiment, the magnetic field application in the upward direction shown in FIG. 9B and the magnetic field application in the downward direction shown in FIG. 9C or sedimentation due to the weight of the magnetic fine particles 9 may be alternately repeated. Since the effect is the same as that described in the second embodiment, the description thereof is omitted.

  According to the present embodiment, the sensing area 101 is positioned above the magnetic fine particles 9, and a magnetic field is applied to the magnetic fine particles 9 by the magnetic field application unit 11. Therefore, at the same time as the magnetic fine particles 9 are attracted to the sensing area 101, the sedimentary contaminant 17 can be allowed to settle downward. Thereby, the contaminant 17 can be naturally moved outside the evanescent light region in the vicinity of the sensing area 101. As a result, the measurement accuracy can be further improved without removing the contaminant 17 by filtration or the like in advance.

  In this embodiment, the same effects as those of the first and second embodiments can be obtained.

Next, the magnetic field application unit will be further illustrated.
FIG. 10 is a schematic diagram for illustrating the configuration of the magnetic field application unit.
FIG. 10A is a schematic diagram for illustrating the configuration of the magnetic field application unit, FIG. 10B is a view taken along the line AA in FIG. 10A, and FIG. 10C is a schematic perspective view.
The magnetic field application unit 40 illustrated in FIG. 10 can be used for both the magnetic field application unit 10 and the magnetic field application unit 11 described above.
As shown in FIG. 10, the magnetic field application unit 40 is provided with a coil 41 and a core 42.
The coil 41 has a bobbin 41a and an insulated wire 41b. The bobbin 41a has a cylindrical portion 41a1 around which the insulated wire 41b is wound, and flanges 41a2 provided at both ends of the cylindrical portion 41a1.
Since the bobbin 41a has a simple cylindrical shape, when the insulated wire 41b is wound in the state of the bobbin 41a alone, high-speed winding can be performed. Therefore, it is possible to shorten the time of the winding process. Moreover, since the bobbin 41a around which the insulated wire 41b is wound can be simply assembled by inserting it into the core 42, it is possible to reduce the time of the assembly process.
The bobbin 41a is not always necessary and can be provided as appropriate. For example, the coil 41 can be formed by winding an insulated wire 41 b around the core 42.

The core 42 includes a first core portion 42a provided with the coil 41, a second core portion 42b provided outside the coil 41, one end portion of the first core portion 42a, and a second core portion 42b. A connecting portion 42c for magnetically connecting the one end of the second portion. In the magnetic field application unit 40 illustrated in FIG. 10, two second core portions 42 b are provided at symmetrical positions with the first core portion 42 a interposed therebetween.
In this case, the first core part 42a, the second core part 42b, and the connection part 42c can be separated.
By doing so, it is possible to improve the assemblability when the coil 41 is assembled to the core 42.
Further, as shown in FIG. 10B, the side 42b1 on the gap side (first core part 42a side) of the end surface of the second core part 42b on the sensing area 101 side, or the first core part 42a. A side 42 a 1 on the gap side (second core portion 42 b side) of the end surface on the sensing area 101 side is parallel to the direction in which light propagates inside the optical waveguide 3.
By doing so, since a uniform magnetic field can be applied, the distribution of the magnetic fine particles 9 in the sensing area 101 can be made uniform.
In addition, in the direction in which light propagates inside the optical waveguide 3, the lengths of the end surfaces on the sensing area 101 side of the first core portion 42 a and the second core portion 42 b are equal to or greater than the length of the sensing area 101. Yes.
By doing so, since the magnetic field can be applied to the entire sensing area 101, the magnetic fine particles 9 can be moved in the entire sensing area 101.

The end surface on the sensing area 101 side of the first core portion 42a and the end surface on the sensing area 101 side of the second core portion 42b are flat surfaces.
If the end surface on the sensing area 101 side is a flat surface, the distance between the first core part 42a and the sensing area 101 and the distance between the second core part 42b and the sensing area 101 can be reduced.
Although not shown, the first core portion 42a and the second core portion 42b may have a configuration in which the cross-sectional area decreases as the sensing area 101 is approached.
That is, the 1st core part 42a and the 2nd core part 42b can have a form where the front end side becomes thin.
By doing so, since the magnetic flux can be concentrated, a strong magnetic field can be applied.
Moreover, although illustration is abbreviate | omitted, the 2nd core part 42b can have the form which inclined in the direction in which the edge part by the side of the sensing area 101 adjoins mutually.
By doing so, the generation of the magnetic field can be facilitated.

The core 42 can be formed from a material having a smaller residual magnetization than carbon steel.
For example, the core 42 can be formed from pure iron or the like.
By making the material of the core 42 in this way, it is possible to suppress the influence of the residual magnetic field of the core 42 on the movement of the magnetic fine particles 9 described above when the application of the magnetic field is stopped by the control unit 20 or the like. The core 42 may have a structure in which an insulating coated thin magnetic plate (for example, a silicon steel plate) is laminated in a direction parallel to the magnetic field.
The thin plate-like core 42 can be processed easily and at high speed using press processing. Furthermore, when laminating simultaneously with pressurization, the laminated cores 42 can be connected and fixed by projecting a part of the cores 42 in the thickness direction. Therefore, the number of processing steps can be reduced.
Further, the magnetic flux in the core 42 has anisotropy in the flow direction of the magnetic flux, which is easy to flow in the plane direction and difficult to flow in the thickness direction. Therefore, if the stacking direction is parallel to the direction in which light propagates inside the optical waveguide 3, leakage of magnetic flux outside the sensing area is reduced, so that a desired magnetic field can be generated with less electromagnetic force. As a result, the efficiency of the optical waveguide type measurement system can be improved.
The core 42 may have a structure in which an insulating coated magnetic powder (for example, a fine powder made of a ferromagnetic material such as carbonyl iron) is pressure-molded.
If the structure of the core 42 is as described above, eddy current loss can be reduced.
In FIG. 10, the coil 41 is provided in the first core portion 42a, but the present invention is not limited to this. For example, the coil 41 may be provided in the second core part 42b, or the coil 41 may be provided in the first core part 42a and the second core part 42b. Moreover, you may provide the coil 41 in the connection part 42c.
Further, the coils 41 may be separately provided on the second core part 42b on the right side in FIG. 10 and the second core part 42b on the left side in FIG. At this time, the energization current can be controlled independently so that only the right side coil can be energized, only the left side coil can be energized, and the left and right coils can be energized simultaneously. Good. By doing so, the movement of the magnetic fine particles 9 in the vertical and horizontal directions can be controlled. For this reason, the chance of bringing the magnetic fine particles 9 and the measurement target substance 14 into contact with each other can be increased, so that the detection accuracy can be improved.

Next, a magnetic field application unit according to another embodiment is illustrated.
FIG. 11 is a schematic diagram for illustrating the configuration of the magnetic field application unit.
11A is a schematic diagram for illustrating the configuration of the magnetic field application unit, FIG. 11B is a view taken along the line BB in FIG. 11A, and FIG. 11C is a schematic perspective view.
Note that the magnetic field application unit 50 illustrated in FIG. 11 can be used for both the first magnetic field application unit 10 and the magnetic field application unit 11 described above.
As shown in FIG. 11, the magnetic field application unit 50 is provided with a coil 41 and a core 52.
The coil 41 is the same as that described above, and a description thereof will be omitted.

The core 52 includes two core portions 52a where the coil 41 is provided, and a connection portion 52c that magnetically connects one end portions of the core portion 52a.
In this case, the core portion 52a and the connection portion 52c can be separated.
By doing so, it is possible to improve the assemblability when the coil 41 is assembled to the core 52.
Further, as shown in FIG. 11B, the gap side 52 a 1 on the end surface of the core portion 52 a on the sensing area 101 side is parallel to the direction in which light propagates inside the optical waveguide 3.
By doing so, since a uniform magnetic field can be applied, the distribution of the magnetic fine particles 9 in the sensing area 101 can be made uniform.
In addition, the length of the end surface on the sensing area 101 side of the core portion 52 a is greater than or equal to the length of the sensing area 101 in the direction in which light propagates inside the optical waveguide 3.
By doing so, since the magnetic field can be applied to the entire sensing area 101, the magnetic fine particles 9 can be moved in the entire sensing area 101.

The end surface of the core portion 52a on the sensing area 101 side is a flat surface.
If the end surface on the sensing area 101 side is a flat surface, the distance between the core portion 52a and the sensing area 101 can be reduced.
Although not shown, the core portion 52a may have a configuration in which the cross-sectional area decreases as the sensing area 101 is approached.
That is, the core part 52a can have a form in which the tip end side is narrowed. By doing so, since the magnetic flux can be concentrated, a strong magnetic field can be applied.
Moreover, the core part 52a shall have the form which inclined in the direction in which the edge part by the side of the sensing area 101 adjoins mutually.
By doing so, the generation of the magnetic field can be facilitated.
In addition, a non-magnetic spacer (not shown) such as resin or copper can be provided in the gap between the ends on the sensing area 101 side. By doing so, it becomes easy to manage the size of the gap. Further, when a magnetic field is applied to the core 52, a magnetic attractive force acts in a direction in which the gap becomes smaller. However, by providing a spacer, the gap size can be maintained. Therefore, it is possible to suppress a change in the magnetic flux distribution in the sensing area.

The material and configuration of the core 52 can be the same as those of the core 42 described above.
For example, the core 52 can be formed of a material having a smaller residual magnetization than carbon steel such as pure iron.
If the material of the core 52 is made in this way, the influence of the residual magnetic field of the core 52 on the movement of the magnetic fine particles 9 described above when the application of the magnetic field is stopped by the control unit 20 or the like can be suppressed. In FIG. 11, the coils 41 are provided in the two core portions 52a, but the present invention is not limited to this. For example, the coil 41 may be provided in one core part 52a, or the coil 41 may be provided in the connection part 52c.
In this case, if the coils 41 are provided in the two core portions 52a, the height of the magnetic field application unit 50 can be kept low. Further, if one coil 41 is provided in one core portion 52c, the number of coils 41 can be reduced, so that the number of winding steps and the number of assembly steps can be reduced in addition to the reduction in the number of components.

Next, the operation and effect of the magnetic field application unit will be further illustrated.
FIG. 12 is a graph for illustrating the operation and effect of the magnetic field application unit.
FIG. 12 is a graph for illustrating the operation and effect of the magnetic field application unit 10 in the optical waveguide measurement system 30 shown in FIG. 1 as an example.
FIG. 12A shows a step of removing magnetic fine particles 9 that may become noise by the magnetic field application unit 10, and FIG. 12B agitates the dispersion of the magnetic fine particles 9 and the sample solution by the magnetic field application unit 10. In this case, the method further includes the step of:
As shown in FIG. 12A, when the dispersion of magnetic fine particles 9 and the sample solution are mixed and introduced into the sensing area 101, the detection signal intensity is increased in accordance with the increase in fine particle density in the vicinity of the sensing area 101 due to sedimentation of the magnetic fine particles 9. The ratio decreases. After that, when the upper magnetic field is applied by the magnetic field application unit 10, the magnetic fine particles 9 that can become noise adsorbed to the sensing area 101 are removed, so that the detection signal intensity ratio rises again and is lower than the initial detection signal intensity ratio. Saturates.

On the other hand, as shown in FIG. 12 (b), after the dispersion of the magnetic fine particles 9 and the sample solution are mixed and introduced into the sensing area 101, the upper magnetic field is applied in a pulse form by the magnetic field application unit 10. For example, the upper magnetic field is applied every 10 seconds by the magnetic field application unit 10. Thereafter, when the magnetic fine particles 9 are allowed to settle, the detection signal intensity ratio decreases as the fine particle density in the vicinity of the sensing area 101 increases. Next, when the upper magnetic field is applied by the magnetic field application unit 10, the magnetic fine particles 9 that can become noise adsorbed to the sensing area 101 are removed, so that the detection signal intensity ratio rises again and is lower than the initial detection signal intensity ratio. Saturates at.
Here, in the case shown in FIG. 12B, the detection signal intensity ratio after removing the magnetic fine particles 9 that may be noise is lower than in the case of FIG. That is, in the case of FIG. 12B, the amount of the magnetic fine particles 9 bonded to the sensing area 101 via the measurement target substance 14 is increased.

When the upper magnetic field is applied in a pulse form by the magnetic field application unit 10, the magnetic fine particles 9 and the sample solution can be stirred. Therefore, the reaction rate between the second substance 13 immobilized on the fine particles 12 of the magnetic fine particles 9 and the measurement target substance 14 is increased, and the amount of the magnetic fine particles 9 coupled to the sensing area 101 via the measurement target substance 14 is increased. This is thought to be due to the increase.
If the reaction rate between the second substance 13 immobilized on the fine particles 12 of the magnetic fine particles 9 and the measurement target substance 14 can be increased, the detection sensitivity of the measurement target substance 14 can be improved with higher accuracy.
In addition, although the case where the upper magnetic field was applied was illustrated as an example, it is not necessarily limited to this. The present invention can be applied to magnetic field application for moving the magnetic fine particles 9 away from the sensing area 101. For example, in the case shown in FIG. 8, the lower magnetic field may be applied in a pulse shape.
Further, the upper magnetic field and the lower magnetic field may be applied in a pulse shape by shifting the application timing. By doing so, the magnetic fine particles 9 and the sample solution can be further agitated.

FIG. 13 is a graph for illustrating the operation and effect of the magnetic field application unit.
FIG. 13 is a graph for illustrating the operation and effect of the magnetic field application units 10 and 11 in the optical waveguide measurement system 30a shown in FIG. 6 as an example.
13A shows the case where only the upper magnetic field is applied by the magnetic field application unit 10, and FIGS. 13B and 13C show the application of the lower magnetic field by the magnetic field application unit 11 and the upper magnetic field by the magnetic field application unit 10. Is applied in combination with the application of.
In the case shown in FIG. 13A, after the dispersion of magnetic fine particles 9 and the sample solution are mixed and introduced into the reaction space 102, the magnetic fine particles 9 are allowed to naturally settle. Then, after the detection signal intensity ratio is reduced to a predetermined value, an upper magnetic field is applied to remove magnetic fine particles 9 that may become noise adsorbed to the sensing area 101.

In the case shown in FIGS. 13B and 13C, the dispersion of the magnetic fine particles 9 and the sample solution are mixed and introduced into the reaction space 102, and then the lower magnetic field is applied to place the magnetic fine particles 9 in the sensing area 101. Move it closer to Thereafter, the application of the lower magnetic field is stopped, and the magnetic fine particles 9 are allowed to settle naturally. Then, after the detection signal intensity ratio is reduced to a predetermined value, an upper magnetic field is applied to remove magnetic fine particles 9 that may become noise adsorbed to the sensing area 101.
As can be seen from FIGS. 13A to 13C, when the lower magnetic field is applied and the magnetic fine particles 9 are moved in the direction approaching the sensing area 101, the upper magnetic field can be applied. Can be shortened. Therefore, the measurement time can be shortened.
FIG. 14 is a process diagram showing a method for measuring a substance to be measured in a sample solution.
14A to 14D illustrate a method of measuring the concentration of the measurement target substance 14 using the measurement system 30a.
14A to 14D, the state in the reaction space 102 is represented in order to avoid complication.
14 (a) is the same as FIG. 7 (a), FIG. 14 (c) is the same as FIG. 7 (b), and FIG. 14 (d) is the same as FIG. 7 (c). ). Therefore, the description regarding FIG. 14 (a), (c), (d) is abbreviate | omitted.
FIG. 14B-1 shows a case where the lower magnetic field application by the magnetic field application unit 11 is not performed. Therefore, the magnetic fine particles 9 in the sample solution are settled (natural sedimentation) toward the sensing area 101 by gravity.
14B-2, the magnetic field application unit 11 applies the lower magnetic field in the settling direction (the direction of the optical waveguide 3, for example, the lower direction in FIG. 14) when viewed from the magnetic fine particles 9. Thereby, the magnetic fine particles 9 are attracted to the sensing area 101 by natural sedimentation due to gravity and attraction by applying a lower magnetic field. However, as shown in FIG. 14B-2, in this state, most of the magnetic fine particles 9 are retained along the lines of magnetic force, so that the binding reaction with the first substance 6 does not proceed. Therefore, as shown in FIG. 14 (c), it is necessary to make the external magnetic field zero once and to proceed the reaction by natural diffusion.
14B and 14B, a part of the magnetic fine particles 9 that have settled toward the sensing area 101 is coupled to the sensing area 101.

As an example, the case of the optical waveguide type measurement system 30a shown in FIG. 6 is illustrated, but the present invention is not limited to this. For example, in the case of the optical waveguide type measurement system 30b shown in FIG. 8, the magnetic fine particles 9 that can become noise adsorbed on the sensing area 101 by applying the lower magnetic field by the magnetic field applying unit 10 and compared with the case of natural sedimentation. Can be removed in a short time. Therefore, the measurement time can be shortened.
The application of the magnetic field illustrated in FIGS. 12 and 13 can be controlled by the control units 20, 20a, and 20b described above.

  As mentioned above, although several embodiment of this invention was illustrated, these embodiment is shown as an example and is not intending limiting the range of invention. These novel embodiments can be implemented in various other forms, and various omissions, replacements, changes, and the like can be made without departing from the spirit of the invention. These embodiments and modifications thereof are included in the scope and gist of the invention, and are included in the invention described in the claims and equivalents thereof. Further, the above-described embodiments can be implemented in combination with each other.

1: substrate, 2a: grating, 2b: grating, 3: optical waveguide, 4: protective film, 5: frame, 6: first material, 7: light source, 8: light receiving element, 9: magnetic fine particles, 10: magnetic field application Part: 11: magnetic field application part, 12: fine particles, 13: second substance, 14: substance to be measured, 15: cap, 17: contaminant substance, 20: control part, 20a: control part, 20b: control part, 20b1: Control unit, 20b2: control unit, 30: optical waveguide type measurement system, 30a: optical waveguide type measurement system, 30b: optical waveguide type measurement system, 40: magnetization applying unit, 41: coil, 42: core, 42a: first Core part, 42b: second core part, 42c: connection part, 50: magnetization application part, 52: core, 52a: core part, 52c: connection part, 100: optical waveguide sensor chip, 101: sensing area, 102 Reaction space, 102a: reaction space

Claims (25)

An optical waveguide having a sensing area in which a first substance that specifically binds to a measurement target substance is immobilized;
A second substance that specifically binds to the substance to be measured is immobilized, has a particle size of 0.2 μm or more and 20 μm or less, is provided to cover the first core and the first core, and is magnetic A magnetic fine particle comprising a nanoparticle and a shell having a refractive index higher than that of the first core;
A magnetic field application unit for generating a magnetic field for moving the magnetic fine particles;
A light source for making light incident on the optical waveguide;
A light receiving element for receiving light emitted from the optical waveguide;
An optical waveguide type measurement system comprising: The magnetic field application unit has a first magnetic field application unit,
2. The optical waveguide measurement system according to claim 1, wherein the first magnetic field application unit applies a magnetic field that moves the magnetic fine particles in a direction away from the optical waveguide. 3. The optical waveguide type measurement system according to claim 2, wherein the first magnetic field application unit applies a magnetic field having a magnetic field intensity such that the magnetic fine particles are separated from the sensing area by a distance satisfying the following expression.
L> λ / {2π (n ₁ ² sin ² θ−n ₂ ² ) ^1/2 }
Here, L is the distance that the magnetic fine particles are separated from the sensing area, λ is the wavelength of light used for measurement, n ₁ is the refractive index of the optical waveguide, n ₂ is the refractive index of the dispersion medium that disperses the magnetic fine particles, θ is the total reflection angle. The magnetic field application unit has a second magnetic field application unit,
4. The optical waveguide type measurement system according to claim 1, wherein the second magnetic field application unit applies a magnetic field that moves the magnetic fine particles in a direction approaching the optical waveguide. 5.   The optical waveguide type measurement system according to claim 4, wherein the first magnetic field application unit and the second magnetic field application unit alternately apply a magnetic field.   The optical waveguide measurement system according to claim 1, wherein the magnetic field application unit includes an electromagnet.   The optical waveguide type measurement according to any one of claims 1 to 6, further comprising a first control unit that controls at least one of a timing and a length of time for applying a magnetic field in the magnetic field application unit. system.   The optical waveguide type measurement system according to any one of claims 1 to 7, further comprising a second control unit that controls a magnetic field strength of a magnetic field applied by the magnetic field application unit.   The optical waveguide measurement system according to claim 1, wherein the magnetic nanoparticle has superparamagnetism. The magnetic fine particles, an optical waveguide type measuring system according to any one of claims 1 to 9 having a positive or negative charge. The optical waveguide type measurement system according to any one of claims 1 to 10 , wherein a surfactant is added to the magnetic fine particles.   6. The optical waveguide measurement system according to claim 1, wherein the magnetic field application unit includes a second core and a coil provided in the second core. 7. The second core includes a plurality of core portions, and a connection portion that magnetically connects ends of the plurality of core portions opposite to the sensing area side ends,
The optical waveguide type measurement system according to claim 12 , wherein a gap is provided between end faces on the sensing area side of the plurality of core portions. The optical waveguide type measurement system according to claim 13 , wherein the side on the gap side of the end surface on the sensing area side is parallel to a direction in which light propagates inside the optical waveguide. The optical waveguide type according to claim 13 or 14 , wherein, in a direction in which light propagates inside the optical waveguide, a length of an end surface on the sensing area side of the plurality of core portions is not less than a length of the sensing area. Measuring system. The optical waveguide type measurement system according to any one of claims 13 to 15 , wherein end portions of the plurality of core portions on the sensing area side are inclined in directions close to each other. The optical waveguide measurement system according to any one of claims 13 to 16 , wherein the plurality of core portions have a form in which a cross-sectional area decreases as the sensing area is approached. The optical waveguide measurement system according to any one of claims 13 to 17 , wherein end surfaces of the plurality of core portions on the sensing area side are flat surfaces. The second core structure and the plate having magnetism and an insulating coating is laminated on the magnetic field in a direction parallel, or claims 12 to claim 18 having a structure in which the powder was pressure-molded with magnetic insulated cover The optical waveguide type measurement system according to any one of the above. The optical waveguide measurement system according to any one of claims 13 to 19 , wherein the plurality of core portions and the connection portion are separable. The optical waveguide measurement system according to any one of claims 12 to 20 , wherein the second core includes a material having a remanent magnetization smaller than that of carbon steel. The optical waveguide type measurement system according to any one of claims 7 to 21 , wherein the first control unit controls the magnetic field application unit to apply a magnetic field in a pulse shape. The first control unit, after introducing the magnetic microparticles and the sample solution into the sensing area, controls the magnetic field application unit to apply a magnetic field that moves the magnetic microparticles in a direction approaching the optical waveguide. The optical waveguide type measurement system according to any one of claims 7 to 22 . An optical waveguide having a sensing area in which a first substance that specifically binds to a measurement target substance is immobilized;
A second substance that specifically binds to the substance to be measured is immobilized, has a particle size of 0.2 μm or more and 20 μm or less, is provided to cover the first core and the first core, and is magnetic An optical waveguide sensor chip comprising: magnetic fine particles including a nano particle and a shell having a refractive index higher than that of the first core. The sample solution containing the measurement target substance and the second substance that specifically binds to the measurement target substance are immobilized, the particle size is 0.2 μm or more and 20 μm or less, the first core, and the first core A magnetic particle having a magnetic nanoparticle and a shell having a refractive index higher than the refractive index of the first core provided on the optical waveguide sensor chip. A step of bringing the first substance that specifically binds to the target substance into contact with the immobilized sensing area;
Applying a magnetic field to the optical waveguide sensor chip;
Measuring the light intensity of light emitted from the optical waveguide sensor chip as a first light intensity before application of the magnetic field;
Measuring the light intensity of light emitted from the optical waveguide sensor chip as the second light intensity after application of the magnetic field;
Measuring the substance to be measured based on a difference between the first light intensity and the second light intensity.

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