WO2020090283A1 - Method for recognizing detection abnormality and detection device - Google Patents

Abstract

This method for recognizing a detection abnormality includes: using a pipette tip, which is attached to a pipette nozzle and is for sucking up or discharging a liquid, to move a sample in a reagent accumulation area to a reaction area and perform detection of a substance to be detected in the sample in the reaction area; using the pipette tip to suck up the sample remaining in the sample accumulation area and then measuring the pressure when the sample in the pipette tip is discharged; and assessing the result of the detection by comparing information obtained from the measured pressure and a reference value.

Description

Method and detection device for detecting abnormal detection

The present invention relates to a method and a detection device for detecting a detection abnormality.

In clinical tests and the like, if a minute amount of a substance to be detected such as protein or DNA can be detected with high sensitivity and quantitatively, the condition of the patient can be quickly grasped and treatment can be performed. Therefore, there is a demand for a method capable of detecting a trace amount of a substance to be detected with high sensitivity and quantitatively. In order to detect a trace amount of a substance to be detected with high sensitivity and quantitatively, it is necessary to accurately measure a sample containing the substance to be detected and a reagent used for detection.

In order to accurately measure a liquid such as a sample or a reagent, a method is known in which gas is taken in and out from the tip of the pipette tip to detect the tip position of the pipette tip (for example, see Patent Document 1).

Patent Document 1 discloses a pipette tip when gas is sucked or discharged from the tip of the pipette tip in a state where the tip of the pipette tip is separated from the surface of the liquid stored in the storage portion and a state where these are brought close to each other. A detection method is described in which the pressure inside each is measured and the tip position of the pipette tip with respect to the liquid surface is detected based on the measured pressure difference. In the technique described in Patent Document 1, the pipette tip is moved based on the detected tip position of the pipette tip to suck the liquid. Then, the substance to be detected in the aspirated sample is detected.

International Publication No. 2016/132793

However, in the technique described in Patent Document 1, if the liquid contains bubbles, the upper end of the bubbles may be erroneously detected as the surface of the liquid. As a result, even if the amount of the sucked liquid is extremely smaller than the specified amount, it is erroneously recognized that the specified amount of the sample is being sucked. If the device operates as it is with a very small amount of sample, it may not be possible to obtain a normal signal.

An object of the present invention is to provide a method and a detection device that can appropriately detect the amount of a sample and detect an abnormality in the detection result and can detect an abnormality in the detection.

As one aspect for solving the above-mentioned problems, a method for detecting an abnormal detection of the present invention is such that a sample in a reagent storage area is reacted with a pipette tip for sucking or discharging a liquid attached to a pipette nozzle. The step of detecting the substance to be detected in the specimen in the reaction area while moving to the area, and the pressure when discharging the specimen in the pipette tip after aspirating the specimen remaining in the reagent storage area with the pipette tip The method includes a step of measuring and a step of determining the detection result of the step of detecting by comparing information obtained by the measured pressure and a reference value.

As one aspect for solving the above-mentioned problems, a detection device of the present invention is a reaction having a pipette tip attached to a pipette nozzle, a liquid storage region for storing a sample, and a reaction region for reacting the sample. A pipette including a tip holder for holding the reaction tip and a pipette nozzle to which the pipette tip is attachable and detachable, which is a detection device for detecting a substance to be detected in a sample in the reaction area by using a tip. And an air pressure sensor for measuring the pressure at the time of discharging the sample in the pipette tip after aspirating the sample remaining in the reagent storage area with the pipette tip, and a control for controlling the pipette and the air pressure sensor And a control unit, the control unit compares information obtained by the measured pressure with a reference value to detect an object to be detected. Judges of the detection result.

According to the present invention, the amount of the sample can be appropriately detected and the abnormality in the detection result can be detected, so that the reliability of the detection result can be improved.

FIG. 1 is a schematic diagram of a detection device. FIG. 2 is a schematic diagram of the detection chip. FIG. 3 is a flowchart showing the operation of the detection device. FIG. 4A is a flowchart showing the contents of the process of acquiring the position information, and FIG. 4B is a flowchart showing the contents of the determination of the detection result. FIG. 5 is a schematic graph showing the relationship between the amount of the sample and the pressure when the sample is discharged.

An embodiment of the present invention will be described below.

First, an example of the detection device 100 for implementing the method for detecting a detection abnormality will be described.

FIG. 1 is a diagram showing the configuration of the detection device 100. FIG. 2 is a schematic diagram of the detection chip 200. As shown in FIG. 1, the detection device 100 includes a liquid delivery unit 110, a transport unit 120, a position information acquisition unit 130, a detection unit 140, and a control unit 150. The detection device 100 is used with the detection chip 200 attached.

(Structure of detection chip)
As shown in FIG. 2, the detection chip 200 has a reaction region 210 and a reagent storage region 220. The detection chip 200 may be a reproducible chip or a disposable chip. In the present embodiment, the detection chip 200 is a disposable chip. Examples of the liquid stored in the reagent storage area 220 are labeled with a sample containing a substance to be detected (for example, blood, serum, plasma, urine, nostril, saliva, semen), or a fluorescent substance. A labeling liquid containing a capturing body and a washing liquid are included.

The reaction area 210 is an area where a reaction or the like is performed in order to detect a substance to be detected in a sample. In the reaction region, the flow channel 212 may be arranged and the well may be arranged. In the present embodiment, a flow path 212 is formed in the reaction area 210.

The reagent storage area 220 is an area in which a sample, a labeling liquid, a cleaning liquid, etc. are stored. The reagent storage area 220 has a plurality of recesses 222. The number of recesses 222 is not particularly limited. In the present embodiment, the number of recesses 222 is two. A sample, a labeling liquid, a cleaning liquid, etc. are stored in the recess 222.

(Structure of detector)
Next, each component of the detection device 100 according to the present embodiment will be described. As described above, the detection device 100 includes the liquid delivery unit 110, the transport unit 120, the position information acquisition unit 130, the detection unit 140, and the control unit 150. The detection chip 200 may be held by the chip holder 121 of the transport unit 120.

The liquid sending unit 110 includes a pipette 111, a pipette moving unit 112, and a liquid sending pump drive mechanism 113. The liquid sending unit 110 moves a liquid such as a sample, a labeling liquid, or a washing liquid stored in the recess 222 of the reagent storage region 220 of the detection chip 200 held by the chip holder 121 into the flow channel 212 of the reaction region 210. To do. In addition, the liquid sending unit 110 also discharges the liquid from the flow channel 212 and also agitates the liquid in the flow channel 212. The liquid sending unit 110 is used with the pipette nozzle 170 of the pipette 111 mounted with the pipette tip 170. As the pipette tip 170, any commercially available pipette tip can be used. From the viewpoint of preventing impurities from mixing in, it is preferable that the pipette tip 170 be replaceable.

The pipette 111 sucks and discharges the liquid when injecting the liquid into the flow channel 212 or removing the liquid from the flow channel 212. The pipette 111 includes a syringe 114, a plunger 115 capable of reciprocating in the syringe 114, and a pipette nozzle 116 connected to the syringe 114. Further, the pipette 111 can quantitatively suction and discharge the liquid by the reciprocating motion of the plunger 115. As a result, the pipette 111 can inject the liquid into the channel 212 or remove the liquid from the channel 212. Further, the pipette 111 can agitate the liquid in the flow path 212 by repeating suction and discharge of the liquid.

The pipette moving unit 112 moves the pipette nozzle 116 for sucking the liquid into the pipette tip 170 or discharging the liquid from the pipette tip 170. The pipette moving unit 112 freely moves, for example, the pipette nozzle 116 in the axial direction (for example, the vertical direction) of the pipette nozzle 116. The pipette moving unit 112 includes, for example, a solenoid actuator and a stepping motor.

The liquid feed pump drive mechanism 113 moves the plunger 115 to suck an external liquid into the pipette tip 170 or discharge the liquid in the pipette tip 170 to the outside. The liquid feed pump drive mechanism 113 includes a device such as a stepping motor for reciprocating the plunger 115. The stepping motor is preferable from the viewpoint of managing the amount of residual liquid in the reagent storage area 220 because it can control the amount of liquid delivered by the pipette 111 and the liquid delivery speed.

As described above, the liquid delivery unit 110 sucks various liquids from the recess 222 and injects them into the flow path 212 of the detection chip 200. At this time, the reciprocating operation of the plunger 115 with respect to the syringe 114 is repeated while the tip of the pipette tip 170 is close to the bottom surface of the flow channel 212 in the flow channel 212, so that the liquid in the flow channel 212 in the detection chip 200 is increased. Is stirred. As a result, it is possible to make the liquid concentration uniform and promote the reaction in the flow channel 212.

The transport unit 120 transports the detection chip 200 to the detection position or the liquid feeding position and holds the detection chip 200. Here, the “detection position” is the position where the detection unit 140 detects the substance to be detected in the sample. Further, the “liquid feeding position” is a position where the liquid feeding unit 110 injects the liquid into the channel 212 of the detection chip 200 or removes the liquid from the channel 212 of the detection chip 200. The transfer unit 120 includes a chip holder 121 and a transfer stage 122.

The chip holder 121 is fixed to the transfer stage 122 and detachably holds the detection chip 200. The shape of the chip holder 121 is not particularly limited as long as it can hold the detection chip 200 and does not interfere with the detection of the substance to be detected. In the present embodiment, the shape of the chip holder 121 is configured so that the detection chip 200 can be held from the side direction.

The transfer stage 122 moves the chip holder 121 in one direction and the opposite direction (left and right direction on the paper surface of FIG. 1). The transport stage 122 also has a shape that does not interfere with the detection of the substance to be detected. The transfer stage 122 is driven by, for example, a stepping motor or the like.

The position information acquisition unit 130 acquires position information regarding the position of the tip of the pipette tip 170 with respect to the liquid surface (hereinafter, also simply referred to as “position information”). The position information acquisition unit 130 includes an air pressure sensor 131. The air pressure sensor 131 is connected between the pipette nozzle 116 and the syringe 114. The type of the air pressure sensor 131 is not particularly limited as long as the air pressure (pressure) inside the pipette tip 170 can be measured. Examples of types of the air pressure sensor 131 include a mechanical sensor that uses a Bourdon tube, an electronic sensor that uses a semiconductor, and the like.

The detection unit 140 detects the substance to be detected in the flow path 212 of the reaction region 210. The configuration of the detection unit 140 is not particularly limited as long as the substance to be detected can be detected. The configuration of the detection unit 140 is appropriately selected depending on the method of detecting the substance to be detected. For example, when the detection device 100 is a surface plasmon resonance excitation enhanced fluorescence spectroscopy (SPFS) device, the detection unit 140 may be configured to be able to detect the generated plasmon scattered light or fluorescence.

The control unit 150 controls the liquid feed pump drive mechanism 113, the transfer stage 122, the air pressure sensor 131, and the like. The control unit 150 is configured by, for example, a known computer or microcomputer including an arithmetic device, a control device, a storage device, an input device, and an output device.

(Detection operation of the detection device)
Next, the detection operation of the substance to be detected by the detection device 100 will be described. FIG. 3 is a flowchart showing an example of the operation procedure of the detection device 100. FIG. 4A is a flowchart showing the contents of the step of acquiring the position information (step S120 in FIG. 3), and FIG. 4B is the flowchart showing the contents of the step of determining the detection result (step S150 in FIG. 3).

First, prepare for measurement (step S110). Specifically, the detection chip 200 is prepared, and the detection chip 200 is installed in the chip holder 121 at the set position of the detection chip 200. Further, the pipette tip 170 is attached to the tip of the pipette nozzle 116.

Next, position information is acquired (step S120). First, the first pressure inside the pipette tip 170 is measured (step S121). Specifically, the control unit 150 drives the pipette moving unit 112 to move the tip of the pipette tip 170 to just above the liquid surface of the liquid stored in the recess 222. Then, the controller 150 drives the liquid feed pump drive mechanism 113 to advance the plunger 115 with respect to the syringe 114, and continuously ejects air from the tip of the pipette tip 170 while the air pressure sensor 131 pipettes the pipette. The first pressure in the tip 170 is measured.

Next, the second pressure inside the pipette tip 170 is measured (step S122). Specifically, the control unit 150 drives the pipette moving unit 112 to cause the tip of the pipette tip 170 to be closer to the liquid surface side of the liquid stored in the recess 222 than in the process of measuring the first pressure (process S121). Move to. Then, the control unit 150 drives the liquid feed pump drive mechanism 113 to advance the plunger 115 with respect to the syringe 114 and continuously ejects air from the tip of the pipette tip 170, while the air pressure sensor 131 pipettes the pipette. The second pressure in the tip 170 is measured.

Next, the difference between the first pressure and the second pressure is calculated (step S123). Specifically, the control unit 150 obtains the difference between the first pressure and the second pressure by subtracting the second pressure (first pressure) from the first pressure (second pressure). Then, the control unit 150 detects the position of the tip of the pipette tip 170 with respect to the liquid surface or the height of the liquid surface due to the difference between the first pressure and the second pressure. That is, the control unit 150 acquires position information of the tip of the pipette tip 170 with respect to the liquid surface or the height of the liquid surface by the air pressure sensor 131 detecting the pressure. The process of moving the tip of the pipette tip 170 and measuring the second pressure are repeated until the difference between the first pressure and the second pressure becomes equal to or greater than a predetermined threshold.

In the step of acquiring the position information (step S120), air is continuously or intermittently ejected from the tip of the pipette tip 170 and the tip of the pipette tip 170 is brought close to the liquid surface while the air pressure sensor 131 is used. The pressure within 170 may be measured. In this case, the pressure before moving the pipette tip 170 is the first pressure. Further, the pressure inside the pipette tip 170 measured by the air pressure sensor 131 while the tip of the pipette tip 170 approaches the liquid surface becomes the second pressure.

Next, the substance to be detected in the sample is detected (step S130). In the present embodiment, the sample and the reagent are mixed to detect the substance to be detected contained in the sample. The control unit 150 operates the transport stage 122 to move the recess 222 in which the sample is stored, to directly below the pipette tip 170. Then, based on the acquired position information, the tip of the pipette tip 170 is moved toward the recess 222 in which the specimen is stored, and the specimen is sucked into the pipette tip 170. Then, the control unit 150 drives the pipette moving unit 112 to move the tip of the pipette tip 170 into the channel 212 and inject the sample into the channel 212.

At this time, as shown in FIG. 2, when the bubble B exists on the surface of the sample, the control unit 150 recognizes the upper end of the bubble B as the liquid surface. In this case, since the tip of the pipette tip 170 is not sufficiently moved into the recess 222, it may not be possible to inhale the sample amount required for detection. Then, if the substance to be detected is detected while the amount of the sample is small, an inaccurate detection result will be obtained. Therefore, in the present embodiment, by measuring the amount of the sample in the reagent storage area 220 after the detection step, it is determined whether or not the detection step is properly performed.

Further, the control unit 150 operates the transfer stage 122 to move the recess 222 in which the reagent is stored, directly below the pipette tip 170. Then, based on the acquired position information, the tip of the pipette tip 170 is moved toward the recess 222 in which the reagent is stored, and the reagent is sucked into the pipette tip 170. Then, the control unit 150 drives the pipette moving unit 112 to move the tip of the pipette tip 170 into the channel 212 and inject the reagent into the channel 212.

Next, the detection unit 140 detects the substance to be detected. Specifically, the controller 150 operates the transfer stage 122 to move the detection chip 200 to the detection position. Then, the detection unit 140 detects the substance to be detected in the sample.

Note that the sample may be stored in another container. In this case, the detection chip 200 has an accommodation hole for accommodating the container.

Also, the types of sample and substance to be detected are not particularly limited. Examples of specimens include body fluids such as blood, serum, plasma, urine, nasal fluid, saliva, semen, and diluted solutions thereof. In addition, examples of the substance to be detected include nucleic acids (such as DNA and RNA), proteins (such as polypeptides and oligopeptides), amino acids, sugars, lipids and modified molecules thereof. For example, blood or the like generally has high viscosity, and thus bubbles are likely to occur.

Next, after the sample remaining in the recess 222 is sucked by the pipette tip 170, the pressure when the sample in the pipette tip 170 is discharged is measured (step S140). Specifically, the control unit 150 operates the transport stage 122 to move the flow path 212 in which the sample is stored, directly below the pipette tip 170. Then, the tip of the pipette tip 170 is moved to the bottom of the flow path 212 in which the sample is stored, and all the sample remaining in the flow path 212 is sucked into the pipette tip 170. The height of the bottom of the flow channel 212 is obtained in advance.

The control unit 150 operates the transfer stage 122 to move the flow path 212 directly below the pipette tip 170. Then, the sample in the pipette tip 170 is discharged to the flow channel 212. At this time, the control unit 150 measures the air pressure inside the pipette tip 170 using the air pressure sensor 131.

Then, the detection result of the detection step (step S130) is judged by comparing the information obtained by the measured pressure with the reference value (step S150). In the present embodiment, when the measured pressure is equal to or higher than the reference value (step S151; Yes), the detection result of the detection step (step S130) is determined to be “there is no abnormality in detection” (step S152), When the measured pressure is less than the predetermined value (step S152; No), the detection result of the detection step (step S130) is determined to be "abnormal in detection" (step S153).

When the amount of the residual sample estimated based on the measured pressure is equal to or higher than the reference value, the detection result of the detection step (step S130) is determined to be “no abnormality in detection”, and the estimated residual sample When the amount is less than the reference value, the detection result of the detection step (step S130) may be determined to be “abnormal in detection”.

Here, the concept of the default value will be explained. The specified value is determined corresponding to the minimum amount of the sample that should remain in the recess 222 after the detection step (step S130). As the amount of the sample in the pipette tip 170 increases, the pressure for discharging the sample from the pipette tip 170 increases. Therefore, when the determination is made based on the pressure when the sample is discharged from the pipette tip 170, the pressure when the minimum amount of the sample that should remain in the recess 222 is discharged may be set as the specified value. Further, when the determination is made by the amount of the sample estimated from the pressure when the sample is discharged from the pipette tip 170, the minimum amount of the sample that should remain in the recess 222 may be set as the specified value.

FIG. 5 is a schematic graph showing the relationship between the amount of sample and the pressure when the sample is discharged.

In order to obtain the reference value, first, the relationship between the amount of sample in the pipette tip 170 and the pressure when the sample is discharged is obtained. Specifically, for example, 0 μL, 40 μL, 90 μL, 140 μL, and 150 μL are sucked. Next, after suction, all the specimens are discharged into the flow path, and the maximum pressure at the time of discharge is obtained. By repeating this process 6 times, the average value of the pressure at the time of discharge is obtained. Next, the relationship between the amount of the sample discharged to the flow path 212 and the average value of the pressure when the sample is discharged is obtained. As shown in FIG. 5, the amount of the sample discharged to the recess 222 is proportional to the pressure when the sample is discharged.

For example, when there are no bubbles in the sample, the lower limit value of the sample amount for which the detection result of the detection step (step S130) is determined to be “abnormal in detection” may be about 146 μL. Then, in the protocol that most uses the sample, 110 μL of the sample is used, and from the lower limit amount and the used amount, the minimum liquid amount of the remaining sample amount is 36 μL. The reference value of the residual sample amount in the calculation is 36 μL, but 40 μL is set as the reference value including the variation of the liquid amount. The average value (actually measured value) of the pressure when discharging 40 μL of the liquid is about 1.7 kPa. However, if the variation (3SD) of the discharge pressure is taken into consideration, the lower limit value (reference value) of the pressure is 0.67 kPa.

Note that the method of obtaining the residual sample amount from the discharged pressure may be obtained, for example, from the relationship between the amount of the sample discharged to the recess 222 and the pressure when the sample is discharged. Also in this case, it is necessary to consider the variation in discharge pressure (3SD).

(effect)
As described above, according to the present invention, since the presence or absence of the detection result is determined by comparing the information obtained by the pressure and the reference value, it is possible to appropriately detect the abnormality of the detection result.

This application claims the priority right based on Japanese Patent Application No. 2018-205571 filed on October 31, 2018. The contents described in the application specification and drawings are incorporated herein by reference.

The method for detecting abnormality of detection according to the present invention can improve the reliability of the detection result of the substance to be detected, for example. Therefore, it is also expected to contribute to the development, spread, and development of a detection system for a trace amount of a substance to be detected.

100 Detection Device 110 Liquid Delivery Unit 111 Pipette 112 Pipette Moving Unit 113 Liquid Delivery Pump Drive Mechanism 114 Syringe 115 Plunger 116 Pipette Nozzle 120 Transport Unit 121 Chip Holder 122 Transport Stage 130 Position Information Acquisition Unit 131 Air Pressure Sensor 140 Detection Unit 150 Control Unit 170 Pipette Chip 200 Detection Chip 210 Reaction Area 220 Reagent Storage Area 212 Flow Path 222 Recess

Claims (6)

A step of transferring the sample in the reagent storage area to the reaction area using a pipette tip for sucking or discharging the liquid attached to the pipette nozzle, and detecting the substance to be detected in the sample in the reaction area,
After aspirating the sample remaining in the reagent storage area with the pipette tip, measuring the pressure when discharging the sample in the pipette tip,
By comparing the information obtained by the measured pressure and a reference value, the step of determining the detection result of the detecting step,
A method of detecting a detection abnormality, including: In the determining step, when the measured pressure is equal to or higher than a reference value, it is determined that the detection result of the detecting step is "abnormal in detection", and when the measured pressure is less than the reference value. Determines that the detection result of the detecting step is "abnormal in detection",
A method for detecting an abnormality in detection according to claim 1. In the determining step, when the amount of the residual sample estimated based on the measured pressure is equal to or higher than a reference value, the detection result of the detecting step is determined to be “there is no abnormality in detection” and estimated. If the amount of the remaining sample is less than the reference value, the detection result of the detecting step is determined to be “abnormal in detection”,
A method for detecting an abnormality in detection according to claim 1. Using a pipette chip attached to a pipette nozzle and a reaction chip having a reagent storage region for storing a sample and a reaction region for reacting the sample, a substance to be detected in the sample is detected in the reaction region. A detection device for
A chip holder for holding the reaction chip,
A pipette including a pipette nozzle to which the pipette tip is detachable,
After sucking the sample remaining in the reagent storage area with the pipette tip, an air pressure sensor for measuring the pressure when discharging the sample in the pipette tip,
The pipette, and a control unit for controlling the air pressure sensor,
The control unit compares the information obtained by the measured pressure with a reference value to determine the detection result of the substance to be detected,
Detection device. When the measured pressure is equal to or higher than the reference value, the control unit determines that the detection result of the substance to be detected is “no abnormality in detection”, and if the measured pressure is less than the reference value, The detection result of the substance to be detected is determined to be "abnormal in detection",
The detection device according to claim 4. When the amount of the residual sample estimated based on the measured pressure is equal to or higher than the reference value, the control unit determines that the detection result of the substance to be detected is “no abnormality in detection”, and the estimated residual If the amount of the sample is less than the reference value, the detection result of the substance to be detected is determined to be “abnormal in detection”,
The detection device according to claim 4.

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