Liquid sample analysis system and microchip

Abstract

To secure the stability of feeding of a liquid sample introduced through a pipette tip.SOLUTION: A liquid sample analysis system comprises: a pipette tip for drawing in and discharging a liquid sample; a dispensing pipette to the tip of which the pipette tip is attached, and which has a space for accommodating the liquid sample drawn in through the pipette tip; a connection part for holding the dispensing pipette to which the pipette tip is attached; a microchip in which is formed a first flow path for the liquid sample introduced from the dispensing pipette into a measurement part for measuring the characteristic of the liquid sample to be flowed in; and a control unit connected to the dispensing pipette, for controlling the flow rate of the liquid sample drawn in and discharged through the pipette tip, and controlling feeding to the measurement part of the liquid sample introduced through the first flow path.SELECTED DRAWING: Figure 1

Description

Technical Field

[0001] The present invention relates to an analysis system and a microchip for liquid samples.

Background Art

[0002] Conventionally, in the medical field, biochemical field, etc., tests and experiments have been conducted in which a small amount of a liquid sample such as a reagent or a specimen is caused to flow into a microfluidic chip or the like and react (see, for example, Patent Document 1). The liquid sample used for the reaction is introduced into the microfluidic chip using, for example, a dispensing pipette (also simply referred to as a pipette) having a pipette tip attached to its tip for sucking and discharging the liquid sample.

Prior Art Documents

Patent Documents

[0003]

Patent Document 1

Summary of the Invention

Problems to be Solved by the Invention

[0004] By the way, when the amount of the liquid sample used for tests or the like is small, the stability of the liquid delivery of the liquid sample introduced in the form of droplets through the pipette tip becomes a problem. The present invention has been made in view of such circumstances, and an object thereof is to provide a technique for ensuring the stability of the liquid delivery of the liquid sample introduced through the pipette tip.

Means for Solving the Problems

[0005] To achieve the above object, one embodiment of the present invention adopts the following configuration. That is, a liquid sample analysis system according to one embodiment includes a pipette tip that inhales and discharges a liquid sample, a dispensing pipette having a space for accommodating the liquid sample inhaled through the pipette tip and attached to the tip of the pipette tip, a connection part that holds the dispensing pipette to which the pipette tip is attached, a microchip in which a first flow path is formed for flowing the liquid sample introduced from the dispensing pipette into a measurement part for measuring the characteristics of the liquid sample, and a control unit that is connected to the dispensing pipette, controls the flow rate of the liquid sample inhaled and discharged through the pipette tip, and controls the liquid feeding of the liquid sample introduced through the first flow path to the measurement part.

[0006] Here, the microchip may be provided with a filter that maintains the ventilation pressure of the liquid sample fed to the measurement part through the first flow path within a certain range. Further, the microchip includes a ventilation hole connected to the measurement part by a second flow path and a sensor that detects that the liquid sample has reached a predetermined region between the fed liquid sample reaching the measurement part and reaching the ventilation hole. The control unit controls a mechanism for discharging the liquid sample in the dispensing pipette, etc., so as to stop discharging the liquid sample in the dispensing pipette based on a detection signal output from the sensor indicating that the fed liquid sample has reached the predetermined region.

[0007] Another embodiment of the present invention is a microchip including a connection part that holds a dispensing pipette having a space for accommodating the liquid sample inhaled through the pipette tip and attached to the tip of the pipette tip that inhales and discharges the liquid sample, a flow path for flowing into a measurement part for measuring the characteristics of the liquid sample introduced from the dispensing pipette held by the connection part, and a filter that maintains the ventilation pressure of the liquid sample fed to the measurement part through the flow path within a certain range.

[0008] Further, another embodiment of the present invention is that a pipette tip that inhales and discharges a liquid sample is attached to the tip, and a dispensing pi A microchip comprising a connection part for holding a pet, a first flow path for flowing into a measurement part for measuring the characteristics of a liquid sample introduced from a dispensing pipette held by the connection part, a vent hole connected by the measurement part and a second flow path, and a sensor for detecting that the liquid sample that has been fed has reached a predetermined region between the measurement part and the vent hole.

Advantages of the Invention

[0009] According to the present invention, the stability of the liquid sample feeding introduced through the pipette chip can be ensured.

Brief Description of the Drawings

[0010]

Figure 1

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[0011] Hereinafter, with reference to the drawings, one mode for carrying out the invention (hereinafter also referred to as one embodiment or an embodiment) will be described. The configurations of the following embodiments are examples, and the configurations of the microfluidic chip and the analysis system disclosed in the present embodiment can be appropriately changed according to various conditions. The configurations disclosed in the present embodiment are not intended to limit the technical scope of the invention only to those, and can be combined as much as possible unless otherwise specified.

[0012] In addition, the drawings referred to in the following description only schematically show the shape, size, and positional relationship to the extent that the content of the present invention can be understood. That is, the present invention is not limited only to the shape, size, and positional relationship illustrated in each figure. Also, there may be parts where the dimensional relationships and ratios between the drawings are different from each other.

[0013] [Embodiment 1] FIG. 1 is a diagram showing a schematic configuration of an analysis system 1 according to the present embodiment. The analysis system 1 illustrated in FIG. 1 is a system that measures and analyzes the reaction of a small amount of liquid sample such as a reagent or a specimen introduced into a microfluidic chip 30 by a dispensing pipette 20. As shown in FIG. 1, the analysis system 1 includes a control unit 10, a dispensing pipette (simply referred to as a pipette) 2 0, and a microfluidic chip 30. The control unit 10 and the dispensing pipette 20 are connected through a predetermined interface.

[0014] The control unit 10 controls the inhalation and ejection of a small amount of liquid sample such as a reagent or a specimen housed inside the dispensing pipette 20, and also controls the liquid delivery of the liquid sample introduced in the form of droplets to the microfluidic chip 30. Introducing a reagent or the like into the microfluidic chip 30 by the dispensing pipette 20 is also referred to as "spotting". The control unit 10 is, for example, a microcomputer unit provided with a processor such as a CPU (Central Processing Unit), a memory such as a RAM (Random Access Memory), a ROM (Read Only Memory), and a flash memory, and a communication interface. For example, an operating system (OS: Operating System), various programs, various tables, etc. are stored in the memory. In the present embodiment, the liquid delivery in the microfluidic chip 30 will be described as being controlled by the control unit 10 connected to the dispensing pipette 20, but the control unit 10 and the dispensing pipette 20 may be integrally configured. That is, the dispensing pipette 20 includes a microcomputer chip or the like that executes the function of the control unit 10 inside the housing. And the dispensing pipette 20 may be configured to control the liquid delivery of the liquid sample or the like according to a program stored in advance in a memory or the like in the microcomputer chip.

[0015] The control unit 10 according to the present embodiment controls various parameters such as the flow rate, time, and flow rate related to the liquid delivery according to each operation of the suction operation and the ejection operation of the dispensing pipette 20. In the microfluidic chip 30 into which the liquid sample is introduced, it is possible to achieve stabilization of the liquid delivery to the measurement unit 34, which is a region site for analyzing and evaluating the components, properties, etc. of the liquid sample through the flow path 32. In the analysis system 1 according to the present embodiment, even if the liquid sample used for inspection or the like is a small amount (for example, 5 to 20 μL), stable liquid delivery can be provided.

[0016] The dispensing pipette 20 has a pipette body 21 that houses a liquid sample and a pipette tip 22 attached to the tip of the pipette body 21. The pipette tip 22 has a fixing portion 23 that fixes the outer periphery of the inscribed pipette body 21 and a fitting portion 24. The fitting portion 24 is formed with a convex structure such as a return structure protruding outward from the peripheral surface, and is fitted into a fitting structure 31a such as a fitting groove formed on the inner peripheral surface of the end portion side where the pipette tip 22 of the connecting portion 31 is mounted. The dispensing pipette 20 sucks and discharges a predetermined amount (for example, 5 to 20 μL) of the liquid sample accommodated in the pipette body 21 through the pipette tip 22. The connecting portion 31 may be integrated with the microfluidic chip 30 or may be a separate component from the microfluidic chip 30. The connecting portion 31 may be removable from the microfluidic chip 30.

[0017] The dispensing pipette 20 with the pipette tip 22 attached to the tip of the pipette body 21 is held by the microfluidic chip 30 via a cylindrical connecting portion 31 provided on the microfluidic chip 30. The connecting portion 31 is a structure for mounting and holding the dispensing pipette 20 containing the liquid sample on the microfluidic chip 30. On the inner peripheral surface of the end portion side of the connecting portion 31 where the pipette tip 22 is mounted, for example, a fitting structure 31a such as a fitting groove that fits with a return structure (convex structure) protruding outward from the peripheral surface of the fitting portion 24 is provided. As shown in FIG. 1, the tip portion of the fitting portion 24 of the pipette tip 22 held by the microfluidic chip 30 through the connecting portion 31 penetrates through the cylindrical internal space and extends into the region where the liquid sample is introduced.

[0018] By fitting the convex structure provided on the fitting portion 24 and the fitting structure 31a such as the fitting groove provided on the inner peripheral surface of the connecting portion 31, the relative positioning of the tip position of the pipette tip 22 with respect to the microfluidic chip 30 is appropriately adjusted. Also, in the state where the dispensing pipette 20 is attached to the microfluidic chip 30 through the connecting portion 31, as shown in FIG. 1, the fitting struct The outer peripheral surface of the connection part 31 provided with the fitting part 31a is in close contact with the inner peripheral surface of the pipette tip 22 at the part where the convex structure of the fitting part 24 is provided. By fitting the convex structure provided on the fitting part 24 and the fitting structure 31a provided on the inner peripheral surface of the connection part 31, it becomes possible to seal so that no leakage of air or the like occurs when a liquid sample or the like is introduced into the microfluidic chip 30 through the mounted dispensing pipette 20. In the pipette tip 22, a contamination prevention filter 25 may be provided between the part of the fitting part 24 where the convex structure is provided and the tip part. The contamination prevention filter 25 is composed of, for example, a member that allows air to pass through while not allowing a liquid sample to pass through.

[0019] Figure 2 is a diagram for explaining the usage mode of the pipette tip 22. In Figure 2, the usage mode of the microfluidic chip 30 with the specimen collected through the pipette tip 22 is illustrated. As shown in Figure 2, in the pipette tip 22, a contamination prevention filter 25 is provided in the internal space where the aspirated specimen is accommodated.

[0020] As shown in Figure 2(a), before collecting the specimen Z1, the pipette body 21 of the dispensing pipette 20 is attached to the pipette tip 22. Through the fixing part 23 of the pipette tip 22, the outer periphery of the pipette body 21 is fixed to the pipette tip 22 by the inscribed fixing part 23. The dispensing pipette 20 with the pipette tip 22 attached aspirates a predetermined amount (for example, about 5 to 20 μL) of the specimen Z1 accommodated in the container by a suction operation. The aspirated specimen Z1 is accommodated, for example, in the internal space at the tip part of the pipette tip 22 where the tip and the contamination prevention filter 25 are provided (Figure 2(b)).

[0021] As shown in Fig. 2(c), the dispensing pipette 20 that houses the specimen Z1 in the internal space at the tip of the pipette tip 22 is attached to the microfluidic chip 30 via the connection part 31. By fitting the convex structure provided on the fitting part 24 and the fitting structure 31a such as the fitting groove provided on the inner peripheral surface of the connection part 31, the tip position of the pipette tip 22 with respect to the microfluidic chip 30 is positioned at a predetermined position. The outer peripheral surface of the connection part 31 provided with the fitting structure 31a is in close contact with the inner peripheral surface of the pipette tip 22 at the part where the convex structure of the fitting part 24 is provided, and the joint part between the connection part 31 and the pipette tip 22 is sealed.

[0022] When the inspection and measurement of the specimen Z1 using the microfluidic chip 30 are completed, as shown in Fig. 2(d), the pipette body 21 of the dispensing pipette 20 is detached from the attached pipette tip 22. The pipette body 21 is inserted and removed from the fixing part 23 that fixes the inscribed outer periphery, and in the pipette tip 22 fitted to the connection part 31 via the fitting part 24, the fitting state with the microfluidic chip 30 is maintained.

[0023] When the pipette body 21 is inserted and removed from the fixing part 23, there may be a case where the specimen Z1 that was left without being introduced into the microfluidic chip 30 exists in the internal space at the tip part. The specimen Z1 left in the internal space at the tip part may cause backflow or scattering to the outside due to the negative pressure generated during the insertion and removal of the pipette body 21, resulting in environmental pollution to the surroundings. The pipette tip 22 is provided with a contamination prevention filter (contamination prevention filter 25), thereby being able to prevent environmental pollution caused by the negative pressure generated during the insertion and removal of the pipette body 21 of the specimen Z1. The microfluidic chip 30 is discarded together with the pipette tip 22 fitted to the connection part 31.

[0024] In addition, by operating the dispensing pipette 20 with the pipette tip 22 attached, for example, as a pretreatment of the sample, dilution of the sample and mixing with a diluent, extraction of nucleic acid, mixing with a reagent, etc. are possible. These pretreatments can be appropriately combined with a plurality of pretreatments. For example, after aspirating a 10 μL (microliter) sample through the pipette tip 22, the dispensing pipette 20 further aspirates 90 μL of the diluent, so that the mixing of the sample and the diluent by the aspiration operation can be promoted, and the sample can be diluted 10 times with the diluent. Further, the pipette tip 22 may have, for example, a silica membrane that can specifically bind to nucleic acid in the presence of a high salt concentration. By attaching the pipette tip 22 containing the silica membrane to the dispensing pipette 20, the nucleic acid purified from the aspirated sample for measurement can be introduced into the microfluidic chip 30. Examples of the sample for measurement include saliva, blood, nasal discharge, etc. after being treated with a lysis buffer used for nucleic acid extraction. After discharging the above sample from the dispensing pipette 20 into the microfluidic chip 30, the nucleic acid adsorbed on the silica membrane can be washed by aspirating and discharging again a cleaning liquid containing ethanol or the like.

[0025] In addition, the pipette tip 22 may be configured to include, for example, a sponge or the like coated with a reagent. During the inhalation operation and the discharge operation of the dispensing pipette 20 with the pipette tip 22 attached, mixing of the reagent coated on the sponge or the like with the sample becomes possible. For example, a sponge coated with calcium chloride is filled in the tip portion of the pipette tip 22, blood or plasma anticoagulated with sodium citrate is aspirated, and the dispensing pipette 20 is attached to the microfluidic chip 30. By discharging the above blood or plasma aspirated into the pipette tip 22 from the dispensing pipette 20, calcium-added blood or plasma can be introduced into the microfluidic chip 30. In the microfluidic chip 30, for example, by applying a blood coagulation activation reagent such as tissue factor or contact factor activation reagent (kaolin, celite, etc.), it becomes possible to evaluate the blood coagulation time in the microfluidic chip 30.

[0026] As described above, the liquid sample introduced into the microfluidic chip 30 through the pipette tip 22 includes liquid samples such as reagents, specimens, reaction substances to react with the liquid sample, cleaning liquids, and the like. The liquid sample is not particularly limited as long as it can pass through the microfluidic chip 30. For example, liquid samples obtained from a living body such as blood and urine or their dilutions, extracts from a living body such as plants and animals, naturally occurring water such as rivers, seas, and rainfall, cosmetics, cleaning liquids, waste liquids, and the like can be mentioned. The components in the sample are also not particularly limited, and examples include proteins, nucleic acids, low-molecular compounds, sugars, and the like.

[0027] The microfluidic chip 30 is a device in which a groove of a flow path 32 for sending the liquid sample introduced into a predetermined region of the connection portion 31 to the measurement portion 34 is formed. Such a device is created, for example, by bonding a base material having a groove serving as a flow path formed on the surface and a film material or the base material using an adhesive or the like.

[0028] FIG. 3 is a diagram for explaining the microfluidic chip 30 according to the present embodiment. In FIG. 3, a top view of the substantially rectangular microfluidic chip 30 is illustrated. As shown in FIG. 3, a recessed region 31b, which is a depression for arranging the liquid sample introduced in a droplet state, is formed at a portion of the microfluidic chip 30 that faces the tip of the fitting portion 24 of the dispensing pipette 20 held by the connection portion 31. A substantially circular measurement portion 34 is formed on the end side in the longitudinal direction facing the recessed region 31b of the microfluidic chip 30, and a groove of the flow path 32 connecting the recessed region 31b and the measurement portion 34 is formed between the recessed region 31b and the measurement portion 34. The cross-sectional shape of the groove constituting the flow path 32 may be concave, U-shaped, or V-shaped. For example, a reaction substrate or the like that reacts with the liquid sample introduced into the recessed region 31b by the dispensing pipette 20 is applied to the flow path 32.

[0029] The flow path 32 connecting the recessed area 31b and the measurement unit 34 is an elongated flow path in which a plurality of substantially U-shaped flow paths 32 are connected in a top view. For example, it constitutes a heating unit 33 where heating is performed to accelerate the reaction of the liquid sample. Below the flow path 32 where the heating unit 33 is formed, a heating mechanism such as a heater for heating the introduced liquid sample is provided. The liquid sample introduced into the flow path 32 via the heating unit 33 can be heated in multiple stages according to the evaluation of its characteristics and properties.

[0030] In the flow path 32 constituting the heating unit 33 and the measurement unit 34, bubble removal structures 32a and 34a are provided for capturing bubbles generated during the processing of the liquid sample introduced into the microfluidic chip 30. The measurement unit 34 is, for example, an area site for analyzing and evaluating the components, properties, etc. of the liquid sample that reacts with a reaction substrate or the like. The measurement unit 34 may be provided with, for example, a stirrer for stirring the liquid sample fed through the heating unit 33. The operation of the stirrer is controlled, for example, by a stirring mechanism using magnetic force.

[0031] In the microfluidic chip 30, by providing the bubble removal structure 32a in the flow path 32 constituting the heating unit 33, the mixing of bubbles generated during the processing of the liquid sample fed to the measurement unit 34 can be suppressed. Also, by providing the bubble removal structure 34a in the measurement unit 34, for example, the interference of bubbles generated during the stirring process of the liquid sample flowing into the measurement unit 34 with the inspection and evaluation of the liquid sample can be suppressed. In the microfluidic chip 30, by suppressing the mixing of bubbles, the stability of the feeding of the introduced liquid sample can be enhanced.

[0032] For example, when PCR is performed using the microfluidic chip 30, the liquid sample introduced into the flow path 32 is heated within a temperature range of 90 to 98 degrees through the heating unit 33. Similarly, when the Lamp method is applied, the liquid sample introduced into the flow path 32 is heated within a temperature range of 60 to 70 degrees through the heating unit 33. In any case, it is desirable to generate sufficient bubbles by allowing the heated liquid sample to stay in the region where the heating unit 33 is provided for about 3 to 10 minutes. The bubbles generated during the processing of the liquid sample are captured through the bubble removal structure 32a formed in the flow path 32, and the liquid sample with the bubbles removed is introduced into the measurement unit 34. Also, in the measurement unit 34, by providing a bubble removal structure 34a, the bubbles flowing in through the flow path 32 can be captured. As shown in FIG. 3, since the microfluidic chip 30 is composed of an elongated flow path in which a plurality of substantially U-shaped flow paths 32 that meander are connected, the contact area between the flow path wall surface in the heating unit 33 and the introduced liquid sample can be relatively increased, and the degree of bubble generation can be enhanced. Furthermore, in the region of the heating unit 33, by providing a plurality of repeating structures for capturing bubbles, such as the meandering flow path 32, the bubble removal structure 32a, the meandering flow path 32, and the bubble removal structure 32a, the capture rate of bubbles can be improved.

[0033] FIG. 4 is a diagram for explaining the bubble removal structure 32a provided in the flow path 32. FIG. 4(a) illustrates an enlarged top view of a partial region of the flow path 32 in which the bubble removal structure 32a is formed. FIG. 4(b) illustrates a cross-sectional view taken along line B-B# of FIG. 4(a), and FIG. 4(c) illustrates a cross-sectional view taken along line C-C# of FIG. 4(a). As shown in FIG. 4(a), in the region of the heating unit 33, the bubble removal structure 32a is formed in the linear flow path 32 that connects between the meandering flow paths 32 that are substantially U-shaped.

[0034] As shown in FIGS. 4(b) and 4(c), the bubble-removing structure 32a is formed as a convex structure protruding from the surface side (the side where the connecting portion 31 is provided) of the microfluidic chip 30 in the region of the heating portion 33. The protruding height of the convex structure protruding from the surface side of the bubble-removing structure 32a linearly inclines such that the protruding height becomes relatively higher with respect to the flow direction from the upstream side to the downstream side of the flow path 32. By providing such a structure, it becomes possible to enhance the accumulation efficiency of bubbles generated during the heating process.

[0035] Returning to FIG. 3, a cross-sectional view of the measurement portion 34 taken along line A-A# is illustrated in the ejection Z1. As shown in the ejection Z1, the bubble-removing structure 34a is formed at the peripheral edge of the measurement portion 34 as a convex structure protruding from the surface side of the microfluidic chip 30. Inside where the bubble-removing structure 34a is formed, a flat portion 34b, which is a measurement region for measuring characteristics such as the turbidity and absorbance of the introduced liquid sample, is formed. The bubble-removing structure 34a provided in the measurement portion 34 protrudes from the surface side surrounding the flat portion 34b and is formed at the peripheral edge.

[0036] Regarding the removal of bubbles generated during the processing of the liquid sample introduced into the microfluidic chip 30, usage forms in which the microfluidic chip 30 in which the flow path 32 for introducing the liquid sample is formed has an inclination angle with respect to the horizontal plane and a usage form in which it is vertically inverted are assumed. For example, it is an inclined usage in which the short side of the microfluidic chip 30 provided with the connecting portion 31 is downward in the vertical direction and the short side provided with the measurement portion 34 is upward, with an inclination angle with respect to the horizontal plane, and a vertical usage form in which it is vertically inverted. In such forms, the liquid sample introduced into the microfluidic chip 30 is fed through the flow path 32 from the lower side in the vertical direction toward the upper side. The structure for capturing bubbles has a structure different from the form in which the microfluidic chip 30 is horizontally installed as described in FIGS. 3 and 4.

[0037] FIG. 5 is a diagram for explaining a microfluidic chip 30 configured to enable the above-described inclined use and vertical use. In FIG. 5, a form of the microfluidic chip 30 capable of multi-channel (3 channels × 4 blocks) reaction measurement is illustrated. In the microfluidic chip 30 of FIG. 5, a flow path 32 is formed from the recessed region 31b to each measurement unit 34 provided in multiple channels. On each of the lower side (recessed region 31b side) and the upper side (measurement unit 34 side) of the flow path 32, heating units 33a and 33b are provided in two stages. Each of the heating units 33a and 33b enables heating of the liquid sample introduced into the flow path 32 at an individual temperature. In each of the flow path 32 connecting the heating unit 33a and the heating unit 33b and the flow path 32 connecting the heating unit 33b and the multi-channel measurement units 34, a bubble removal structure 32b for capturing bubbles generated during the processing of the introduced liquid sample is provided.

[0038] FIG. 6 is a diagram for explaining a bubble removal structure provided in the microfluidic chip 30 configured to enable the inclined use and vertical use. FIG. 6(a) is an enlarged view of the circled Z3 in FIG. 5, and the bubble removal structure 34c provided in the upper region of each measurement unit of the 3 channels is illustrated. FIG. 6(b) is an enlarged view of the circled Z2 in FIG. 5, and the bubble removal structures 32b and 32c formed in the flow path 32 of the heating unit 33b are illustrated.

[0039] When the microfluidic chip 30 is inclined or vertical, air bubbles will accumulate on the upper side in the vertical direction of the flow path 32 that meanders in a substantially U shape and on the upper side in the vertical direction of the measurement unit 34. Therefore, as shown in Fig. 6(b), in the flow path 32 that meanders in a substantially U shape, by providing a bubble-removing structure 32c that widens the flow path space between the upper and lower flow path walls in the vertical direction, the accumulation efficiency of air bubbles can be increased. The same applies to the bubble-removing structure 32b provided in the flow path 32 connecting the heating unit 33a and the heating unit 33b. The bubble-removing structure 32b can accumulate air bubbles on the upper flow path wall side, for example, by making the upper flow path wall in the vertical direction of the flow path 32 expand upward into a substantially triangular shape. Also, in the measurement unit 34, a bubble-removing structure 34c that accumulates air bubbles on the upper flow path wall side in the vertical direction can be formed by configuring the shape viewed from the surface side of the measurement unit 34 to be substantially droplet-shaped.

[0040] A filter 35 is provided in the measurement unit 34 to maintain the ventilation pressure of the flow path 32 from the recessed area 31b to the measurement unit 34 within a certain pressure range. The filter 35 is provided, for example, at a position in the measurement unit 34 that faces the inlet through which the liquid sample that has passed through the heating unit 33 flows in. Since the filter 35 allows air to pass through but does not allow the liquid sample to pass through, the ventilation pressure between the recessed area 31b and the measurement unit 34 is maintained within a certain range. That is, when the liquid sample reaches the filter 35 provided at the position facing the inlet through which the liquid sample flows in the measurement unit 34, the liquid delivery of the flow path 32 from the recessed area 31b to the measurement unit 34 stops.

[0041] In the form of the microfluidic chip 30 shown in Fig. 6, which enables multi-channel inspection and testing, it is required to uniformly introduce the liquid sample into each of the multiple measurement units provided. For example, the three measurement units 34 of each block arranged on the upper side in the vertical direction, and the lower It is required to uniformly flow a liquid sample into the three measurement units 34 of each block arranged on the side. By providing the filter 35 in the measurement unit 34, the liquid feed can be stopped when the liquid sample reaches the measurement unit 34. That is, when the liquid sample reaches the measurement unit 34 arranged on the lower side in the vertical direction, the liquid feed to the measurement unit can be stopped, so that the liquid sample can be continuously fed in sequence to each non-reached channel connected to the branched path. As a result, even in the microfluidic chip 30 having branched flow paths to a plurality (for example, 10 or more) of measurement units 34, the liquid sample can be uniformly flowed into each measurement unit 34.

[0042] In addition, in FIG. 3 and the like, the configuration includes the filter 35, but instead of the filter 35, a sensor for detecting that the fed liquid sample has reached a predetermined region (for example, the measurement unit 34) may be provided. Examples of such a sensor include a liquid sensor or an electrode sensor for detecting the liquid level, a pressure sensor for detecting the ventilation pressure of the flow path 32 from the recessed region 31b to the measurement unit 34, and the like.

[0043] The control unit 10 of the analysis system 1 detects, for example, that the liquid sample fed through the flow path 32 has reached a predetermined region by using a liquid sensor or an electrode sensor. The control unit 10 may control to stop the liquid feed of the liquid sample based on the reached state indicated by the binary status signal representing the active status or the non-active status output from the liquid sensor or the electrode sensor. For example, when the status signal output from the electrode sensor indicates a non-active status of the non-reached state, the dispensing pipette 20 is controlled to continue the liquid feed, and otherwise, to stop the liquid feed. In the present embodiment, by using a liquid sensor or an electrode sensor, it becomes possible to stop the liquid feed of the fed liquid sample at the target location (the desired location of the flow path 32 in the microfluidic chip 30).

[0044] In addition, the control unit 10 of the analysis system 1 can detect, for example, the ventilation pressure of the flow path 32 from the recessed area 31b to the measurement unit 34 using a pressure sensor, and control the liquid feeding to stop when the detected pressure reaches a certain pressure (threshold value). In the present embodiment, by using a pressure sensor, it is possible to sense a constant pressure increase in the ventilation pressure of the flow path 32 from the recessed area 31b to the measurement unit 34 and stop the liquid feeding to the microfluidic chip 30.

[0045] (Example 1) Next, the results of the liquid feeding test by the analysis system 1 according to the present embodiment will be described. FIG. 7 is a table showing an example of the control sequence for the dispensing pipette 20 during the test, and FIG. 8 is an example of a graph showing the measurement results of absorbance. In the liquid feeding test, a liquid sample was plasma and a diluent, and the above liquid sample was introduced into the microfluidic chip 30 using the dispensing pipette 20 equipped with the pipette tip 22. Then, the liquid feeding of the liquid sample was controlled, and the change in the absorbance of the liquid sample fed to the measurement unit 34 was measured.

[0046] As shown in the table Tb1 of FIG. 7, plasma was aspirated using the dispensing pipette 20 equipped with the pipette tip 22. The aspiration of plasma was operated for 1 second at a flow rate of 3 μL / second to aspirate a flow rate of 3 μL (step1). The aspiration of the diluent was operated for 3 seconds at a flow rate of 50 μL / second to aspirate a flow rate of 150 μL (step2). Then, the dispensing pipette 20 equipped with the pipette tip 22 was fixed to the connection part 31 of the microfluidic chip 30, and the measurement was started.

[0047] After the start of the measurement, the diluted plasma sample was introduced into the recessed area 31b through the pipette tip 22 of the dispensing pipette 20 fixed to the connection part 31, and the liquid feeding to the measurement unit 34 was started through the flow path 32 formed in the microfluidic chip 30. The discharge of the diluted plasma sample was performed at a flow rate of 50 μL / second to discharge the total amount (153 μL) of the flow rate (step3).

[0048] The diluted plasma sample is sent through the flow path 32 from the recessed area 31b, reaches the heating unit 33 with the heating temperature set at 37°C, and is heated. Three minutes after the liquid feeding stop in step 3, the air in the dispensing pipette 20 is discharged through the dispensing pipette. The air was discharged at a flow rate of 50 μL / second with a flow volume of 153 μL (steps 4 and 5). By the discharging operation in step 5, the plasma sample heated by the heating unit 33 provided in the flow path 32 of the microfluidic chip 30 is sent and injected into the measuring unit 34.

[0049] In the measuring unit 34, latex agglutination measurement beads and a stirrer (φ0.5 mm × length 1 mm) are installed. The plasma sample injected into the measuring unit 34 is stirred at 500 rpm by the stirrer. After stirring, the change in absorbance at a wavelength of 600 nm is measured. The change in absorbance is measured, for example, as a change in transmittance using anti-D dimer antibody-immobilized latex beads. The volume of the measuring unit is 203 μL.

[0050] Figure 8 shows an absorbance curve. The change in absorbance is the result of the binding between the plasma sample and the anti-D dimer antibody immobilized on the latex beads, and is exemplified as the pseudo reaction curve Z10. In the graph shown in Figure 8, the vertical axis represents absorbance and the horizontal axis represents time (minutes). As shown in the pseudo reaction curve Z10, the absorbance (transmittance) of the plasma sample by the latex beads changes from around 1.1 to around 1.43 during the 5 minutes from the start of measurement. During the 5 minutes from 5 minutes to 10 minutes, the change in absorbance becomes gentle and transitions from around 1.43 to around 1.5. After 10 minutes, the change in absorbance is a slight increase, and it is confirmed that it transitions around 1.5.

[0051] (Example 2) The microfluidic chip 30 of Example 1 is configured to maintain the ventilation pressure of the flow path 32 from the recessed region 31b to the measurement unit 34 within a certain range by means of a filter 35 provided at a site facing the inlet through which the liquid sample of the measurement unit 34 flows in. In Example 2, the microfluidic chip 30 is provided with an air vent hole 36, which is a vent hole, instead of the filter 35, and a sensor 37 is installed in the flow path 38 between the measurement unit 34 and the air vent hole 36. The sensor 37 is, for example, a liquid sensor or an electrode sensor that detects that the fed liquid sample has reached a predetermined region. Even with such a configuration, the control unit 10 of the analysis system 1 can control to stop the discharge of the liquid sample from the dispensing pipette 20 based on the binary status signal (detection signal) output from the sensor 37. As a result, in the form of Example 2, it is possible to ensure the stability of liquid feeding in the microfluidic chip 30, suppress the member cost on the chip side, and achieve accurate liquid volume control. In the form of the analysis system 1 of Example 2, it is possible to perform sample measurement with improved accuracy.

[0052] FIG. 9 is a diagram showing a schematic configuration of the analysis system 1 according to Example 2, and FIG. 10 is a top view for explaining the microfluidic chip 30 according to Example 2. As described above, in the form of Example 2, the microfluidic chip 30 is provided with an air vent hole 36 instead of the filter 35, and a sensor 37 is installed in the flow path 38 between the measurement unit 34 and the air vent hole 36, which is different. Also, the control unit 10 of the analysis system 1 according to Example 2 is different in that it controls the liquid feeding and stopping of the liquid sample discharged from the dispensing pipette 20 based on the binary status signal output from the sensor 37. Hereinafter, the analysis system 1 according to Example 2 will be mainly described with the differences. Note that the flow path 32 is an example of the "first flow path", and the flow path 38 is an example of the "second flow path". The first flow path may have a storage unit, a pretreatment unit, an observation unit, etc. Also, the second flow path may have a waste liquid storage unit, etc.

[0053] As shown in FIG. 9, a binary status signal output from the sensor 37 is input to the control unit 10. The control unit 10 indicates, for example, that the status signal output from the sensor 37 has not reached the flow path 38 between the measurement unit 34 and the air vent hole 36. In the case of the non-active status, the dispensing pipette 20 is controlled to continue discharging the liquid sample. On the other hand, in the case of the active status where the status signal output from the sensor 37 indicates that the status signal has reached the flow path 38 between the measurement unit 34 and the air vent hole 36, the control unit 10 controls the dispensing pipette 20 to stop discharging the liquid sample and controls to stop the liquid feeding in the microfluidic chip 30.

[0054] As shown in FIGS. 9 to 10, the air vent hole 36 is formed on the longitudinal end side facing the recessed region 31b of the substantially rectangular microfluidic chip 30. The sensor 37 is installed in the flow path 38 between the measurement unit 34 and the air vent hole 36. The liquid sample fed from the dispensing pipette 20 to the microfluidic chip 30 reaches the measurement unit 34 while pressing the air in the flow path 32. The gas such as air present in the flow path 32 pressed by the liquid sample is discharged to the outside of the microfluidic chip 30 through the air vent hole 36. When the discharge of the liquid sample from the dispensing pipette 20 continues, the liquid sample that has reached the measurement unit 34 reaches the installation position of the sensor 37 installed in the flow path 38 between the measurement unit 34 and the air vent hole 36. The sensor 37 detects the liquid sample that has reached the installation position and shifts the status signal from the non-active status to the active status. When the status signal shifts to the active status, the control unit 10 controls the dispensing pipette 20 to stop discharging the liquid sample. In the microfluidic chip 30 according to the second embodiment, the liquid feeding of the liquid sample from the recessed region 31b through the flow path 32 to the measurement unit 34 is stopped.

[0055] Also in the form of Example 2, a liquid feeding test was conducted in the same manner as in Example 1. FIG. 11 is a table showing an example of a control sequence for the dispensing pipette 20 in Example 2. In Example 2, since a sensor 37 for detecting that the liquid sample has reached a predetermined region is provided, latex particles (50 μL) were previously encapsulated in the heating unit 33. Since the latex particles encapsulated in the heating unit 33 are encapsulated in the elongated flow path 32, they stably exist in the flow path without causing movement or scattering during the storage and transportation of the microfluidic chip.

[0056] In the table Tb2 of FIG. 11, the processes of step 1 to step 3 are the same as those in Example 1. That is, plasma (3 μL) was aspirated using the dispensing pipette 20 with the pipette tip 22 attached, then the diluent (150 μL) was aspirated, and the dispensing pipette 20 with the pipette tip 22 attached was fixed to the connection part 31 of the microfluidic chip 30, and the measurement was started. After the start of the measurement, the dispensing pipette 20 introduced the diluted plasma sample into the concave region 31b and started the liquid feeding to the measurement unit 34 through the flow path 32 formed in the microfluidic chip 30. The discharge of the diluted plasma sample was operated for approximately 3 seconds at a flow rate of 50 μL / second to discharge a flow rate of 153 μL.

[0057] The diluted plasma sample was fed through the flow path 32 from the concave region 31b, reached the heating unit 33 whose heating temperature was set to 37°C, and was heated (step 4). At this time, the latex particles encapsulated in the heating unit 33 moved to the measurement unit 34 together with the plasma sample. Then, 3 minutes after the liquid feeding stop in step 3, the diluted plasma sample (153 μL) was discharged through the dispensing pipette 20 (step 5). The plasma sample heated in the heating unit 33 was injected into the measurement unit 34, mixed with the latex particles, and the reaction was started. The sensor 37 installed in the flow path 32 between the measurement unit 34 and the air vent hole 36 detected the plasma sample that had reached the region, and the liquid feeding of the plasma sample in the microfluidic chip 30 was stopped (step 6). In Example 2, the liquid feeding of the liquid sample in the microfluidic chip 30 may be controlled in combination with a pressure sensor, a valve, etc.

[0058] As described above, in the analysis system 1 according to this embodiment, the liquid transfer from the dispensing pipette 20 to which the pipette tip 22 is attached to the microfluidic chip 30 can be controlled using parameters such as flow rate, time, and flow rate. The delivery of the liquid sample that has reached the measurement unit 34 through the channel 32 can be stopped using a binary output signal output from a filter 35 or a sensor 37 such as a liquid sensor or an electrode sensor. In this embodiment, even if the amount of liquid sample used for testing or the like is small, stable delivery of the liquid sample introduced through the channel formed in the microfluidic chip to a predetermined region can be provided.

[0059] Example 3 From the nasal swab, RNA was extracted using an RNA extraction kit (Loopamp viral RNA extraction kit; Sakae An RNA extract was prepared using a Loopamp (Kenkagaku) ​​microfluidic chip. The dispensing pipette 20 aspirated the RNA extract (specimen) through the pipette tip 22, and introduced the aspirated RNA extract into the recessed area 31b of the fitted microfluidic chip 30. The specimen was then sent to the heating section 33 through the flow path 32, and stopped for three minutes. The heating section 33 was heated to 65°C by a heater. After three minutes had elapsed, the dispensing pipette 20 was controlled to start sending the liquid through the microfluidic chip 30. The specimen, which had been heated to a predetermined temperature (65°C) through the heating section 33, passed through an air bubble removal structure 32a provided in the flow path 32, and the specimen from which air bubbles had been removed was introduced into reaction section A (measurement section 34, channel A) and reaction section B (measurement section 34, channel B). A Lamp method influenza test reagent (Loopamp) was used in reaction sections A, B, and C (measurement section 34, channel C). Influenza B virus detection kit (Eiken Chemical Co., Ltd.), A is the measurement, B is the positive control C is the negative control.

[0060] Each of reaction parts A, B, and C is heated to 60 degrees. Further downstream of reaction parts A, B, and C, a flow path 32 connected to a filter 35 continues. The filter 35 allows air to pass through but does not allow liquid to pass through. By liquid feeding through the dispensing pipette 2, the reaction parts A, B, and C are filled with the specimen and reacted with the reagent applied to each reaction part. The turbidity of reaction parts A, B, and C was evaluated using a semiconductor laser with a wavelength of 600 nm. In the case of a positive result, an increase in turbidity is detected for reaction part A and reaction part B.

[0061] (Example 4) Using a DNA extraction kit (DNeasy Blood & Tissue kit: Qiagen) from a blood sample Subsequently, a DNA extraction solution was prepared. In the same manner as in Example 3, the dispensing pipette 20 sucked the DNA extraction solution (specimen) through the pipette tip 22 and introduced the sucked DNA extraction solution into the recessed region 31b of the microfluidic chip 30 to which the pipette tip was fitted. The microfluidic chip 30 used in Example 4 has, for example, the form shown in FIG. 12. By controlling the discharging operation of the dispensing pipette 20, the specimen was fed through the flow path 32 to the heating section 33c and stopped for 3 minutes. The heating section 33c is heated to 95 degrees by a heater. After 3 minutes elapsed, the dispensing pipette 20 was controlled to start the liquid feeding in the microfluidic chip 30. The specimen heated to a predetermined temperature (95 degrees) through the heating section 33c passed through the bubble removal structure 32a provided in the flow path 32, and the specimen with the bubbles removed was stopped at the heating section A (33d). After 1 minute elapsed, the specimen was fed to the heating section B (33e) and stopped for 30 seconds. The heating section B (33e) is heated to 55 degrees by a heater. After 30 seconds elapsed, the specimen was fed to the heating section C (33f) and stopped. The heating section B (33e) is heated to 72 degrees by a heater. Thereafter, the sucking operation of the dispensing pipette 20 was controlled to draw back the specimen that had reached the heating section C (33f) again into the flow path 32 of the heating section A (33d). By appropriately controlling the sucking operation and the discharging operation of the dispensing pipette 20 and repeating the above cycle about 20 to 30 times, the PCR reaction was advanced. The amplification of PCR by each cycle can be monitored as the fluorescence intensity, for example, by a fluorescence detection section 80 provided in the flow path 32 between the heating section A (33d) and the heating section B (33e).

[0062] In addition, for the detection of PCR amplification, for example, as shown in FIG. 13, by using a microfluidic chip 30 incorporating a test strip 83 of a nucleic acid chromatograph, a lateral flow Visual inspection becomes possible. In that case, after performing nucleic acid amplification by PCR using the dispensing pipette 20 as described above, the sample is sent to the reagent mixing section (measurement section 34) using the dispensing pipette 20 and stopped in the reagent mixing section (measurement section 34) for a certain period of time. In the reagent mixing section (measurement section 34), the sample may be stirred through the inserted stirrer 81, or it may be stopped for several minutes to allow natural diffusion of the sample. In the reagent mixing section (measurement section 34), after the sample is mixed with the reagent, it is further sent and reaches the test strip reagent reaction section 82. Thereafter, when the sample reaches the test strip 83 of the nucleic acid chromatograph, the test result can be evaluated using the nucleic acid chromatograph.

[0063] (Modification example) In Embodiment 1, the microfluidic chip 30 was described as an instrument having a substantially rectangular shape and having a groove of a flow path 32 for sending a liquid sample from the concave region 31b of the connection portion 31 for fixing the dispensing pipette 20 to which the pipette chip 22 is attached to the measurement section 34. For example, as disclosed in Japanese Patent Application Laid-Open No. 2021-159011, Japanese Patent Application Laid-Open No. 2019-088332, Japanese Patent Application Laid-Open No. 2018-014966, etc., in PCR, immunoassay, etc., a disk-shaped microchip is used. In the analysis system 1 of the modification example, a disk shape is applied to the shape of the microfluidic chip 30 in which liquid feeding is performed by the dispensing pipette 20.

[0064] FIG. 14 is a diagram for explaining the microfluidic chip 50 according to the modification example. FIG. 14(a) shows a top view of the microfluidic chip 50 according to the modification example as viewed from above, and (b) shows a side view of the microfluidic chip 50 as viewed from the side. As shown in FIGS. 14(a) and 14(b), the microfluidic chip 50 has a disk shape, and a connection portion 51 for fixing the dispensing pipette 20 to which the pipette chip 22 is attached is formed at the center. Below the connection portion 51, for example, a reagent application for reacting with a liquid sample, a plasma separation filter, a nucleic acid extraction filter, etc. can be installed.

[0065] A groove of the flow path 52 that linearly extends in the circumferential direction from the central portion provided with the connection portion 51 is radially formed, and the groove of the flow path 52 is connected to the measurement portion 54 formed at the peripheral portion. Note that the form illustrated in FIG. 8(a) is an example of a form in which grooves of four flow paths 52 that orthogonally cross in a cross shape are formed at the central portion.

[0066] The dispensing pipette 20 to which the pipette tip 22 is attached is attached to the connection portion 51 provided at the central portion of the disk-shaped microfluidic chip 50 in the same manner as in the first embodiment and the second embodiment, and liquid samples are injected into the flow path 52 and stepwise liquid feeding to the measurement portion 54 is performed. Thereafter, the attached dispensing pipette 20 is detached from the connection portion 51, and two-stage liquid feeding becomes possible by advancing the liquid feeding by centrifugation. That is, in the form of the modification, by adopting the disk-shaped microfluidic chip 50, it becomes possible to perform two-stage liquid feeding, that is, liquid feeding by the dispensing pipette 20 in which the flow velocity, time, flow rate, etc. are controlled and liquid feeding by centrifugation. In the form of the modification, control by multi-stage precise liquid feeding becomes possible.

[0067] In the same manner as in the first embodiment and the second embodiment, also in the modification, the analysis system 1 can be combined with a valve, a liquid sensor, an electrode sensor, a pressure sensor, etc. to enable complicated liquid feeding control. In the liquid feeding in the modification, for example, mixing of a sample and a reagent, plasma separation from a blood sample, nucleic acid extraction from a sample as a pretreatment for PCR, etc. are possible. In the case of mixing a sample and a reagent, for example, a reagent is applied to the flow path 52, and the sample is injected up to the portion where the reagent is applied. Then, mixing becomes possible by repeating suction and discharge of the sample injected into the flow path 32 with the dispensing pipette 20. It is also possible to perform pretreatment such as performing plasma separation from whole blood as a pretreatment and then performing liquid feeding by centrifugation.

Description of Reference Numerals

[0068] 1 ··· Analysis system, 10 ··· Control unit, 20 ··· Dispensing pipette, 21 ··· Pipette Body, 22... Pipette tip, 23... Fixing part, 24... Fitting part, 25... Contamination prevention filter, 30, 50... Microfluidic chip, 31, 51... Connection part, 31a... Fitting structure, 31b... Concave region, 32, 38, 52... Flow path, 32a, 32b, 34a, 34c... Bubble removal structure, 33, 33a, 33b, 33c, 33d, 33e, 33f... Heating part, 34, 54... Measuring part, 34b... Flat part, 35... Filter, 36... Air vent hole, 37... Sensor

Claims

1. A pipette tip for sucking and discharging a liquid sample, A dispensing pipette having a space for accommodating the liquid sample sucked through the pipette tip, with the pipette tip attached to its tip, A connection part for holding the dispensing pipette to which the pipette tip is attached, and a microchip in which a first flow path for flowing the liquid sample introduced from the dispensing pipette into a measurement part for measuring the characteristics of the liquid sample is formed, A control unit connected to the dispensing pipette, controlling the flow rate of the liquid sample sucked and discharged through the pipette tip, and controlling the liquid feeding of the introduced liquid sample to the measurement part through the first flow path, A liquid sample analysis system comprising the above.

2. The liquid sample analysis system according to claim 1, wherein the microchip includes a filter for maintaining the ventilation pressure of the liquid sample fed to the measurement part through the first flow path within a certain range.

3. The microchip includes a ventilation hole connected to the measurement part by a second flow path, And a sensor for detecting that the fed liquid sample has reached a predetermined region between the measurement part and the ventilation hole, The control unit controls to stop the discharge of the liquid sample accommodated in the dispensing pipette based on a detection signal output from the sensor indicating that the fed liquid sample has reached the predetermined region. The liquid sample analysis system according to claim 1.

4. A connection part for holding a dispensing pipette having a space for accommodating the liquid sample sucked through the pipette tip, with the pipette tip for sucking and discharging the liquid sample attached to its tip, A flow path for flowing the liquid sample introduced from the dispensing pipette held by the connection part into a measurement part for measuring the characteristics of the liquid sample, A filter for maintaining the ventilation pressure of the liquid sample fed to the measurement part through the flow path within a certain range, A microchip comprising the above.

5. A connection part for holding a dispensing pipette having a space for accommodating the liquid sample sucked through the pipette tip, with the pipette tip for sucking and discharging the liquid sample attached to its tip, A first flow path for flowing the liquid sample introduced from the dispensing pipette held by the connection part into a measurement part for measuring the characteristics of the liquid sample, A ventilation hole connected to the measurement part by a second flow path, A sensor that detects that the supplied liquid sample has reached a predetermined region between the measurement unit and the vent hole; A microchip comprising the same.

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