JP2012233888A - Analytical method for simultaneously detecting or quantitatively determining multiple types of target substance - Google Patents

Abstract

PROBLEM TO BE SOLVED: To localize signal light from probe beads for simultaneously detecting or quantitatively determining multiple types of target substances.SOLUTION: In the analytical method for simultaneously detecting or quantitatively determining multiple types of target substances, a plurality of carriers which are stored in a single container and in which a substance for capturing the multiple types of target substances, a capture substance, and a capture substance are immobilized respectively, emit light with their immersed in liquid. The light emission is measured from the outside of the container so as to simultaneously detect or quantitatively determine the multiple types of target substances. In the method, the light transmittance of the liquid for an emission wavelength band is suppressed or adjusted for suppressing optical intersection of the light emitting signals from the plurality of carriers. An inspection kit and a device used for the method are also provided.

Description

  The present invention relates to an analysis method for simultaneously detecting or quantifying a plurality of types of target substances.

  BIST (Beads array In Single Tip) has been developed as a technique for simultaneously detecting a plurality of analysis objects (Non-patent Document 1). BIST is a cylindrical tip (hereinafter referred to as “capillary”) with a probe bead with a diameter of 1 mm to which a biomolecule such as an antibody, antigen, or DNA fragment that binds to an analyte such as an antigen or gene is immobilized. It is a device encapsulated side by side. Since this BIST can be fitted to the nozzle of a magnetization device, the same automation device as nucleic acid extraction (for example, Magtration (registered trademark) System 6GC, Magtration (registered trademark) System 12GC Plus (manufactured by Precision System Science)) ) Can be used for hybridization reaction or antigen-antibody reaction and washing operation. The generated signal is detected by a scanner (for example, BISTnner (manufactured by Precision System Science)).

  The biggest technical feature of BIST is that multiple probe beads for capturing different target substances are arranged in the capillary, allowing simultaneous detection of multiple items. It is practically used for quantitative testing of thyroid stimulating hormone (TSH), FT3 (Free Triiodo thyronine), and FT4 (Free thyroxine), which are highly clinically useful by being tested at the same time. The required detection concentration range covers a wide range (TSH: 0.05-50 uIu / mL, FT3: 0.5-20 pg / mL, FT4: 0.1-10 ng / dL, about 103 in the dynamic range). For this reason, in order to realize multi-item simultaneous inspection, signal light from each probe bead (light whose intensity increases in proportion to the concentration of the target substance) is localized and signal light from other probe beads is localized. Interference must be suppressed. However, so far, the localization of signal light has not been sufficient.

Simplified genetic polymorphism measurement method published by CMMC "Monthly Bioindustry September 2008"

  An object of the present invention is to localize signal light from probe beads in order to simultaneously detect or quantify a plurality of types of target substances.

  As a result of diligent efforts, the present inventors have found that signal light diffusion is significantly improved by taking the following measures, and have completed the present invention.

1. Fix probe beads to capillaries in the reaction and detection process.
2. Keep the temperature low when aspirating the substrate.
3. Increase the light shielding properties of the reaction solution itself.
4). Increase the transparency of capillaries.

The gist of the present invention is as follows.
(1) A plurality of carriers housed in a single container and fixed with substances for capturing a plurality of types of target substances are light-emitted while immersed in a liquid, and this light emission is emitted from the outside of the container. An analysis method for simultaneously detecting or quantifying a plurality of types of target substances by measuring, wherein light transmission of the liquid in the emission wavelength band is performed in order to suppress optical crossing of emission signals from the plurality of carriers. The method, wherein the rate is suppressed or adjusted.
(2) The method according to (1), wherein the emission is chemiluminescence or fluorescence, and the emission wavelength band is 340 to 900 nm.
(3) The method according to (1) or (2), wherein the light transmittance of the liquid is suppressed or adjusted by adding a light absorber.
(4) The method according to (3), wherein the light absorber is fine particles.
(5) The method according to any one of (1) to (4), wherein the light transmittance of the liquid is suppressed or adjusted so as to be 80% or less per 1 mm of the optical path length.
(6) The optical path length between the photometry unit and each of the plurality of carriers is geometrically defined by the relative positional relationship between the container, the surface shape of the carrier and the photometry unit located outside the container, so that the closest to the photometry unit. The method according to any one of (1) to (5), wherein the light emission signal from the carrier is selectively received.
(7) The method according to any one of (1) to (6), wherein the carrier is spherical.
(8) The method according to any one of (1) to (7), wherein the carriers are arranged one-dimensionally or two-dimensionally.
(9) The method according to any one of (1) to (8), wherein the constituent material of the container is 20% or less in terms of a haze value having a thickness of 1 mm.
(10) The method according to any one of (1) to (9), wherein the refractive index of the container in the wavelength band of light emission or radiation is 1.6 or less.
(11) A test kit for use in the method according to any one of (1) to (10), comprising a plurality of carriers on which substances for capturing a plurality of types of target substances are fixed.
(12) A substance for capturing a target substance is fixed to a carrier accommodated in a container, and the substance is reacted with a substance having a light-emitting ability contained in a liquid added from the outside. A method for detecting or quantifying a target substance by emitting or emitting a light-emitting substance, the method comprising adjusting the temperature of the liquid and / or the container when the liquid is added .
(13) The substance having luminous ability is at least one compound selected from the group consisting of luminol, diphenyl oxalate, ruthenium complex, oxalyl chloride, luciferin, AMPPD, indole, polyphenol, dioxetane, and derivatives of these substances. The method according to (12).
(14) The method according to (12) or (13), wherein the temperature of the liquid and / or the container is adjusted to 15 ° C. or lower.
(15) The method according to any one of (12) to (14), wherein the carrier is fixed to the container when the liquid is added.
(16) comprising: means for suppressing or adjusting the light transmittance of a liquid in a wavelength band of light emission or emission; and means for optically detecting and / or quantifying a target substance by light emission or radiation phenomenon. (10) The apparatus used for the method in any one of.
(17) The method according to any one of (12) to (15), comprising: means for adjusting the temperature of the liquid; and means for detecting or quantifying the target substance when the substance having a light-emitting ability emits or emits light. Equipment used for.
(18) The device according to (17), further comprising means for adjusting the temperature of the container.
(19) The method according to any one of (12) to (15), comprising: means for adjusting the temperature of the container; and means for detecting or quantifying the target substance by emitting or emitting a substance having a light-emitting ability. Used for equipment.
(20) The device according to (19), further comprising means for adjusting the temperature of the liquid.

  According to the present invention, in a system for simultaneously detecting or quantifying a plurality of types of target substances, the signal light from the probe beads to which the substance for capturing the target substance is fixed is localized and the signal light from other probe beads is localized. Interference with can now be suppressed.

Signal light distribution of TSH measurement in BIST when using a capillary made of polypropylene and mixing by inverting after aspirating the substrate at room temperature. Comparison of signal light distribution between BIST capillaries made of polypropylene and polycarbonate containing spherical light emitters inside the cylinder. Change in signal light distribution due to the difference in light transmittance of the liquid filled in the cylinder of polycarbonate BIST capillary. Change in signal light distribution due to the difference in light transmittance of the liquid filled in the cylinder of polypropylene BIST capillary. Changes in the signal light distribution of the BIST capillary due to the difference in the degree of agitation after substrate aspiration. Signal light distribution of BIST capillary when substrate is aspirated at room temperature. Signal light distribution of BIST capillaries due to differences in temperature conditions during substrate aspiration. Comparison of optical diffusion and signal light diffusion by agitation suppression after substrate aspiration. It is a perspective view which removes and shows a part of the housing | casing of the sample test apparatus which concerns on the 1st Embodiment of this invention. FIG. 10 is a perspective view showing a state in which a part of the sample testing apparatus in FIG. 9 is removed and the test cartridge container is pulled out. It is an expansion perspective view of the components incorporated in the optical measurement part shown in FIG.9 and FIG.10. FIG. 11 is an enlarged perspective view of the inspection cartridge container shown in FIGS. 9 and 10. It is a perspective view which shows the various chips accommodated in the test | inspection cartridge container shown in FIG. FIG. 11 is a process flowchart of the sample testing apparatus shown in FIGS. 9 and 10. It is a perspective view which takes out from the housing | casing and shows the main components of the sample test apparatus which concerns on the 2nd Embodiment of this invention. It is a perspective view which takes out from the housing | casing and shows the main components of the sample test apparatus which concerns on the 3rd Embodiment of this invention. FIG. 17 is an enlarged perspective view showing the light measurement unit and the temperature controller shown in FIG. It is a perspective view which takes out from the housing | casing and shows the main components of the sample test apparatus which concerns on the 4th Embodiment of this invention. FIG. 19 is an enlarged perspective view showing the light measurement unit and the temperature controller shown in FIG. FIG. 20 is an enlarged perspective view showing the lid shown in FIG. 19. It is a perspective view which notches and shows a part of lid shown in FIG. It is a schematic diagram which takes out from the housing | casing and shows the main components containing the four test | inspection cartridge containers of the sample test apparatus which concerns on the 5th Embodiment of this invention.

  Hereinafter, embodiments of the present invention will be described in more detail.

  In the present invention, a plurality of carriers, each of which is fixed in a substance for capturing a plurality of types of target substances (hereinafter referred to as “capture substances”), contained in a single container, are immersed in a liquid. An analysis method for simultaneously detecting or quantifying a plurality of types of target substances by measuring the luminescence from the outside of the container and suppressing optical crossing of luminescence signals from the plurality of carriers. The light transmission rate of the liquid in the emission wavelength band is suppressed or adjusted, and the method is provided.

  The container may be anything as long as light emission can be measured from the outside of the container, but is preferably translucent and is preferably a material that transmits light, such as resin or glass. The shape of the container is not particularly limited, but may be a shape such as a thin tube or pipette. For example, in the case of BIST, the container is a cylindrical chip capable of sucking and discharging the liquid contained therein.

In the method of the present invention, the diffusion of signal light can be reduced by increasing the transparency of the container. The haze value of the container having a thickness of 1 mm is preferably 20% or less, preferably 10% or less, and more preferably 1% or less. Examples of the material of the container having a haze value of 20% or less with a thickness of 1 mm include polypropylene, polycarbonate, polystyrene, polymethyl methacrylate, cyclic polyolefin, polymethylpentene and the like.
The haze value is a value representing the degree of transparency, and is defined as a ratio of diffuse transmittance to total light transmittance in JIS K7136.

  Further, the refractive index of the container in the wavelength band of light emission or radiation is preferably 1.6 or less, preferably 1.55 or less, and more preferably 1.50 or less. Examples of the material of the container having a refractive index of 1.6 or less include polypropylene, polycarbonate, polystyrene, polymethyl methacrylate, cyclic polyolefin, polymethylpentene, and the like.

  The target substance can be a substance to be analyzed included in biological samples such as blood, serum, urine, lymph, oral mucosa, nails, sputum, and samples such as river water, lake water, sea water, and soil. Examples of target substances include biological substances such as nucleic acids, proteins, antigens, antibodies, enzymes, sugars, bacteria such as Salmonella and Escherichia coli, metals such as cobalt, iron, molybdenum, copper, zinc, arsenic, cadmium, and vanadium. can do.

  The target substance may be labeled with a measurable label, and examples of the measurable label include a fluorescent label, a chemiluminescent label, an electrochemiluminescent label, an enzyme label, a radioactive label, and a magnetic label. Examples of labeling substances include Marine Blue, Cascade Blue, Cascade Yellow, Fluorescein, Rhodamine, Phycoerythrin, CyChrome, PerCP, Texas Red, Allophycocyanin, PharRed, Oregon Green-488, and Cy dyes (for example, Cy2, Cy3, Cy3.5, Cy5, Cy7 etc.), Alexa dyes (eg Alexa-488, Alexa-532, Alexa-546, Alexa-633, Alexa-680, Alexa-700, Alexa-750 etc.), BODIPY dyes (eg BODIPY FL , BODIPY TR-, etc.), luminol, acridine, adamantyldiokitacene, chemiluminescent materials such as oxalate, lophine, lucigenin, isolminol derivatives, tris-bipyridine- (4-methylsulfone) NHS ester ruthenium (II) complexes, electrochemiluminescent substances such as diphenylanthracene, tetraphenylanthracene, POD (Hydrogen peroxidase), AP (alkaline phosphatase), β-galactosidase, glucose oxidase, Course oxidase, and the like can be exemplified enzymes such as glucose-6-phosphate dehydrogenase. In addition, a labeling substance such as a radioactive substance such as a radioisotope (for example, 3H, 14C, 32P, 33P, 35S, 125I) may be used.

  The target substance may be directly labeled or indirectly labeled. For example, in the examples described later, the sandwich ELISA method is used. Specifically, after a target substance that is an antigen is bound to a primary antibody fixed to a carrier, a biotin-labeled antibody that is a secondary antibody is bound to the target substance. Furthermore, streptavidinated HRP that catalyzes chemiluminescence binds to this secondary antibody, and labels the target substance.

  The substance for capturing the target substance may be any substance that binds to the target substance, and examples thereof include antibodies, antigens, DNA fragments, metal complexes, and other low molecular weight compounds, but are not limited thereto. is not.

  The carrier may have any shape such as irregular particles, spheres (beads), threads (strands), magnetic beads, etc., but is preferably spherical. When the carrier takes the form of a sphere (bead) or magnetic bead, a substance for capturing a different target substance may be fixed to each of the sphere (bead) or magnetic bead. When the carrier takes the shape of a thread, it is preferable that a substance for capturing a different target substance is fixed to each of the different positions of the thread. In the case of BIST, the carrier takes the form of a sphere (bead).

  The sphere (bead) is preferably about 1 mm in size and is preferably made of a material such as plastic or ceramic. Such a sphere (bead) is disclosed in JP 2000-346842 A and JP 14-534657 A. It is described in gazettes.

  The magnetic beads are preferably tens of μm in size and are made magnetic by mixing iron powder or the like into plastic or ceramic. Such magnetic beads are described in JP-A-8-62224 (Patent No. 3115501), International Publication WO96 / 29602, International Publication WO97 / 44671, and the like.

  The yarn (strand) has a diameter of about 0.1 mm and is preferably made of a resin-based material. Such yarn is disclosed in JP 2006-214759 A, International Publication WO01 / 53831 Pamphlet, International Publication WO2003. It is described in / 7901 pamphlet.

  The carriers may be arranged one-dimensionally or two-dimensionally. The one-dimensional arrangement of the carrier means a state in which the positional information necessary for specifying the carrier from which the observed emission signal is derived is one-dimensional, for example, BIST It can be said that the carriers (beads) are arranged one-dimensionally. The carrier is arranged two-dimensionally means a state in which the positional information necessary for specifying the carrier from which the observed emission signal is derived is two-dimensional, for example, A measurement kit in which a carrier is spread in a single layer on a plate can be said to be a carrier that is two-dimensionally arranged.

  A substance for capturing a target substance is fixed to the carrier by covalent bond, chemisorption, physical adsorption, electrical interaction, hydrophobic interaction, van der Waals force, hydrogen bond, and the like.

  The analysis of the target substance can be performed by bringing a substance for capturing the target substance into contact with the target substance in a state immersed in a liquid, and then measuring the emitted light.

  The liquid may be any liquid that can immerse the carrier and can measure luminescence from the outside of the container, and examples thereof include water, an organic solvent, and a buffer solution, but are not limited thereto.

  In the method of the present invention, the diffusion of signal light can be reduced by suppressing or adjusting the light transmittance of the liquid. The light transmittance of the liquid is preferably suppressed or adjusted so as to be 80% or less per 1 mm of the optical path length. Preferably, the light transmittance of the liquid is suppressed or adjusted so that it is 60 to 80% per 1 mm of the optical path length, and more preferably 50% or less per 1 mm of the optical path length.

  The luminescence can be chemiluminescence, fluorescence or electrochemiluminescence, but is chemiluminescence or fluorescence, and the wavelength band of luminescence is preferably 340-900 nm. Luminescence can be measured using a known technique such as a fluorometer, spectrophotometer, scintillation counter, photodiode, CCD, CMOS, photomultiplier tube or the like.

  In the method of the present invention, plural types of target substances are detected or quantified simultaneously. The plural kinds are at least two kinds, for example, 2, 3, 4, 5 kinds or more. Substances for capturing a plurality of types of target substances are respectively fixed to a plurality of carriers. For example, in the case of BIST, about 50 beads including probe beads (beads on which materials for capturing a target substance are fixed) and light-shielding beads can be arranged in a capillary having a total length of 50 mm. The probe beads may include blanks and internal standards. A blank is a carrier on which a capture substance is not fixed. The internal standard is a carrier on which a substance that cannot be contained in a specimen and that captures a standard substance having a known concentration added before analysis is fixed.

  In order to detect or quantify multiple types of target substances at the same time, the capture substance captures the target substance specifically regardless of the presence or absence of other capture substances. A plurality of types of target substances can be detected simultaneously by placing the carrier in the same container and bringing a liquid specimen into contact therewith.

In this specification, “optical crossing of light emission signals from a plurality of carriers” refers to a state in which light emission signals from a plurality of carriers are simultaneously received by a detector and the light emission signals from the respective carriers cannot be separated.

  The optical intersection of the emission signals from the plurality of carriers can be suppressed by suppressing or adjusting the light transmittance of the liquid in the emission wavelength band.

  For example, the light transmittance of the liquid can be suppressed or adjusted by adding a light absorber. Examples of the light absorber include fine particles (for example, carbon black, fine particles composed of inorganic or organic substances), dyes that dissolve inorganic or organic substances, and the like, but are not limited thereto.

  In the method of the present invention, the optical path length between the photometric unit and each of the plurality of carriers is geometrically defined by the relative positional relationship between the container, the surface shape of the carrier, and the photometric unit located outside the container. The light emission signal from the nearest carrier may be selectively received. Specifically, in the BIST example, in the BIST chip, spherical carriers having a diameter of about 1 mm are one-dimensionally arranged inside a cylindrical capillary having an inner diameter of 1.1 to 1.2 mm. Of the light emission signal emitted from the carrier surface and reaching the outside of the capillary, the shortest distance passing through the liquid part including the light absorption band is from the outermost part of the carrier (closest to the capillary inner wall of the carrier) to the cylindrical inner wall. It is a light beam emitted vertically. At the same time, as the traveling direction of the light emission signal moves away from the perpendicular to the inner wall of the cylinder, the distance passing through the liquid portion including the light absorption band increases. Therefore, when the light emission signal from the carrier is received from the outside of the cylinder, the signal light from the nearest carrier can be selectively received.

The present invention comprises a plurality of kinds of means comprising means for suppressing or adjusting the light transmittance of a liquid in a wavelength band of light emission or radiation, and means for optically detecting and / or quantifying a target substance by light emission or radiation phenomenon. An apparatus for simultaneously detecting or quantifying a target substance is provided. The apparatus of the present invention emits light in a state in which a plurality of carriers housed in a single container and fixed with substances (capturing substances) for capturing a plurality of types of target substances are immersed in a liquid. An analysis method for simultaneously detecting or quantifying a plurality of types of target substances by measuring the luminescence from the outside of the container, wherein the emission wavelength is suppressed in order to suppress optical crossing of luminescence signals from the plurality of carriers. The light transmittance of the liquid in the band can be suppressed or adjusted, and the method can be used.

  This apparatus has a function of dispensing and mixing specimens and reagents, a function of adjusting the temperature of specimens and reagents, and a function of detecting signal light.

  In addition, the present invention provides a test kit including a plurality of carriers in which substance capturing substances for capturing a plurality of types of target substances, which are accommodated in a single container, are fixed. This kit emits light in a state where multiple carriers housed in a single container and fixed with substances for capturing multiple types of target substances (capture substances) are immersed in a liquid. An analytical method for simultaneously detecting or quantifying a plurality of types of target substances by measuring luminescence from the outside of the container, and in order to suppress optical crossing of luminescence signals from the plurality of carriers, in the emission wavelength band It can be used in the method, wherein the light transmittance of the liquid is suppressed or adjusted. The container, the substance for capturing the target substance, and the carrier are described above.

  Furthermore, the present invention is such that a substance for capturing a target substance is immobilized on a carrier housed in a container, and the substance reacts with a substance having a light-emitting ability contained in a liquid added from the outside. A method of detecting or quantifying a target substance by emitting or emitting light having a light-emitting ability, and adjusting the temperature of the liquid and / or the container when the liquid is added. Including said method. This method may be used in combination with the above-described method for suppressing or adjusting the light transmittance of the liquid in the emission wavelength region. By doing so, signal light can be localized more effectively. The container, the substance for capturing the target substance, and the carrier are described above.

  The liquid is the same as the above liquid, but the liquid contains a substance having a light emitting ability and is added from the outside of the container. Substances having luminous ability include luminol, diphenyl oxalate, ruthenium complex, oxalyl chloride, luciferin, AMPPD (3- (2'-spiroadamantane) -4-methoxy-4- (3 "-phosphoryloxy) phenyl-1 , 2-dioxetane disodium salt), indole, polyphenol, dioxetane, derivatives of these substances, and the like, but are not limited thereto.

  For example, luminol reacts with hydrogen peroxide by a catalytic action of iron or the like to generate an unstable reaction intermediate. When this reaction intermediate changes to a phthalate dianion, light emission occurs. Sandwich ELISA is used to detect the target substance in BIST, but the secondary antibody is labeled with horseradish peroxidase containing heme, thereby catalyzing the luminescence of luminol.

  In the method of the present invention, when a liquid containing a substance having a light-emitting ability is added, the temperature of the liquid and / or the container to which the liquid is added is adjusted and optimized, thereby spreading the signal light. Can be reduced. By keeping the temperature of the liquid and / or the container at a low temperature, the diffusion of the signal light can be reduced. The temperature of the liquid and / or container may be adjusted to 15 ° C. or lower, preferably 10 to 2 ° C., more preferably 6 to 4 ° C. The temperature of the liquid and / or the container can be adjusted by a Peltier element, a heating wire, liquid nitrogen, or the like.

  In the method of the present invention, the carrier may be fixed to the container when the liquid containing the substance having a light emitting ability is added. The diffusion of signal light can be reduced by fixing the carrier to the container. Examples of the method for fixing the carrier to the container include a method for shortening the distance between the carriers, a method for caulking at a position between the carriers, and a method for using positioning pins.

  The present invention provides an apparatus for detecting or quantifying a target substance, comprising means for adjusting the temperature of a liquid, and means for detecting or quantifying a target substance by luminescence or emission of a substance having a light-emitting ability. . In the apparatus of the present invention, a substance for capturing a target substance is fixed to a carrier accommodated in a container, and the substance reacts with a substance having a light-emitting ability contained in a liquid added from the outside. A method of detecting or quantifying a target substance by emitting or emitting light having a light-emitting ability, and adjusting the temperature of the liquid and / or the container when the liquid is added. It can be used for the above-mentioned methods. The apparatus of the present invention may further include means for adjusting the temperature of the container.

  Furthermore, the present invention provides an apparatus for detecting or quantifying a target substance, comprising means for adjusting the temperature of the container, and means for detecting or quantifying the target substance by luminescence or emission of a substance having a light-emitting ability. I will provide a. In the apparatus of the present invention, a substance for capturing a target substance is fixed to a carrier accommodated in a container, and the substance reacts with a substance having a light-emitting ability contained in a liquid added from the outside. A method of detecting or quantifying a target substance by emitting or emitting light having a light-emitting ability, and adjusting the temperature of the liquid and / or the container when the liquid is added. It can be used for the above-mentioned methods. The apparatus of the present invention may further include means for adjusting the temperature of the liquid.

  Next, the sample testing apparatus 10 according to the first embodiment of the present invention will be described with reference to FIGS.

  The sample testing apparatus 10 is surrounded by a book-like casing 12 having a length of about 250 to 400 mm (X-axis direction), a width of 70 to 100 mm (Y-axis direction), and a height of about 300 to 500 mm (Z-axis direction). It is. In the housing 12, a plurality of (10 in this example) wells 22 and a plurality of types of test instruments that can contain or contain a sample and one or more reagent solutions used for testing the sample are stored. The chip storage unit 20 in which chips of three types (in this example) are stored is arranged in a line, and the sample information for identifying or managing the sample and the test information indicating the test contents are used as a visible recording medium. A test cartridge container 14 displayed on the seal 24 and formed of a translucent member, and a sample contained in the test cartridge container 14 and the reagent are reacted to obtain light emission in a predetermined optical state. An automatic inspection unit (15, 19), a light measurement unit 17 for measuring the luminescence generated as a result of the inspection by the automatic inspection unit, and the inspection camera including the specimen information and the inspection information A digital camera 28 that captures the image displayed on the cartridge container 14 to obtain image data, a thermal transfer printer mechanism 21 that can print the inspection result in a blank of the seal 24 of the inspection cartridge container 14, and the automatic inspection unit (15, 19), a light measurement unit 17, a digital camera 28, and a board 52 provided with an integrated circuit such as a CPU for controlling the thermal transfer printer mechanism 21.

  The inspection cartridge container 14 is detachably loaded in a loading box 18 that is connected to the fitting plate 16 that can be manually pulled out of the casing 12 from the casing 12.

  The chamber in which the automatic inspection section (15, 19), the inspection cartridge container 14 and the light measurement section 17 are provided and the chamber in which the board 52 is provided are separated by a partition plate 51, and are sucked and discharged. The circuit is prevented from being damaged or contaminated by splashing liquid. A ventilation fan 54 is provided so as to penetrate the partition plate 51, and another ventilation fan 56 is provided so as to penetrate the housing 12 of the chamber in which the board 52 is provided.

  The automatic inspection unit (15, 19) includes a nozzle head 15 of a dispenser, and a moving mechanism 19 that allows the nozzle head 15 to move with respect to the inspection cartridge container 14 accommodated in the housing 12. Have

  The nozzle head 15 of the dispenser can be moved by the moving mechanism 19 in the X-axis direction corresponding to the longitudinal direction with respect to the inspection cartridge container 14 accommodated in the housing 12. 11 and a Z-axis moving body 13 provided so as to be movable while being guided by the guide column 41 in the vertical direction with respect to the X-axis moving body 11. A nut portion connected to the Z-axis moving body 13 is screwed into the X-axis moving body 11, and a Z-axis moving ball screw 43 (to be described later) that moves the Z-axis moving body 13 in the vertical direction is rotatable. In addition to being attached, the guide column 41 and a support plate 39 attached via the guide column 41 are attached.

  The nozzle head 15 is attached to the Z-axis moving body 13 and communicates with a cylinder that sucks and discharges gas via an air rubber tube 31 provided so as to protrude from the side surface, A motor 40 for driving the piston and a ball screw 42 rotatably mounted.

  Further, the support plate 39 attached to the X-axis moving body 11 rotatably supports the ball screw 42, and on the lower side thereof, a chip such as the carrier enclosing chip 26 is detached from the nozzle 30. Therefore, a tip detachment plate 48 in which a U-shaped hole that is larger than the diameter of the nozzle 30 and smaller than the outer diameter of the thickest portion of the tip is formed is movably supported. On the upper side, a motor 38 for driving the chip attaching / detaching plate 48 in the front-rear direction is attached to the X-axis moving body 11.

  The digital camera 28 is attached to the X-axis moving body 11 via a camera support plate 29, and the specimen information and examination information on the seal 24 of the examination cartridge container 14 housed in the housing 12 are stored. The nozzle head 15 is moved to a position where the entire image can be photographed.

  A moving mechanism 19 for moving the nozzle head 15 of the dispenser with respect to the inspection cartridge container 14 accommodated in the housing 12 is engaged with the X-axis moving body 11 of the nozzle head 15 to perform the inspection. A rail 44 that guides along the longitudinal direction of the cartridge container 14, that is, the X-axis direction, an X-axis moving motor 58 that moves the nozzle head 15 along the X-axis direction, and the Z-axis moving body 13 in the vertical direction. That is, it has the said guide pillar 41 guided along a Z-axis direction, the said Z-axis movement ball screw 43, and the Z-axis movement motor. The cylinder, the ball screw 42, and the motor 40 correspond to a suction / discharge mechanism. The guide pillar 41, the Z-axis moving ball screw 43, and the Z-axis moving motor correspond to the Z-axis moving mechanism in the moving mechanism 19.

  The light measurement unit 17 includes a chip insertion unit 34 and a photoelectric unit 32 having at least one photoelectric element such as a photomultiplier tube that converts received fluorescence into a predetermined electrical signal.

  The thermal transfer printer mechanism 21 is connected to the light measurement unit 17 via the board 52 and receives an electrical signal corresponding to the measurement result of the light measurement unit 17 to print on the seal 24 of the inspection cartridge container 14. To do. The thermal transfer printer mechanism 21 is positioned so as not to contact the inspection cartridge container 14 when the inspection cartridge container 14 is inserted into the housing 12, and stores the inspection cartridge container 14. For example, it is preferable that the print head 21 a of the thermal transfer printer mechanism 21 is provided so that it is lowered by a cam mechanism and is located in a predetermined blank portion on the seal 24 of the inspection cartridge container 14. The print head 21a is formed of a predetermined number of digital numbers, and a predetermined number of digital numbers of the print head 21a is heated to automatically convert the digital numbers corresponding to the seal 24 formed of a thermal medium. Is what you write.

  FIG. 10 shows a state in which the test cartridge container 14 of the sample test apparatus 10 is manually pulled out from the housing 12. The thermal transfer printer mechanism 21 is removed for convenience of explanation.

  The loading box 18 in which the inspection cartridge container 14 is loaded has a guide member 18a extending along the longitudinal direction of the loading box 18, that is, along the X-axis direction. The inspection cartridge container 14 is guided by the guide rail 23 so as to be manually movable in the X-axis direction, whereby the inspection cartridge container 14 can be completely accommodated in the housing 12.

  The guide member 18a and the thermal transfer printer mechanism 21 are preferably provided with the cam mechanism to interlock the insertion / extraction of the container 14 and the vertical movement of the mechanism 21.

  The nozzle 30 of the nozzle head 15 is detachably mounted with a carrier enclosing tip 26 in which particles 26c as a plurality of carriers are accommodated.

  The light measurement unit 17 further includes a measurement block 36 having a semicircular hole 36a formed at the edge and fixed to the photoelectric unit 32, and a lower side of the measurement block 36 and above the chip insertion unit 34. And a measurement plate 35 that is formed at the edge and can be moved back and forth along the long axis direction (X-axis direction) of the long hole 35a by an electromagnet. The tip insertion portion 34 provided below the measurement plate 35 has a small diameter tube 26a of the carrier enclosing tip 26 descending through a gap portion combined by the semicircular hole 36a and the long hole 35a. It is formed in a box shape so that it can be inserted through the square hole 34a. The measurement plate 35, the measurement block 36, and the photoelectric unit 32 are fixed to the housing 12 at the time of measurement, and the carrier enclosing chip 26 is raised or lowered with respect to the housing 12. And scanning a plurality of particles 26c.

  FIG. 11 shows an optical system built in the light measuring unit 17. The optical system is an apparatus suitable for measuring chemiluminescence, for example, and includes three sets of optical fibers 37a, 37b, and 37c, and light receiving ends 33a and 33b including a lens provided at the tip of the optical fiber. , 33c. The light receiving ends 33a and 33b are disposed on the side wall of the elongated hole 35a of the measurement plate 35, and the light receiving end 33c is disposed on a side wall of the semicircular hole 36a of the measurement block 36. The ends 33a, 33b and 33c radially surround the small diameter tube 26a of the carrier enclosing tip 26 from three directions. The measurement plate 35 is moved forward using the magnetic force of an electromagnet when the carrier-enclosed chip 26 is inserted, and the horizontal cross-sectional area of the gap portion formed by the long hole 35a and the semicircular hole 36a is enlarged, At the time of measurement, the measurement plate 35 is moved backward to approach the carrier enclosing tip 26 inserted into the long hole 35a to narrow the horizontal sectional area.

  FIG. 12 shows the inspection cartridge container 14 in an enlarged manner.

  The substrate 14 a of the test cartridge container 14 is provided with an opening of the chip housing portion 20 and an opening of the well 22. The capacity of each well 22 is, for example, about 1 cc to several cc, for example, 2 cc. In this example, three tips, in this example, a dispensing tip 25, a carrier enclosing tip 26, and a perforating tip 27 are attached to the tip accommodating portion 20 so that they can be attached by lowering and inserting the nozzle 30. The openings are placed in the cylinders 20a, 20b, and 20c with the corresponding depths. Ten wells 22 contain a specimen and one or more reagent solutions used for the examination of the specimen, and the opening is closed with a single film that can be perforated by the perforating tip 27. The opening of the chip accommodating portion 20 is closed with a seal that can be peeled by hand, and the seal is peeled off during use.

  As described above, the specimen information (24a, 24b) and the examination information (24c, 24d, 24e) indicating the examination contents are stored in the seal attaching region 14b as the medium attachment portion of the substrate 14a of the examination cartridge container 14. A visually displayed seal 24 is detachably attached. Here, in the sample information (24a, 24b), for example, a column 24a for entering and displaying a patient name by hand and a column 24b for displaying a patient identification number are provided, and examination information (24c, 24d, 24e) is provided. ) Includes, for example, a column 24c for displaying an inspection item, the manufacturing location, the manufacturing date, the expiration date, the number of manufactured reagents, the storage location, the quality, etc. of one or more reagents stored in the inspection cartridge container 14 in advance. As the LOT number column 24d for displaying the LOT number indicating the management information and the remarks column 24e, for example, a column for entering and displaying the inspection result measured by the light measuring unit 17 is provided. Examples of the examination items include examinations for TSH (thyroid stimulating hormone), inflammation in the body, allergies, and the like. As shown in FIG. Reference numeral 24f denotes a knob for peeling off the seal 24 from the substrate 14a.

  FIG. 13 shows three types of chips (25, 26, 27) housed in the chip housing portion 20 of the inspection cartridge container 14.

  As shown in FIG. 13A, the dispensing tip 25 accommodates the liquid in the tip by sucking the liquid, moves between the wells 22 and discharges the contained liquid, thereby transferring the liquid between the wells 22. Used for etc. The dispensing tip 25 has a small-diameter tube 25a having a thickness that allows the tip to be inserted into the well 22, and a mounting opening that communicates with the small-diameter tube 25a and can be attached to the nozzle 30 at the rear end. It has a large diameter tube 25b and a plurality of protrusions 25d provided in parallel to the axial direction at the rear end of the large diameter tube 25b.

  As shown in FIG. 13 (B), the carrier enclosing tip 26 includes a plurality of (43 in this example) particles 26c as the carrier, and the small diameter tube 26a having a thickness that can be inserted into the well 22. A binding substance that can bind to the target substance is fixed to each particle, and the small diameter pipe 26a is caulked at positions 26d and 26e to be enclosed inside. . By shortening the distance between the positions 26d and 26e, the particles 26c can be fixed to the carrier enclosing tip 26. Alternatively, the particles 26c may be fixed to the carrier enclosing tip 26 by caulking at a position between the particles of the plurality of particles 26c. The small-diameter tube 26a communicates with the large-diameter tube 26b via a filter portion 26f provided with a filter that allows only air to pass therethrough, and the opening of the large-diameter tube 26b is provided so as to be attachable to the nozzle 30. . Around the large-diameter pipe 26b, a plurality of protrusions 26g are provided in parallel to the axial direction. The carrier enclosing tip 26 may have a light transmittance of 70% or more. The refractive index of the carrier enclosing tip 26 is preferably 1.6 or less. The material of the carrier enclosing tip 26 is preferably polycarbonate, polystyrene, polymethyl methacrylate, cyclic polyolefin, or polymethylpentene.

  As shown in FIG. 13C, the perforating chip 27 has a sharp tip 27a and a rear end 27b for punching a film that closes the opening of the well 22 of the test cartridge container 14. A certain opening can be attached to the nozzle 30, and a plurality of protrusions 27 c are provided on the outer periphery of the rear end portion 27 b in parallel to the axial direction. These chips have a small diameter tube or a tip having a length of, for example, 1 cm to 10 cm, a large diameter tube having a length of, for example, 1 cm to 10 cm, and the particle diameter of, for example, 0.1 mm. 3mm. Then, the inner diameter of the small diameter tube 26a is such a size that the particles can be held in a line, and has an inner diameter of about 0.2 mm to 6 mm, for example.

  Subsequently, the operation of the sample testing apparatus 10 according to the first embodiment will be described with reference to FIG.

  As shown in FIG. 14A, in step S1, the fitting plate 16 of the housing 12 of the sample testing apparatus 10 is pulled out by hand. As shown in FIG. 14B, the loading box 18 is developed outside the housing 12 in step S2. As shown in FIG. 14C, in step S 3, the test cartridge container 14 in which the specimen to be tested, the test reagent, and the chip are stored in advance is loaded into the loading box 18. At that time, the name of the patient belonging to the sample information is written in the seal 24 of the test cartridge container 14 by hand, and the test information indicating the test contents is described in advance. As shown in FIG. 14D, in step S4, the loading box 18 and the loaded inspection cartridge container 14 are manually inserted into the housing 12 and stored.

  In the state of FIG. 14D, the following processing is performed.

  In step S <b> 5, the nozzle head 15 moves to the chip accommodating portion 20 of the inspection cartridge container 14, and the nozzle 30 is positioned just above the punching chip 27. The nozzle 30 is lowered along the Z-axis direction, and the tip of the nozzle 30 is inserted into the opening of the drilling tip 27 and pushed in.

  In step S 6, the nozzles 30 to which the perforating tips 27 are attached are sequentially positioned directly above the wells 22 of the test cartridge container 14 and then lowered to cover the ten wells 22. Perforate the film.

  When the perforation of all the wells 22 is completed in step S7, the nozzle 30 moves to a position where the perforating chip 27 is accommodated in the chip accommodating part 20, and the U-shaped groove of the chip attaching / detaching plate 48 is reached. Is moved close to the nozzle 30 and then the nozzle 30 is moved in the upward direction (Z-axis direction) to detach the drilling tip 27 from the cylindrical body 20c of the tip accommodating portion 20.

  In step S8, the nozzle 30 is moved to a position just above the position where the dispensing tip 25 (or the carrier-filled tip 26) of the tip accommodating portion 20 is accommodated, and the nozzle 30 is lowered along the Z-axis direction. Then, the tip of the nozzle 30 is inserted into the opening of the dispensing tip 25 (or the carrier-filled tip 26) and is pushed in.

  For example, the process of the sample test apparatus 10 when performing an allergy test on a subject will be described.

  Various allergen substances, for example, several kinds of allergen substances (antibodies) obtained from cedar pollen, ragweed, egg white, soybean, house dust, mites, molds, etc., are fixed to the particles 26c of the carrier-encapsulating chip 26. The particles 26c to which each allergen substance is fixed are encapsulated in advance in an arrangement position corresponding to the type of the fixed allergen substance. Further, the particles 26c to which any kind of allergen substance is not fixed are arranged between the particles 26c to which the allergen substance is fixed in the carrier enclosing tip 26.

  The well 22a of the test cartridge container 14 contains serum collected from a subject as a specimen, the well 22b contains a peroxidase solution of a labeling enzyme, and the well 22c contains luminol as a chemiluminescent substrate solution. / Store hydrogen peroxide solution. Further, for example, a cleaning solution such as a phosphate buffer solution or a tris buffer solution is stored in the wells 22d to 22i. The light absorber containing liquid may be accommodated in any one or more of the wells 22d to 22i. Alternatively, a light absorber may be added to the chemiluminescent substrate solution. The arrangement of the particles 26c, the types of reagents, and the like are displayed as examination information, and the subject information is displayed as sample information.

  In step S9, the dispensing tip 25 is attached to the nozzle 30, the attached dispensing tip 25 is positioned in the well 22b, the peroxidase solution is aspirated, moved, and the well 22a in which serum is contained. And is maintained at room temperature for a certain period of time. Thereby, the human IgE antibody in the serum is labeled with the peroxidase.

  In step S10, the dispensing tip 25 is moved to the position where the dispensing tip 25 is accommodated in the tip accommodating portion 20 of the inspection cartridge container 14, and the U-shaped groove of the tip attaching / detaching plate 48 is moved. After the nozzle 30 is moved close to the nozzle 30, the dispensing tip 25 is detached from the tip body 20 toward the cylindrical body 20a by moving the nozzle 30 along the upward direction.

  In step S11, the nozzle 30 is moved to a position just above the position where the carrier-enclosed chip 26 is accommodated in the chip accommodating portion 20, and the nozzle 30 is lowered along the Z-axis direction. The tip of 30 is inserted into the opening of the carrier enclosing tip 26 and pushed in.

  In step S12, the carrier enclosing tip 26 is moved to the well 22d, and the cleaning liquid is cleaned by, for example, sucking and discharging 100 μL.

  In step S13, the carrier-enclosed chip 26 is moved to the well 22e, and the cleaning liquid accommodated in the well 22e is further sucked and discharged to perform further cleaning.

  In step S14, the carrier-encapsulating chip 26 attached to the nozzle 30 is moved to the position of the well 22a, and the serum containing the human IgE antibody labeled with the peroxidase contained in the well 22a is encapsulated in the carrier. The large-diameter tube 26b is aspirated to contact the particle 26c so as to fill the small-diameter tube 26a of the chip 26, and the human IgE antibody in serum and the allergen substance are reacted for about 30 minutes.

  In step S15, the carrier-enclosed chip 26 is moved to the well 22f of the test cartridge container 14, and the washing liquid contained in the well 22f is washed by repeating, for example, about 100 μl of suction and discharge 10 times. Further, the carrier enclosing tip 26 is moved to the well 22g of the test cartridge container 14 and transferred to the well 22g to repeat washing.

  In step S16, the carrier-enclosed chip 26 is transferred to the well 22c of the test cartridge container 14, and the luminal / hydrogen peroxide solution as the substrate solution is sucked to react with the peroxidase as the labeling substance. Then, the carrier-enclosed chip 26 that emits light is positioned just above the semicircular hole 36a of the light measurement unit 17. A light absorber may be added to the chemiluminescent substrate solution. Alternatively, the light absorber-containing liquid is stored in any one or more of the wells 22d to 22i, the carrier-encapsulating chip 26 is moved to the wells, and the light absorption stored in the wells is absorbed. The body-containing liquid may be aspirated. In addition, the temperature of the chemiluminescent substrate solution or the well 22c that accommodates it may be controlled. Alternatively, the temperature of the carrier enclosing tip 26 may be controlled. The temperature control can be performed by a temperature controller described later.

  In step S17, the small diameter tube 26a of the carrier enclosing tip 26 is inserted into a gap formed by the semicircular hole 36a and the long hole 35a. At that time, the measurement plate 35 is moved backward to move the elongated hole 35a along its axial direction so that the small diameter tube 26a of the carrier enclosing tip 26 is lowered and the fine plate 35a is lowered. The particle 26c is scanned by accommodating the diameter tube 26a in the chip insertion portion 34, and the light emission state of each particle 26c is measured.

  In step S18, the presence or absence of light emission is measured for each particle 26c. Each particle 26c is associated with allergens in advance in the order of arrangement, and the allergen substance bound to the labeled antibody is identified by its luminescence. The measurement result is analyzed by the control unit of the board 52, the measurement result is sent to the thermal transfer printer mechanism 21, and the measurement result is added to the remarks column of the seal 24 by the print head 21a. Will be displayed as a number.

  In step S <b> 19, the digital camera 28 shoots image data including sample information and test information on the seal 24 of the test cartridge container 14 in accordance with an instruction signal from the board 52. At this time, the analysis unit of the control unit searches for data that can be analyzed from the image data, and when the two-dimensional barcode data indicating the inspection content included in the inspection information is found in the image data, the 2 The analysis data is obtained by analyzing the dimensional barcode data, and the data combination unit of the control unit stores the analysis data and the image data in a memory that can be output in combination.

  In step S20, the carrier enclosing tip 26 attached to the nozzle 30 moves to the tip accommodating portion 20, and the carrier enclosing tip 26 is just above the position where the carrier enclosing tip 26 is accommodated. After moving the U-shaped groove of the chip attaching / detaching plate 48 to the nozzle 30, the nozzle 30 is moved upward to move the carrier into the cylinder 20 b of the chip accommodating portion 20. The enclosing tip 26 is removed.

  When the specimen test is completed in step S21, the loading box 18 loaded with the test cartridge container 14 is manually pulled out from the housing 12, and the seal 24 attached to the test cartridge container 14 is peeled off. The test cartridge container 14 itself is discarded by being attached to a management board or the like prepared separately, but a new test cartridge container 14 is loaded in the housing 12 to test a new sample. Can be performed.

  Next, the processing performed by the sample testing apparatus 10 will be described for testing for the presence of food allergens.

  For example, seven items of ingredients that are mandatory for food in Japan, eggs, milk, wheat, buckwheat, peanuts, shrimp, crab, and further, for example, item 18 recommended for display (peach, pork) The allergen substance selected from allergen substances (antibodies) obtained from chicken, beef, etc. is fixed to the particles 26c of the carrier-encapsulating chip 26. The particles 26c to which each allergen substance is fixed are sealed in advance after blocking the arrangement according to the type of the fixed allergen substance. Further, the particles 26c to which any kind of allergen substance is not fixed are arranged between the particles 26c to which the allergen substance is fixed in the carrier enclosing tip 26.

  The well 22a of the test cartridge container 14 contains an extract (antigen) extracted from food as a specimen, and the well 22b contains a solution of various labeled antibodies labeled with the chemiluminescent substance HRP enzyme. To house. The well 22c accommodates TMB as a chemiluminescent substrate solution. A washing solution or a buffer solution is stored in the wells 22d to 22j. The light absorber containing liquid may be accommodated in any one or more of the wells 22d to 22i. Alternatively, a light absorber may be added to the chemiluminescent substrate solution. The arrangement of the particles 26c, the type of reagent, and the like are displayed as examination information, and the information on the subject is displayed as sample information by handwriting or the like.

  In step S9 ′, the nozzle 30 is moved to a position directly above the position where the carrier enclosing tip 26 is accommodated in the tip accommodating portion 20, and the nozzle 30 is lowered along the Z-axis direction, The tip of the nozzle 30 is inserted into the opening of the carrier enclosing tip 26 and pushed in.

  In step S10 ′, the carrier enclosing tip 26 is moved to the well 22d, and the cleaning liquid is cleaned by, for example, sucking and discharging 100 μL.

  In step S11 ′, the carrier-enclosed chip 26 is moved to the well 22a, and 20 μL of the food extract as a specimen is aspirated and discharged to the carrier-enclosed chip 26 to come into contact with the particles.

  In step S12 ′, the carrier-enclosed chip 26 is moved to the well 22e, and the buffer solution is aspirated and discharged 300 times for 80 μL, thereby performing incubation for 30 minutes at room temperature. At that time, if necessary, it is preferable to use a buffer solution whose temperature is controlled by a temperature controller described later.

  In step S13 ′, the carrier-enclosed chip 26 is moved to the well 22f, and the cleaning liquid is cleaned by sucking and discharging 100 μL. Similarly, the carrier is enclosed in the wells 22g and 22h containing the cleaning liquid. Washing is performed three times in total by moving the tip 26 and sucking and discharging 100 μl.

  In step S14 ′, the washed carrier-enclosed chip 26 is moved to the well 22b, and the labeled antibody accommodated in the well 22b is aspirated and discharged 300 times for 100 μL, and incubated for 30 minutes.

  In step S15 ′, the carrier-enclosed chip 26 is moved to the well 22i, and the washing liquid is washed by sucking and discharging 100 μL. Similarly, the carrier-enclosed chip 26 is moved to the well 22j, and the washing liquid is 100 μL. Then, the washed carrier-filled chip 26 is moved to the well 22c, and 60 μL of the substrate solution is sucked and discharged to cause chemiluminescence. A light absorber may be added to the chemiluminescent substrate solution. Alternatively, the light absorber-containing liquid is stored in any one or more of the wells 22d to 22i, the carrier-encapsulating chip 26 is moved to the wells, and the light absorption stored in the wells is absorbed. The body-containing liquid may be aspirated. In addition, the temperature of the chemiluminescent substrate solution or the well 22c that accommodates it may be controlled. Alternatively, the temperature of the carrier enclosing tip 26 may be controlled. The temperature control can be performed by a temperature controller described later.

  Since the following processing is the same as the above-described step S17 to step S21, the description thereof is omitted.

  Subsequently, a sample testing apparatus 70 according to the second embodiment is shown in FIG.

  The sample testing apparatus 70 is different in that a light measuring unit 77 is used instead of the light measuring unit 17 used in the sample testing apparatus 10 according to the first embodiment.

  The light measurement unit 77 has the photoelectric unit 32 provided with at least one photoelectric element, and a hole 76 into which the small diameter tube 26a of the carrier-enclosed chip 26 can be inserted, and is inserted through the hole 76. The light receiving ends 33a, 33b, and 33c of the optical fibers 37a, 37b, and 37c connected to the photoelectric unit 32 provided so as to surround the small-diameter tube 26a of the carrier-sealed chip 26 pass through the hole 76. And a scanning measurement unit 74 provided so as to be movable along the axial direction of the inserted small-diameter tube 26a. That is, at the time of measurement, the light receiving ends 33a, 33b, 33c are not fixed with respect to the housing 12, but are relatively movable, and the optical measurement according to the first embodiment. The part 17 is different.

  FIG. 16 shows a specimen testing apparatus 80 according to the third embodiment.

  The sample test apparatus 80 can exert and remove magnetic force mainly on the small diameter tube 25a of the dispensing tip 25, unlike the sample test apparatuses 10 and 70 according to the first and second embodiments. A magnetic force means 79 having a magnet 106 provided so as to be able to come in contact with and separate from the small-diameter tube 25, a temperature controller 82 for controlling the temperature of a well 96 described later provided in the inspection cartridge container 84, and The difference is that a lid moving mechanism 86 for closing the well 96 with the lid 92 is provided.

  The sample testing apparatus 80 is incorporated in the housing 12 in the same manner as the sample testing apparatuses 10 and 70 according to the first and second embodiments. In the housing 12, a chip storage unit 20 containing a plurality of types of chips (in this example, two types of dispensing tips 25 and 125 having different capacities and three types of drilling tips 27), a specimen, A plurality (10 in this example) of wells 22 that can contain or contain one or two or more reagent solutions used for the examination of the specimen, and wells 96 that are provided apart from the wells 22 and that perform temperature control are arranged in a line. A test cartridge container 84 that is provided in an array and has test information for identifying or managing the sample and test information indicating the test content displayed on a seal 94 as a visible recording medium and formed of a translucent member; An automatic inspection unit (85, 19) for obtaining a predetermined light emission by reacting the specimen contained in the inspection cartridge container 84 with the reagent, and a result of the inspection by the automatic inspection unit. A light measuring unit 177 for measuring the emitted light, a digital camera 28 for obtaining image data by photographing contents displayed on the test cartridge container 84 including the specimen information and the test information, and the test cartridge container 84 The thermal transfer printer 21 (see FIG. 9) as a writing mechanism capable of printing the inspection result in the blank of the seal 94, the magnetic force means 79, the temperature controller 82, the lid moving mechanism 86, and the automatic inspection unit (85, 19), an integrated circuit such as a CPU for controlling the light measuring unit 177, the digital camera 28, the thermal transfer printer mechanism 21, the magnetic force means 79, the temperature controller 82, and the lid moving mechanism 86. Board 52.

  As shown in FIGS. 9 and 10, the inspection cartridge container 84 is provided so that it can be manually pulled out of the housing 12 from the housing 12. The capacity of the well 96 that controls the temperature of the inspection cartridge container 84 is, for example, 0.2 cc.

  The automatic inspection unit (85, 19) is a moving mechanism 119 that can move the nozzle head 85 relative to the nozzle head 85 of the dispenser and the inspection cartridge container 84 housed in the housing 12. And have.

  The nozzle head 85 of the dispenser can be moved in the X-axis direction corresponding to the longitudinal direction with respect to the inspection cartridge container 84 housed in the housing 12 by the moving mechanism 119. A movable body 81 and a Z-axis movable body 83 provided so as to be movable while being guided by the guide pillar 111 in the vertical direction with respect to the X-axis movable body 81 are provided. A nut portion connected to the Z-axis moving body 83 is screwed into the X-axis moving body 81, and a Z-axis moving ball screw 113 (to be described later) that moves the Z-axis moving body 83 in the vertical direction is rotatable. At the same time, the guide pillar 111 and the support plate 89 attached via the guide pillar 111 are attached.

  The nozzle head 85 is attached to the Z-axis moving body 83 and drives a cylinder that sucks and discharges gas and a nozzle 100 that communicates via an air rubber tube 101 provided on a side surface, and a piston in the cylinder. Motor 110 and a ball screw 112 rotatably mounted.

  Further, the support plate 89 attached to the X-axis moving body 81 supports the ball screw 113 in a rotatable manner, and on the lower side thereof, the tip such as the dispensing tip 25 is detached from the nozzle 100. Therefore, a tip detachment plate 118 in which a U-shaped hole that is larger than the diameter of the nozzle 100 and thinner than the outer diameter of the thickest portion of the tip is formed, and the dispensing tip 25 attached to the nozzle 100 is thin. A magnet 106 which is provided so as to be able to contact with and separate from the diameter tube 25a, and which can exert a magnetic force from the outside and remove it in the small diameter tube 25a is movably supported in the front-rear direction. On the upper side of 89, a motor 108 for driving the chip attaching / detaching plate 118 and a motor 109 for driving the magnet 106 are attached to the X-axis moving body 81. The magnet 106 and the motor 109 correspond to the magnetic force means 79.

  The digital camera 28 is attached to the X-axis moving body 81 via a camera support plate 99, and the entire specimen information and examination information on the seal 94 of the examination cartridge container 84 housed in the housing 12. The nozzle head 85 is moved to a position where it can be photographed.

  A moving mechanism 119 for moving the nozzle head 85 of the dispenser with respect to the inspection cartridge container 84 housed in the housing 12 is engaged with the X-axis moving body 81 of the nozzle head 85 so as to perform the inspection. A rail 44 that guides along the longitudinal direction of the cartridge container 84, that is, the X-axis direction, an X-axis movement motor 58 (see FIG. 9) that moves the nozzle head 85 along the X-axis direction, and the X-axis movement It has the guide column 111 for guiding the body 83 in the vertical direction, that is, the Z-axis direction, the Z-axis moving ball screw 113, and the Z-axis moving motor. The ball screw 112 and the motor 110 correspond to a suction / discharge mechanism. The guide pillar 111, the Z-axis moving ball screw 113, and the Z-axis moving motor correspond to the Z-axis moving mechanism in the moving mechanism.

  Note that the sample inspection apparatus 80 according to the present embodiment also includes a thermal transfer printer mechanism 21 as a writing mechanism. The thermal transfer printer mechanism 21 is as described above.

  The lid moving mechanism 86 is provided with a lid 92 for covering the opening of the well 96, the lid 92 at one end, and the other end pivotally supported on a rotating shaft so that the rotating shaft can rotate 90 degrees. It has an arm 93 and a rotation drive unit 95 provided with a motor for driving the rotation shaft.

  Further, the specimen testing apparatus 80 further uses the moving mechanism 119 including the Z-axis moving mechanism to move the lid 92, which closes the opening of the well 96 of the test cartridge container 84, to the Z-axis direction, the X-axis direction, and the Y-axis. Using the nozzle 100 that can be pressed, shaken or moved along the direction, the lid 92 can be pressed, vibrated or moved. In other words, the nozzle 100 driven by the moving mechanism 119 including the Z-axis moving mechanism corresponds to a lid closing mechanism. At this time, it is preferable that the lid 92 is supported by being urged by an elastic force in the Z-axis direction with respect to the rotating shaft.

  As shown in FIG. 17, the temperature controller 82 has a temperature at which a tapered fitting hole having a shape and a size that fits with the well 96 of the test cartridge container 84 is formed in the center as the well receiving hole. A control block 98; a Peltier element part 97 provided with a Peltier element as the heating and cooling part provided in contact with the temperature control block 98; and a fin 103 provided below the Peltier element part 97 And an optical fiber for irradiation 74a that extends from the bottom of the fitting hole through the Peltier element portion 97 and through the fin 103, and the six optical fibers 74a for irradiation. The light receiving optical fiber 74b and one end of the irradiation optical fiber 74a are connected to the excitation light source 75b, and one end of the light receiving optical fiber 74b is connected to the photomultiplier tube 72. The other ends 74c of the optical fibers 74a and 74b are bundled around the irradiation optical fiber so that the tip is located at the bottom of the fitting hole as the well receiving hole. It has been.

  Here, the optical fibers 74a and 74b pass through the fiber accommodating portion 174 of the light measuring portion 177 and are connected to the excitation light source 72a and the photomultiplier tube 72b built in the photoelectric / light source portion 72. Yes.

Subsequently, an operation of the specimen testing apparatus 80 according to the third embodiment will be described.
In steps S31 to S38, except that the nozzle head 85 is used instead of the nozzle head 15, the nozzle 100 is used instead of the nozzle 30, and the inspection cartridge container 84 is used instead of the inspection cartridge container 14. As described in steps S1 to S8.

In the state of FIG. 14D, the following processing is performed.
Here, instead of performing the allergy test described in the first embodiment, the operation in the case of performing PCR processing by controlling the temperature of DNA or genome will be described.

For example, a specimen such as an oral mucosa collected from a subject is accommodated in the well 22a of the test cartridge container 84. The well 22b contains a reagent for genome extraction.
The well 22c contains a magnetic particle suspension. The well 22d contains a separation liquid. The well 22e is empty. The wells 22f to 22i contain a primer-containing solution and a cleaning solution as PCR reagents. The well 22j contains mineral oil. In addition, two types of dispensing tips 25 and 125 and a drilling tip 27 are accommodated in the tip accommodating portion 20. The light absorber-containing liquid may be stored in any one or more of the wells 22f to 22i. Or you may add a light absorber to a primer containing liquid.

  In step S39, the nozzle 100 is moved to the position of the dispensing tip 25 accommodated at the end of the tip accommodating portion 20, and lowered to attach the nozzle 100 for genome extraction, and the dispensing tip 25 is moved to the well 22b by the moving mechanism 119, and the corresponding extraction reagent is aspirated by using the suction / discharge mechanism. The dispensing tip 25 is moved to the well 22a in which the specimen is accommodated, and the liquid sucked into the dispensing tip 25 is discharged into the well 22a. Further, the dispensing tip 25 is moved to the well 22c, the magnetic particle suspension is sucked, the dispensing tip 25 is moved to the well 22a and discharged, and there are other reagents necessary for extraction. For example, they are transferred to the well 22a using the dispensing tip 25 and discharged. These mixed solutions accommodated in the well 22a are reacted by stirring and incubating by repeating suction and discharge, and the extracted DNA is bound to the surface of the magnetic particles and captured.

  In step S40, by using the magnetic force means 79, the magnet 106 is brought close to the small diameter tube 25a of the dispensing tip 25 to apply a magnetic field to the inner wall of the small diameter tube 25a. DNA is separated by adsorption.

  In step S41, the dispensing chip 25 for genome extraction is moved by the moving mechanism 119 while the magnetic particles capturing the DNA are adsorbed on the inner wall, and is positioned in the well 22d that contains the detachment liquid. The tip of the dispensing tip 25 is inserted into the well 22d, and the DNA is separated from the magnetic particles by repeating suction and discharge while adsorbing the magnetic particles to the inner wall. After the DNA solution containing DNA deviated from the magnetic particles is discharged and stored in the empty well 22e, the chip-accommodating portion is kept with the magnetic particles adsorbed on the inner wall of the dispensing chip 25 for genome extraction. It is transferred to the original storage position of 20 and is detached using the chip detaching plate 118.

  In step S42, the nozzle head 85 is moved, and the nozzle 100 of the nozzle head 85 is moved to the new dispensing tip 125 for PCR accommodated in the middle position of the chip accommodating portion 20. Thereafter, the nozzle 100 is lowered by the Z-axis moving mechanism, so that the nozzle 100 is inserted and mounted in the mounting opening of the accommodated dispensing tip 125 for PCR.

  In step S43, the nozzle head 85 is moved to rotate the arm 93 by 90 degrees as shown in FIG. 17, so that the lid 92 opens the opening of the well 96 and then the opening of the well 96. Is exposed to the outside. Next, a PCR reagent, for example, a primer-containing solution labeled with a fluorescent substance, contained in the wells 22f to 22i is aspirated using the PCR dispensing chip 125, and the well 96 is aspirated. It is accommodated by dispensing inside. The above steps are repeated until the necessary reagents are dispensed.

  In step S44, after the dispensing tip 125 is washed, the nozzle head 85 is moved, and the extracted DNA solution accommodated in the well 22e is sucked and dispensed into the well 96. Thereafter, the dispensing tip 125 is used to move to the well 22j, and the mineral oil is sucked and discharged into the well 96 for introduction.

  In step S45, the lid 92 is rotated 90 degrees to cover the opening of the well 96.

  In step S46, the nozzle 100 is lowered and the lid 92 is pressed using the Z-axis moving mechanism.

  In step S47, the temperature control is performed on the well 96 by the temperature controller 82 according to the PCR method. The temperature control according to the PCR method is such that the temperature of the well 96 is set at 94 ° C. in order to denature the input double-stranded sample DNA into single-stranded DNA, and then the single-stranded DNA and In order to perform annealing or hybridization with the primer, the temperature of the well 96 is set to 50 ° C. to 60 ° C. Next, in order to synthesize a DNA strand complementary to the single strand, the operation of setting the temperature to 74 ° C. and incubating is set as one cycle, and is repeated a predetermined number of times, for example, for about several minutes. I do. A light absorber may be added to the primer-containing liquid. Alternatively, the light absorber-containing liquid is accommodated in any one or more of the wells 22f to 22i, the carrier-enclosed chip 125 is moved to the wells, and the light absorption accommodated in the wells is absorbed. The body-containing liquid may be aspirated.

  At that time, excitation light is irradiated using optical fibers 74a and 74b provided in fitting holes as well receiving holes of the temperature control block 98, and the generated fluorescence intensity is received through the optical fiber 74b. The fluorescence intensity is measured by being changed into an electric signal by the multiplier 72b.

  In step S48, the measurement result is analyzed by the control unit of the board 52, the measurement result is sent to the thermal transfer printer mechanism 21, and the measurement result is added to the remarks column of the seal 24 by the print head 21a. It is printed as one piece of information and displayed by numbers.

  In step S <b> 49, the digital camera 28 shoots image data including specimen information and examination information on the seal 94 of the examination cartridge container 84 according to an instruction signal from the board 52. At this time, the analysis unit of the control unit searches for data that can be analyzed from the image data, and when the two-dimensional barcode data indicating the inspection content included in the inspection information is found in the image data, the 2 The analysis data is obtained by analyzing the dimensional barcode data, and the data combination unit of the control unit stores the analysis data and the image data in a memory that can be output in combination.

  In step S50, the dispensing tip 125 attached to the nozzle 100 moves to the tip accommodating portion 20, and the dispensing tip 125 is just above the position where the dispensing tip 125 was accommodated. After moving the U-shaped groove of the chip attaching / detaching plate 118 to the nozzle 100, the nozzle 100 is moved upward to move the nozzle 100 into the cylindrical body 20b of the chip accommodating portion 20. The tip 125 is removed.

  When the specimen test is completed in step S51, the loading box 18 loaded with the test cartridge container 84 is manually pulled out from the housing 12, and the seal 94 attached to the test cartridge container 84 is peeled off. The test cartridge container 84 itself is discarded by being attached to a management board or the like prepared separately, but the new test cartridge container 84 is loaded in the casing 12 to test a new specimen. Can be performed. According to the present embodiment, the lid 92 can be pressed using the moving mechanism, so that the opening of the well 96 is reliably closed, and condensation is prevented to open the lid 92. It can be done easily.

18 and 19 show a specimen testing apparatus 180 according to the fourth embodiment.
In addition, about the thing same as the specimen test | inspection apparatus 80 shown in FIG. 16, description is abbreviate | omitted with or without attaching | subjecting the same code | symbol.

  This sample testing apparatus 180 is different from the sample testing apparatus 80 according to the third embodiment shown in FIG. 16 in that the nozzle head 185 communicates with a cylinder that performs gas suction and discharge via an air rubber tube 201. The nozzle 200 to which the chip 25 or the like can be attached and the Z-axis moving body 83 that can move in the Z-axis direction are interlocked with the nozzle support 183 to which the nozzle 200 is attached, and the nozzle support 183 is attached to the inspection. An end of the light receiving optical fiber 174a and an end of the irradiating optical fiber 174b are provided inside to perform light measurement from above the light-transmitting lid 192 that covers the opening of the well 96 of the cartridge container 184. It differs in that it has a measuring rod 172 (see FIG. 19).

  The sample testing apparatus 180 is different from the sample testing apparatus 80 according to the third embodiment in addition to the above, and instead of having the lid moving mechanism 86, the lid 192 is placed in the chip housing portion 120 of the test cartridge container 184. Instead of the carrier enclosing tip 26, the nozzle 200 and the measuring rod 172 are attached to the tip of the nozzle 200 or the tip of the measuring rod 172 by lowering the nozzle 200 and the measuring rod 172 by the Z-axis moving mechanism. It is used at the time of pressing, etc. or at the time of measurement. Therefore, the inspection cartridge container 184 is also different in that the lid 192 can be accommodated in the chip accommodating portion 120.

As shown in FIG. 19, the light measurement unit 277 and the temperature controller 182 are also different from the light measurement unit 177 and the temperature controller 82 according to the third embodiment.
The light measuring unit 277 is provided with an end of the light receiving optical fiber 174a and an end of the irradiation optical fiber 174b in the measuring rod 172, and the other end of the light receiving optical fiber 174a is: Connected to the photoelectric element 172a, the other end of the irradiation optical fiber 174b is connected to the light source 172b.

  The temperature controller 182 includes a temperature control block 198 in which a tapered fitting hole having a shape and a size that fits into the well 96 of the test cartridge container 184 is formed in the center as a well accommodation hole; A Peltier element part 197 provided with a Peltier element as the heating / cooling part provided in contact with the temperature control block 198, a fin 203 provided on the lower side of the Peltier element part 197, It has only the fan housing frame 102 provided on the side, and the end of the optical fiber is not provided at the bottom of the fitting hole, and the optical fiber does not pass through the fins 203 and the like.

  20 and 21 show the lid 192. FIG. The lid 192 includes a mounting opening 193 into which the measuring rod 172 and the nozzle 200 can be mounted, and a fitting portion 194 that fits into the opening of the well 96. According to the apparatus according to the present embodiment, the opening of the well 96 can be closed with the lid without providing a lid moving mechanism, and thus the structure of the apparatus can be simplified. Further, since the lid that may be contaminated by the specimen is housed in the test cartridge container, it can be disposed of together with the test cartridge container like a chip after the test is completed, so that highly safe management can be performed.

Next, a sample testing apparatus according to the fifth embodiment will be described with reference to FIG.
The specimen testing apparatus 280 according to the present embodiment is, for example, in a casing having a length of 250 to 400 mm (X axis direction), a width of 140 to 200 mm (Y axis direction), and a height of about 300 to 500 mm (Z axis direction). The provided chip accommodating portions 220a, 220b, and 220c accommodating a plurality of types (three types in this example) of chips as a sample and one or more test instruments used for testing the sample are arranged in a line. Two test cartridges 284 in which the sample information for identifying or managing the sample and the test information indicating the test contents are displayed on the seal 224 as a visible recording medium and arranged in parallel, the sample and the sample A plurality of (in this example, 10) wells 322, each containing one or two or more reagent solutions or the like used for testing, are provided in a line and identify or manage the specimen. Two test cartridge containers 384 in which the specimen information and the test information indicating the test contents are formed of a translucent member displayed on the seal 324 as a visible recording medium and arranged in parallel, and the two test cartridge containers An automatic inspection section (285, 289) for obtaining a predetermined optical state (for example, luminescence) by reacting the specimen contained in 384 with the reagent, and the optical generated as a result of the inspection by the automatic inspection section A light measuring unit for measuring the state, a digital camera 228, a thermal transfer printer mechanism capable of printing the inspection result in blanks of the seals 224, 324 of the inspection cartridge containers 284, 384, and the automatic inspection unit (285, 289) An integrated circuit such as a CPU for controlling the light measurement unit, digital camera 228, and thermal transfer printer mechanism. Board. Individually, reference numeral 285a mainly indicates a unit provided with a Z-axis moving mechanism for moving the nozzle 230 in the Z-axis direction.

  Here, the two cartridge containers 284 are a plurality of types (three types in this example) of chips as the inspection instrument, and a dispensing chip 225, a carrier-enclosed chip 226, and a drilling chip 227 each have a chip accommodating portion 220a. , 220b, 220c. Since the dispensing tip 225 is already attached to the nozzle 230 of the nozzle head 285, the accommodating portion 220a is empty.

  The two cartridge containers 284 are provided with openings of chip accommodating portions 220a, 220b, and 220c in the substrate 284a. A seal 224 provided with a specimen information column 224a and a test information column 224b indicating the test content is detachably pasted on a seal sticking region as a medium attachment portion of the substrate 284a. Here, the specimen information field 224a is pre-printed with a QR code and a handwritten entry field is provided, and the examination information field 224b is pre-printed with examination information, and a handwritten entry field and a blank space for printing. Is provided. Similarly, the two cartridge containers 384 are provided with wells 322a-322j containing ten reagent solutions and specimen solutions on the substrate 384a. A seal 324 provided with a specimen information column 324a and an examination information column 324b indicating examination contents is detachably attached to a sticker attaching region as a medium attachment portion of the substrate 384a. Here, a QR code is printed and a handwritten entry field is provided in the specimen information field 324a, and a test information field is printed and a handwritten entry field and a blank space for printing are provided in the examination information field 324b. Yes.

  Note that all cartridge containers 284 and 384 arranged in the sample testing apparatus have the same contents for the test information when used for the same test. In addition, the test cartridge containers 284, 384 arranged in one row (along the X-axis direction) have common sample information, but the test cartridge containers 284, 384 in the other rows have different samples. In this case, the sample information is different from the sample information.

  As the automatic inspection unit (285, 289), two nozzles 230, 230 are provided, and a dispensing tip 225 is detachably attached to each nozzle 230, and each dispensing tip 225 is arranged in the two rows. The cartridge containers 284 and 384 are movably provided. Reference numerals 244a and 244b are rails for moving the nozzle head 285 in the X-axis direction and belong to the moving mechanism 289.

  Here, the digital camera 228 covers the two rows of test cartridge containers 284 and 384 by providing the digital camera 228 so as to be rotatable at a constant angle by a rotation mechanism 228a having a rotation axis along the X-axis direction. Can do. In addition, the light measuring unit, the thermal transfer printer mechanism, and the light measuring unit are also provided so as to be movable in the Y-axis direction, so that one unit can correspond to two rows of inspection cartridge containers, and the scale of the apparatus. Is compactly formed. According to this embodiment, since a plurality of inspections can be processed in parallel, efficient and quick processing can be performed.

  Each of the embodiments described above is specifically described for better understanding of the present invention, and does not limit other embodiments. Therefore, changes can be made without changing the gist of the invention. For example, in the above-described embodiment, only the case of allergen substances, foods, and DNA has been described. However, the present invention can naturally be applied to tests of other proteins, sugar chains, DNA substances, RNA, and the like. Further, the numerical values, the number of times, the shape, the number, the amount, and the like used in the above description are not limited to these cases.

  Examples of the types of chips, lids, rods, and the like to be accommodated as the configuration of the test cartridge container, the structure or number of chips, lids, rods, the number or capacity of wells, or the contents of sample information and test information These can be appropriately changed according to the specimen and the contents of the examination.

  In addition, each of the above components, for example, each nozzle head, various chips, each lid, each nozzle, each temperature controller, each light measuring unit, each inspection cartridge container, or magnetic force means, etc. can be arbitrarily modified Can be combined.

For example, the test cartridge container having a well whose temperature is controlled and the temperature controller can be used while using the carrier-enclosed chip. In addition, the above-described reagents, specimens, and processing steps are examples, and it is of course possible to use other reagents, specimens, and processing steps.
In the above example, only the case where one or two rows of test cartridge containers are loaded into the sample testing apparatus has been described. However, the present invention is not limited to this case, and can be applied to three or more rows. Not too long. Also, when loading and using two rows of test cartridge containers, the present invention is not limited to this example, and it is of course possible to use the test cartridge containers used in the first embodiment side by side.

  EXAMPLES The present invention will be specifically described below with reference to examples, but the scope of the present invention is not affected by these.

In the present embodiment, the TSH detection by capillary total length of about 50 mm, for securing the dynamic range ^103, the fundamental verification of signal light localization was performed.

(Summary)
A chemiluminescence system using luminol, hydrogen peroxide, and enzyme (horseradish peroxidase (hereinafter “HRP”)) was used to detect the target substance. In this system, the causes of signal light diffusion are classified as follows.

A, optical diffusion
A-1, Diffusion involving capillary walls
A-2, Diffusion due to propagation in the reaction liquid (water)
A-3, measuring instrument resolution
A-4, other
B, material diffusion
B-1, Diffusion of the light emitter itself (activated luminol, etc.)
B-2, Diffusion of substances that catalyze luminescence (activated hydrogen peroxide, HRP, etc.)
B-3, Others In this example, A-1 and A-2 were verified for correlation with signal light diffusion. As a result, all the verified elements are factors of signal light diffusion, and quantitatively, B (probably B-2) was quantitatively the largest factor. Furthermore, for each factor,
B Immobilization of probe beads to capillaries during reaction and detection processes
B-2, Keeping the temperature low during substrate aspiration
A-2, Increasing the light shielding property of the reaction solution itself
A-1, Increase the transparency of the capillary
It was suggested that significant improvements in signal light diffusion can be expected by taking measures such as these.

(experimental method)
3-1 Verification experiment on optical diffusion
(1) The surface of φ1mm ZrO2 beads was coated with Asahi Pen Co., Ltd. nocturnal paint (light green). This was repeated three times, and burrs and surplus parts were removed as necessary.
(2) The beads were filled into a capillary (φ1.1 mm, L50 mm) made of polypropylene (1 mm haze value 25%) or polycarbonate (1 mm haze value 0.1%) depending on the experimental conditions. At this time, from the capillary holder side, 20 light shielding beads / 1 night probe bead / 10 light shielding beads were arranged in this order.
(3) The capillary holder side and the suction port side were caulked, and liquid was sucked according to the experimental conditions.
(4) Light was irradiated as needed to accumulate the luminous probe beads.
(5) This was mounted on LuBEA (Precision System Science), and the detection-only protocol was executed to measure the emission intensity distribution.

3-2 Verification experiment on substance diffusion Preparation of antibody solid layer and BIST on probe beads (1) Preparation of beads supporting TSH antibody A spherical carrier (bead) is immersed in a buffer containing an appropriate amount of TSH antibody. Thus, the antibody was adsorbed to the beads. After the washing step, the beads carrying the antibody were immersed in a buffer solution containing an appropriate amount of a commercially available blocking agent to perform a blocking treatment. Antibody-bearing beads were prepared by washing several times with a buffer solution.

(2) Production of light-shielding beads The light-shielding beads were immersed in a buffer solution containing an appropriate amount of a commercially available blocking agent to perform a blocking treatment. The light-shielding beads were prepared by washing several times with a buffer solution.

(3) Production of BIST
After caulking the lower end side of the BIST capillary, 10 light-shielding beads, 1 antibody solid layer bead, 10 light-shielding beads or 10 light-shielding beads were filled in this order, and the upper end side was caulked. The caulking position on the upper end side was a position where the beads did not move in the bead arrangement 40-1-10, and the caulking position in the case of the bead arrangement 10-1-10 was almost the same as that of 40-1-10.

Reaction conditions
(1) As shown in Table 1, the reagent was added to the GC series cartridge and set in the SX-12GC. BIST was set together with Sheath DN-100 on Hole-2 of SX-12GC combo rack. (“BIST-ELISA_ver.5BS01.pcl” is used for “well mixed” and “Not mixed”, and “BIST-ELISA_ver.5BS01-fixedbead.pcl” is used for “Fixed bead”). The reaction is completed in about 110 minutes.
(2) After the reaction was completed, the well 1 solution was filled in the capillary and stored until measurement. The solution in the capillary was removed immediately before the measurement, and then 80 ul of a detection reagent (SuperSignal West Femto Maximum Sensitivity Substrate manufactured by Thermo Fisher Scientific) was aspirated. Thereafter, the measurement was immediately performed with LuBEA, or mixed after inverting and rotating 3 times and then measured.

(result)
4-0 Control The signal light intensity distribution of BIST was measured according to the procedure of 3-2 using TSH (13uIU / mL) with a light emission intensity of about 10,000 counts. In this experiment, capillary: made of polypropylene, probe beads: ZrO2 (lot ID 4), and light shielding beads: SiC were used.

In Fig. 1 (a), in order to compare the diffusion of signal light at a certain distance from the probe beads, the peak emission intensity is normalized to 10,000 counts, and at the same time, the raw data is shown in Fig. 1 (b). Described in full scale. The horizontal axis in the figure schematically overlaps the outline of the arrangement of actual probe beads and light shielding beads.

In FIG. 1 (a), since is normalized to the emission intensity of the signal light to 10,000 counts, in order to ensure a 10 ³ dynamic range, it is one that can be satisfactorily detected emission intensity 10 counts in the same capillary in It becomes a standard. In contrast, the current diffusion of signal light was at a level that exceeded 10 counts of emission intensity for both the baseline and noise level (variation of the baseline). Also, in the capillary, the intensity of the detected light was higher in the region where the beads were not present (non-bead region) than in the region where the beads were present (bead region).

4-1 Verification of optical diffusion 4-1-1 Light diffusion involving capillary walls (A-1)
Two types of capillaries with different transparency (polypropylene (PP): relatively low transparency) and polycarbonate (in order to verify the degree of light diffusion involving the capillary wall, such as light propagation in the capillary wall and reflection on the capillary wall, and polycarbonate ( PC): comparatively high transparency), the BIST signal light intensity distribution was measured by the procedure of 3-1, which can ignore the diffusion of the substance. In this experiment, light shielding beads: SiC was used.

  In order to compare the diffusion of signal light at a certain distance from the probe bead in Fig. 2, the peak emission intensity is normalized to 10,000 counts, and at the same time the actual probe bead is shown on the horizontal axis. And the outline of the arrangement of the light shielding beads are schematically overlapped.

  As a result, in both PP and PC capillaries, the signal light diffusion was at a low level in comparison with the data in FIG. 1, which is expected to include material diffusion. In addition, PC signal light diffusion was lower than that of PP.

4-1-2, Verification of Light Diffusion (A-2) due to Propagation in Reaction Solution (Water) To verify the degree of signal light diffusion that propagates in the reaction solution (water) that fills the capillary, In the procedure of 3-1, which can ignore the diffusion,
・ Differences in signal light intensity distribution due to the presence or absence of water in the capillary ・ Effects of light transmittance suppression due to coloring of the reaction solution were evaluated.

In this experiment, capillary: PC and light shielding beads: SiC were used, and no frequency divider was used to collect data from PMT. Moreover, as the coloring reaction solution, an aqueous solution obtained by diluting “Kitake for Kiyosho” (black ink) by Kuretake Co., Ltd. 200 times was used.

  In order to compare the diffusion of signal light at a certain distance from the probe bead, Fig. 3 shows the peak emission intensity normalized to 10,000 counts. At the same time, the horizontal axis in the figure shows the actual probe bead. And the outline of the arrangement of the light shielding beads are schematically overlapped.

  As a result, when the water in the capillary was discharged (air), the signal light diffusion was suppressed to a lower level than when the capillary was filled with water (water). Furthermore, the improvement of the light shielding property by dyeing the reaction solution in black (Black water) was most effective for the localization of the signal light.

4-1-3 Summary of verification of optical diffusion From basic verification of signal light diffusion of 4-1-1, 4-1-2, 1) Increase the transparency of the capillary 2) Reaction solution (water) It was suggested that suppressing the diffusion of the signal light propagating through the inside is effective for the localization of the signal. With PP & Water in Fig. 4, it is difficult to detect 10 counts of luminescence intensity in the same capillary (probe bead interval, within about 20), but it was detectable with PC & Air and PC & Black water.

4-2 Verification of substance diffusion 4-2-1 Influence of bead movement As shown in Fig. 1 (a), which is seen when detecting relatively high concentrations of TSH whose detected light intensity exceeds 10,000 counts. As a cause of turbulent diffusion of signal light, diffusion of a substance (a luminescent material or a substance that catalyzes luminescence) can be considered. Therefore, we focused on the movement of the probe beads inside the capillaries in the reaction process as one factor that promotes the diffusion of substances. In the current BIST configuration, beads can move freely within a certain range in the capillary. In order to verify the correlation between the movement of the beads during the reaction process and the diffusion of signal light, the BIST is adjusted by the procedure in 3-2 and the three protocols shown in Table 2, and the distribution of signal light during TSH detection is adjusted. Measured. In order to compare the diffusion of signal light at a certain distance from the probe beads in Fig. 5, the peak emission intensity is normalized to 10,000 counts, and at the same time the actual probe beads are shown on the horizontal axis. And the outline of the arrangement of the light shielding beads are schematically overlapped.

As a result, the current standard protocol “well mixed” showed a turbulent diffusion of signal light exceeding 50 counts in the non-bead region, whereas “Not mixed” and “Fixed bead” This portion was greatly reduced regardless of the bead region and non-bead region.

4-2-2 Effect of sucking reaction substrate TSH detection in a relatively high concentration range to confirm repetitive reproducibility of the reduction effect of signal light diffusion due to substance diffusion by suppressing bead motion in the capillary An experiment to be repeated was tried. The experiment was performed according to the procedure in 3-2, and the degree of signal light diffusion was measured at three levels of TSH concentrations of 13 uIU / mL (n = 5), 50 uIU / mL (n = 6), and 100 uIU / mL (n = 3). Verified. In order to compare the diffusion of signal light at a certain distance from the probe beads, Fig. 6 shows the peak emission intensity normalized to 10,000 counts. At the same time, the horizontal axis shows the actual probe beads. The outline of the arrangement of the light shielding beads was schematically overlapped.

 As is clear from FIG. 6, no signal light diffusion was observed at a position extremely away from the peak as seen in the experiment of 4-2-1 in which stirring was performed by inversion mixing. However, it has been clarified that by overlapping a plurality of data, fine characteristic noise appears sporadically on the left slope of the signal light peak (a region where z_puls is young).

  This result suggests that the suction of the reaction substrate may cause the illuminant diffusion. In the first place, in this detection system, chemiluminescence is enzymatically induced by the reaction between the enzyme immobilized on the probe beads and the substrate, and TSH is detected. In FIG. 6, since the reaction substrate is sucked from the right side of the peak (the side with the larger z_puls) to the left, all the reaction substrates located on the left side of the probe beads (the side with the smaller z_puls) are in contact with the probe beads. Will be. There may be a possibility that the illuminant is diffused only to the left side of the peak due to the momentary contact between the substrate and the probe beads during the substrate aspiration.

  In order to verify the hypothesis of noise generation due to instantaneous contact between the substrate and probe beads during substrate aspiration, the following experiment was attempted focusing on the temperature that greatly affects the enzyme activity during substrate aspiration. Substrate aspiration was performed at around 40 ° C where the enzyme activity on the probe beads was maximum and around 4 ° C where the enzyme activity decreased, and the occurrence of characteristic noise was compared. The results are shown in FIG. As in Fig. 6, in order to compare the diffusion of signal light at a certain distance from the probe bead, the peak emission intensity is standardized to 10,000 counts, and at the same time, the horizontal axis in the figure shows the actual probe bead and The outline of the arrangement of the light shielding beads was schematically overlapped.

  In FIG. 7, the red line represents data when the substrate is aspirated at 40 ° C. and the blue line is 4 ° C. As is clear from the figure, in the substrate suction at 40 ° C., significant noise was detected on the left side of the peak, whereas no noise was detected in the substrate suction at 4 ° C. This result supports the above-mentioned hypothesis and also shows that temperature control during substrate aspiration has a significant effect on noise reduction.

(Discussion)
From the verification experiments of 4-1, 4-2, it was suggested that the factor of signal light diffusion seen in the TSH detection system is roughly divided into optical diffusion and material diffusion. In addition, from the verification of 4-2, it was shown that the diffusion of the signal light could be significantly improved by suppressing the movement of the beads in the capillary and optimizing the temperature when sucking the substrate. However, if the effective signal intensity in the measurement is defined as 10 times the S / N ratio, even in the improvement of 4-2, it is possible to detect the emission intensity of 10 counts in the same capillary. I don't think so.

Therefore, in order to estimate the influence of the optical diffusion component verified in 4-1 among the remaining signal light diffusion factors in FIGS. 5 to 7, the “PP & water” in FIGS. The signal light distribution in a system in which PP capillaries are filled with luminescent beads and light-shielding beads and the interior is filled with water is superimposed on FIGS. 5 to 7 (FIG. 8).

In order to compare the diffusion of signal light at a certain distance from the probe beads, Fig. 8 shows the peak emission intensity normalized to 10,000 counts. At the same time, the horizontal axis in the figure shows the actual probe beads. And the outline of the arrangement of the light shielding beads are schematically overlapped.

  As shown in Fig. 8, the remaining signal light diffusion (z_puls = 50-200 region) in “Not mixed” and “Fixed bead”, which suppresses the movement of beads in the capillary and greatly suppresses the diffusion of the substance, is optical. It is in good agreement with “PP & water”, which has only local diffusion. From this, it is considered that the signal light diffusion derived from the diffusion of the substance is largely reduced by the suppression of the bead movement in the capillary. In addition, as verified in 3-2, A-1) optical diffusion derived from capillaries, A-2) suppression of optical diffusion propagating in the reaction solution greatly increases the dynamic range of BIST measurement. Can be increased.

  The method and apparatus of the present invention can be used for detection and quantitative analysis of a plurality of biomolecules and small molecules that bind to biomolecules.

10, 70, 80, 180, 280 Sample inspection device
14, 84, 184, 284, 384 Inspection cartridge container 15, 85, 185, 285 Nozzle head 17, 77, 177, 277 Light measuring unit 24, 94, 224 Seal 25, 125, 225 Dispensing tip 26, 226 Carrier enclosed Chip (chip with built-in solid phase)
28, 228 Digital camera 30, 100, 200, 230 Nozzle 92, 192 Lid

Claims (20)

A plurality of carriers housed in a single container, each of which is fixed with a substance for capturing a plurality of types of target substances, emit light while being immersed in a liquid, and measure this light emission from the outside of the container. An analysis method for simultaneously detecting or quantifying a plurality of types of target substances, wherein the light transmittance of the liquid in the emission wavelength band is suppressed in order to suppress optical crossing of emission signals from the plurality of carriers. Or said method being adjusted. The method according to claim 1, wherein the emission is chemiluminescence or fluorescence, and the emission wavelength band is 340-900 nm. The method according to claim 1 or 2, wherein the light transmittance of the liquid is suppressed or adjusted by the addition of a light absorber. The method according to claim 3, wherein the light absorber is a fine particle. The method according to claim 1, wherein the light transmittance of the liquid is suppressed or adjusted so as to be 80% or less per 1 mm of the optical path length. By geometrically defining the optical path length between the photometric unit and each of the plurality of carriers based on the relative positional relationship between the surface shape of the container and the carrier and the photometric unit located outside the container, The method according to claim 1, wherein the light emission signal is selectively received. The method according to claim 1, wherein the carrier is spherical. The method according to claim 1, wherein the carriers are arranged one-dimensionally or two-dimensionally. The method according to any one of claims 1 to 8, wherein a constituent material of the container is 20% or less in terms of a haze value having a thickness of 1 mm. The method according to any one of claims 1 to 9, wherein the refractive index of the container in the wavelength band of light emission or radiation is 1.6 or less. The test kit for using for the method in any one of Claims 1-10 containing the some support | carrier which each accommodated the substance for capturing the multiple types of target substance accommodated in the single container. A substance for capturing the target substance is fixed to a carrier accommodated in the container, and the substance emits light through a reaction with a substance having a light-emitting ability contained in a liquid added from the outside. A method for detecting or quantifying a target substance by emitting or emitting a substance having a function, which comprises adjusting the temperature of the liquid and / or the container when the liquid is added. The substance having a light-emitting ability is at least one compound selected from the group consisting of luminol, diphenyl oxalate, ruthenium complex, oxalyl chloride, luciferin, AMPPD, indole, polyphenol, dioxetane, and derivatives of these substances. 12. The method according to 12. The method according to claim 12 or 13, wherein the temperature of the liquid and / or the container is adjusted to 15 ° C or lower. The method according to any one of claims 12 to 14, wherein the carrier is fixed with respect to the container when the liquid is added. 11. The apparatus according to claim 1, comprising means for suppressing or adjusting light transmittance of a liquid in a wavelength band of light emission or radiation, and means for optically detecting and / or quantifying a target substance by light emission or radiation phenomenon. The apparatus used for the method of crab The apparatus used for the method in any one of Claims 12-15 provided with the means to adjust the temperature of a liquid, and the means to detect or quantify a target substance by light-emitting or radiating | emitting the substance which has a luminescent ability. The apparatus of claim 17, further comprising means for adjusting the temperature of the container. The apparatus used for the method in any one of Claims 12-15 provided with the means to adjust the temperature of a container, and the means to detect or quantitate a target substance by the light emission or emission of the substance which has a luminescent ability. The apparatus of claim 19 further comprising means for adjusting the temperature of the liquid.