Reaction measurement device

Abstract

A reaction measurement device of the present invention includes: a reaction chamber for generating a reaction product of a substance immobilized in an array plate as a spot, a substance contained in a sample liquid, and a fluorescent label by supplying, to the array plate, a reagent liquid containing both the sample liquid and the fluorescent label; and a measurement chamber for irradiating the spot with excitation light and measuring the fluorescence emitted from the reaction product. The reaction chamber comprises: a reaction chamber placement part where the array plate is placed; a sample liquid placement part; a reagent liquid placement part; a buffer liquid placement part; an observation liquid placement part; and a liquid supply part which supplies, to the array plate, the sample liquid, the reagent liquid, a buffer liquid, and an observation liquid. The measurement chamber comprises: a measurement chamber placement part where the array plate is placed; and an optical measurement part which irradiates the array plate with excitation light and measures the fluorescence emitted. The observation liquid placement part is thermally coupled to the internal atmosphere of the measurement chamber or to the measurement chamber placement part.

Description

Reaction measurement device

[0001] The present disclosure relates to a reaction measurement device that measures the fluorescence of a reaction product produced on an array plate.

[0002] Array plates such as protein arrays, peptide arrays, and DNA arrays are known, in which a large number of substances such as proteins, peptides, and nucleic acids are fixed as array spots on a substrate. By using an array plate, it is possible to simultaneously observe interactions between the large number of substances fixed on the array plate and substances in a specimen. Therefore, by using an array plate, interactions between the large number of substances fixed as array spots and various substances contained in biological specimens such as blood, cell extracts, saliva, and interstitial fluid can be comprehensively analyzed.

[0003] Patent Document 1 discloses a sample analyzer. Patent Document 1 describes an inspection technology that uses a plate having multiple spots containing biological substances, performs a reaction process between the labeled spots and substances in the sample, and then performs a measurement process in which the pattern of labeled spots on the plate is optically acquired. In Patent Document 1, the surface of the plate having multiple spots is surrounded by a bank, and the reagent inside the bank used in the reaction process is replaced with an observation liquid. Then, while the observation liquid remains inside the bank, excitation light is irradiated from the back side of the plate (the side opposite the surface having the multiple spots), and fluorescence from the labeled spots is observed.

[0004] JP 2023-12426 A

[0005] It is generally known that the fluorescence luminescence efficiency (quantum yield) of fluorescent substances depends on temperature and decreases at high temperatures. Patent Document 1 does not provide any special temperature management for the observation solution, which poses a problem of fluctuations in fluorescence intensity between measurements, for example, when the environmental temperature fluctuates or the array plate temperature differs. The present disclosure aims to provide a reaction measurement device that suppresses fluctuations in the temperature of the measurement target, thereby suppressing fluctuations in fluorescence intensity between measurements.

[0006] The present disclosure relates to a reaction measurement device including a reaction chamber for supplying a sample liquid and a reagent liquid containing a fluorescent label to one surface of an array plate having a plurality of substances fixed as an array of spots on the one surface of a plate substrate, to generate reaction products on the spots between the substances contained in the sample liquid, the fluorescent labels in the reagent liquid, and the substances fixed as spots, and a measurement chamber for measuring fluorescence emitted from the reaction products by irradiating the spots with excitation light, wherein the array of spots is fixed to a plate substrate that is transparent to the excitation light and fluorescence, and the array of spots is fixed to the plate substrate, and the array of spots has a storage section that can store the sample liquid and reagent liquid supplied so as to be in contact with the array of spots, and the reaction chamber includes a reaction chamber mounting section on which the array plate is mounted, a sample liquid mounting section on which the sample liquid to be supplied to the storage section is mounted, a reagent liquid mounting section on which the reagent liquid to be supplied to the storage section is mounted, and a measurement chamber for measuring fluorescence emitted from the storage section. the measurement chamber comprises a measurement chamber mount section on which the array plate transported from the reaction chamber mount section is mounted, and an optical measurement section that measures the emitted fluorescence when excitation light is applied to the surface of the array plate mounted on the measurement chamber mount section opposite to the one surface of the array plate; and the observation liquid mount section is thermally coupled to a heat transfer member that is thermally coupled to at least one of the interior atmosphere of the measurement chamber and the measurement chamber mount section.

[0007] The present disclosure also provides a reaction measurement device including a reaction chamber for supplying a sample liquid and a reagent liquid containing a fluorescent label to one surface of an array plate having a plurality of substances fixed as an array of spots on the one surface of a plate substrate, to generate reaction products on the spots between the substances contained in the sample liquid, the fluorescent labels in the reagent liquid, and the substances fixed as spots, and a measurement chamber for measuring fluorescence emitted from the reaction products by irradiating the spots with excitation light, wherein the array of spots is fixed to a plate substrate that is transparent to the excitation light and fluorescence, and the array of spots is fixed to the plate substrate, and the array of spots has a storage section that can store the sample liquid and reagent liquid supplied so as to be in contact with the array of spots, and the reaction chamber includes a reaction chamber mounting section on which the array plate is mounted, a sample liquid mounting section on which the sample liquid supplied to the storage section is mounted, and a measurement chamber for measuring fluorescence emitted from the reaction products on the spots. the measurement chamber comprises a measurement chamber mount section on which the array plate transported from the reaction chamber mount section is mounted, and an optical measurement section that irradiates the array plate mounted on the measurement chamber mount section with excitation light from a surface opposite to the one surface of the array plate, and measures the emitted fluorescence; and the observation liquid mount section comprises an observation liquid cooling section that cools the observation liquid.

[0008] According to the present disclosure, a reaction measurement device is provided in which the temperature fluctuation is suppressed by thermally managing the optical measurement target, thereby suppressing fluctuations in fluorescence brightness between measurements.

[0009] FIG. 2B is a diagram conceptually illustrating the configuration of a reaction measurement device according to the present disclosure. FIG. 2C is a schematic top view of a reaction measurement device according to Example 1. FIG. 2D is a schematic cross-sectional view showing the 2B-2B cross section of FIG. 2A. FIG. 2E is a diagram illustrating a heat transfer member according to Example 1. FIG. 2F is a schematic top view of a reaction measurement device according to Example 2. FIG. 2G is a diagram illustrating a cooling mechanism for an observation liquid mounting section according to Example 3. FIG. 2H is a diagram illustrating a dew condensation prevention mechanism according to Example 4.

[0010] (Reaction measurement device) Fig. 1 is a diagram conceptually illustrating the configuration of the reaction measurement device of the present disclosure. An overview of the configuration of the reaction measurement device of the present disclosure will be described using Fig. 1. The reaction measurement device of the present disclosure has a reaction chamber 1 and a measurement chamber 3. In the reaction chamber 1, a sample liquid to be tested is supplied to an array plate on which a plurality of substances are fixed, and a reaction operation is performed in which the substances fixed to the array plate react with substances contained in the sample liquid. Meanwhile, in the measurement chamber 3, an optical measurement operation is performed on the array plate on which the reaction operation has been performed to confirm the presence or absence of a reaction product. The reaction chamber 1 includes a reaction chamber mounting section 11 for mounting an array plate, a specimen liquid mounting section 13 for mounting a specimen liquid to be supplied to the array plate, a reagent liquid mounting section 15 for mounting a reagent liquid to be supplied to the array plate together with the specimen liquid during the reaction operation, a buffer liquid mounting section 17 for mounting a buffer liquid to be supplied to the array plate to (intermediately) replace the liquid remaining after the reaction and stabilize the reaction product, an observation liquid mounting section 19 for mounting an observation liquid to (finally) replace the buffer liquid and prepare a test array plate to be subjected to optical measurement, and a liquid supply section 21 that supplies at least one of the specimen liquid, reagent liquid, buffer liquid, and observation liquid to the array plate. The measurement chamber 3 includes a measurement chamber mounting section 31 for mounting a test array plate, and an optical measurement section 33 that irradiates the test array plate with light to perform optical measurement. A passageway 51 is provided between the reaction chamber 1 and the measurement chamber 3, serving as a passageway for moving the test array plate prepared in the reaction chamber to the measurement chamber.

[0011] (Array Plate) An array plate has multiple spots containing various types of biological materials on a substrate and is used for comprehensive analysis of samples. Array plates are also called microchips, microarrays, protein chips, DNA chips, etc. The indication that identifies each spot on the array plate is called an address within the array plate. The address may be determined by the physical location of each spot on the array plate. In this case, it can be determined by the distance from each edge of the array plate or by coordinates.

[0012] The array plate used in the reaction measurement device of the present disclosure typically has a flat substrate (plate base) with a thickness of approximately 0.1 mm to 2.5 mm, on one side of which multiple spots containing various individual substances are fixed in an array. These substances are often biological substances such as proteins, peptides, nucleic acids, cells, and microorganisms. The biological substances referred to here may be naturally occurring, synthetic, or recombinantly produced. More specifically, examples of biological substances include antibodies, antigens, phosphorylated proteins, dephosphorylated proteins, receptors, enzymes, enzyme substrates, kinase substrates, allergens, cytokines, hormones, bacteria, viruses, DNA, RNA, cDNA, cells themselves, cell membrane components, cancer markers, disease markers, low molecular weight compounds derived from living organisms, extracts from living organisms, blood, blood-derived substances, food, food-derived substances, natural products, substances derived from natural products, and culture medium-derived substances. The multiple spots may contain different biological substances or the same biological substance. Alternatively, the device may be configured to have a plurality of spot groups each consisting of a plurality of spots containing the same biological material, with spots belonging to different spot groups containing different proteins.

[0013] The array plate used in the reaction measurement device of the present disclosure has a reservoir that can store sample liquid and a reagent liquid containing a fluorescent label when supplied to a surface on which spots are fixed in an array so that the liquid contacts the spots. When the liquid is stored in the reservoir, a reaction product is generated on each spot due to a reaction between the substance fixed on each spot, a substance contained in the sample liquid, and the fluorescent label in the reagent liquid. The reservoir can be formed, for example, by providing a bank surrounding the area on one side where the spots are fixed in an array. In this case, the liquid supplied to the array plate is stored in the area surrounded by the bank and contacts the spots.

[0014] The array plate used in the reaction measurement device of the present disclosure is used to irradiate excitation light from the side opposite the side on which the spots are fixed in an array, and detect fluorescence emitted from reaction products containing fluorescent labels on the opposite side. Therefore, the plate substrate is made of a material that is transparent to the wavelengths of the excitation light and fluorescence. Examples of such materials include glass, synthetic quartz, quartz, and borosilicate glass. Other examples include resins such as polystyrene, polypropylene, acrylic resin, methacrylic resin, polyamide, polyimide, melamine resin, ABS resin, polyphenylene oxide urethane, silicone, epoxy resin, and polydimethylsiloxane. Alternatively, membranes such as nitrocellulose or polyvinylidene fluoride, or gels such as agarose or acrylamide can also be used.

[0015] (Reaction Chamber Mounting Section) The array plate described above is mounted on the reaction chamber mounting section 11. The reaction chamber mounting section 11 may have a heating section (heater) that heats the array plate to promote the generation of reaction products. For the generation of reaction products simulating reactions in the human body, it is preferable to heat the array plate to a temperature of 30 to 37 degrees Celsius. The heating section may be a known heating means (heat source) incorporated into the mounting section in a known mounting format. Examples of suitable heating means include resistance heating mechanisms such as ribbon heaters, nichrome wires, and panel heaters; semiconductor electrothermal conversion mechanisms including Peltier elements; induction heating mechanisms; and microwave heating mechanisms. Known mounting formats for such heating means include providing a heat transfer member such as a heat spreader or heat pipe between the mounting section and the heating means, or incorporating the heat transfer member into the mounting section. The heating unit (heater) has a heat spreader (not shown) for equalizing the temperature of the array plate and the stored liquid and for homogenizing the reaction on the array plate.

[0016] (Specimen Liquid Placement Section) A specimen liquid placed in a container such as a centrifuge tube is placed on the specimen liquid placement section 13. The specimen liquid is prepared into a liquid form by diluting, suspending, or extracting a specimen, which may be in a solid, liquid, or gaseous state, with water, physiological saline, a buffer solution, or the like as appropriate. The specimen liquid may contain surfactants, preservatives, nonspecific adsorption inhibitors, and other additives. The specimen is, for example, a lysate of biological cells.

[0017] (Reagent Liquid Placement Unit) Reagent liquids placed in containers such as centrifuge tubes are placed on the reagent liquid placement unit 15. Examples of reagent liquids include a reaction liquid that generates a desired reaction product on the spot, a reaction stop liquid that stops the reaction, an antibody liquid that binds a primary antibody to the reaction product, and a secondary antibody liquid that binds a secondary antibody containing a predetermined fluorescent label. The reagent liquid is selected and used so that the substance fixed as a spot on the array plate, the substance to be measured contained in the sample liquid, and the substance contained in the reagent liquid react with each other to generate a reaction product containing a fluorescent label on the spot. Furthermore, the reagent liquid placement unit 15 may be configured to include both or either a reagent liquid placement unit with a refrigeration function to prevent deterioration of reagents that deteriorate at room temperature, and a reagent liquid placement unit without a refrigeration function for reagents that can be stored at room temperature.

[0018] The use of fluorescent labels in the present disclosure offers advantages not found in the use of other labeling substances, such as stable emission, region-specific labeling without the need for substrates, making them suitable for two-dimensional analysis, and the ability to perform multiple labeling using fluorescent substances with different wavelengths. Fluorescent labels are selected appropriately to suit the reaction system and optical system. Fluorescent labels include fluorescein, cyanine, rhodamine, Texas Red, coumarin dyes, fluorescent proteins, quantum dots, and the like. Those skilled in the art can select fluorescent labels with appropriate excitation wavelengths, fluorescence wavelengths, and properties to suit the optical system and experimental system.

[0019] (Buffer Solution Mounting Section) A buffer solution placed in a container such as a centrifuge tube is mounted on the buffer solution mounting section 17. The buffer solution is used to temporarily replace a previous reagent (intermediate replacement) when different reagents are sequentially supplied to the reservoirs on the array plate to prevent components of the previous reagent from contaminating the subsequent reagent. Therefore, it is not preferable for the buffer solution to contain a substance that reacts with the substances fixed as spots on the array plate. Suitable buffer solutions include, for example, Tris-buffered saline (TBST) containing a nonionic surfactant or Tris-buffered saline (TBS).

[0020] (Observation Liquid Placement Unit) An observation liquid placed in a container such as a centrifuge tube is placed on the observation liquid placement unit 19. The observation liquid is supplied to a reservoir on the array plate to replace the reagent liquid or buffer solution supplied to the array plate at the end of the process of generating a reaction product between the substance fixed as the spot, the substance in the sample liquid, and the substance in the reagent liquid, preventing the spot from drying out during fluorescence measurement, as described below. The observation liquid preferably has a refractive index close to that of the plate substrate (e.g., glass) to prevent excitation light incident from the plate substrate from reflecting at the interface between the observation liquid and the plate substrate (the upper surface of the plate substrate). Specifically, the observation liquid is preferably a liquid with a refractive index of 1.3 or higher, more preferably 1.33 to 1.60, and even more preferably 1.40 to 1.46. The observation liquid is similar to a buffer solution in that it replaces the liquid stored in the reservoir and is inactive against the spot array, primary antibodies, and secondary antibodies. In other words, it can be considered a type of buffer solution. The final replacement with the observation liquid ultimately determines the liquid composition of the liquid stored in the reservoir during observation of the array plate (optical measurement step).

[0021] Furthermore, since fluorescence generated by the observation solution in response to excitation light becomes stray light, it is desirable for the observation solution to be fluorescently inert at the wavelength of the excitation light. It is also desirable to be able to prevent oxidation of the spot and reaction products. Furthermore, it is desirable for the observation solution to have a specific heat capacity greater than that of the plate substrate so as to minimize temperature changes on the spot surface during fluorescence measurement. Specifically, for example, glycerol solutions, particularly water containing 40 vol% to 90 vol% glycerol, physiological saline, phosphate buffer, Tris-HCl buffer, etc., are suitable. Furthermore, if the buffer used at the end of the process of generating reaction products between the substance fixed as the spot, the substance in the sample solution, and the substance in the reagent solution can be used as the observation solution, it may be used as is.

[0022] (Liquid Supply Unit) The liquid supply unit 21 dispenses a predetermined amount of at least one of the specimen liquid, reagent liquid, buffer solution, and observation liquid from the liquid placement unit and supplies it to a storage unit on the array plate. The liquid supply unit is configured, for example, by an electric pipette equipped with a pipette tip. The liquid supply unit 21 may also be used to remove (drain) the specimen liquid, reagent liquid, buffer solution, observation liquid, etc. stored in the storage unit on the array plate from the storage unit. Of course, a drainage unit may be provided separately from the liquid supply unit. Such a drainage unit may be configured by an electric pipette, as with the liquid supply unit, but may also be configured by a suction nozzle connected to a pump.

[0023] (Measurement chamber mounting section) An array plate is mounted on the measurement chamber mounting section 31, which has undergone a process of generating reaction products between substances fixed as spots, substances in the sample liquid, and substances in the reagent liquid, and in which observation liquid is stored in the storage section.

[0024] (Optical measurement unit) The optical measurement unit 33 irradiates excitation light from the plate substrate side onto the array plate placed on the measurement chamber mounting unit 31, and measures the fluorescence generated from the reaction products. That is, the optical measurement unit 33 has an irradiation unit that irradiates excitation light and a light receiving unit that receives fluorescence, and is preferably disposed below the measurement chamber mounting unit.

[0025] (Passage Port) The passage port 51 is a window provided in a partition wall provided between the reaction chamber and the measurement chamber. In the reaction measurement device of the present disclosure, the array plate that has undergone the process of generating reaction products between the substances fixed as spots, the substances in the sample liquid, and the substances in the reagent liquid and in which the observation liquid is stored in the storage section is configured to move from the reaction chamber mounting section 11 to the measurement chamber mounting section 31 through the passage port 51. The reaction measurement device of the present disclosure preferably has an inter-chamber transport section that transports the array plate from the reaction chamber mounting section to the measurement chamber mounting section.

[0026] (Heat Transfer Member) The heat transfer member 41 is thermally coupled to the interior atmosphere of the measurement chamber 3 or the measurement chamber mounting portion 31, and also to the observation liquid mounting portion 19 and the buffer solution mounting portion 17. Here, "two members or spaces being thermally coupled" means that when there is a temperature difference between them, heat can be transferred at a sufficient rate from the higher temperature to the lower temperature. The presence of this heat transfer member 41 ensures that the temperatures of the observation liquid mounting portion 19 and the buffer solution mounting portion 17 are always close to the interior atmosphere of the measurement chamber 3 or the temperature of the measurement chamber mounting portion 31. Therefore, when an array plate that has completed the process of generating reaction products in the reaction chamber mounting portion 11 and has the observation liquid stored in its reservoir is moved from the reaction chamber mounting portion 11 to the measurement chamber mounting portion 31, temperature changes are suppressed or mitigated. A more detailed configuration and its effects will be apparent from the following examples.

[0027] Example 1 A reaction measurement device according to this example will be described using Figures 2A and 2B. Figure 2A is a schematic top view, and Figure 2B is a schematic cross-sectional view taken along line 2B-2B. The space below the central dashed line extending horizontally in Figure 2A is the reaction chamber 1. Reaction chamber mounting section 11, which includes plate holder 101 and heater 103 provided on its upper surface, is disposed within reaction chamber 1. Furthermore, refrigerated reagent solution holder 105, which is disposed within reaction chamber 1, holds sample liquid 107, reaction liquid 109, primary antibody liquid 111, and fluorescently labeled secondary antibody liquid 113, each contained in a centrifuge tube. Meanwhile, room-temperature reagent solution holder 115 holds reaction stop solution 117 contained in a centrifuge tube. 1 , the refrigerated reagent solution holder 105, which has a cooling unit, doubles as the specimen solution placement unit 13 and the reagent solution placement unit 15, while the room-temperature reagent solution holder 115, which does not have a cooling unit, is part of the reagent solution placement unit 15. Furthermore, the buffer solution observation solution holder 119 holds a buffer solution 121 and an observation solution 123, each placed in a centrifuge tube. That is, the buffer solution observation solution holder 119 doubles as the buffer solution placement unit 17 and the observation solution placement unit 19 in FIG. 1 . Additionally, a pipette tip rack 125 is disposed within the reaction chamber 1, and pipette tips 127 are placed on it. Furthermore, the reaction chamber 1 is equipped with a liquid supply unit 21 (see FIG. 1 ), which is composed of an X-axis stage 129, a Y-axis stage 131, a Z-axis stage 133, and a pipetter 135. By driving the X-axis stage 129 , the Y-axis stage 131 and the Z-axis stage 133 , the pipetter 135 can be moved three-dimensionally within the reaction chamber 1 .

[0028] The space above the central dashed line extending horizontally in Figure 2A is the measurement chamber 3. A measurement chamber mounting section 31 is disposed within the measurement chamber 3, and an excitation light irradiation section 32, which is part of the optical measurement section 33 (see Figure 1), is disposed below the measurement chamber mounting section 31. The measurement chamber mounting section 31 is movable in the Y and Z directions by a transfer stage 151 and a measurement chamber Z-axis stage 153. The measurement chamber mounting section 31, transfer stage 151, and measurement chamber Z-axis stage 153 constitute an inter-chamber transfer section. In addition, a heat transfer member 161 made of a metal plate that contacts the bottom of the buffer solution observation solution holder 119 disposed within the reaction chamber 1 is provided, extending from the reaction chamber 1 into the atmosphere within the measurement chamber 3.

[0029] Inter-chamber transport and fluorescence measurement will be described with reference to FIG. 2B . A partition 201 is provided between the reaction chamber 1 and the measurement chamber 3 to spatially separate them, and a passage port 51 is formed in the partition 201. A reaction chamber mounting section 11, which is configured by providing a heater 103 on the upper surface of a plate holder 101, is disposed within the reaction chamber 1, and an array plate 203 is mounted thereon, which has a reservoir 206 surrounded by a bank 205 on its upper surface. A base plate 207 connected to the array plate 203 and the bank 205 has an opening corresponding to the shape of the heater 103, so that the heater 103 and the array plate 203 are in close proximity. When the array plate 203 is transported from the reaction chamber mounting section 11 to the measurement chamber mounting section 31, the observation liquid 123 is stored in the reservoir.

[0030] The measurement chamber mounting part 31 has two arms. The transfer stage 151 and the measurement chamber Z-axis stage 153 are driven to insert the arms of the measurement chamber mounting part 31 under the base plate 207. Then, the measurement chamber Z-axis stage 153 is driven to lift and pull out the base plate 207 to a position where it does not interfere with the heater 103, thereby moving the array plate 203 toward the measurement chamber 3. The base plate 207 and the arms of the measurement chamber mounting part 31 may be provided with positioning mating pins and mating holes.

[0031] The excitation light irradiation unit 32 includes a guide 209, a stage 211, a mirror holder 213 incorporating a prism mirror, and a lens holder 215 incorporating an objective lens. The stage 211, mirror holder 213, and lens holder 215 are connected and movable in the X direction along the guide 209. The excitation light propagates in the X direction, is reflected in the Z direction by the prism mirror in the mirror holder 213, and is focused by the objective lens in the lens holder 215. The focused position of the excitation light is configured to coincide with the surface of the array plate where the spot is located. The excitation light irradiation unit 32 scans the excitation light in the X direction and the transport stage 151 scans the array plate 203 in the Y direction, causing the excitation light spot to two-dimensionally scan the surface of the array plate where the spot is located in the X and Y directions. Fluorescence generated from the spot by irradiation with excitation light propagates in the opposite direction to the propagation of the excitation light by the objective lens and prism mirror, is separated from the optical path of the excitation light by a dichroic mirror, and is detected by a photodetector (not shown).

[0032] The heat transfer member will be described with reference to FIG. 3 . The heat transfer member 161 is disposed below the partition wall 201, spanning the reaction chamber 1 and the measurement chamber 3. The buffer solution observation solution holder 119 is thermally coupled to the heat transfer member 161. As a result, the observation solution 123 contained in the centrifuge tube 124 is maintained at a temperature close to the ambient temperature of the measurement chamber 3. The heat transfer member 161 may be a thermal conductor containing a metal or diamond having a thermal conductivity λ of 20 W / (m·K) or more at room temperature (300 K), or a heat pipe that utilizes latent heat. The thermal conductivity λ of the heat transfer member 161 is preferably 50 W / (m·K) or more, and more preferably 100 W / (m·K) or more. Alternatively, the connector of the housing that structurally connects the reaction chamber 1 and the measurement chamber 3 may constitute the heat transfer member. The heat transfer portion 161 may be referred to as a heat transfer member 161 .

[0033] The steps for generating reaction products and measuring fluorescence are as follows: Attach the array plate 203, equipped with the base plate 207 and bank portion 205, to the plate holder 101. Supply the mixture of specimen liquid and reaction liquid to the reservoir 206 of the array plate 203, and heat it to a predetermined temperature (e.g., 30°C) using the heater 103 (reaction step). Discharge the mixture of specimen liquid and reaction liquid. Supply a stop solution. Replace the stop solution with a buffer solution. Supply a primary antibody solution and heat it to a predetermined temperature (e.g., 30°C) (primary antibody step). Replace the primary antibody solution with a buffer solution. Supply a secondary antibody solution and heat it to a predetermined temperature (e.g., 30°C) (secondary antibody step). Replace the secondary antibody solution with a buffer solution. Supply an observation solution to replace the buffer solution. Transport to the measurement chamber. Fluorescence measurement is performed.

[0034] Assume that for sample A, the temperatures for the reaction process, primary antibody process, and secondary antibody process are all set to 35°C to accelerate the reaction, while for sample B, the temperatures for the reaction process, primary antibody process, and secondary antibody process are all set to 25°C to limit the reaction. If non-temperature-controlled buffer solution and observation solution are supplied to the reservoirs, they will follow the set temperature of the array plate, resulting in a higher temperature for sample A during fluorescence measurement. Generally, higher temperatures result in lower fluorescence intensity, which is the opposite of what would be expected for sample A, resulting in reduced reliability of the instrument. On the other hand, if temperature-controlled buffer solution and observation solution are supplied to the reservoirs, the array plate temperature can be kept nearly constant during fluorescence measurement regardless of the set temperature of the array plate, improving instrument reliability. Furthermore, even if the ambient temperature changes between measurements, supplying temperature-controlled buffer solution and observation solution to the reservoirs reduces fluctuations in fluorescence intensity between measurements, resulting in more stable results. Additionally, providing a mechanism for cooling the measurement chamber (measurement chamber cooling unit) can also improve fluorescence intensity. For example, a mechanism for circulating cooled air within the measurement chamber may be provided.

[0035] (Example 2) A reaction measurement device according to this example will be described using Figure 4. The same components as those in Figures 2A and 2B are given the same numbers, and their description will be omitted. In this example, the device is configured so that four array plates can be installed at the same time.

[0036] The plate holder 101 and heater 103 constitute the reaction chamber mounting sections 11-1, 11-2, 11-3, and 11-4, respectively. The reaction chamber mounting sections 11-1, 11-2, 11-3, and 11-4 are mounted on a mounting section mounting table 303, which is configured to be movable in the X-axis direction along the reaction chamber X-axis stage 301. This configuration allows the array plates mounted on each reaction chamber mounting section to be moved to a position where they can be transported by the inter-chamber transport section (measurement chamber mounting section 31, transport stage 151, and measurement chamber Z-axis stage 153). Four sample solutions (107-1, 107-2, 107-3, and 107-4) can be placed in the refrigerated reagent solution holder 105, depending on the number of array plates that can be mounted. The process of generating reaction products and measuring fluorescence is as described in Example 1.

[0037] In this example, when the reaction product generation process is performed at different set temperatures for each array plate, or when the environmental temperatures during fluorescence measurement are different for the first and fourth array plates, it is possible to suppress fluctuations in fluorescence brightness between the array plates and obtain more stable results. In this example, the number of mounted array plates is four, but this is not limited to four, and the number of mounted array plates may be, for example, ten.

[0038] Example 3 By utilizing the phenomenon that the higher the temperature, the lower the fluorescence intensity, it is possible to lower the temperature of the spot surface and improve the fluorescence intensity. This example aims to achieve this effect. The buffer solution observation solution holder of the reaction measurement device according to the present disclosure will be described with reference to Figure 5.

[0039] The buffer solution observation solution holder 119 is installed and thermally coupled to a Peltier unit 401 that can be set to a predetermined temperature. The temperature of the Peltier unit 401 can be set in several ways, such as: - Set to match the temperature of the reaction chamber - Set to a temperature that is a predetermined temperature lower than the temperature of the reaction chamber (for example, -10 degrees) - Set to a constant temperature (for example, 15 degrees) regardless of the temperature of the reaction chamber The optimum setting can be made depending on the object to be measured.

[0040] In this way, by adding a cooled or thermostated buffer solution or observation solution, the temperature of the spot surface can be kept constant during fluorescence measurement. In particular, when a cooled buffer solution or observation solution is added, it is possible to improve the fluorescence intensity from the reaction product, so it is preferable to provide a buffer solution cooling unit or observation solution cooling unit.

[0041] In this embodiment, the buffer solution and the observation solution are controlled by a buffer solution observation solution holder that is integrated into one unit. However, this is not a limitation. The buffer solution holder and the observation solution holder may be provided independently, each with their own temperature control mechanism (cooling unit). In particular, since the timing of fluorescence measurement and the timing of introducing the observation solution are close in time, it is preferable to provide a temperature control mechanism in the observation solution holder. In addition, providing a mechanism for cooling the measurement chamber (measurement chamber cooling unit) can also improve the fluorescence brightness. For example, a mechanism for circulating cooled air within the measurement chamber may be provided.

[0042] Example 4: When the temperature of the buffer solution or observation solution is lowered, condensation may occur on the underside of the array plate. Condensation on the underside of the array plate may scatter the excitation light or the generated fluorescence, potentially reducing detection efficiency. An effective way to prevent condensation is to blow anti-condensation air onto the underside of the array plate. For example, a fan may be installed at a position on the reaction chamber mounting section where the air blows onto the array plate. Alternatively, anti-condensation air may be generated using the X-direction movement mechanism of the excitation light irradiation unit 32 described in FIG. 2B. An example of this is described with reference to FIG. 6.

[0043] The excitation light irradiation unit 32 includes a guide 209, a stage 211, a mirror holder 213 incorporating a prism mirror, and a lens holder 215 incorporating an objective lens (see FIGS. 2B and 6). The stage 211, mirror holder 213, and lens holder 215 are connected and can move in the X direction along the guide 209. When blades 403 connected to the mirror holder 213 reciprocate in the X direction, air is generated toward the underside of the array plate 203. Because the blades 403 are constantly moving during fluorescence measurement, the constantly generated air can prevent condensation from forming on the underside of the array plate 203.

[0044] The present disclosure is not limited to the above-described embodiments, and various modifications and variations are possible without departing from the spirit and scope of the present disclosure. Therefore, the following claims are appended to apprise the public of the scope of the present disclosure.

[0045] This application claims priority based on Japanese Patent Application No. 2024-98325, filed June 18, 2024, the entire contents of which are incorporated herein by reference.

[0046] 1: Reaction chamber 3: Measurement chamber 11: Reaction chamber mounting section 13: Sample liquid mounting section 15: Reagent liquid mounting section 17: Buffer solution mounting section 19: Observation liquid mounting section 21: Liquid supply section 31: Measurement chamber mounting section 32: Excitation light irradiation section 33: Optical measurement section 41: Heat transfer member 51: Passage opening 101: Plate holder 103: Heater 105: Refrigerated reagent liquid holder 107: Sample liquid 111: Primary antibody liquid 113: Secondary antibody liquid 115: Room temperature reagent liquid holder 117: Reaction stop solution 119: Buffer solution observation liquid holder 121: Buffer solution 123: Observation liquid 124: Centrifuge tube 125: Pipette tip rack 127: Pipette tip 129: X-axis stage 131: Y-axis stage 133: Z-axis stage 135: Pipetter 151: Transfer stage 153: Measurement chamber Z-axis stage 161: Metal plate (heat transfer member) 201: Partition wall 203: Array plate 205: Bank portion 206: Storage portion 207: Base plate 209: Guide 211: Stage 213: Mirror holder 215: Lens holder 301: Reaction chamber X-axis stage 303: Mounting portion installation base 401: Peltier unit 403: Blade

Claims

1. A reaction measurement device including a reaction chamber for supplying a specimen liquid and a reagent liquid containing a fluorescent label to one surface of an array plate having a plurality of substances fixed as an array of spots on the surface of a plate substrate, and for generating reaction products on the spots between the substances contained in the specimen liquid, the fluorescent labels in the reagent liquid, and the substances fixed as spots, and a measurement chamber for irradiating the spots with excitation light and measuring fluorescence emitted from the reaction products, wherein the array of spots is fixed to a plate substrate that is transparent to the excitation light and fluorescence, and the array of spots is fixed to the plate substrate, and the array of spots has a storage section that can store the supplied specimen liquid and reagent liquid so as to be in contact with the array of spots, the reaction chamber comprises a reaction chamber mounting section on which the array plate is mounted, a specimen liquid mounting section on which the specimen liquid to be supplied to the storage section is mounted, a reagent liquid mounting section on which the reagent liquid to be supplied to the storage section is mounted, a buffer liquid mounting section on which a buffer solution for intermediate replacement of the liquid stored in the storage section is mounted, an observation liquid mounting section on which a fluorescence-inactive observation liquid for final replacement of the liquid stored in the storage section is mounted, and a liquid supply section that sucks up the liquid mounted on at least one of the specimen liquid mounting section, the reagent liquid mounting section, the buffer liquid mounting section, and the observation liquid mounting section, and supplies it to the storage section; the measurement chamber comprises a measurement chamber mounting section on which the array plate transported from the reaction chamber mounting section is mounted, and an optical measurement section that irradiates the array plate mounted on the measurement chamber mounting section with excitation light from the surface opposite to the one surface, and measures emitted fluorescence; The reaction measurement device includes a heat transfer member that thermally couples at least one of the interior atmosphere of the measurement chamber and the measurement chamber mounting portion to the observation liquid mounting portion and the buffer solution mounting portion.

2. A reaction measurement device as described in claim 1, wherein the measurement chamber has a measurement chamber cooling section that cools the interior atmosphere, and the heat transfer member and the measurement chamber mounting section are thermally connected to the measurement chamber cooling section.

3. A reaction measuring device according to claim 1 or 2, wherein the heat transfer member has a thermal conductivity of 20 W / (m·K) or more at 300K.

4. A reaction measurement device according to any one of claims 1 to 3, wherein the observation liquid placement section has an observation liquid cooling section for cooling the observation liquid.

5. A reaction measurement device including a reaction chamber for supplying a specimen liquid and a reagent liquid containing a fluorescent label to one surface of an array plate having a plurality of substances fixed as an array of spots on one surface of a plate substrate, and for generating reaction products on the spots between the substances contained in the specimen liquid, the fluorescent labels in the reagent liquid, and the substances fixed as spots, and a measurement chamber for irradiating the spots with excitation light and measuring fluorescence emitted from the reaction products, wherein the array of spots is fixed to a plate substrate transparent to the excitation light and fluorescence, and the array of spots is fixed to the plate substrate, and the array of spots has a storage section that can store the supplied specimen liquid and reagent liquid so as to be in contact with the array of spots, the reaction chamber comprises a reaction chamber mounting section on which the array plate is mounted, a specimen liquid mounting section on which the specimen liquid to be supplied to the storage section is mounted, a reagent liquid mounting section on which the reagent liquid to be supplied to the storage section is mounted, a buffer solution mounting section on which a buffer solution for intermediate replacement of the liquid stored in the storage section is mounted, an observation liquid mounting section on which a fluorescence-inactive observation liquid for final replacement of the liquid stored in the storage section is mounted, and a liquid supply section that aspirates the liquid mounted in at least one of the specimen liquid mounting section, the reagent liquid mounting section, the buffer solution mounting section, and the observation liquid mounting section, and supplies it to the storage section; the measurement chamber comprises a measurement chamber mounting section on which the array plate transported from the reaction chamber mounting section is mounted, and an optical measurement section that irradiates the array plate mounted on the measurement chamber mounting section with excitation light from the surface opposite to the one surface, and measures emitted fluorescence; and the observation liquid mounting section has an observation liquid cooling section that cools the observation liquid.

6. A reaction measurement device according to claim 4 or 5, wherein the buffer solution placement section has a buffer solution cooling section for cooling the buffer solution.

7. A reaction measurement device according to claim 4 or 5, wherein the measurement chamber has a measurement chamber cooling section for cooling the atmosphere within the chamber.

8. The reaction measurement device of claim 7, wherein the measurement chamber is configured such that the measurement chamber cooling section cools the interior atmosphere so that the array plate placed on the measurement chamber mounting section and the objective lens of the optical measurement section do not condense.

9. A reaction measurement device as described in claim 8, wherein the measurement chamber is configured to form a flow in which air in contact with the surface opposite to the one surface of the array plate or the surface of the objective lens facing the array plate moves toward the surface in contact.

10. A reaction measurement device according to any one of claims 1 to 9, wherein the reaction chamber mounting section has a heating section for heating the array plate.

11. A reaction measurement device according to any one of claims 1 to 10, wherein the liquid supply unit supplies a buffer solution for cooling the array plate to the reservoir unit.

12. A reaction measurement device according to any one of claims 1 to 11, wherein the liquid supply unit supplies an observation liquid having a specific heat greater than that of the plate substrate of the array plate to the reservoir unit.

13. A reaction measurement device according to any one of claims 1 to 12, wherein the sample liquid placement section has a cooling section for cooling the sample liquid.

14. A reaction measurement device according to any one of claims 1 to 13, wherein the reagent liquid placement section has a cooling section for cooling the reagent liquid.

15. A reaction measurement device according to any one of claims 1 to 14, wherein the reagent liquid placement section is configured to place a reagent liquid containing a secondary antibody as the reagent liquid containing a fluorescent label.

16. The reaction measurement device according to claim 15, wherein the reagent solution placement section is configured to further place a reagent solution containing either a primary antibody or a reaction stop solution thereon.

17. A reaction measurement device according to any one of claims 1 to 16, wherein the observation liquid is fluorescently inactive at the wavelength of the excitation light.

18. A reaction measurement device according to any one of claims 1 to 17, wherein the reaction chamber mounting section has a configuration capable of mounting a plurality of the array plates.

19. The reaction measurement device according to claim 18, wherein the sample liquid placement section is configured to be capable of placing a plurality of sample liquids thereon.

20. The reaction measurement device according to any one of claims 1 to 19, further comprising an inter-chamber transport unit that transports the array plate between the reaction chamber and the measurement chamber.

21. The reaction measurement device according to any one of claims 1 to 20, further comprising a drainage section for removing the liquid stored in the storage section.

22. The reaction measurement device according to claim 21, wherein the liquid supply section also serves as the liquid drainage section.

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