WO2014168043A1 - Measuring device and measuring method - Google Patents

Abstract

In one example of an embodiment, a measuring device forms solution particles with a diameter of 20 μm or less from a solution obtained by mixing a fluorescent substance that is specifically adsorbed to a substance to be detected and a sample. In the example of the embodiment, the measuring device irradiates the formed particles with light. In the example of the embodiment, the measuring device measures the fluorescence intensity of the particles irradiated with the light. Further, in the example of the embodiment, the measuring device forms the particles using at least one of an electrospray, a two fluid nozzle, a nebulizer, a piezoelectric element, an ultrasonic wave, and decompression treatment.

Description

Measuring apparatus and measuring method

Various aspects and embodiments of the present invention relate to a measuring apparatus and a measuring method.

Conventionally, there is a detection method for detecting a detection target substance as a detection target. As a detection method, for example, there is a method using a fluorescent substance that specifically adsorbs to a detection target substance to be detected. In the method using a fluorescent substance, for example, the virus in the gas is diffused into the drug solution by bringing the drug solution containing the fluorescent antibody that specifically adsorbs to the virus into contact with the gas to be inspected, and the virus in the gas is diffused. The fluorescence intensity of the mist group of the chemical solution is measured, and the virus is detected according to the fluorescence intensity.

International Publication No. 2012/056641

Here, it is required to improve the detection accuracy of the detection target substance.

In one embodiment, the disclosed measuring method includes a particle forming unit that forms particles of the solution having a diameter of 20 μm or less from a solution in which a fluorescent substance that specifically adsorbs to a detection target substance and a sample are mixed. And a fluorescence measuring unit that irradiates the particles formed by the particle forming unit with light and measures the fluorescence intensity of the particles irradiated with the light.

According to one aspect of the disclosed measurement method, the detection accuracy of the detection target substance can be improved.

FIG. 1 is a diagram illustrating an example of a configuration of a measurement apparatus according to the first embodiment. FIG. 2 is a longitudinal sectional side view showing a part of the microfluidic chip used in the first embodiment. FIG. 3 is a diagram showing noise due to an unreacted fluorescent material. FIG. 4 is a diagram showing the difference in fluorescence intensity when the particle size is small. FIG. 5 is a longitudinal sectional side view showing the fluorescence measurement unit used in the first embodiment. FIG. 6 is a diagram showing the diffusion of the virus in the chemical solution, the adsorption of the fluorescent antibody, and the particle formation from the solution in the diffusion unit of the first embodiment. FIG. 7 is a diagram showing diffusion of atmospheric dust in the chemical solution, adsorption of fluorescent antibodies, and particle formation from the solution in the diffusion unit of the first embodiment. FIG. 8 is a flowchart illustrating an example of a processing flow in the first embodiment. FIG. 9 is a diagram illustrating an example of a measurement apparatus that does not have a microfluidic chip. FIG. 10 is a diagram showing an example of a measuring apparatus provided with a chemical solution supply pump in the middle of the chemical solution supply pipe. FIG. 11 is a diagram illustrating an example of a measurement apparatus that uses an aeration tank as a diffusion unit. FIG. 12 is a diagram illustrating an example of the particle forming unit and the fluorescence measuring unit. FIG. 13 is a diagram illustrating an example of a configuration of a measurement apparatus when particles are formed using electrospray. FIG. 14 is a diagram illustrating an example of the configuration of a measurement apparatus in the case where particles are formed using a lectrospray. FIG. 15 is a diagram illustrating an example of the configuration of the measurement apparatus according to the second embodiment.

Hereinafter, embodiments of the disclosed measuring apparatus and measuring method will be described in detail based on the drawings. The invention disclosed by this embodiment is not limited. Each embodiment can be appropriately combined as long as the processing contents do not contradict each other.

(First embodiment)
In one embodiment, the measuring apparatus according to the first embodiment is a particle that forms particles of a solution having a diameter of 20 μm or less from a solution in which a fluorescent substance that specifically adsorbs to a detection target substance and a sample are mixed. A forming unit, and a fluorescence measuring unit that irradiates the particles formed by the particle forming unit with light and measures the fluorescence intensity of the particles irradiated with the light.

In the measurement apparatus according to the first embodiment, in one embodiment, the particle forming unit is at least one of electrospray, a two-fluid nozzle, a nebulizer, a piezoelectric element (registered trademark), an ultrasonic wave, and a decompression process. To form particles.

In the measurement apparatus according to the first embodiment, in one embodiment, the particle formation unit further includes a particle guide path that guides the particle to a position where light is irradiated, and the fluorescence measurement unit includes the particle guide. Light is irradiated to particles guided through the road.

In addition, the measurement method in the first embodiment includes a particle forming step of forming particles having a diameter of 20 μm or less from a solution in which a fluorescent substance that specifically adsorbs to a detection target substance and a sample are mixed, and light is applied to the particles. And a fluorescence measurement step of measuring the fluorescence intensity of the particles irradiated with light.

Further, in the measurement method according to the first embodiment, the particle forming step forms particles using at least one of electrospray, two-fluid nozzle, nebulizer, piezoelectric element, ultrasonic wave, and reduced pressure treatment.

In the measurement method according to the first embodiment, the particle formation step further includes a guide step for guiding the particles to a position where light is irradiated via the particle guide path, and the irradiation step is performed via the guide path. The guided particles are irradiated with light.

In addition, in one embodiment, the measuring apparatus according to the first embodiment supplies a solution in which a fluorescent substance that specifically adsorbs to a detection target substance and a sample are mixed, and forms a particle of the solution. And an electrode installed so as to face the emission part, and a fluorescence measurement part that measures the fluorescence intensity of the particle by irradiating the particle with light, and the light irradiation is performed between the emission part and the electrode. Is called.

In addition, in one embodiment, the measurement apparatus according to the first embodiment further includes a laminar flow forming mechanism that forms a laminar flow that flows from the discharge portion to the electrode.

(Configuration of the measuring apparatus according to the first embodiment)
FIG. 1 is a diagram illustrating an example of a configuration of a measurement apparatus according to the first embodiment. In the example illustrated in FIG. 1, a case where the particle forming unit 4 of the measurement apparatus 100 according to the first embodiment forms particles using a two-fluid nozzle will be described as an example. However, it is not limited to this. For example, the particle forming unit 4 may form particles of 20 μm or less using at least one of electrospray, a two-fluid nozzle, a nebulizer, a piezoelectric element, ultrasonic waves, and a reduced pressure treatment. The particle forming unit 4 preferably forms particles of 20 μm or less using electrospray or a two-fluid nozzle. By forming the particles using an electrospray or a two-fluid nozzle, the particles can be formed even if the amount of the solution is small. In addition, the particle | grains of 20 micrometers or less do not need the diameter of all the produced | generated particles to be 20 micrometers or less. For example, 50% or more of particles having a size of 20 μm or less may be included among the formed particles.

Further, in the example illustrated in FIG. 1, a case where a sample to be measured by the measurement apparatus 100 is included in the atmosphere will be described as an example. In other words, the measuring apparatus 100 forms a particle from the solution after the chemical solution and the atmosphere come into contact with each other after the chemical solution containing the fluorescent substance that specifically adsorbs to the detection target substance is brought into contact with the atmosphere. A case where the fluorescence intensity is measured will be described as an example. However, the present invention is not limited to this, and the specimen may be a liquid or a solid. In this case, the specimen is mixed in advance with a chemical solution containing a fluorescent substance that specifically adsorbs to the detection target substance, and the measuring apparatus 100 forms particles from the solution in which the specimen is mixed, and measures the fluorescence intensity of the particles. May be. Further, in the case described below, a case where the detection target substance is detected based on the fluorescence intensity of the particles will be described as an example, but the present invention is not limited to this. For example, the measuring apparatus 100 may be limited to measuring the fluorescence intensity. In this case, the user or another device determines whether or not the detection target substance is contained in the sample based on the fluorescence intensity measured by the measurement device 100. The detection target substance is, for example, a virus, a bacterium, or a cell. However, the detection target substance is not limited to viruses, bacteria, and cells. For example, it can also be used to detect pollen and other allergenic substances, any food that you do not want to ingest, antibodies to specific diseases, trace proteins such as cytokines and hormones, metabolite biomarkers such as serotonin, and other harmful substances can do.

In addition, when providing the solution with which the sample was mixed beforehand with respect to the measuring apparatus 100, after omitting the sample solution provision part 200 which is the part enclosed with the dotted line among the measuring apparatuses 100 shown in FIG. The solution may be supplied as it is from the solution supply path 40 to the particle forming unit 4.

Returning to the explanation of FIG. In the example shown in FIG. 1, the measuring apparatus 100 includes a dust removing unit 1, a main pipe 8, a particle forming unit 4, a fluorescence measuring unit 5, a chemical solution collecting unit 6, a suction pump 7, and a sample solution providing unit. 200.

A brief description of the positional relationship of each part. The main pipe 8 is an airflow guideway. The dust removing unit 1 is disposed on the upstream side of the airflow guided by the main pipe 8. The suction pump 7 forms an air flow inside the main pipe 8 and is disposed on the downstream side of the air flow guided by the main pipe 8. In other words, the suction pump 7 forms an airflow that flows from the dust removing unit 1 to the suction pump 7 in the main pipe 8.

The main pipe 8 includes a particle forming unit 4, a fluorescence measuring unit 5, and a chemical solution collecting unit 6 between the dust removing unit 1 and the suction pump 7. The particle forming unit 4, the fluorescence measuring unit 5, and the chemical solution collecting unit 6 are provided in the order of the particle forming unit 4, the fluorescence measuring unit 5, and the chemical solution collecting unit 6 from the upstream side.

Further, the sample solution providing unit 200 provides the solution to the particle forming unit 4. In other words, the sample solution providing unit 200 provides the particle forming unit 4 with a solution in which a specimen to be measured by the measuring apparatus 100 is mixed.

1 will be further described. The dust removal unit 1 has an airflow resistance necessary to allow the virus V to pass therethrough and form particles in the main pipe 8. The dust removing unit 1 captures relatively large particles.

The sample solution providing unit 200 will be described. As shown in FIG. 1, the sample solution providing unit 200 includes a chemical solution storage tank 2, a microfluidic chip 3, an intake pump 11, an air flow rate adjusting unit 12, a branch pipe 13, a chemical solution supply pump 21, and a chemical solution. The flow rate adjusting unit 22, the pipe 23, and the solution supply path 40 are included.

Here, as shown in FIGS. 1 and 2, the microfluidic chip 3 includes a raised portion 30, a diffusion channel 31, a lid 32, a plate-like substrate 33, a gas channel 34, and a liquid flow. The passage 35 has an air inlet 36, a chemical liquid inlet 37, an exhaust port 38, and a chemical liquid outlet port 39. FIG. 2 is a longitudinal sectional side view showing a part of the microfluidic chip used in the first embodiment.

As shown in FIG. 2, a diffusion channel 31 is formed on the upper surface of the substrate 33. The diffusion channel 31 is covered with the lid 32 and functions as the diffusion channel 31. As shown in FIG. 2, the diffusion channel 31 has a cross-sectional shape in which two semicircular lower chords arranged side by side overlap each other. As a result, the diffusion channel 31 is formed with a raised portion 30 at the center of the channel. The diffusion channel 31 is divided into a gas channel 34 and a liquid channel 35 by the raised portion 30. The dimensions of the diffusion channel 31 are, for example, a channel width W of 1 mm or less, a channel depth H of 0.5 mm, and a height of a gap between the raised portion 30 and the lid 32 of 0.2 mm. However, it is not limited to this, and may be any value. As shown in FIG. 1, the diffusion flow path 31 meanders in order to increase the contact time and contact area between the atmosphere and the chemical solution. Both ends of the diffusion channel 31 are bifurcated at the branching portion and reach the end of the microfluidic chip 3 as it is. One end side (upstream end) of the diffusion flow path 31 corresponds to the air inlet 36 and the chemical liquid inlet 37, and the other end side (downstream end) corresponds to the exhaust port 38 and the chemical liquid outlet port 39.

As shown in FIG. 1, the branch pipe 13 branched from the main pipe 8 between the dust removing section 1 and the particle forming section 4 is airtightly connected to the air inlet 36 of the microfluidic chip 3. In the branch pipe 13, an intake pump 11 and an air flow rate adjusting unit 12 which are gas introduction mechanisms are interposed in this order from the upstream side. The chemical solution inlet 37 is connected to a pipe 23 from the chemical solution storage tank 2 in which a chemical solution that is an aqueous solution containing the fluorescent antibody F is stored. In the pipe 23, a chemical supply pump 21 and a chemical flow rate adjusting unit 22 which are liquid introduction mechanisms are interposed in this order from the chemical storage tank 2 side. The exhaust port 38 of the microfluidic chip 3 is connected to the outside of the measuring apparatus 100. The exhaust port 38 exhausts the air flowing into the microfluidic chip 3. The chemical solution outflow port 39 of the microfluidic chip 3 is connected to the particle forming unit 4 via a solution supply path 40 that is a guide path. The solution supply path 40 sends the solution to the particle forming unit 4.

The particle forming unit 4 will be described. The particle forming unit 4 forms particles of a solution having a diameter of 20 μm or less from a solution in which a fluorescent substance that specifically adsorbs to a detection target substance and a specimen are mixed. The particle forming unit 4 in the first embodiment forms solution particles having a diameter of 20 μm or less using a two-fluid nozzle.

In the example shown in FIG. 1, the particle forming unit 4 includes a narrowed portion 81 in which the diameter of the main pipe 8 is sharply narrowed, and a particle guide path 82 for guiding particles from the portion 81 to the fluorescence measuring unit 5. Further, the portion 81 is connected to the solution supply path 40. In the particle forming unit 4, particles of 20 μm or less are formed by mixing the airflow flowing from the dust removing unit 1 to the suction pump 7 and the solution supplied by the solution supply path 40. In other words, in the particle formation part 4, the part 81 becomes a two-fluid nozzle and forms particles of 20 μm or less.

Here, the speed of the airflow flowing from the dust removing unit 1 to the suction pump 7 and the amount and speed of the solution supplied by the solution supply path 40 are such that the diameter of the particles supplied to the fluorescence measuring unit 5 is 20 μm or less. Set to a value.

The point that the particle forming unit 4 forms particles of 20 μm or less will be described. In a method using a fluorescent substance that specifically adsorbs to a detection target substance to be detected, the unreacted fluorescent substance becomes noise. FIG. 3 is a diagram showing noise due to an unreacted fluorescent material. As shown in the particle 301 in FIG. 3, the fluorescent substance emits fluorescence even if it is not adsorbed to the detection target substance to be detected. In addition, when the fluorescent substance specifically adsorbs the detection target substance to be detected and the fluorescent substance, the fluorescence intensity is higher than that of the fluorescent substance not adsorbed to the detection target substance to be detected. In the example shown in FIG. 3, the color intensity indicates the intensity of the fluorescence intensity. In the example illustrated in FIG. 3, the case where the particle 301 does not include the fluorescent substance adsorbed on the detection target substance to be detected is illustrated as an example. In addition, the particle 302 includes a fluorescent material in which the portion 303 which is a part of the particle 302 is not adsorbed to the detection target material, and the part 304 which is a part of the particle 302 is the detection target material. An example is shown in which a fluorescent substance adsorbed on is contained. In this case, the difference between the fluorescence intensity of the particle 301 and the fluorescence intensity of the particle 302 may not be distinguished due to the fluorescence of the fluorescent substance that is not adsorbed to the detection target substance that is the detection target. In other words, the fluorescence intensity is equivalent, and it may be difficult to measure the difference in fluorescence intensity. In this case, the detection target substance that is the detection target cannot be detected. The particles may include not only a fluorescent substance and a detection target substance to be detected, but also a solvent component in which the fluorescent substance and the specimen are mixed.

Here, according to the first embodiment, even if the particle forming unit 4 does not remove the unreacted fluorescent substance by forming particles that are 20 μm or less, the detection target substance that is a detection target Can be measured with high accuracy. Further, as a result of the improvement in measurement sensitivity, the detection accuracy of the detection target substance can be improved. In addition, the detection target substance can be detected in real time. For example, viruses and bacteria can be detected in real time with high accuracy.

In the prior art, there is a problem that the fluorescence derived from the unreacted fluorescent material may be measured, and the measurement accuracy may be poor. Here, a method of measuring the fluorescence intensity after separating the unreacted fluorescent material can be considered, but it takes time and is considered difficult to measure continuously. On the other hand, according to the first embodiment, it is possible to easily and continuously measure without separating the unreacted fluorescent material.

That is, as a rule of thumb, in fluorescence correlation spectroscopy, the volume of the substance to be measured can be reduced to 7.2 femtoliters (fL) by focusing the laser. Here, in the fluorescence correlation spectroscopy, when the volume of the substance to be measured is reduced to 7.2 femtoliters, even if the unreacted fluorescent substance is not removed, the measurement of the detection target substance to be detected is performed. Was possible.

Based on this, the particle forming unit 4 sets the particle diameter measured by the fluorescence measuring unit 5 to 20 μm or less, which is a diameter corresponding to 7.2 femtoliters. As a result, according to the first embodiment, the measuring apparatus 100 can measure the detection target substance to be detected with high accuracy even if the fluorescent substance in the reaction is not removed.

FIG. 4 is a diagram showing the difference in fluorescence intensity when the particle size is small. As shown in the particle 311 and the particle 312 in FIG. 4, by reducing the diameter of the particle, the fluorescence intensity by the particle formed of the fluorescent material that is not adsorbed to the detection target material to be detected, and the detection target The difference from the fluorescence intensity due to the particles formed of the fluorescent substance adsorbed on the detection target substance becomes large, and the detection target substance to be detected can be detected with high accuracy. In the example shown in FIG. 4, the particle 311 does not include a fluorescent material that is not adsorbed on the detection target substance to be detected, and the particle 312 is a fluorescent substance that is adsorbed on the detection target substance that is the detection target. The case where it included in the part 313 was shown as an example. In the example of the particle 312, the portion 314 includes a fluorescent substance that is not adsorbed to the detection target substance to be detected.

Further, it is difficult to make the laser diameter thinner, and in the fluorescence correlation spectroscopy, it is difficult to make the volume of the substance to be measured smaller than 7.2 femtoliters (fL). Similarly, it is difficult to reduce the diameter of the light emitted from the light emitting unit 51 in the fluorescence measuring unit 5. In other words, there is a limit to reducing the amount of particles measured at a time by reducing the diameter of the laser.

On the other hand, according to the first embodiment, the volume of the substance measured at a time can be reduced by reducing the diameter of the particles formed by the particle forming unit 4 without reducing the diameter of the laser. The measurement sensitivity can be improved.

In addition, according to the first embodiment, the particle forming unit 4 has the particle guide path 82 that guides the particle to a position where light is irradiated by the light emitting unit 51 of the fluorescence measuring unit 5. Here, when the solvent component is volatilized from the particles when passing through the particle guide path 82, the particle diameter can be further reduced. As a result, by further using the particle guide path 82, the volume of the substance measured at a time can be further reduced as compared with a method not using the particle guide path 82, and the measurement sensitivity can be improved. Further, by adjusting the length of the particle guide path 82, the size of the diameter can be easily adjusted.

FIG. 5 is a longitudinal sectional side view showing the fluorescence measuring unit used in the first embodiment. A fluorescence measuring unit 5 that is a measuring unit is provided on the downstream side of the particle forming unit 4. As shown in FIG. 5, the fluorescence measurement unit 5 includes, for example, a rectangular case body 56 that is connected to the particle guide path 82 and forms a flow space for airflow including particles formed by the particle formation unit 4. . Light transmitting windows 52a and 52b made of quartz that are parallel to each other are disposed on the upper and lower (or left and right) surfaces of the case body 56 that face each other. Outside the one light transmission window 52a, a light emitting unit 51 that irradiates the case body 56 with laser light having a wavelength deviated from the wavelength of fluorescence emitted from the fluorescent antibody F is provided. Outside the other light transmission window 52b, an optical filter 53 that blocks light having a wavelength deviating from the wavelength of fluorescence emitted from the fluorescent antibody F is provided. Further outside, a light receiving unit 54 that receives the fluorescence of the fluorescent antibody F and converts it into an electrical signal is provided. The light receiving unit 54 outputs, for example, a current having a signal level corresponding to the intensity of light received from the optical filter 53 to the light receiving output measuring unit 55. For example, the light reception output measuring unit 55 converts current into voltage, compares the voltage signal Ia indicating the converted voltage with a preset threshold Is, and the voltage signal Ia is larger than the threshold Is. When it is determined, a virus detection alarm is notified or displayed on a display unit (not shown).

Since the voltage signal Ia is a signal corresponding to the received light intensity, the threshold value Is is determined as follows. That is, the threshold value Is is the fluorescence intensity when the virus V is not present in the atmosphere, and the particle forming unit 4 in a state where the virus V is contained in the atmosphere and the fluorescent antibody F is adsorbed to the virus V. Is set to a value between the fluorescence intensity when the particles formed by the above pass through the case body 56. The fluorescence intensity when the virus V is not present in the atmosphere is the fluorescent antibody F attached to the dust D contained in the atmosphere passing through the inside of the case body 56 or the fluorescent antibody contained in the particles formed by the particle forming unit 4 of the chemical liquid. Corresponds to the intensity of fluorescence from F. The fluorescent antibody F specifically adsorbs to the virus V. As a result, in general terms, due to the presence of virus V, the density of fluorescent antibody F becomes higher than when virus V does not exist, and a difference in fluorescence intensity corresponding to the presence or absence of virus V occurs.

At the downstream side of the fluorescence measuring unit 5, a chemical solution collecting unit 6 made of, for example, a mesh body for capturing particles formed by the particle forming unit 4 is provided. A suction pump 7 is provided on the downstream side of the chemical solution recovery unit 6, and the separated gas is exhausted to the outside of the measuring apparatus 100 via a filter for removing a virus (not shown), for example.

The operation in the first embodiment will be described. FIG. 6 is a diagram showing the diffusion of the virus in the chemical solution, the adsorption of the fluorescent antibody, and the particle formation from the solution in the diffusion unit of the first embodiment. FIG. 7 is a diagram showing diffusion of atmospheric dust in the chemical solution, adsorption of fluorescent antibodies, and particle formation from the solution in the diffusion unit of the first embodiment. 6 and 7, the case where the presence or absence of a virus is detected has been described as an example. However, the present invention is not limited to this, and the detection target substance may be arbitrary.

6 and 7, “V” indicates a detection target substance, “F” indicates a fluorescent substance contained in the chemical solution, “M” indicates a particle formed by the particle forming unit 4, and “D” "" Indicates dust contained in the atmosphere.

First, the air (outside air) is taken into the main pipe 8 via the dust removing unit 1 by the suction pump 7, and an air flow is formed in the order of the particle forming unit 4, the fluorescence measuring unit 5, and the chemical solution collecting unit 6. 7 and a filter not shown. At this time, the dust removing unit 1 removes coarse dust in the atmosphere that may block the diffusion flow path 31 of the microfluidic chip 3 or interfere with fluorescence detection in the fluorescence measuring unit 5. Part of the air taken into the main pipe 8 is sent to the air inlet 36 of the microfluidic chip 3 by the intake pump 11. The chemical solution containing the fluorescent antibody F is sent from the chemical solution storage tank 2 to the chemical solution inlet 37 of the microfluidic chip 3 by the chemical solution supply pump 21.

The flow rate of the atmosphere sent to the atmospheric inlet 36 and the flow rate of the chemical sent to the chemical inlet 37 are set to appropriate values obtained in advance by experiments by the atmospheric flow rate adjusting unit 12 and the chemical flow rate adjusting unit 22. The atmosphere and the chemical solution sent to the microfluidic chip 3 form a boundary surface on the rising portion 30 of the diffusion flow path 31 and flow in parallel. That is, the atmosphere flows through the gas flow path 34 in the diffusion flow path 31 toward the exhaust port 38, and the chemical liquid flows through the liquid flow path 35 in the diffusion flow path 31 toward the chemical liquid outflow port 39. When the atmosphere and the chemical solution are flowing in the microfluidic chip 3, the virus V in the atmosphere diffuses into the chemical solution through the boundary surface, and the fluorescent antibody F in the chemical solution is specifically adsorbed by the virus V. The atmosphere and the chemical liquid are separated at a branch portion near the outlet in the diffusion flow path 31, and the atmospheric air is exhausted outside the measuring apparatus 100 via the exhaust port 38, via the chemical liquid outflow port 39 and the solution supply path 40 that is a guide path. Then, the solution after the atmosphere and the chemical solution are mixed is sent to the particle forming unit 4.

In the particle forming unit 4, the chemical solution sent from the microfluidic chip 3 through the solution supply path 40 is discharged from the solution in the particle forming unit 4 by the air flow speeded up by the rapid constricted portion 81 of the main pipe 8. It is formed. That is, the solution is drawn into the high-speed air stream from the outlet of the solution supply path 40 and is torn, becomes a particle group formed by the particle forming unit 4, rides on the air stream, and is downstream of the particle forming unit 4 in the main pipe 8. It is guided to the fluorescence measuring unit 5 by a particle guide path 82 as a part.

The fluorescence measuring unit 5 irradiates the particles formed by the particle forming unit 4 with light, and measures the fluorescence intensity of the particles irradiated with the light. Specifically, the fluorescence measuring unit 5 irradiates the particles guided through the particle guide path 82 with light and measures the fluorescence intensity. Further, thereafter, for example, the fluorescence measurement unit 5 determines whether or not the detection target substance is contained in the specimen by comparing the measured fluorescence intensity with a threshold value. In other words, the fluorescence measuring unit 5 detects the detection target substance from the specimen.

For example, in the fluorescence measuring unit 5, the light emitting unit 51 irradiates the case body 56 through which the solution flows with ultraviolet laser light. Here, the fluorescent antibody F in the solution is fluorescent by ultraviolet laser light. Thereafter, the ultraviolet laser light is shielded by the optical filter 53, and the light receiving unit 54 selectively detects light having a fluorescence wavelength. The received light intensity detected by the light receiving unit 54 is proportional to the volume density of the fluorescent antibody F in the particles formed by the particle forming unit 4 of the chemical solution.

When the virus V is present in the particles formed by the particle forming unit 4, as shown in FIG. 6, the fluorescence intensity detected by the light receiving unit 54 becomes larger than the threshold value Is, and the received light output measuring unit 55. A virus V detection alarm is issued.

Further, when the virus V does not exist in the particles formed by the particle forming unit 4, fine dust D in the atmosphere is taken into the particles formed by the particle forming unit 4 as shown in FIG. Even if the fluorescent antibody F adheres to the dust D, the density of the fluorescent antibody F is much smaller than the density of the fluorescent antibody F adsorbed to the virus V. For this reason, the received light intensity detected by the light receiving unit 54 is smaller than a preset threshold value Is.

The particles formed by the particle forming unit 4 that has passed through the fluorescence measuring unit 5 are separated into gas and liquid by the chemical solution collecting unit 6, and the chemical solution is collected. On the other hand, the gas is exhausted out of the measuring apparatus 100 by the suction pump 7 provided on the downstream side of the chemical solution recovery unit 6.

(Processing flow in the first embodiment)
FIG. 8 is a flowchart illustrating an example of a processing flow in the first embodiment. In the example shown in FIG. 8, when the processing timing comes (Yes in Step S101), the measuring apparatus 100 forms particles having a diameter of 20 μm or less from the solution in which the fluorescent substance and the specimen are mixed (Step S102). For example, the fluorescence measuring unit 5 forms particles of 20 μm or less using a two-fluid nozzle.

In the measuring apparatus 100, the fluorescence measuring unit 5 irradiates the particles formed by the particle forming unit 4 with light (step S103). For example, the fluorescence measurement unit 5 irradiates the particles guided through the particle guide path 82 with light. And the fluorescence measurement part 5 measures the fluorescence intensity of the particle | grains irradiated with light (step S104).

According to the first embodiment, the measuring apparatus 100 diffuses the virus V in the atmosphere to be inspected into a chemical solution (aqueous solution) containing the fluorescent antibody F that adsorbs to the specific virus V, and forms particles from the chemical solution. To monitor the fluorescence intensity. When the virus V is present, the fluorescent antibody F is specifically adsorbed to increase the number of fluorescent antibodies F in the particles formed by the particle forming unit 4 and is emitted from the particles formed by the particle forming unit 4. The intensity of the fluorescence is greater than the intensity of the fluorescence emitted from the particles formed by the particle forming unit 4 when the virus V is not present. For this reason, the laser light is shielded by the optical filter 53, the fluorescence intensity transmitted through the optical filter 53 is monitored, and the fluorescence intensity (threshold value) corresponding to the particles formed by the particle forming unit 4 when the virus V is not present. ), It is possible to accurately detect the virus V contained in the gas in real time. Further, since virus detection can be automated, virus V can be constantly monitored. Therefore, the measuring apparatus 100 of the present invention is very effective in that it can quickly detect the virus V by installing it at an airport or the like, and can take a countermeasure quickly.

(Other embodiments)
Although the measurement apparatus 100 and the measurement method according to the first embodiment have been described, the present invention is not limited to this, and the measurement apparatus 100 and the measurement method may be realized in various embodiments.

For example, in the first embodiment, the air that forms an air flow in the main pipe 8 and the air that is brought into contact with the chemical solution in the microfluidic chip 3 are both supplied from the same system that has passed through the dust removing unit 1. However, the present invention is not limited to this, and a dust removing unit may be provided separately from the dust removing unit 1 in the main pipe 8, and air may be supplied to the microfluidic chip 3 from a pipe separate from the main pipe 8. .

For example, in the first embodiment, the microfluidic chip 3 is provided, but the microfluidic chip 3 may not be provided. FIG. 9 is a diagram illustrating an example of a measurement apparatus that does not have a microfluidic chip. As shown in FIG. 9, without providing the microfluidic chip 3, one end side of the solution supply path 40 is immersed in the chemical solution storage tank 2, and the other end side of the solution supply path 40 is mainly used as in the previous embodiment. The particle forming unit 4 may be configured by being provided so as to enter the throttle portion 81 of the pipe 8. In this case, the air flow in the main pipe 8 formed by the suction pump 7 causes the other end side of the solution supply path 40 to have a negative pressure, and the chemical solution in the chemical solution storage tank 2 passes through the solution supply path 40 in the main pipe 8. To form particles. The virus V in the atmosphere is torn from the other end side of the solution supply path 40 and taken into the chemical solution when passing through the particle forming unit 4 from the main pipe 8. Accordingly, since the virus V is diffused into the chemical solution by the particle forming unit 4, it can be said that the particle forming unit 4 also serves as the diffusion unit in this example. Also in this example, the same effect as the above-described embodiment can be obtained.

For example, as shown in FIG. 9, in the example where the particle forming unit 4 also serves as the diffusion unit, a chemical solution supply pump 21 may be provided in the middle of the solution supply path 40. FIG. 10 is a diagram illustrating an example of a measuring apparatus 100 in which a chemical supply pump is provided in the middle of the chemical supply pipe. As shown in FIG. 10, a chemical solution supply pump 21 may be provided in the middle of the solution supply path 40, and the chemical solution in the chemical solution storage tank 2 may be sent to the particle forming unit 4 by the liquid supply operation of the chemical solution supply pump 21. .

Further, for example, in the first embodiment, the microfluidic chip 3 is used as the diffusion unit, but an aeration tank 90 may be used as the diffusion unit. FIG. 11 is a diagram illustrating an example of a measurement apparatus 100 that uses an aeration tank as a diffusion unit. As shown in FIG. 11, an aeration tank 90 is used as a diffusing unit, and the atmosphere and the chemical solution are brought into contact with the chemical solution in the aeration tank 90 by the diffuser 91, thereby diffusing the virus V in the atmosphere into the chemical solution. May be. In FIG. 11, 93 is a ventilation port. In FIG. 11, 92 is an intake port. In this case, one end of the solution supply path 40 may be immersed in the aeration tank 90, and particles may be formed by drawing the chemical solution from the other end side of the solution supply path 40 by an air flow generated by suction of the suction pump 7. Similarly to the above, a chemical liquid flow rate adjusting unit may be provided in the middle of the solution supply path 40.

FIG. 12 is a diagram illustrating an example of a particle forming unit and a fluorescence measuring unit. For example, as shown in FIG. 12, the measuring apparatus 100 restricts the distal end opening portion of the inner tube 94 of the double tube 96 including the inner tube 94 and the outer tube 95, and the distal end side of the double tube 96 is the fluorescence measuring unit. For example, the suction pipe 97 may be connected to a surface of the case body 56 that faces the tip of the double pipe 96. In this case, the chemical solution is supplied to the inner tube 94 of the double tube 96 and the atmosphere is passed to the outer tube 95. For example, the chemical liquid may be sent to the inner tube 94 by a chemical liquid flow rate adjusting unit (not shown). By driving the suction pump 7, the atmosphere is drawn into the outer tube 95, and the chemical solution from the inner tube 94 is made into particles by the air flow of the atmosphere, and the particles are scattered in the case body 56, and from the light emitting unit 51. The particle group passes through the light-transmitting region of the laser beam. In this example, the distal end portion of the double tube 96 serves as both the diffusion portion and the particle formation portion.

Further, for example, in the first embodiment, the atmosphere may be the outside air or a breath exhaled by a person. When using a breath exhaled by the person as the atmosphere, for example, one end of a pipe for taking in the atmosphere may be widened in a trumpet shape, and a person's breath may be introduced by bringing the mouth close to the trumpet-shaped portion.

For example, in the first embodiment, the case where the substance to be measured is detected from the gas has been described as an example. However, the present invention is not limited to this, and the substance to be measured may be a liquid. . To explain with a more detailed example, the substance to be measured itself may be a liquid, or a solution in which the substance to be measured is already mixed may be used as the measurement object. In this case, if it demonstrates using the example shown in FIG. 1, the measuring apparatus 100 should just be able to provide a solution by the particle | grain formation part 4, and it is good also as the structure which makes air | atmosphere and chemical | medical solutions, such as the microfluidic chip 3, contact.

For example, in the first embodiment, the case where the particle forming unit 4 forms particles using a two-fluid nozzle has been described as an example. However, as described above, the present invention is not limited to this, and the particle forming unit 4 may form particles using any method as long as particles of 20 μm or less can be formed. Further, depending on the measurement object and measurement purpose, particles of 50 μm or less may be formed. For example, the particle forming unit 4 forms particles of 20 μm or less using at least one of electrospray, a two-fluid nozzle, a nebulizer, a piezoelectric element (for example, bubble jet (registered trademark)), an ultrasonic wave, and a decompression process. You can do it. The minimum particle diameter that can be formed by the above-described particle forming means is about 3 nm. In particular, when particles are formed using electrospray, particles of 3 nm to 50 μm can be formed with good controllability.

For example, the particle forming unit 4 may form particles by vaporizing a solution by ultrasonic waves or reduced pressure treatment, or may form particles by atomizing a nebulizer. Further, for example, the particle forming unit 4 may form particles by ejecting a solution from, for example, a nozzle filled with a solution using a piezoelectric element that converts a voltage into a pressure.

The particle forming unit 4 preferably forms particles of 20 μm or less using an electrospray or a two-fluid nozzle. By forming the particles using an electrospray or a two-fluid nozzle, the particles can be formed even if the amount of the solution is small.

FIG. 13 and FIG. 14 are diagrams showing an example of the configuration of a measuring apparatus when particles are formed using electrospray. In the example illustrated in FIG. 13, the measuring apparatus 100 b includes an electrospray 400 as the particle forming unit 4 and a sample providing unit 410 that provides a solution to the electrospray 400.

In the example illustrated in FIG. 13, the sample providing unit 410 includes a vial 411 that holds a solution in which a specimen and a chemical solution are mixed, and a pressure unit 412 that applies pressure to the vial 411. In the example shown in FIG. 13, the solution held in the vial 411 contains one end of a capillary 401 that supplies the solution held in the vial 411 to the discharge unit 402.

In the example shown in FIG. 13, the electrospray 400 includes a capillary 401, a solution discharge portion 402 provided at the other end of the capillary 401, an air intake port 403, and an electrode 404 attached to the capillary 401. , An electrode 405 provided in a radiation direction in which the solution is discharged from the discharge portion 402, and a voltage supply portion 406.

As shown in FIG. 14, the voltage supply unit 406 supplies a voltage to the electrode 404 and the electrode 405. For example, the voltage supply unit 406 supplies a positive voltage to the electrode 404 and supplies a negative voltage to the electrode 405. As a result, a strong electric field is generated in the emission part 402 which is the tip part of the capillary 401.

Here, when the particle forming unit 4 forms particles, the pressure unit 412 applies pressure, so that the solution is supplied from the vial 411 to the electrospray 400 through the capillary 401. Thereafter, a strong electric field is generated in the emitting portion 402, and charged ions gather on the surface of the solution to form a cone. This cone is also called Tylor Cone. Thereafter, particles are formed and emitted from the emission part 402.

In the example shown in FIG. 14, the electrode 405 has a hole in the middle. As a result, among the particles emitted by the emission unit 402, the particles that have passed through the hole in the middle of the electrode 405 are sent to the particle guide path 82 and sent to the fluorescence measurement unit 5. Moreover, the particle | grain diameter discharged | emitted from the discharge | release part 402 becomes small because a volatile solvent evaporates after that.

Note that the configuration of the measuring apparatus 100b having the electrospray 400 is not limited to the examples shown in FIGS. 13 and 14, and may be any configuration. For example, in the example shown in FIG. 13 and FIG. 14, the case where the solution held in the vial 411 is supplied to the electrospray 400 by applying pressure to the electrospray 400 has been described as an example. Absent. For example, a solution may be supplied by providing a separate pump, and any method may be used. In the example shown in FIGS. 13 and 14, the case where the atmosphere is taken in from the atmosphere intake port 403 is shown as an example, but the present invention is not limited to this. For example, clean air may be provided, or clean air and CO 2 may be combined and placed in the electrospray 400. Further, in the example illustrated in FIGS. 13 and 14, the case where the electrode 405 is provided in the radial direction in which the solution is discharged from the discharge unit 402 is illustrated as an example, but the present invention is not limited thereto. For example, the electrode 405 may be provided along the radial direction in which the solution is discharged from the discharge portion 402. For example, in the example shown in FIG. 13, the inner wall of the electrospray 400 may be used as the electrode 405. Further, the fluorescent substance that specifically adsorbs to the detection target substance is not bound by the fluorescent antibody. For example, sugar chains or proteins may be used as fluorescent labels.

(Second Embodiment)
In one embodiment, the measurement apparatus according to the second embodiment forms particles that form particles of a solution from a solution in which a fluorescent substance that specifically adsorbs to a detection target substance and a sample are mixed, and emits the particles from the emission unit. And a fluorescence measuring unit that irradiates light at a position within a predetermined distance from the position where the particles are emitted by the particle forming unit, and measures the fluorescence intensity of the particles irradiated with the light.

Further, in the measuring apparatus according to the second embodiment, in one embodiment, the particle forming unit is an electrospray. Further, the electrospray has a discharge portion and an electrode provided in a radiation direction in which the solution is discharged from the discharge portion. The fluorescence measurement unit irradiates light to the space between the emission unit and the electrode, and measures the fluorescence intensity of the particles emitted from the emission unit.

In addition, in one embodiment, the measurement apparatus according to the second embodiment further includes a laminar flow forming mechanism that forms a laminar flow that flows from the discharge portion to the electrode.

(Configuration of the measuring apparatus according to the second embodiment)
FIG. 15 is a diagram illustrating an example of the configuration of the measurement apparatus according to the second embodiment. As shown in FIG. 15, the measuring apparatus 500 includes a particle forming unit 510 and a fluorescence measuring unit 520.

The particle forming unit 510 forms solution particles from a solution in which a fluorescent substance that specifically adsorbs to the detection target substance and the sample are mixed, and emits the particles from the emitting unit. In the example shown in FIG. 15, the particle forming unit 510 includes a discharge unit 511 that forms and discharges particles of a solution from a solution in which a fluorescent substance that specifically adsorbs to the detection target substance and the sample are mixed, and a detection target substance. A capillary 512 for supplying a solution in which a fluorescent substance specifically adsorbed to the sample and a specimen are mixed to the discharge unit 511, an electrode 513 provided in a radial direction in which the solution is discharged from the discharge unit 511, and the discharge unit 511 and the electrode 513 and a voltage supply unit 514 for supplying a voltage.

The voltage supply unit 514 supplies a positive voltage to the electrode 513 and supplies a negative voltage to the emission unit 511, for example. As a result, a strong electric field is generated in the emission portion 511 that is the tip of the capillary 512. Note that the voltage supply unit 514 may not supply a voltage to the emission unit 511 as long as it can generate a strong electric field in the emission unit 511. For example, the voltage supply unit 514 supplies a voltage to the electrode provided in the capillary 512 and the electrode 513. May be.

The fluorescence measuring unit 520 irradiates light at a position within a predetermined distance from the position where the particles are emitted by the particle forming unit 510, and measures the fluorescence intensity of the particles irradiated with the light. Specifically, the fluorescence measurement unit 520 irradiates light within a unit in which particles are formed by the particle formation unit 510 and measures the fluorescence intensity. In other words, the particles formed by the particle forming unit 510 are irradiated with light as soon as they are formed, and the fluorescence intensity is measured. For example, the fluorescence measurement unit 520 irradiates the space between the emission unit 511 and the electrode 513 with light, and measures the fluorescence intensity of the particles emitted from the emission unit 511.

In addition, about the other point of the fluorescence measurement part 520, it is good also as a structure similar to the fluorescence measurement part 5 in the above-mentioned 1st Embodiment.

The measuring apparatus 500 further includes a laminar flow forming mechanism that forms a laminar flow that flows from the discharge portion 511 to the electrode 513. In the example illustrated in FIG. 15, the measuring apparatus 500 includes a laminar flow intake port 531 and a laminar flow exhaust port 532 as a laminar flow forming mechanism.

The laminar flow intake port 531 and the laminar flow exhaust port 532 may be provided at arbitrary positions as long as a laminar flow flowing from the discharge portion 511 to the electrode 513 can be formed. The laminar flow inlet 531 is provided, for example, at a position farther from the electrode portion 513 than the discharge portion 511 in the wall surface of the particle forming portion 510. The laminar exhaust port 532 is provided, for example, in a direction in which particles are emitted by the emission unit 511 on the wall surface of the particle forming unit 510 and at a position farther from the emission unit 511 than the electrode 513.

In the example shown in FIG. 15, the air taken in through the laminar flow inlet 531 is discharged from the laminar air outlet 532, thereby forming a laminar flow that flows from the discharge portion 511 to the electrode 513. In addition, the gas taken in by the laminar flow intake port 531 is not limited to the atmosphere, and may be any gas.

Also, the laminar flow outlet 532 separates and collects the particles formed by the particle forming unit 510, for example, when discharging the laminar flow, and discharges only the gas. The laminar exhaust port 532 corresponds to, for example, the chemical solution recovery unit 6 and the suction pump 7 in the first embodiment.

Here, by making the electrode 513 mesh, it is possible to efficiently flow a laminar flow formed by a laminar flow forming mechanism.

As described above, according to the second embodiment, the particle forming unit that forms particles of the solution from the solution in which the fluorescent substance that specifically adsorbs to the detection target substance and the sample are mixed and emits the particles from the emitting unit 511. 510 and a fluorescence measuring unit 520 that irradiates light at a position within a predetermined distance from the position where the particles are emitted by the particle forming unit 510 and measures the fluorescence intensity of the particles irradiated with the light. As a result, it is possible to quickly irradiate the particles formed by the particle forming unit 510 with light and measure the fluorescence intensity. Further, for example, the measurement apparatus 500 can be downsized by integrating the particle forming unit 510 and the fluorescence measurement unit 520.

In the measurement apparatus 500 according to the second embodiment, the particle forming unit 510 is an electrospray, and the electrospray is provided in the emission direction of the emission unit 511 and the emission direction of the solution from the emission unit 511. And have. In addition, the fluorescence measurement unit 520 irradiates the space between the emission unit 511 and the electrode 513 with light, and measures the fluorescence intensity of the particles emitted from the emission unit 511. As a result, it is possible to quickly irradiate the particles formed by the particle forming unit 510 with light and measure the fluorescence intensity.

For example, when measuring fluorescence with a fluorescence measuring unit provided separately with particles formed by electrospray, it is stabilized by neutralizing the charge of particles formed by electrospray and then provided separately. It is conceivable to lead to the measured fluorescence measurement unit. Compared to such a method, according to the second embodiment, the fluorescence intensity of particles formed using electrospray is measured in the particle forming unit 510. As a result, particle loss can be reduced. For example, before the fluorescence intensity is measured, it is possible to prevent the loss of particles due to the particles adhering to the electrode 513 provided to face the emission portion 511.

In addition, the measuring apparatus 500 in the second embodiment further includes a laminar flow forming mechanism that forms a laminar flow that flows from the discharge portion 511 to the electrode 513. As a result, it is possible to prevent particles adhering to the electrode 513 from peeling off and moving to the emitting portion 511 side, and it is possible to maintain high detection accuracy. For example, even if particles are peeled off from the electrode 513, it is possible to maintain high detection accuracy by being discharged in a laminar flow.

(Other embodiments)
Although the measurement apparatus 500 according to the second embodiment has been described, the present invention is not limited to this, and the measurement apparatus 500 may be realized in various embodiments.

For example, in the second embodiment, the particle size is not particularly mentioned, but as in the first embodiment, the particle forming unit 510 may form particles of the solution having a diameter of 20 μm or less. .

In addition, for example, in the measurement apparatus 500 according to the second embodiment, a part of the configuration of the measurement apparatus in the above-described embodiment may be used as long as no contradiction occurs.

For example, in the second embodiment, the particle forming unit 510 is an electrospray, and the fluorescence measuring unit 520 measures the fluorescence intensity by irradiating the space between the emitting unit 511 and the electrode 513. Although described using cases, the present invention is not limited to this. That is, the fluorescence measurement unit 520 may measure the fluorescence intensity by irradiating light to a position within a predetermined distance from the position where the particles are emitted by the particle forming unit 510, and may have an arbitrary configuration.

4 Particle Forming Unit 5 Fluorescence Measuring Unit 51 Light Emitting Unit 54 Light Receiving Unit 82 Particle Guide Path 92 Intake Port 100 Measuring Device

Claims (8)

A particle forming unit for forming particles of the solution having a diameter of 20 μm or less from a solution in which a fluorescent substance that specifically adsorbs to a detection target substance and a sample are mixed;
A measurement apparatus comprising: a fluorescence measurement unit configured to irradiate the particles formed by the particle formation unit with light and measure fluorescence intensity of the particles irradiated with the light. The measurement apparatus according to claim 1, wherein the particle forming unit forms the particles using at least one of electrospray, a two-fluid nozzle, a nebulizer, a piezoelectric element, ultrasonic waves, and a decompression process. . The particle forming unit further includes a particle guide path for guiding the particle to a position where light is irradiated.
The measurement apparatus according to claim 1, wherein the fluorescence measurement unit irradiates the particles guided through the particle guide path with light. A particle forming step of forming particles having a diameter of 20 μm or less from a solution in which a fluorescent substance that specifically adsorbs to a detection target substance and a sample are mixed;
An irradiation step of irradiating the particles with light;
And a fluorescence measurement step of measuring the fluorescence intensity of the particles irradiated with light. 5. The measurement method according to claim 4, wherein in the particle forming step, the particles are formed using at least one of electrospray, a two-fluid nozzle, a nebulizer, a piezoelectric element, ultrasonic waves, and a reduced pressure treatment. . The particle forming step further includes a guide step of guiding the particle through a particle guide path to a position where light is irradiated.
6. The measurement method according to claim 4, wherein the irradiation step irradiates the particles guided through the particle guide path with light. Supplying a solution in which a fluorescent substance that specifically adsorbs to a detection target substance and a specimen are mixed, and a discharge section that forms particles of the solution;
An electrode installed to face the emission part;
A fluorescence measuring unit that measures the fluorescence intensity of the particles by irradiating the particles with light, and
The light irradiation is performed between the emitting part and the electrode. The measuring apparatus according to claim 7, further comprising a laminar flow forming mechanism for forming a laminar flow flowing from the discharge portion to the electrode.

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