JP6768621B2 - Fluorescence detection device using surface plasmon resonance and its operation method - Google Patents

Description

The present invention relates to a fluorescence detection device using surface plasmon resonance and a method of operating the same.

A fluorescence detection device using a fluorescence method is known in bio-measurement and the like (see, for example, Patent Document 1). The fluorescence method is a method of irradiating a fluorescent label bound to a test substance with excitation light, detecting fluorescence generated from the fluorescent label, and analyzing the test substance. The fluorescence detection device described in Patent Document 1 uses a surface plasmon resonance phenomenon to enhance the fluorescence emitted by a fluorescent label to analyze a test substance. Such a fluorescence detection device is also called a measurement system using surface plasmon resonance excitation enhanced fluorescence spectroscopy (SPFS) (SPFS: Surface Plasmon field-enhanced Fluorescence Spectroscopy).

In such a fluorescence detection device, an analysis chip provided with a flow path through which a sample solution containing a test substance flows is used. The analysis chip is provided with a flow path through which the sample solution containing the test substance flows, a reaction region in which the antibody that specifically reacts with the test substance is fixed and the sample solution contacts, and a reaction region. It has a metal film to be formed and a main surface in contact with the back surface opposite to the front surface having a reaction region in the metal film, and has a dielectric material transparent to excitation light.

An injection port for injecting the sample solution is provided at one end of the flow path. A nozzle for injecting the sample solution is attached to the injection port, and a pump for discharging and sucking the sample solution via a gas is connected to the nozzle.

In the fluorescence detection device described in Patent Document 1, the sample solution is reciprocated in the flow path by repeatedly discharging and sucking the sample solution with a pump. The reciprocation of the sample solution is performed with the sample solution in constant contact with the reaction region. When a gas such as air comes into contact with the reaction region, the reaction amount decreases. By keeping the sample solution in constant contact with the reaction region, a decrease in the reaction amount can be suppressed, and the test substance can be detected with high accuracy even when the sample solution is small.

Japanese Patent No. 5640980

However, as described in Patent Document 1, when the sample solution is reciprocated in the flow path, the sample solution is kept in constant contact with the reaction region simply by controlling the discharge amount and the suction amount of the pump. Sometimes it was difficult. This is because the sample solution has individual differences in viscosity, etc., and due to the individual differences, the sample solution may not reach the target position even if the discharge amount and suction amount of the pump are accurately controlled. On the contrary, it may pass the target position. For example, even if the pump alone tries to control the gas-liquid interface that is the boundary between the sample solution and air so that it does not pass through the reaction region, the gas-liquid interface passes through the reaction region and the reaction region is exposed to air. Sometimes things happened.

If the measurement conditions vary, such as when the reaction area is exposed to air when testing one sample solution, or when the reaction area is not exposed to air when testing another sample solution, the reliability of the measurement results becomes unreliable. It may decrease. Therefore, when such a situation occurs, it may be preferable to treat it as a liquid feeding abnormality.

The present invention provides a fluorescence detection device using surface plasmon resonance and an operating method thereof that can ensure the reliability of measurement results when the sample solution is reciprocated in the flow path of the analysis chip by using a pump. The purpose is to provide.

The present invention provides an analysis chip having a flow path through which a sample solution containing a test substance flows, and a flow path in which an antibody that specifically reacts with the test substance is fixed and a reaction region in contact with the sample solution is arranged. In a fluorescence detection device that uses surface plasmon resonance to detect fluorescence generated from a fluorescent label bound to a test substance, the sample solution is discharged and sucked in the flow path in the flow path in the opposite direction to the first direction. A pump that reciprocates in the second direction and a pump control unit that controls the discharge amount and suction amount of the pump. The sample solution is in contact with the sample solution and the reaction region through the control of the discharge amount and the suction amount. A gas-liquid interface detection sensor used to detect that the gas-liquid interface, which is the interface between the sample solution and the gas, has passed through at least one preset position in the flow path, and a pump control unit that reciprocates the gas. Based on the output of the gas-liquid interface detection sensor, there is a liquid supply abnormality determination unit that determines whether or not a liquid supply abnormality has occurred in the sample solution, and a warning unit that warns that fact if it is determined that a liquid supply abnormality has occurred. The flow path has a cross-sectional area change portion in which the cross-sectional area perpendicular to the flow direction of the sample solution changes, and the gas-liquid interface sensor is located in the cross-sectional area change portion on the upstream side of the reaction region. The passage of the gas-liquid interface is detected at at least one of the first position where it is located, the second position located at the cross-sectional area change portion on the downstream side of the reaction region, and the third position in the reaction region.

The pump discharges and sucks the sample solution through the gas,
The pump control unit preferably controls the gas discharge amount and the suction amount.

The pump control unit preferably reciprocates the sample solution in a state where the sample solution and the reaction region are in constant contact with each other.

At least one position is a first position located upstream of the reaction region in the flow path and a position downstream of the reaction region, with reference to one of the first and second directions. It is preferably either the second position to be used or the third position in the reaction region.

The flow path has a cross-sectional area change portion in which the cross-sectional area of the cross section orthogonal to the flow direction of the sample solution changes at at least one of the first position and the second position, and the gas-liquid interface detection sensor has a gas-liquid interface detection sensor. It is preferable to include a pressure sensor for detecting the pressure fluctuation in the flow path that occurs when the sample solution passes through the cross-sectional area change portion.

The pressure sensor is preferably connected to a branch path that branches off from the pipeline between the flow path and the pump.

The gas-liquid interface detection sensor preferably includes an optical sensor that optically detects that the gas-liquid interface has passed at least one of the first position, the second position, and the third position.

The optical sensor includes a first optical sensor that detects the passage of the gas-liquid interface at the first position, a second optical sensor that detects the passage of the gas-liquid interface at the second position, and a third that detects the passage of the third position. It preferably includes at least one of the optical sensors.

The measuring unit that detects fluorescence may be used as at least one of the first optical sensor, the second optical sensor, and the third optical sensor.

When the liquid-liquid interface detecting unit detects that the gas-liquid interface has passed at least one of the first position and the second position based on the output of the gas-liquid interface detection sensor, a liquid feeding abnormality occurs. It is preferable to determine that

It is preferable that the liquid feed abnormality determination unit determines that a liquid feed abnormality has occurred when it detects that the gas-liquid interface has passed the third position based on the output of the gas-liquid interface detection sensor.

From the time when the gas-liquid interface detection sensor detects that the gas-liquid interface has passed through one of the first position and the second position toward the reaction region, the flow direction of the sample solution is reversed and the same position is again obtained. It is equipped with a timer that measures the round-trip time until it detects that the gas-liquid interface has passed, and the liquid-feeding abnormality determination unit detects that the liquid-feeding abnormality occurs when the round-trip time exceeds a preset scheduled time. It is preferable to determine that it has occurred.

The present invention is a flow path through which the sample solution containing the analyte, the analysis chip having a flow path of the reaction region antibody which reacts specifically with the analyte contacts the sample solution is fixed is disposed In the method of operating the fluorescence detection device using the surface plasmon resonance that detects the fluorescence generated from the fluorescent label bound to the test substance, the sample solution is discharged and sucked to make the sample solution the first direction in the flow path. It is a pump control step that controls the discharge amount and the suction amount of the pump that reciprocates in the opposite second direction, and the sample is in contact with the sample solution and the reaction region through the control of the discharge amount and the suction amount. A pump control step for reciprocating the solution, a gas-liquid interface detection step for detecting that the gas-liquid interface, which is the interface between the sample solution and the gas, has passed at at least one preset position in the flow path, and a gas. Based on the detection of passage through the liquid interface, there are a liquid feeding abnormality determination step for determining whether or not a liquid feeding abnormality has occurred in the sample solution, and a warning step for warning when it is determined that a liquid feeding abnormality has occurred. The flow path has a cross-sectional area change portion in which the cross-sectional area perpendicular to the flow direction of the sample solution changes, and the gas-liquid interface sensor is located in the cross-sectional area change portion on the upstream side of the reaction region. The passage of the gas-liquid interface is detected at at least one of the first position, the second position located in the cross-sectional area change portion on the downstream side of the reaction region, and the third position in the reaction region.

According to the present invention, the reliability of the measurement result can be ensured when the sample solution is reciprocated by using a pump in the flow path of the analysis chip.

It is an external view of the fluorescence detection apparatus of this invention. It is a block diagram of a fluorescence detection device. It is a schematic diagram which shows an example of the analysis chip used for a fluorescence detection apparatus. It is a schematic diagram which shows a mode that a sample is extracted from a sample container by a sample processing part using a nozzle tip. It is a schematic diagram which shows how the sample in a nozzle tip is injected into a reagent cell and agitated by a sample processing part. It is explanatory drawing which shows the positional relationship between a flow path and a measuring part, and the moving method of a measuring part. It is explanatory drawing explaining the outline of fluorescence detection using a measuring part. It is explanatory drawing of the incident angle of the excitation light and the plasmon strengthening. It is explanatory drawing of the structure which detects the gas-liquid interface of the sample solution flowing in a flow path. It is a flowchart which shows the procedure of a measurement process. It is explanatory drawing which shows the state transition of the sample solution in the flow path of the normal pattern in which the liquid feeding abnormality is not detected. FIG. 11A shows a state immediately after injecting the sample solution, FIG. 11B shows a state in which the pump is sucking, and FIG. 11C shows a state in which the pump is discharging. It is explanatory drawing which shows the state transition of the sample solution in the flow path of the abnormality pattern in which the liquid feeding abnormality is detected. FIG. 12A shows a state immediately after injecting the sample solution, FIG. 12B shows a state in which the pump is sucking, and FIG. 12C shows a state in which the pump is discharging. It is explanatory drawing of the 2nd Embodiment which uses an optical sensor as a gas-liquid interface detection sensor. FIG. 13A shows a state in which the pump is discharging, and FIG. 13B shows a state in which the pump is sucking. It is explanatory drawing which shows the modification of 2nd Embodiment. It is explanatory drawing which shows the example which uses the measuring part as an optical sensor. FIG. 15A shows a state in which the pump is discharging, and FIG. 15B shows a state in which the pump is sucking. It is explanatory drawing of the example which determines the liquid feeding abnormality by the round-trip time of a sample solution. FIG. 16A shows a state in which the pump is sucking, FIG. 16B shows a state in which the pump is further sucking, and FIG. 16C shows a state in which the pump is discharging.

[First Embodiment]
The fluorescence detection device 100 shown in FIG. 1 is a fluorescence detection device 100 that utilizes surface plasmon resonance, and more specifically, it is a device used for analysis including inspection of a biological substance by an antigen-antibody reaction or the like.

When the analysis is performed using the fluorescence detection device 100, a sample container CB containing the sample shown in FIG. 1, a nozzle tip NC used for extracting the sample and the reagent, a reagent cell and a microchannel are formed. The analysis chip 10 is set in the fluorescence detection device 100. The sample container CB, the nozzle tip NC, and the analysis tip 10 are all disposable items that are discarded once they are used. Then, the fluorescence detection device 100 injects the sample into the flow path 15 of the analysis chip 10 to perform quantitative or qualitative analysis on the test substance in the sample.

The sample is, for example, blood, more specifically serum, plasma, or whole blood. The sample may be other than blood, and may be urine, nostril fluid, saliva, stool, body cavity fluid, or the like. Examples of the test substance contained in the sample include nucleic acids, proteins, amino acids, carbohydrates, lipids, or modified molecules and complexes thereof. The complex may be, for example, a tumor marker, a signal transduction substance, a hormone, or the like.

As shown in FIG. 2, the fluorescence detection device 100 includes a mounting unit 101, a sample processing unit 20, a measuring unit 30, a control unit 40, and the like. The analysis chip 10 is mounted on the mounting portion 101. The sample processing unit 20 extracts a sample from the sample container CB (see FIG. 1) using the nozzle tip NC, and mixes the extracted sample with the reagent to generate a sample solution. Further, the sample processing unit 20 injects the generated sample solution into the analysis chip 10.

Specifically, the sample processing unit 20 includes a nozzle moving mechanism 21, a pump 22, a pressure gauge 23, and the like. The nozzle moving mechanism 21 is a mechanism for moving the nozzle tip NC in the vertical direction and the horizontal direction. The pump 22 is connected to the nozzle tip NC via a pipe 26, and discharges and sucks a liquid such as a sample via a gas. The pipe 26 is branched, and a pressure gauge 23 is connected to the branch path. The pressure gauge 23 measures the pressure in the pipe 26.

The measuring unit 30 measures the reaction status of the test substance in the sample on the analysis chip 10 to measure the concentration of the test substance in the sample and the like. The measuring unit 30 includes a light irradiation unit 31, an incident angle adjusting mechanism 33, a fluorescence detection unit 32, and the like.

The light irradiation unit 31 irradiates the analysis chip 10 with the excitation light L. The light irradiation unit 31 irradiates the excitation light L. The incident angle adjusting mechanism 33 adjusts the incident angle of the excitation light L to irradiate the analysis chip 10. The fluorescence detection unit 32 detects the fluorescence emitted by the fluorescence label excited by the excitation light L in the analysis chip 10 and outputs a fluorescence detection signal to the control unit 40. The fluorescence detection unit 32 includes a photodiode, a photomultiplier, a CCD (Charge Coupled Device) image sensor, a CMOS (Complementary Metal Oxide Semiconductor Image Sensor) image sensor, and the like.

The measuring unit moving mechanism 36 is a moving mechanism that moves the measuring unit 30. As will be described later, the analysis chip 10 is provided with a plurality of regions to be measured, and the measuring unit moving mechanism 36 analyzes the measuring unit 30 so that the plurality of regions of the analysis chip 10 can be measured. Move relative to chip 10.

The control unit 40 comprehensively controls each unit of the fluorescence detection device 100. An operation unit 51 and a display unit 52 are connected to the control unit 40. Further, the control unit 40 has a built-in timer for performing various timekeeping. The operation unit 51 is composed of buttons, cross keys, and the like, and inputs an operation instruction such as a measurement start instruction to the control unit 40. In addition, input of patient information related to the sample is also performed through the operation unit 51. The display unit 52 is composed of, for example, a liquid crystal panel or the like, and displays messages such as a measurement result, a status indicating an operating state, and a warning.

The control unit 40 controls the sample processing unit 20 in accordance with the measurement start instruction from the operation unit 51 to inject the sample solution into the analysis chip 10. Then, the measuring unit moving mechanism 36 and the measuring unit 30 are operated to perform measurement. In the measurement, the control unit 40 performs data analysis based on the fluorescence detection signal acquired from the fluorescence detection unit 32, and outputs the measurement result including the analysis result to the display unit 52.

FIG. 3 is a schematic view showing an example of the analysis chip 10. The analysis chip 10 has a structure in which an injection port 12, a discharge port 13, sample cells 14A and 14B, and a flow path 15 are formed in a main body 11 formed of a dielectric material such as a light-transmitting resin. The injection port 12 communicates with the discharge port 13 via the flow path 15. The nozzle tip NC is inserted into the injection port 12. The sample processing unit 20 injects the sample solution into the flow path 15 via the nozzle tip NC. The sample cells 14A and 14B are containers for containing the fluorescent reagent to be mixed with the sample in the sample container CB. The fluorescent reagent is subjected to pretreatment such as adsorbing to a protein in the sample to dissociate the target for pH adjustment, for example. The openings of the sample cells 14A and 14B are sealed by a sealing member, and the sealing member is perforated when the sample and the fluorescent reagent are mixed.

Further, a reaction region 16 for detecting the test substance in the sample is provided in the flow path 15. A test region TR and a control region CR are formed in the reaction region 16. In the flow path 15, when the side where the injection port 12 is located is the upstream side of the reaction region 16, the control region CR is provided on the downstream side of the test region TR.

The first antibody is immobilized on the test region TR, and the antibody labeled by the so-called sandwich method is captured. Further, the reference antibody is fixed in the control region CR, and the reference antibody captures the fluorescent substance by flowing the sample solution on the control region CR. Two control regions CR are formed, a so-called negative control region CR for detecting non-specific adsorption and a so-called positive control region CR for detecting the difference in reactivity due to sample difference. Is formed.

Then, when the measurement start is instructed, the sample processing unit 20 sucks the sample from the sample container CB using the nozzle tip NC as shown in FIG. After that, the sample processing unit 20 punches the seal member of the sample cell 14A as shown in FIG. 5, mixes and stirs the sample with the reagent in the sample cell 14A, and then sucks the sample solution again using the nozzle tip NC. .. This operation is also performed for the sample cell 14B. The reagent is a second antibody B2 labeled with a fluorescent label F. The second antibody B2 specifically binds to the test substance (antigen) A present in the sample. Therefore, by mixing and stirring the sample and the reagent, the second antibody B2 and the fluorescent label F are attached to the surface of the test substance (antigen) A by binding between the second antibody B2 and the test substance (antigen) A. A modified sample solution is produced.

Then, the sample processing unit 20 inserts the nozzle tip NC containing the sample solution into the injection port 12. Then, by operating the pump 22 to perform a discharge operation of discharging the sample solution from the nozzle tip NC, the sample solution in the nozzle tip NC is injected into the flow path 15. As described above, the atmosphere may be sucked from the discharge port 13 in synchronization with the discharge operation from the injection port 12. In this way, the sample solution can be injected more smoothly into the flow path 15.

Although the case where the sample processing unit 20 supplies the sample solution in which the sample and the reagent are mixed into the flow path 15 is illustrated, the sample processing unit 20 fills the flow path 15 with the reagent in advance. Only the sample may flow in from the inlet 12.

FIG. 6 is an explanatory diagram showing a test region TR and a control region CR of the analysis chip 10 and a state in which the measurement unit 30 moves with respect to the analysis chip 10. The test region TR and the two control regions CR are arranged along the flow direction (X direction) of the sample solution in the flow path 15. The main body 11 is provided with a prism 11A having an incident surface on which the excitation light L is incident, corresponding to the test region TR and the two control regions CR.

In the measuring unit 30, the light irradiation unit 31 is arranged at a position facing the incident surface of the prism 11A of the analysis chip 10 when the analysis chip 10 is mounted on the mounting unit 101. On the other hand, the fluorescence detection unit 32 is arranged above the flow path 15 of the analysis chip 10 at a position facing the test region TR and the control region CR, and is arranged at a position where fluorescence from each region TR and CR can be detected. ing.

The measuring unit moving mechanism 36 linearly moves the light irradiation unit 31 and the fluorescence detecting unit 32 along the flow direction (X direction) of the flow path 15, that is, the arrangement direction of the test region TR and the control region CR. As a result, the measuring unit 30 can selectively move to a position facing each of the test region TR and the two control regions CR, and measure the reaction status of each region.

FIG. 7 is an explanatory view of the relationship between the reaction region 16 of the analysis chip 10 and the light irradiation unit 31 and the fluorescence detection unit 32 as viewed from the X direction. In FIG. 7, the test area TR will be focused on, but the same applies to the control area CR. The main body 11 of the analysis chip 10 has a dielectric plate 17. In the dielectric plate 17, the front surface 17A constitutes the bottom surface of the flow path 15, and the prism 11A is provided on the back surface 17B. A metal film 18 constituting the test region TR and the control region CR is formed. The material of the metal film 18 is gold in this example. The dielectric plate 17 and the prism 11A are integrally molded, and the prism 11A is also a dielectric.

In the dielectric plate 17, the front surface 17A corresponds to the main surface of the metal film 18 in contact with the back surface opposite to the front surface on which the test region TR corresponding to the reaction region is provided.

The light irradiation unit 31 causes the excitation light L to enter the surface 17A from the back side of the front surface 17A of the dielectric plate 17 in contact with the back surface of the metal film 18 via the prism 11A. The incident angle θ of the optical axis with respect to the surface 17A is an angle equal to or higher than the critical angle that satisfies the total reflection condition. As a result, the excitation light L is applied to the back surface of the metal film 18 in the test region TR and the control region CR. The light irradiation unit 31 is composed of, for example, an LD (Laser Diode) which is a light source that emits excitation light L, a reflection mirror that reflects the excitation light L, and the like. The reflection mirror can be rotationally moved, and the light irradiation unit 31 can change the incident angle θ of the excitation light L by rotationally moving the reflection mirror. The incident angle adjusting mechanism 33 adjusts the incident angle of the excitation light L without changing the irradiation position of the excitation light L on the back surface of the metal film 18 by rotating and moving the reflection mirror by using a lens in combination.

When the excitation light L is incident on the back surface of the metal film 18 at a specific incident angle equal to or higher than the critical angle by the light irradiation unit 31, the evanescent wave Ew exudes onto the metal film 18, and the evanescent wave Ew causes the metal. Surface plasmons are excited on the surface of the film 18. This surface plasmon creates an electric field distribution on the surface of the metal film 18, and an electric field enhancement region is formed. Then, the fluorescent label F bound to the first antibody B1 fixed on the metal film 18 is excited by the evanescent wave Ew to generate enhanced fluorescent Lf. The fluorescence detection unit 32 receives the enhanced fluorescence Lf and outputs a fluorescence detection signal according to the amount of light of the received fluorescence Lf.

Here, a specific incident angle θ at which surface plasmon resonance occurs and the enhanced fluorescence Lf is maximized is called a resonance angle. The resonance angle changes depending on the type of the sample solution SL that comes into contact with the surface of the metal film 18. Therefore, the incident angle θ of the excitation light L is adjusted by the incident angle adjusting mechanism 33.

FIG. 8 shows the relationship between the plasmon intensification of the fluorescence Lf when plasma is used as the sample and the reflectance of the reflected light RL of the excitation light L with respect to the incident angle θ. The profile of FIG. 8 is an example in which the wavelength of the excitation light L is 658 nm, the thickness of the metal film 18 is 36 nm, the material of the metal film 18 is gold, and the material of the prism 11A is PMMA (polymethylcrylic). Here, the plasmon increasing intensity is an index indicating how many times the amount of light of the fluorescent Lf after the enhancement is multiplied by the reference value, using the amount of light of the fluorescence Lf when there is no enhancement as a reference value. The plasmon enhancement has a proportional relationship with the amount of light of the fluorescence Lf detected by the fluorescence detection unit 32, and even if the vertical axis is the amount of light of the fluorescence Lf in FIG. 8, the relationship with the incident angle θ has a similar profile.

In FIG. 8, the incident angle θ at which the plasmon increasing intensity of the fluorescence Lf shows a peak value and becomes maximum is specified as a resonance angle. In the example shown in FIG. 8, the resonance angle is 73.6 degrees. Since the excitation light L consumes energy for plasmon enhancement, the reflected light RL of the excitation light L is greatly attenuated in the vicinity of the resonance angle, and the reflectance shows a minimum value, contrary to the plasmon enhancement of the fluorescence Lf. By adjusting the incident angle, as shown in FIG. 8, the resonance angle at which the plasmon increasing intensity of the fluorescence Lf shows a maximum value is specified.

FIG. 9 shows how the sample solution SL is injected into the flow path 15 to bring the sample solution SL into contact with the reaction region 16. In the fluorescence detection device 100, the control unit 40 operates the pump 22 with the nozzle tip NC inserted into the injection port 12 even after the sample solution SL is injected into the flow path 15. By operating the pump 22, the control unit 40 reciprocates the sample solution SL in the flow path 15 in the first direction and the second direction opposite to the first direction. As described above, when the sample solution SL is injected or reciprocated, a pump may be connected to the discharge port 13 side to perform suction and discharge in synchronization with the pump 22.

Here, the first direction is the direction in which the sample solution SL flows from the injection port 12 side toward the discharge port 13 side in the flow path 15, and the second direction is the opposite direction.

The control unit 40 is a pump control unit that controls the gas discharge amount and the suction amount of the pump 22. Specifically, the control unit 40 controls the gas discharge amount and the suction amount of the pump 22 to reciprocate the sample solution SL, and the sample solution SL is brought into constant contact with the sample solution SL and the reaction region 16. It flows in the first direction and the second direction to reciprocate in the flow path 15. That is, the control unit 40 controls the gas discharge amount and the suction amount of the pump 22 so that the sample solution SL reciprocates in the flow path 15 in a state of being in constant contact with the reaction region 16.

The reason why the sample solution SL and the reaction region 16 are kept in constant contact with each other during this reciprocation is to prevent variations in measurement conditions that affect the measurement results. That is, when the front end and the rear end of the sample solution SL, that is, the gas-liquid interface which is the interface with the atmosphere, pass through the reaction region 16 when the sample solution SL reciprocates, the reaction region 16 is exposed to air. For example, if the measurement conditions vary, such as the reaction region 16 being exposed to air in the inspection of one sample solution SL, or the reaction region 16 not being exposed to air in the inspection of another sample solution SL. It also affects the amount of binding (reaction amount) between the test substance (antigen) A and the second antibody B2 in the sample. If the reaction amount changes due to variations in measurement conditions, the reliability of the measurement results may decrease. When such a situation occurs, it may be preferable to treat it as a liquid feeding abnormality.

Therefore, the control unit 40 controls the gas discharge amount and the suction amount of the pump 22 so that the gas-liquid interface BS of the sample solution SL does not pass through the reaction region 16 when the sample solution SL reciprocates. There is. However, the control of the pump 22 alone may not be sufficient to reliably prevent the gas-liquid interface BS of the sample solution SL from passing through the reaction region 16. This is because the sample solution SL has individual differences such as viscosity differences. If there is such an individual difference, even if the discharge amount and the suction amount of the pump 22 are the same, the gas-liquid interface BS of the sample solution SL may advance beyond the target position or may not reach the target position. there were.

Therefore, the control unit 40 detects that the sample solution SL and the gas-liquid interface BS have passed through at least one preset position in the flow path 15, so that the reaction region 16 is exposed to air. It is determined whether or not an abnormality has occurred in the delivery of the sample solution SL. In this example, the pressure gauge 23 is used to detect the passage of the gas-liquid interface BS at a predetermined position.

As shown in FIG. 9, in the flow path 15, a certain section including a section in which the reaction region 16 is provided is narrower than other sections, and specifically, a certain section including the reaction region 16. The cross-sectional area S1 of the cross section orthogonal to the flow direction (X direction) of the sample solution SL is smaller than the cross-sectional area S2 at both ends thereof. Here, the cross-sectional area changing portions 15A and 15B whose cross-sectional area changes are designated as the first position and the second position, respectively. The first position is a position located upstream of the reaction region 16 in the flow path 15 with respect to the first direction, and the second position is a position located downstream of the reaction region 16. is there. In this example, the first and second positions correspond to at least one position preset in the flow path 15 as a detection position for detecting the passage of the gas-liquid interface when determining a liquid feeding abnormality. To do.

When the gas-liquid interface BS located at the front end or the rear end of the sample solution SL in the flow path 15 passes through the cross-sectional area changing portions 15A and 15B, a pressure fluctuation occurs in the flow path 15 at the time of passage. As described above, the pressure gauge 23 is connected to the branch path of the pipe 26 between the flow path 15 and the pump 22. The pressure gauge 23 corresponds to a pressure sensor for detecting such pressure fluctuations in the flow path 15.

The pressure fluctuation when the gas-liquid interface BS of the sample solution SL passes through the cross-sectional area change portions 15A and 15B is a spike-like pressure fluctuation such that the pressure rises sharply at the time of passing and falls immediately after passing. Appears as. A slight pressure change occurs when the sample solution SL flows through the flow path 15 or when the flow direction is reversed, but the spike-like pressure fluctuation when the gas-liquid interface BS passes through the cross-sectional area change portions 15A and 15B Does not occur.

The control unit 40 acquires the pressure value measured by the pressure gauge 23 in real time. The control unit 40 monitors the pressure value output by the pressure gauge 23, and detects that the gas-liquid interface BS has passed the first position or the second position when a spike-like pressure fluctuation occurs. In this way, the control unit 40 determines whether or not the liquid feeding abnormality of the sample solution SL has occurred based on the pressure value which is the output of the pressure gauge 23. That is, the control unit 40 functions as a liquid feeding abnormality determination unit. The pressure gauge 23 corresponds to a gas-liquid interface detection sensor used for detecting that the gas-liquid interface BS has passed the first position or the second position.

In addition, the control unit 40 functions as a warning unit that warns when it is determined that a liquid feeding abnormality has occurred while the sample solution SL is being reciprocated. Specifically, a message indicating that a liquid transfer abnormality has occurred in which the gas-liquid interface BS passes through the first position or the second position is displayed on the display unit 52. The warning may be given by voice.

Hereinafter, the operation of the above configuration will be described with reference to the flowchart shown in FIG. 10 and the state transition diagram of the sample solution SL during the reciprocating operation shown in FIGS. 11 and 12. FIG. 11 shows an example of a normal pattern that is not determined to be a liquid feeding abnormality, and FIG. 12 shows an example of an abnormal pattern that is determined to be a liquid feeding abnormality.

When performing analysis using the fluorescence detection device 100, first, step (S) 1001 is executed, and the analysis chip 10 and the sample container CB containing the sample are set in the fluorescence detection device 100. The analysis chip 10 is mounted on the mounting portion 101.

Next, sample information such as patient information is input through the operation unit 51, and the control unit 40 receives the input sample information (S1002). The control unit 40 waits for the input of the measurement start instruction from the operation unit 51 (S1003). When the measurement start instruction is input (Y in S1003), the control unit 40 starts the operation of the sample processing unit 20. As shown in FIG. 4, the sample processing unit 20 sucks the sample from the sample container CB with the nozzle tip NC. Then, as shown in FIG. 5, the sucked sample is injected into the sample cell 14A from the nozzle tip NC, and the fluorescent reagent containing the second antibody B2 and the fluorescent label F and the sample solution SL are mixed to prepare the sample solution. Generate SL.

Then, the sample processing unit 20 inserts the tip of the nozzle tip NC into the injection port 12 of the analysis tip 10 and injects the generated sample solution SL into the flow path 15 (S1004). After injecting the sample solution SL into the flow path 15, the sample processing unit 20 operates the pump 22 to start the reciprocating operation of the sample solution SL (S1005). When the control unit 40 starts the reciprocating operation, the control unit 40 operates a timer to start timing. Then, the control unit 40 starts monitoring whether or not the gas-liquid interface BS of the sample solution SL has passed the first position or the second position based on the pressure value from the pressure gauge 23 (S1006).

Here, if the liquid feeding abnormality does not occur, the sample solution SL is reciprocated as shown in FIG. In FIG. 11A, the sample solution SL is injected into the flow path 15, and the sample solution SL flows from the upstream side to the downstream side of the reaction region 16 by the discharge operation of the pump 22, passes through the second position, and further. It shows a state of reaching near the discharge port 13.

In FIG. 11A, the graph showing the time change of the pressure value output by the pressure gauge 23 shows the time change after the gas-liquid interface BS is in the state shown in FIG. 11A, and the sample solution SL is injected into the flow path 15. The pressure value at the time is not shown. In this state, the pressure value is stable and spike-like pressure fluctuation does not occur.

When the suction operation of the pump 22 is performed from the state shown in FIG. 11A, the sample solution SL is sucked into the nozzle tip NC, and the sample solution SL flows in the second direction. As a result, the gas-liquid interface BS near the discharge port 13 approaches the second position. As shown in FIG. 11C, when the pump 22 starts the discharge operation and the flow direction of the sample solution SL is reversed to the first direction before the gas-liquid interface BS reaches the second position, the gas-liquid interface BS is again generated. Heads to the outlet 13. As shown in FIG. 11, as long as the sample solution SL reciprocates, the gas-liquid interface BS does not pass through the second position or the first position.

In this case, as the graphs of the pressure values in FIGS. 11B and 11C show, spike-like pressure fluctuation does not occur. Therefore, the control unit 40 determines in FIG. 10 that the gas-liquid interface BS has not passed through the first position or the second position (N in S1006), and proceeds to S1007. In S1007, if a predetermined time has not elapsed since the reciprocating operation was started, the process returns to step S1006 again.

On the other hand, in the pattern shown in FIG. 12, the control unit 40 performs error processing (S1008). In FIG. 12, the state shown in FIG. 12A is the same as that in FIG. 11A. From this state, as shown in FIG. 12B, when the pump 22 starts the suction operation, the sample solution SL flows in the second direction. For example, if the viscosity of the sample solution SL in the case of FIG. 12 is lower than that in the case of FIG. 11, the gas-liquid interface BS of the sample solution SL is as shown in FIG. 12B even if the suction amount of the pump 22 is the same. , Beyond the second position and approach the reaction region 16. Alternatively, it reaches the first position beyond the reaction region 16 and the reaction region 16.

Then, as shown in FIG. 12C, when the operation of the pump 22 is switched to the discharge operation, the flow direction of the sample solution SL is reversed from the second direction to the first direction, and the gas-liquid interface BS again moves to the first position. It passes and heads for the discharge port 13.

In this case, as the graph of the pressure value in FIG. 12B shows, spike-like pressure fluctuation occurs. In FIG. 10, when the control unit 40 detects such spike-like pressure fluctuations based on the pressure value from the pressure gauge 23, it determines that the gas-liquid interface BS has passed the first position or the second position (S1006). And Y). In this case, the control unit 40 determines that a liquid feeding abnormality has occurred, and proceeds to the error processing of S1008. In S1008, the control unit 40 outputs a message to the effect that a liquid feeding abnormality has occurred to the display unit 52 to warn. Then, when such a liquid feeding abnormality occurs, the control unit 40 stops the measurement because it causes the measurement conditions to vary between inspections.

Further, the control unit 40 ends the reciprocating operation of the sample solution SL when a predetermined time elapses (Y in S1007) without causing a liquid feeding abnormality (S1009). By reciprocating the sample solution SL with respect to the reaction region 16 in this way, the reaction amount of the test substance (antigen) A and the first antibody B1 can be increased with a small amount of the sample solution SL.

The control unit 40 adjusts the incident angle of the light irradiation unit 31 after completing the reciprocating operation of the sample solution SL (S1010). The control unit 40 adjusts the incident angle θ of the light irradiation unit 31 to a preset reference angle. Then, in a state where the excitation light L is irradiated from the light irradiation unit 31, the incident angle θ is changed in the plus direction and the minus direction within a predetermined range with reference to the reference angle. During this time, the fluorescence detection unit 32 detects fluorescence according to the amount of reaction. The control unit 40 identifies the resonance angle at which the plasmon increasing intensity is maximized based on the fluorescence detection signal detected by the fluorescence detection unit 32.

After the incident angle adjustment (S1010) is completed, the measurement is performed (S1011). The light irradiation unit 31 irradiates the excitation light L at a specified resonance angle for a predetermined time, during which the fluorescence detection unit 32 detects fluorescence. Such fluorescence detection is performed for the test region TR and the control region CR while moving the measuring unit 30.

The control unit 40 analyzes the fluorescence detection signal detected by the fluorescence detection unit 32 in each of the test region TR and the control region CR. Then, the reaction amount of the test substance (antigen) A and the second antibody B2 is calculated, the concentration of the test substance (antigen) A in the sample is measured, and this is output as the measurement result (S1012). The measurement result is displayed on the display unit 52 and recorded as data in the memory. Further, the measurement result may be printed out.

As described above, in this example, since a warning is given when a liquid feeding abnormality that causes variation in measurement conditions occurs, it is possible to notify the user that a liquid feeding abnormality has occurred. As a result, the measurement result in which the measurement condition varies is excluded from the evaluation target, so that the measurement condition can be made uniform among a plurality of inspections, and as a result, the reliability of the measurement result is ensured.

Further, in this example, a pressure gauge 23 (pressure sensor) is used as the gas-liquid interface detection sensor. The pressure gauge 23 is a standard equipment for the fluorescence detection device 100 that utilizes the analysis chip 10 having the flow path 15. In this example, since the gas-liquid interface can be detected by using such standard equipment, the increase in cost can be suppressed.

[Modification 1-1]
In the above example, when the gas-liquid interface BS passes through the first position or the second position even once when determining the liquid feeding abnormality, it is determined that the liquid feeding abnormality is performed, but the number of passages is preset. If the number of times is exceeded, it may be determined that the liquid supply is abnormal. That is, the gas-liquid interface BS may be allowed to pass through the first position or the second position up to a preset number of times.

[Modification 1-2]
In the above example, a warning that a liquid feeding abnormality has occurred is given and the measurement is also stopped. However, only the warning is given and the measurement does not have to be stopped. In this case, it is preferable to include a record that a liquid feeding abnormality has occurred in the measurement result. In this way, although the measurement itself is performed, the user sees the result of the liquid feeding abnormality and excludes the measurement result from the evaluation target, or takes into account that the liquid feeding abnormality has occurred. The measurement result can be evaluated.

[Second Embodiment]
The second embodiment shown in FIG. 13 is an example in which an optical sensor 61 that optically detects that the gas-liquid interface BS has passed through the first position or the second position is used as the gas-liquid interface detection sensor. Since other configurations and processing procedures are the same as those in the first embodiment, the same components are designated by the same reference numerals and the description thereof will be omitted.

As shown in FIG. 13, optical sensors 61 are arranged at the first position and the second position, respectively. The optical sensor 61 is, for example, a transmissive type composed of a light emitting unit 61A and a light receiving unit 61B. The light emitting unit 61A and the light receiving unit 61B are arranged at positions facing each other with the flow path 15 interposed therebetween. The light emitting unit 61A is, for example, a photodiode, and the light receiving unit 61B is, for example, an LED (Light Emitting Diode). The optical sensor 61 is connected to the control unit 40, and its operation is controlled by the control unit 40. The control unit 40 receives the output (light amount signal) of the optical sensor 61, and determines the liquid feeding abnormality based on the light amount signal.

The amount of light I that the light emitted by the light emitting unit 61A passes through the flow path 15 changes depending on whether the sample solution SL is present in the flow path 15 or not. This is due to the difference in refractive index between air and the sample solution SL. Which of the transmitted light amount increases with or without the sample solution SL depends on the difference in the refractive index between the refractive index of the transparent material constituting the flow path 15 and the refractive index of the sample solution SL or air. It depends on the size. In this example, an example will be described in which the amount of light I is larger when the sample solution SL is present in the flow path 15 than when it is not present.

Further, in the second embodiment, the first position or the second position where the optical sensor 61 is arranged is set to a position closer to the reaction region 16 from the portion where the cross-sectional area of the flow path 15 changes. When the optical sensor 61 is used, if the cross-sectional area changing portions 15A and 15B (see FIG. 9) are set to the first position and the second position as in the first embodiment, light scattering becomes large, so that the second As the position for setting the first position or the second position, it is preferable to avoid the cross-sectional area changing portions 15A and 15B (see FIG. 9).

FIG. 13A shows the case where the sample solution SL is present at the first position or the second position where the optical sensor 61 is arranged. FIG. 13B shows a state in which the sample solution SL flows in the second direction and passes through the second position from the state of FIG. 13A. In FIG. 13B, since the gas-liquid interface BS passes through the second position where the optical sensor 61 is arranged, there is no sample solution SL at the second position. Therefore, as shown in the graph of FIG. 13B, the amount of light I detected by the light receiving unit 61B at the second position is lower than that of FIG. 13A. The control unit 40 determines whether or not the gas-liquid interface BS has passed the first position or the second position based on the change in the light intensity signal.

Further, in the second embodiment, the detection positions for detecting the passage through the gas-liquid interface BS are two positions, the first position and the second position, but the detection positions are the first position and the second position. In addition to the position, a third position within the reaction region 16 may be provided. In this case, a position in the reaction region 16 where the metal film 18 is not provided, specifically, a third position is set between the test region TR and the control region CR, and the optical sensor 61 is set at this third position. To place. By arranging the optical sensor 61 at the third position in this way, it may be possible to detect that the gas-liquid interface BS has passed each of the first position, the second position, and the third position. The larger the number of detection positions, the better the detection accuracy of passing through the gas-liquid interface BS.

Further, at least one of the three positions of the first position, the second position, and the third position may be set as the detection position.

[Modification 2-1]
In the above example, the passage of the gas-liquid interface BS on the discharge port 13 side has been described, but as shown in FIG. 14, the passage of the gas-liquid interface BS on the injection port 12 side may be detected. Further, the passage of both the gas-liquid interface BS near the injection port 12 and the gas-liquid interface BS near the injection port 12 may be detected.

[Modification 2-2]
Further, as shown in FIG. 15, the measuring unit 30 may also be used as an optical sensor for detecting whether or not the gas-liquid interface BS has passed through the third position in the reaction region 16. When the measuring unit 30 is used, the amount of transmitted light in the flow path 15 is not detected, but the fluorescence emitted by the fluorescent label F on the metal film 18 constituting the test region TR and the control region CR is detected. Therefore, as a premise that the measurement unit 30 can also be used as an optical sensor, it is necessary that the amount of light of the fluorescence Lf detected by the fluorescence detection unit 32 changes depending on whether the gas-liquid interface BS is in the reaction region 16 or not. is there. By using the measuring unit 30 as an optical sensor, it is possible to detect at a third position in the reaction region 16 without using a dedicated optical sensor.

Further, the measuring unit 30 is used not only as a third optical sensor for detecting the passage of the gas-liquid interface BS in the third position in the reaction region 16, but also in the first position or the second position outside the reaction region 16. It may also be used as an optical sensor for detecting the passage of the liquid interface BS. In this case, a metal film 18 is provided at the first position or the second position so that the measuring unit 30 can detect fluorescence even at the first position or the second position outside the reaction region 16, and the gas-liquid interface BS is provided. It is necessary to provide a dummy area for detecting the passage of.

[Other]
In the example of FIG. 15, the optical sensor 61 and the measuring unit 30 are combined, but the passage of the gas-liquid interface BS may be detected by using only the measuring unit 30 without providing the optical sensor 61. ..

Further, when the optical sensor 61 is used, the first position or the second position, which is the detection position, may be set anywhere within the flow path 15. For example, although the example described in the case where the flow path 15 is provided in a section where the cross-sectional area near the reaction region 16 is narrow, the first is closer to the injection port 12 and the discharge port 13 in the section where the cross-sectional area is relatively wide. A position or a second position may be set. As described above, when the optical sensor 61 is used, there is an advantage that the degree of freedom of arrangement is high as compared with the case where the pressure gauge 23 is used.

[Third Embodiment]
The third embodiment shown in FIG. 16 is a mode in which one optical sensor 61 and a timer are combined to detect a liquid feeding abnormality. As the timer, for example, a timer built in the control unit 40 is used. In the third embodiment, the optical sensor 61 is provided at a second position set near, for example, the discharge port 13.

In the third embodiment, in the control unit 40, first, as shown in FIG. 16A, the sample solution SL flows in the second direction, and the gas-liquid interface BS passes through the second position toward the reaction region 16. Is detected. Then, the control unit 40 starts timing the built-in timer when the gas-liquid interface BS passes through the second position, and as shown in FIG. 16B, the sample solution SL flows in the second direction, and then FIG. 16C. As shown in the above, the reciprocating time until it is detected that the flow direction of the sample solution SL is reversed in the first direction and the gas-liquid interface BS has passed through the same second position again is measured.

The control unit 40 determines that a liquid feeding abnormality has occurred when the measured round-trip time exceeds a preset scheduled time. For example, in FIG. 16B, when the sample solution SL is flowing in the second direction, the gas-liquid interface BS reaches the vicinity of the injection port 12 beyond the reaction region 16 and the flow direction is reversed in the first direction from there. Then, the round-trip time until the gas-liquid interface BS returns to the position shown in FIG. 16C is compared with the round-trip time until the gas-liquid interface BS reverses the flow direction in front of the reaction region 16 and returns to the position shown in FIG. 16C. In this case, it is considered that the round trip time is longer when the vehicle reaches the injection port 12. Therefore, when the round-trip time exceeds the scheduled time, the control unit 40 determines that a liquid feeding abnormality has occurred and warns.

In this way, the liquid feeding abnormality may be determined based on the reciprocating time of the sample solution SL from passing through the predetermined detection position to returning to the same detection position. According to this method, the optical sensor can be arranged at a position away from the reaction region 16. It may be difficult to place the optical sensor near the reaction region 16. This example is effective in such a case.

Although the preferred embodiments of the present invention have been described above, the present invention is not limited to the above embodiments, and various aspects such as combinations of the respective embodiments and the respective modifications are not deviated from the gist of the present invention. Of course, the configuration can be adopted.

In each of the above embodiments, for example, the hardware structure of the processing unit that executes the processing of the electric signal such as the control unit 40 is various processors as shown below.

Various processors include a CPU, a programmable logic device (PLD), a dedicated electric circuit, and the like. As is well known, a CPU is a general-purpose processor that executes software (program) and functions as various processing units. PLD is a processor such as FPGA (Field Programmable Gate Array) whose circuit configuration can be changed after manufacturing. A dedicated electric circuit is a processor having a circuit configuration specially designed for executing a specific process such as an ASIC (Application Specific Integrated Circuit).

One processing unit may be composed of one of these various processors, or may be composed of a combination of two or more processors of the same type or different types (for example, a plurality of FPGAs or a combination of a CPU and an FPGA). May be done. Further, a plurality of processing units may be configured by one processor. As an example of configuring a plurality of processing units with one processor, first, there is a form in which one processor is configured by a combination of one or more CPUs and software, and this processor functions as a plurality of processing units. .. Secondly, as typified by System On Chip (SoC) and the like, there is a form in which a processor that realizes the functions of the entire system including a plurality of processing units with one IC chip is used. As described above, the various processing units are configured by using one or more of the above-mentioned various processors as a hardware-like structure.

Further, the hardware structure of these various processors is, more specifically, an electric circuit (circuitry) in which circuit elements such as semiconductor elements are combined.

In addition to the above, it goes without saying that various improvements and modifications may be made without departing from the gist of the present invention.

11 Main body 11A Prism 12 Injection port 13 Discharge port 14A, 14B Sample cell 15 Flow path 15A, 15B Cross-sectional area change part 16 Reaction area 17 Dielectric plate 17A Front side surface 17B Back side surface 18 Metal film 20 Sample processing unit 21 Nozzle movement Mechanism 22 Pump 23 Pressure gauge 26 Piping 30 Measuring unit 31 Light irradiation unit 32 Fluorescence detection unit 33 Incident angle adjustment mechanism 36 Measuring unit movement mechanism 40 Control unit 51 Operation unit 52 Display unit 61 Light sensor 61A Light emitting unit 61B Fluorescence detection unit 100 Device 101 Mounting part A Test substance (antigen)
B1 1st antibody B2 2nd antibody BS Gas-liquid interface CB Specimen container CR Control area Ew Evanescent wave F Fluorescent label I Light amount L Excitation light Lf Fluorescent NC Nozzle tip RL Reflected light S1, S2 Cross-sectional area SL Specimen solution TR Test area θ Incident Horn

Claims (12)

An analysis chip having a flow path through which a sample solution containing a test substance flows and a reaction region in which an antibody that specifically reacts with the test substance is fixed and a reaction region in contact with the sample solution is arranged is used. In a fluorescence detection device using surface plasmon resonance that detects fluorescence generated from a fluorescence label bound to the test substance.
A pump that discharges and sucks the sample solution and reciprocates the sample solution in the flow path in the second direction opposite to the first direction.
A pump control unit that controls the discharge amount and suction amount of the pump, and reciprocates the sample solution in a state where the sample solution and the reaction region are in contact with each other through the control of the discharge amount and the suction amount. Pump control unit and
A gas-liquid interface detection sensor used to detect that a gas-liquid interface, which is an interface between a sample solution and a gas, has passed through at least one preset position in the flow path.
Based on the output of the gas-liquid interface detection sensor, a liquid feed abnormality determination unit that determines whether or not a liquid feed abnormality has occurred in the sample solution, and a liquid feed abnormality determination unit.
It is equipped with a warning unit that warns when it is determined that the liquid supply abnormality has occurred .
The flow path has a cross-sectional area changing portion in which the cross-sectional area of the cross section orthogonal to the flow direction of the sample solution changes.
The gas-liquid interface sensor
The first position located in the cross-sectional area change portion on the upstream side of the reaction region, the second position located in the cross-sectional area change portion on the downstream side of the reaction region, or the third position in the reaction region. A fluorescence detection device using surface plasmon resonance that detects the passage of the gas-liquid interface in at least one of them . The pump discharges and sucks the sample solution through a gas.
The fluorescence detection device using surface plasmon resonance according to claim 1, wherein the pump control unit controls the discharge amount and the suction amount of the gas. The fluorescence detection device using surface plasmon resonance according to claim 1 or 2, wherein the pump control unit reciprocates the sample solution in a state where the sample solution and the reaction region are in constant contact with each other. The gas-liquid interface detection sensor according to any one of claims 1 to 3 , further comprising a pressure sensor for detecting a pressure fluctuation in the flow path generated when the sample solution passes through the cross-sectional area change portion. Fluorescence detection device using surface plasmon resonance. The fluorescence detection device using surface plasmon resonance according to claim 4 , wherein the pressure sensor is connected to a branch path branched from a pipeline between the flow path and the pump. The gas-liquid interface detection sensor from claim 1, including the first position, the light sensor for optically detecting said that the gas-liquid interface at least one position of the second position and the third position has passed A fluorescence detection device using the surface plasmon resonance according to any one of 3 . The optical sensor includes a first optical sensor that detects the passage of the gas-liquid interface at the first position, a second optical sensor that detects the passage of the gas-liquid interface at the second position, and the third position. The fluorescence detection device using surface plasmon resonance according to claim 6 , which comprises at least one of a third photosensor that detects passage. The fluorescence detection device using surface plasmon resonance according to claim 7 , wherein the measuring unit for detecting fluorescence is also used as at least one of the first optical sensor, the second optical sensor, and the third optical sensor. .. When the liquid feed abnormality determination unit detects that the gas-liquid interface has passed at least one of the first position and the second position based on the output of the gas-liquid interface detection sensor, The fluorescence detection device using surface plasmon resonance according to any one of claims 1 to 8 , which determines that the liquid feeding abnormality has occurred. The claim that the liquid feeding abnormality determination unit determines that the liquid feeding abnormality has occurred when it detects that the gas-liquid interface has passed through the third position based on the output of the gas-liquid interface detection sensor. The fluorescence detection device using the surface plasmon resonance according to any one of Items 1 to 9 . The flow direction of the sample solution is reversed from the time when the gas-liquid interface detection sensor detects that the gas-liquid interface has passed through one of the first position and the second position toward the reaction region. Then, it is equipped with a timer that measures the round-trip time until it detects that the gas-liquid interface has passed through the same position again.
The surface plasmon resonance according to any one of claims 1 to 9 , wherein the liquid feeding abnormality determining unit determines that the liquid feeding abnormality has occurred when the reciprocating time exceeds a preset scheduled time. Fluorescence detection device used. An analysis chip having a flow path through which a sample solution containing a test substance flows and a reaction region in which an antibody that specifically reacts with the test substance is fixed and a reaction region in contact with the sample solution is arranged is used. In the method of operating a fluorescence detection device using surface plasmon resonance that detects fluorescence generated from a fluorescence label bound to the test substance,
A pump control step for controlling the discharge amount and suction amount of a pump that discharges and sucks the sample solution and reciprocates the sample solution in the first direction and the second direction opposite to the first direction in the flow path. A pump control step of reciprocating the sample solution in a state where the sample solution and the reaction region are in contact with each other through control of the discharge amount and the suction amount.
A gas-liquid interface detection step for detecting that a gas-liquid interface, which is an interface between the sample solution and a gas, has passed at at least one preset position in the flow path,
A liquid feeding abnormality determination step for determining whether or not a liquid feeding abnormality has occurred in the sample solution based on the detection of passage through the gas-liquid interface, and a liquid feeding abnormality determination step.
It is provided with a warning step to warn when it is determined that the liquid feeding abnormality has occurred .
The flow path has a cross-sectional area changing portion in which the cross-sectional area of the cross section orthogonal to the flow direction of the sample solution changes.
The gas-liquid interface sensor
The first position located in the cross-sectional area change portion on the upstream side of the reaction region, the second position located in the cross-sectional area change portion on the downstream side of the reaction region, or the third position in the reaction region. A method of operating a fluorescence detection device using surface plasmon resonance that detects the passage of the gas-liquid interface in at least one of the above .

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