WO2007018187A1

WO2007018187A1 - Measurement instrument, measurement kit using the same, measurement method, measurement device, and piezoelectric oscillator reproducing method

Info

Publication number: WO2007018187A1
Application number: PCT/JP2006/315614
Authority: WO
Inventors: Haruki Tsunoda; Akihito Tomita; Takaaki Nozaki
Original assignee: Kyowa Medex Co., Ltd.; Citizen Holdings Co., Ltd.
Priority date: 2005-08-05
Filing date: 2006-08-07
Publication date: 2007-02-15

Abstract

When a piezoelectric oscillator is arranged at a detection portion of a reaction vessel, a piezoelectric oscillator sensor film can be easily generated and the sensor can easily be reproduced, thereby enabling re-use. A measurement instrument (1) includes a reaction vessel (2) capable of supplying a measurement object (10) in a sample. The reaction vessel (2) includes a piezoelectric oscillator (6) arranged at a detection portion (5) capable of detecting a measurement object (10); a magnetic force generation material (7) capable of selectively and reversibly forming a magnetic field in the piezoelectric oscillator (6); and a carrier (11) formed by magnetic particles (11a) and a trapper (11b) capable of specifically binding the measurement object (10) or formed by the magnetic particles (11a) and an object (11c) similar to the measurement object and magnetically carried on the surface of the piezoelectric oscillator (6) when a magnetic field is applied to the piezoelectric oscillator (6). There are also provided a measurement kit using the measurement instrument (1), a measuring method and device for measuring the concentration of a measurement object, and a piezoelectric oscillator reproducing method.

Description

Specification

Measuring instrument, measuring kit using the same, measuring method, measuring device, and method of regenerating piezoelectric vibrator

Technical field

[0001] The present invention relates to a measuring instrument for measuring a measurement object contained in a sample to be analyzed such as a living body, food, or soil, and in particular, has a piezoelectric vibrator at a detection site of a reaction vessel, BACKGROUND ART Related to a measuring instrument effective in an aspect in which a reaction with an object to be measured is performed on the surface portion of the piezoelectric vibrator, a measuring kit, a measuring method, a measuring apparatus using the same, and a method for regenerating a piezoelectric vibrator

Conventionally, quantitative analysis and qualitative analysis using a piezoelectric vibrator such as a quartz vibrator are known. For example, as a method of loading a target substance on the electrode surface of a quartz vibrator, SA M (Self Assembled Monolayer ) Method or the like is used (for example, see Non-Patent Document 1). Further, a method of reusing a crystal resonator sensor after supporting a target substance on the crystal resonator is known. For example, as a method of regenerating the sensor film after binding the target substance to the surface of the crystal unit, the target substance is chemically treated using, for example, pyranno, solution (concentrated sulfuric acid: 30% hydrogen peroxide = 3: 1). (For example, see Non-Patent Document 1) and a method of reusing a crystal resonator sensor by separating smoke particles (adsorbed molecules) adsorbed on the crystal resonator using a hot air source (For example, see Patent Document 1) and a method of reusing the crystal resonator sensor by making the surface of the crystal substrate in the crystal resonator sensor flat to facilitate cleaning of this surface (for example, patent (Ref. 2) is known.

[0003] However, the SAM method has a technical problem that, for example, the process of generating a sensor film made of a predetermined target substance on the electrode surface of a crystal resonator is very complicated and time-consuming. . On the other hand, the quartz vibrator sensor uses an expensive gold electrode, so that it is desired to reuse the gold electrode. All of the conventional methods for reusing a quartz crystal sensor are: a sensor film generated on the surface of the crystal oscillator electrode, or thermal treatment by a hot air source. Since the separation process is complicated, for example, by giving a detachment, a simple method for reusing a crystal resonator sensor is desired.

In particular,

In recent years, point-of-care testing (POCT) has been carried out to output sample measurement results in the vicinity of a subject in accordance with the rapid and diversified testing methods and the flow of testing in the environmental and food fields. Therefore, the development of a simple and highly sensitive measuring instrument that can handle POCT, a measuring method and a measuring apparatus using the measuring instrument is desired.

[0004] Non-Patent Document 1: “Biochemistry” American Chemical Society, 1998, 37th, 5666- 5672

Patent Document 1: Japanese Patent Laid-Open No. 2000-275157

Patent Document 2: JP 2000-283905 A

Disclosure of the invention

Problems to be solved by the invention

[0005] The present invention has been made to solve the above technical problem, and in a mode in which a piezoelectric vibrator is disposed at a detection site of a reaction vessel, a piezoelectric vibrator sensor film can be easily generated. Another object of the present invention is to provide a measuring instrument, a measuring kit, a measuring method, and a measuring apparatus that can be reused by easily regenerating the sensor.

Means for solving the problem

That is, the present invention relates to the following (1) to (29).

(1) A carrier formed by binding a trapper that specifically binds to a measurement object or a measurement object analog and a magnetic particle, or a carrier formed by combining a measurement object analog and a magnetic particle. A measurement instrument for measuring a measurement object in a sample using a reaction vessel having a sample addition site and a detection site equipped with a piezoelectric vibrator, and for applying a magnetic field to the piezoelectric vibrator A measuring instrument comprising: a magnetic force generating member, wherein the carrier is magnetically supported on the piezoelectric vibrator by the magnetic force of the magnetic force generating member.

[0007] (2) The measuring instrument according to (1), wherein the magnetic force generation member also has means for releasing the magnetic field applied to the piezoelectric vibrator. (3) In the measuring instrument according to the above (1) or (2), the reaction container is bonded to the measurement object or the measurement object analog, or the binder or the measurement object analog. A measuring instrument comprising a binder or a labeled body supply site capable of supplying a labeled body formed by binding to an insoluble endoplasmic reticulum.

(4) In the measuring instrument according to (1) or (2), the reaction container is a flow cell type in which the measurement object and the support body can move along the flow path, and the flow path of the flow cell type reaction container A measuring instrument, wherein a sample addition site and a detection site are provided.

[0008] (5) In the measuring instrument according to (3), the reaction container is a flow cell type in which the measurement object, the support, and the label can move along the flow path. A measuring instrument comprising a sample addition site, a detection site, and a binder or label supply site in the flow path of the reaction vessel.

(6) The measuring instrument according to (5), wherein the binder or label supply site is provided downstream of the sample addition site in the flow path of the flow cell type reaction vessel.

(7) The measuring instrument according to (5), wherein the sample addition site also serves as a binder or label supply site.

(8) The measuring instrument according to any one of (4) to (7), wherein an absorption part is provided downstream of the detection part.

(9) A measuring instrument according to any one of (1) to (8), wherein a plurality of types of carriers are used.

(10) The measuring instrument according to any one of (1) to (9), wherein a plurality of piezoelectric vibrators are provided at a detection site.

(11) The measuring instrument according to (10), wherein a magnetic force generating member is provided for each piezoelectric vibrator.

(12) The measuring instrument according to any one of (1) to (11), wherein the piezoelectric vibrator is a crystal oscillator.

(13) The measuring instrument according to any one of (4) to (12), wherein the flow cell type reaction container includes a plurality of flow paths. (14) The measuring instrument according to any one of (5) to (13), wherein a noinder or a label is movably held in advance at a binder or label supply site.

(15) The measuring instrument according to any one of (1) to (14), wherein the carrier is magnetically supported on the surface of the piezoelectric vibrator in advance.

[0010] (16) A measuring kit comprising the measuring instrument according to any one of (1) to (13) and a carrier.

(17) A measuring kit comprising the measuring instrument according to any one of (1) to (13), a support, and a binder or a label.

[0011] (18) A reaction vessel having a sample addition site and a detection site equipped with a piezoelectric vibrator, and a trapper that specifically binds to a measurement object or a measurement object analog and magnetic particles are combined. The measurement object in the sample is measured using the support, and the measurement object in the sample and the support are reacted in a reaction container, and the composite including the support is measured. The sample reaction step to be generated, the magnetic force acting on the magnetic particles of the carrier, the carrier carrying step of magnetically carrying the magnetic particles on the piezoelectric vibrator, and the amount of the composite carried on the piezoelectric vibrator is piezoelectric The measurement target in the sample from the frequency measurement process measured as the change in the vibration frequency of the vibrator, and the change in the vibration frequency of the piezoelectric vibrator measured in this frequency measurement process and the calibration curve created in advance. It features a concentration determination process that determines the concentration of the product. Measurement method.

(19) The measurement method according to (18), further comprising a binder that specifically binds to the sample and the measurement object or the measurement object analog in the reaction container, or a small amount insoluble in the binder. A measurement method comprising: reacting a labeled body formed by binding with a cell body to generate a complex including a carrier, a measurement object, and a binder; and a carrier, a measurement object, and a complex including the label. .

(20) A reaction vessel having a sample addition site and a detection site equipped with a piezoelectric vibrator, and a trapper that specifically binds to a measurement object or a measurement object analog and magnetic particles are combined. The measurement object in the sample is measured using the carrier, and the measurement object in the sample and the analog of the measurement object are bound to the insoluble endoplasmic reticulum in the reaction container. The sample reacts with the carrier to form a complex containing the carrier, and the carrier carries the magnetic particles on the piezoelectric vibrator by applying a magnetic force to the magnetic particles of the carrier. A body supporting step, a frequency measuring step for measuring the amount of the composite supported on the piezoelectric vibrator as a change amount of the frequency of the piezoelectric vibrator, and a frequency of the piezoelectric vibrator measured in the frequency measuring step. A measurement method comprising a concentration determination step for determining the concentration of a measurement object in a sample from a change amount of the sample and a calibration curve prepared in advance.

[0014] (21) A reaction vessel having a sample addition site and a detection site equipped with a piezoelectric vibrator, a carrier formed by binding a measurement object analogue and magnetic particles, and a measurement object or measurement object A method for measuring a measurement object in a sample using a binder that specifically binds to an analog or a label formed by binding the binder to an insoluble endoplasmic reticulum. The object to be measured and the object to be measured and the magnetic object are bonded to each other. The carrier and the binder that specifically binds to the object to be measured or the object to be measured or the binder and the insoluble vesicle are reacted. , Support body Sample reaction process for generating a composite containing a binder or a support body-a label body, a support body support that applies magnetic force to the magnetic particles of the support body and magnetically supports the magnetic particles on the piezoelectric vibrator Process, supported on piezoelectric vibrator Frequency measurement step of measuring the amount of the composite as a change amount of the vibration frequency of the piezoelectric vibrator, and the change amount of the vibration frequency of the piezoelectric vibrator measured in this frequency measurement step and a calibration curve prepared in advance And a concentration determination step for determining the concentration of the measurement object in the sample.

[0015] (22) The measurement method according to any one of (18) to (21), further including a cleaning step of cleaning the complex supported on the piezoelectric vibrator. .

(23) The measurement method according to any one of (18) to (22), wherein a support carrying step is performed before the sample reaction step.

(24) In the measurement method according to any one of (18) to (23), the reaction vessel is a flow cell type in which the measurement object and the support can move along the flow path, and the flow cell type reaction is performed. A measurement method characterized in that the sample addition site and the detection site are provided in a flow path of a container.

(25) In the measurement method according to (24), the binder or the label is held in the flow path. And a sample reaction step is started by adding the sample to the sample addition site.

[0016] (26) The amount of the composite carried on the measurement instrument according to any one of (1) to (17) above and the piezoelectric vibrator at the detection unit of the measurement instrument is determined by the vibration of the piezoelectric vibrator. And a frequency measuring means for measuring the change in the number.

(27) The amount of the composite carried on the measurement device according to any one of (1) to (17) above and the piezoelectric vibrator at the detection unit of the measurement device is changed in the frequency of the piezoelectric vibrator. A frequency measuring means for measuring as a quantity, and a concentration determining means for determining the concentration of the measurement object in the sample from the amount of change in the vibration frequency of the piezoelectric vibrator measured by the frequency measuring means and a calibration curve prepared in advance. A measuring apparatus comprising:

(28) A support formed by binding a trapper that specifically binds to a measurement object or a measurement object analog and a magnetic particle, or a measurement object analog and a magnetic particle. A method for reproducing a piezoelectric vibrator in measurement of a measurement object in a sample using a carrier and a piezoelectric vibrator, comprising: a carrier and a measurement object carried on the piezoelectric vibrator by a magnetic force generating means. A method for regenerating a piezoelectric vibrator, comprising: measuring a composite body, and then dissociating the carrier by a magnetic field release means.

(29) The reproducing method according to (28), wherein the piezoelectric vibrator is a quartz crystal vibrator.

The invention's effect

[0017] According to the measuring instrument of the present invention, a support formed by binding a trapper that specifically binds to a measurement object or a measurement object analog and a magnetic particle, or a measurement object analog A measuring instrument for measuring an object to be measured in a sample using a carrier bonded to magnetic particles, the reaction container having a sample addition site and a detection site equipped with a piezoelectric vibrator, and this A magnetic force generating member for selectively applying a magnetic field to the piezoelectric vibrator is provided, and the carrier is magnetically supported on the piezoelectric vibrator by the magnetic force of the magnetic force generating member. By acting or canceling the magnetic field, a carrier functioning as a sensor film can be easily generated on the piezoelectric vibrator and can be removed. For this reason, in a measuring instrument equipped with a piezoelectric vibrator at the detection site, Thus, the piezoelectric vibrator sensor film can be easily generated, and the force can be reused by easily regenerating the sensor.

In addition, if the measurement kit includes a measuring instrument and a carrier according to the present invention, and further includes a noinder or a label, the measurement object can be measured very easily. Furthermore, according to the measuring method and apparatus using this type of measuring instrument, a measuring instrument that can easily generate a piezoelectric vibrator sensor film and regenerate the sensor can be used to measure a large number of items in a sample. It is possible to measure quickly and accurately for each object.

Furthermore, according to the method for reproducing a piezoelectric vibrator according to the present invention, the piezoelectric vibrator sensor can be easily reproduced.

Brief Description of Drawings

[FIG. 1] (a) is an explanatory plan view showing a basic configuration of a measuring instrument according to the present invention, and (b) is an explanatory view showing a detection site thereof.

[FIG. 2] (a) to (c) are explanatory views showing the measurement principle by the measuring instrument of FIG.

3 is an explanatory view showing a state after the reaction of the measuring instrument of FIG. 1 is completed.

[FIG. 4] (a) and (b) are explanatory views showing another embodiment of the measuring instrument according to the present invention and the measurement principle thereof.

FIG. 5 is an explanatory diagram showing a measurement principle of measurement in another aspect by the measurement instrument of FIG. 1.

[FIG. 6] (a) is an explanatory view showing an embodiment in which the present invention is applied to a measuring instrument including a flow cell type reaction vessel, and (b) is an arrow view seen from the M direction in (a).

[Fig. 7] (a) is an explanatory view showing a more preferred embodiment in which the present invention is applied to a measuring instrument including a flow cell type reaction vessel, and (b) is an arrow view seen from the M direction in (a).

[FIG. 8] (a) is an explanatory view showing a measuring method using the measuring instrument according to the present invention, and (b) is an explanatory view showing a measuring apparatus embodying the measuring method.

FIG. 9 is an explanatory view showing Embodiment 1 of a measuring apparatus to which the present invention is applied.

FIG. 10 is a schematic plan view of a measuring instrument used in Embodiment 1.

FIG. 11 is an explanatory diagram showing an equivalent circuit of the crystal resonator used in the first embodiment.

FIG. 12 (a) is an explanatory plan view showing a sensor substrate used in Embodiment 1, and FIG. 12 (b) is an arrow view as viewed from the M direction in (a). FIG. 13 (a) is an explanatory plan view showing the crystal resonator used in the first embodiment, and FIG. 13 (b) is a cross-sectional explanatory view taken along line MM in (a).

[14] FIG. 14 is an explanatory view showing Embodiment 2 of a measuring apparatus to which the present invention is applied.

FIG. 15 is a schematic plan view of a measuring instrument used in Embodiment 2.

16] An explanatory diagram showing a crystal oscillation circuit used in the second embodiment.

圆 17] An explanatory diagram showing a third embodiment of a measuring apparatus to which the present invention is applied.

18] An explanatory diagram showing a crystal oscillation circuit used in the third embodiment.

FIG. 19 is an explanatory view showing a modification of the crystal oscillation circuit used in the third embodiment.

[FIG. 20] (a) is a schematic plan view showing a measuring instrument used in Embodiment 4 of the measuring apparatus to which the present invention is applied, and (b) is an arrow view seen from the M direction in (a). .

FIGS. 21 (a) and 21 (b) are explanatory diagrams showing an operation example of the measuring instrument according to the fourth embodiment.

[FIG. 22] (a) is a schematic plan view showing a variation of the measuring instrument according to Embodiment 4, and (b) is (a

) Medium It is an arrow view seen from the M direction.

Explanation of symbols

1 Measuring instrument

2 reaction vessel (flow cell type reaction vessel)

3 Flow path

4 Sample addition site

5 Detection site

6 Piezoelectric vibrator

7 Magnetic force generating member

8 Binder Z label supply area

9 Absorption site

10 Measurement object

11 Carrier

11a Magnetic particles

l ib

11c Analogue to be measured 12 binder

13a Insoluble endoplasmic reticulum

13b binder

13c Analogue to be measured

13 Marker

15 Frequency measurement means

16 Concentration determination means

BEST MODE FOR CARRYING OUT THE INVENTION

That is, the representative embodiment of the measuring instrument according to the present invention specifically binds to the measurement object 10 (see FIG. 2 (a)) or the measurement object analog as shown in FIGS. L (a) and (b). A measuring instrument for measuring the measurement object 10 in the sample using the carrier 11 formed by binding the trapper l ib and the magnetic particles 11a, the measurement object 10 in the sample (Fig. 2) (see (a)) includes a reaction container 2 that can be supplied, and the reaction container 2 includes a piezoelectric vibrator 6 provided at a detection site 5 where the measurement target 10 can be detected, and the piezoelectric vibrator 6. A magnetic force generating member 7 that selectively and reversibly applies a magnetic field, and a carrier 11 that is magnetically supported on the surface of the piezoelectric vibrator 6 when the magnetic field is applied to the piezoelectric vibrator 6 are provided. It is characterized by.

[0021] Further, another representative embodiment of the measuring instrument according to the present invention is, as shown in FIGS. 4 (a) and 4 (b), a carrier formed by combining a measurement object analogue 11c and magnetic particles 11a. 11 is a measuring instrument for measuring the measurement object 10 in the sample, and includes a reaction vessel 2 to which the measurement object 10 in the sample can be supplied. When the piezoelectric vibrator 6 provided in the detection part 5 that can detect 10, the magnetic force generating member 7 that selectively and reversibly applies a magnetic field to the piezoelectric vibrator 6, and the magnetic field applied to the piezoelectric vibrator 6 And a carrier 11 that is magnetically supported on the surface of the piezoelectric vibrator 6.

Here, the difference between the former and the latter is that the former carrier 11 is a magnetic particle 11a and a trapper l ib, whereas the latter carrier 11 is a magnetic particle 1 la and a measurement object analogue 1 lc. It is a point.

In any embodiment, the trapper l ib or the measurement object analog 11c is used alone. These are not carried directly on the surface of the piezoelectric vibrator 6 but are carried on the surface of the piezoelectric vibrator 6 in the form of a carrier 11 combined with the magnetic particles 1 la.

Further, it is preferable that the magnetic force generating member 7 also has a means for releasing the magnetic field applied to the piezoelectric vibrator 6.

[0022] In such technical means, a typical embodiment of the reaction vessel 2 includes one having a sample addition site 4 where a sample is usually added (see FIGS. 6 (a) and 6 (b)).

In addition, as a typical embodiment of the reaction vessel 2, a labeled body 13 in which a binder 12 or a binder 13b or a target object analog 13c that specifically binds to a measurement object 10 is bound to an insoluble endoplasmic reticulum 13a can be supplied. And a binder Z labeled body supply site 8 (see FIG. 2 (b) (c), FIG. 5, FIG. 7 (a) (b)).

Further, the measuring instrument 1 of the present invention includes not only a flow cell type reaction vessel as long as it includes the reaction vessel 2, but also includes a well plate type reaction vessel.

[0023] In the flow cell type reaction vessel 2, as shown in FIGS. 7 (a) and 7 (b), the binder Z labeled body supply site 8 is the binder 12 or the labeled body 13 (see FIGS. 2 (b) and (c)). ) Can be selected as long as it can be supplied! /.

For example, if the binder 12 or the labeling body 13 is held in advance in the flow path 3 of the flow cell type reaction vessel 2, the binder 12 or the labeling body 13 is held in a part of the flow path 3, It can be moved with the added sample.

In addition, the layout of the binder Z label supply part 8 is as shown in Figs. 7 (a) and (b).

Alternatively, it may be provided downstream of the sample addition site 4 in the flow path 3 of the flow cell type reaction vessel 2.

Alternatively, as shown in FIGS. 6 (a) and 6 (b), the sample addition site 4 may also be used.

[0024] Further, as a preferred embodiment of the measuring instrument 1, as shown in Fig. 6 (a) (b) or Fig. 7 (a) (b), a detection site 5 in the flow path 3 of the flow cell type reaction vessel 2 is used. It is preferable to provide an absorption site 9 on the downstream side.

A typical example of the piezoelectric vibrator 6 is a quartz crystal vibrator.

Furthermore, the magnetic force generating member 7 includes a wide range of members that generate magnetic force, but a permanent magnet, an electromagnet or the like is typically used. [0025] <Element description>

Next, the concept of each element used in the present embodiment will be described.

Sample

The sample that can be used in the present embodiment is not particularly limited, and examples thereof include biological samples such as whole blood, plasma, serum, spinal fluid, saliva, amniotic fluid, urine, sweat, spleen, and tears. In addition, these samples or those derived from stool, food or soil can be used as samples by diluting, concentrating or extracting by adding an aqueous medium.

[0026] Aqueous medium

The aqueous medium is not particularly limited as long as it dissolves the above-described sample or label. Force buffers such as deionized water, distilled water, and buffer solutions are preferable. The buffer used in the buffer is not particularly limited as long as it has a buffering capacity. However, pH 1 to: L 1 For example, lactate buffer, citrate buffer, acetate buffer, succinate buffer, phthalate buffer , Phosphate buffer, triethanolamine buffer, diethanolamine buffer, lysine buffer, barbitur buffer, tris (hydroxymethyl) aminomethane buffer, imidazole buffer, malate buffer, oxalate buffer Agents, glycine buffer, borate buffer, carbonate buffer, glycine buffer, Good buffer, and the like. Examples of good buffering agents include 2-morpholinoethanesulfonic acid (MES), bis (2-hydroxyethyl) iminotris (hydroxymethyl) methane (Bis-Tris), N- (2-acetamido) iminoniacetic acid (ADA), Piperazine-N, N, monobis (2-ethanesulfonic acid) (PIPES), N- (2-acetamido) 2-aminoethanesulfonic acid (ACES), 3 morpholino-2-hydroxypropanesulfonic acid (MOPSO), N , N Bis (2-hydroxyethyl) -2-aminoethanesulfonic acid (BES), 3-morpholinopropanesulfonic acid (MOPS), N- [Tris (hydroxymethyl) methyl] 2 aminoethanesulfonic acid (TES) ), 2- [4 (2 Hydroxyethyl) 1-piperadi-l] ethanesulfonic acid (HEPES), 3- [N, N bis (2 hydroxyethyl) amino] — 2-hydroxypropanesulfonic acid (DIPSO) , N— (Hydroxymethyl) methyl] -2 hydroxy-1-3aminopropane sulfonic acid (TAPSO), piperazine 1 N, N, 1 bis (2 hydroxypropane sulfonic acid) (POPSO), 3- [4- (2-hydroxyethyl) — 1—Piperazyl] — 2 Hydroxypropanesulfonic acid (HEPPSO), 3— [4— (2 —Hydroxyethyl) —1-piperaduryl] propanesulfonic acid [(H) EPPS], N [Tris (hydroxymethyl) methyl] glycine (Tricine), N, N-bis (2-hydroxyethyl) glycine (Bicine), N Tris (hydroxymethyl) methyl-3-aminopropanesulfonic acid (TAPS), N-cyclohexyl 2-aminoethanesulfonic acid (CHES), N-cyclohexyl 3-amino-2-hydroxypropanesulfonic acid (CAPSO), N cyclohexyl 3-aminopropanesulfonic acid (CAPS), and the like. The concentration of the buffer solution is not particularly limited as long as it is suitable for the measurement, but 0.001-2. OmolZL is preferred 0.0.05-: L OmolZL is more preferred 0.001-0. ImolZL Is particularly preferred.

In addition, foods and soils that have been pretreated can be used as samples. Here, the pretreatment of food and soil includes, for example, extraction of ingredients in food and soil with an appropriate solvent, chemical modification, and the like. Examples of the solvent include the above-mentioned aqueous media, organic solvents such as acetolyl, hexane, methanol, ethanol, dichloromethane, chloroform, and acetone.

Examples of the chemical modification include structural conversion of components in food and soil with chemical reagents.

Measurement object

The measurement object in the present embodiment is not particularly limited as long as it binds to a specific substance, for example, a component measured using an antigen-antibody reaction, a component measured using an enzyme reaction, and other specificities. Ingredients measured by a chemical reaction can be mentioned, but a component measured using an antigen antibody reaction is preferred.

Components measured by antigen-antibody reaction include, for example, IgG, IgM, IgA, IgE, apoprotein AI, apoprotein ΑΠ, apoprotein Β, apoprotein Ε, rheumatoid factor, D-dimer, oxidized LDL, glycated LDL , Glycoalbumin, adiponectin, T3, Τ4, drug (anti-tencan, etc.), C-reactive protein (CRP), cytodynamic ins, a-fetoprotein (AFP), cancer fetal antigen (CEA) ゝ CA19 — 9, CA— 125, PIVKA- II (Protein induced by vitamin

K absence-II), parathyroid hormone (PTH), human chorionic gonadotropin (hCG), thyroid stimulating hormone (TSH), insulin, C peptide, estrogen, anti-GAD antibody, pepsinogen, influenza A antigen, influenza B-type antigen, anti-coronavirus Original, HBV antigen, anti-HBV antibody, HCV antigen, anti-HCV antibody, HTLV-I antigen, anti-HTLV I antibody, HIV antibody, tuberculosis antibody, mycoplasma antibody, hemoglobin Alc, atrial sodium diuretic peptide (ANP), brain sodium Diuretic peptide (BNP), troponin T, troponin I, creatine kinase MB (CK—MB), myoglobin, H—FABP (human heart-derived fatty acid binding protein), DON, NIV, mold fungi such as T2, bisphenol Α, nonylphenol, dibutyl phthalate, polychlorinated biphenyls (PCBs), dioxins, ρ, ρ, endocrine disruptors such as dichlorodiphenyltrichloroethane, tryptyltin, fungi such as enterococci, eggs Food allergens such as milk, wheat, buckwheat, and peanuts, allergens such as mites such as cypress mite and toyohyouda, and anti-allergic antibodies. .

Examples of biological components measured using an enzymatic reaction include glucose, 1,5-anhydroglucitol, hemoglobin Alc, glycoalbumin, fucose, urea, uric acid, ammonia, creatine, total cholesterol, free cholesterol, and high-density lipoprotein. Cholesterol in protein (HDL—C), cholesterol in low density lipoprotein (LDL—C), cholesterol in very low density lipoprotein (VLDL—C), cholesterol in remnant-like lipoprotein (RLP—C), Triglyceride, phospholipid, total protein, albumin, globulin, pyrilvin, bile acid, sialic acid, lactic acid, pyruvic acid, free fatty acid, cell mouth plasmin, alanine aminotransferase (ALT), aspartate aminotransferase (A ST), Creatine phosphokinase (CPK) Over peptidase (PK), amylase, lipase, cholinesterase, _I Monteagle Tamil trans peptidase, leucine aminopeptidase Daze, L-lactic acid dehydrogenase (LDH), aldolase, alkaline phosphatase, acid phosphatase, New § cetyl Darco Sami - Daze, Guanaze And monoamine oxidase.

Examples of other components to be measured by specific binding include methods using nucleic acids, lectins and the like. For example, DNA or RNA encoding cancer genes such as ras, cancer suppressor genes such as p53, peptide nucleic acids, and abutama 1, glycoprotein and the like.

Similar object to be measured

The analog of the measurement object is the measurement pair in the sample with respect to the binder or trapper. There is no particular limitation as long as it reacts competitively with the figurine, and examples include the measurement object itself, a substance containing an epitope for a noinder or trapper, and the like.

[0029] Reaction vessel

A typical embodiment of the reaction vessel 2 is a flow cell type.

As used herein, the flow cell type reaction vessel 2 widely includes those having a flow path 3 through which a sample flows, and is applied to immunochromatography, liquid chromatography, microchemical systems, and the like. In immunochromatography, the sample solution added to the sample addition site moves through the membrane by capillary action and is measured at the detection site. In liquid chromatography, a sample containing a measurement object is injected into a sample addition site installed in a flow cell through which liquid flows by a pump, and measurement is performed at a detection site. A micro chemical system is an analytical sensor that integrates chemical devices such as semiconductor integrated circuits by processing minute flow paths on a substrate such as glass or plastic using micro processing technology. When a sample is added to the sample addition site installed above, the sample moves in the channel and is measured at the detection site installed at the end of the channel.

Examples of the material of the flow cell type reaction vessel 2 include plastic, silica, ceramics, glass, metal, graphite, resin, and porous membrane.

As the reaction vessel, a fixed cell type such as a well plate can be selected as appropriate.

[0030] Channel

The flow path 3 installed in the flow cell type reaction vessel 2 holds the measurement object, binder, and label together with the sample, flows without adsorption, and can form at least the sample addition site 4 and the detection site 5 As a material that can be used, plastic, silica, ceramics, glass, metal, graphite, porous membrane, etc. are preferable. Examples of the material of the porous membrane include glass fiber, cellulose, nylon (registered trademark), crosslinked dextran, various chromatographic papers, nitrocellulose, and the like, and nitrocellulose is preferable. The channel diameter is preferably lnm to 10 cm, more preferably lOOnm to lcm, and particularly preferably 1 μm to 2 mm.

In addition, at least one flow path 3 of the flow cell type reaction vessel 2 may be used. From the viewpoint of expanding the range, it is also possible to provide multiple lines of flow paths 3 and provide at least the sample addition site 4 and the detection site 5 in the flow path 3 of each line.

[0031] Sample addition site

The sample addition site 4 is a site for adding the sample to the flow cell type reaction vessel 2. Examples of the material of the sample addition site 4 include glass fiber, cellulose, nylon, cross-linked dextran, various chromatographic papers, and nitrocellulose. Nitrocellulose is preferable.

Binder Z label supply site

The binder Z label supply part 8 is a part for supplying a binder or a label, and the sample addition part 4 is also used as the binder Z label supply part 8, and the binder Z label is supplied from the same part as the sample addition part 4. It is also possible to supply a point force different from the sample addition site 4. In this case, the binder Z label can be added from the noinder Z label supply site 8, but the binder Z label may be held in the member.

Examples of the material of the Norder Z labeled substance supply site 8 include glass fiber, cellulose, nylon, cross-linked dextran, various chromatographic papers, nitrocellulose, and the like, which are the same as or different from those of the sample addition site 4.

[0032] Binder

The binder 12 in the present embodiment is present in a state where it can move in the flow path 3, and can be bound to magnetic particles. For example, antibodies and antibodies that specifically bind to the antigen and the antigen, saccharides and lectins for the saccharides, DNA and DNA complementary to the DNA, etc., each of which is used it can.

[0033] Marker

The labeled body 13 is composed of a measurement object analog or binder 13b and an insoluble vesicle 13a, and transmits information depending on the amount of the complex containing the carrier 11 formed on the piezoelectric vibrator 6. There is no particular limitation as long as it is a material, but a material in which the frequency change of the piezoelectric vibrator 6 is increased by the marker 13 is preferable. The object recognition site on the label 13 The measurement object recognition site in the carrier 11 may be the same, but is preferably different.

[0034] Insoluble endoplasmic reticulum

There is no particular limitation on the insoluble vesicle 13a that binds to the measurement object analog or the binder 13b as long as the complex formed on the piezoelectric vibrator 6 can be detected as the amount of change in the frequency. What can increase the change in the frequency of the piezoelectric vibrator 6 having a large mass is preferable. Examples of insoluble vesicles having a large mass include metal colloids and latex, and examples of metal colloids include gold colloids and silver colloids. In addition, when a flow cell type reaction vessel is used as the reaction vessel, magnetic particles can also be used as the insoluble vesicle. The particle size of the insoluble endoplasmic reticulum is preferably 0.1 to: LOOOO nm 1 to: 5 to 500 nm, more preferably LOOOnm. The concentration of the insoluble endoplasmic reticulum used is preferably 0.001% to 10%, more preferably 0.01% to 5%, and particularly preferably 0.1% to 1%. Insoluble endoplasmic reticulum can also be used in which the surface of the insoluble endoplasmic reticulum is coated with a hydrophilic protein such as bovine serum albumin or a polymer compound such as polyethylene glycol (PEG) or polyvinylpyrrolidone (PVA). .

[0035] A method of binding a binder or an analyte-like analog to an insoluble endoplasmic reticulum

The insoluble endoplasmic reticulum and the binder or the analog to be measured may be physically bonded or may be chemically bonded. Examples of physical bonds include non-covalent bonds such as physical adsorption. Examples of the chemical bond include a covalent bond. Examples of non-covalent bonds include electrostatic bonds, hydrogen bonds, hydrophobic bonds, and coordinate bonds. As a method for preparing a labeled body by covalently bonding an insoluble vesicle and a binder, for example, using a cross-linking agent such as divalent daltaraldehyde, the insoluble vesicle and the binder are passed through the cross-linking agent. The method of combining is mentioned.

[0036] Detection site

The detection part 5 is preferably a quartz crystal vibrator as the piezoelectric vibrator 6 that is not particularly limited as long as it has the piezoelectric vibrator 6 that is a sensor.

-Piezoelectric vibrator (quartz crystal vibrator)

The piezoelectric vibrator 6 is not particularly limited as long as it is made of a crystal having a piezoelectric effect. Examples of crystals include crystals such as crystal, Rossiel salt, and electron stone, lithium tantalate (

LiTaO), lithium niobate (LiNbO), etc., single crystals of acids, zinc oxide (ZnO), etc.

twenty three

Quartz is preferable.

[0037] Carrier

The carrier 11 is composed of magnetic particles 1 la and a trapper 1 lb or a measurement object analogue 1 lc, and is not particularly limited as long as it is supported on the piezoelectric vibrator 6 by a magnetic force, but the trapper 1 When using the support 11 consisting of lb and magnetic particles 1 la, the measurement object recognition site of the trapper l ib constituting the support 11 and the measurement target recognition site of the binder or label I prefer to be the same, but different! /.

[0038] Magnetic particles

The magnetic particles in the present embodiment are ferrite or magnetite (magnetite, Fe SO)

Those having 3 4 as the main component and large mass that reacts efficiently with the magnet are preferred. The particle size of the magnetic particles is preferably 1 to: LOOOOOnm force S, more preferably 10 to: LOOOOnm force S, and particularly preferably 100 to 500 Onm. The concentration of magnetic particles supplied to the flow path as a labeling body is preferably 0.001% to 10%, more preferably 0.01% to 5%, and particularly preferably 0.1% to 1%. That's right. In addition, magnetic particles whose surface is coated with a hydrophilic protein such as bovine serum albumin or a polymer compound such as polyethylene glycol (PEG) or polybulurpyrrolidone (PVA) can be used.

[0039] Trapper

The trapper is not particularly limited as long as the trapper specifically binds to an object to be measured and is fixed on the crystal unit 25 together with magnetic particles. For example, an antigen and an antibody that specifically binds to the antigen And abutama, saccharides and lectins for the saccharides, DNA and DNA complementary to the DNA, etc., one of which can be used.

[0040] Method for binding trapper or measurement object analogue to magnetic particle

Examples of the method for binding the trapper or the analog to be measured to the magnetic particle include physical bonding and chemical bonding. Examples of physical bonds include non-covalent bonds such as physical adsorption. Examples of the chemical bond include a covalent bond. Examples of non-covalent bonds include electrostatic bonds, hydrogen bonds, hydrophobic bonds, and coordinate bonds. Examples of the method of binding the trapper or the measurement object analogue to the magnetic particles by covalent bonding include a method of binding via a crosslinking agent using a crosslinking agent such as bivalent dartalaldehyde.

[0041] Magnetic force generating member

It is preferable that the magnetic force generating member 7 also has means for releasing the magnetic field applied to the piezoelectric vibrator 6.

Here, the magnetic force generating member 7 is not particularly limited, such as a permanent magnet or an electromagnet. However, it is preferable that the magnetic force generating member 7 can generate a magnetic field reversibly so that the magnetic force can be applied or released as necessary. . For example, in the case of a permanent magnet, in addition to making the magnet removable, the magnetic field application range can be changed reversibly by making the position of the magnet relative to the surface of the piezoelectric vibrator 6 variable. It suffices to turn on / off the power supply to the coil.

The arrangement of the magnetic force generation member 7 is not particularly limited as long as the magnetic field can be selectively and reversibly formed on the piezoelectric vibrator 6. In particular, when the reaction vessel 2 is a flow cell type reaction vessel, the magnetic force generating member 7 may be present in the flow path 3, but if the viewpoint is such that the flow of the sample is not impaired, the flow cell type reaction vessel 2 It is preferably provided outside the channel 3.

[0042] Absorption site

The absorption site 9 is a site provided in the flow cell type reaction vessel 2 for absorbing the unreacted sample and label, and is preferably located downstream of the sample addition site 4 and the detection site 5.

The absorption site 9 can absorb unreacted components that have passed through the detection site 5 as long as it can absorb unreacted components (unreacted sample or label). The influence of the component on the detection site 5 can be reduced.

As the absorption site 9, an absorptive polymer compound can be used. Examples of the polymer compound include cellulose, glass fiber, cotton, polyurethane and the like. In addition, the absorption part 9 can use a forced discharge means such as a pump.

[0043] Washing liquid The cleaning liquid is not particularly limited as long as it can wash the sample components and the label that cannot react on the piezoelectric vibrator 6, but an aqueous medium is preferable. In particular, the above-mentioned aqueous medium containing a surfactant is more preferable. The buffer used for the buffer is not particularly limited as long as it has a buffer capacity. The surfactant is not particularly limited as long as it has a surfactant effect, and examples thereof include a cationic surfactant, an anionic surfactant, an amphoteric surfactant, and a nonionic surfactant. Nonionic surfactants are preferred. Nonionic surfactants include polyoxyethylene sorbitan monolaurate (Tween 20) and polyoxyethylene octyl ether (TritonX-lOO). The concentration of the surfactant is not particularly limited, but is preferably 0.001 to 20%, more preferably 0.01 to 10%, and particularly preferably 0.05 to 1%.

[0044] Temperature control in a flow cell type reaction vessel

The method for controlling the temperature of the reaction liquid in the flow cell type reaction vessel 2 is not particularly limited as long as the temperature can be controlled by heating the entire bottom surface force of the flow cell type reaction vessel 2, but for example, the temperature using a Peltier element Control.

[0045] <Operation Principle of Measuring Instrument>

In such a measuring instrument 1, when the carrier 11 is composed of the magnetic particles 11 a and the trapper l ib, as shown in FIG. The carrier 11 is carried on the surface of the piezoelectric vibrator 6 by (magnetic force or magnetic attraction). At this time, in the reaction between the measurement object 10 in the sample and the carrier 11, the measurement object 10 is bonded to the carrier 11 (specifically, the trapper l ib), and consists of the trapper measurement object. A complex is generated.

[0046] Further, in the case of a sandwich reaction in which the measurement object 10 is sandwiched between the support 11 (magnetic particle l la + trapper l ib) and the label 13, as shown in FIGS. 2 (b) and 2 (c). In addition, the support 11 supported on the piezoelectric vibrator 6 is bonded to the measurement object 10, and as a result, a sandwich-type complex (by the sandwich reaction involving the measurement object 10 and the binder 12 or the label 13) ( The carrier-measuring object-binder or marker is produced.

At this time, the measurement object recognition site in the binder 12 or the label 13 may be the same as the measurement object recognition site in the carrier 11 (trapper l ib), but differently. I prefer to do that!

Furthermore, when the reaction at the detection site 5 is completed, it is necessary to remove the reaction product generated on the piezoelectric vibrator 6 after the reaction is completed. In this example, for example, as shown in FIG. 3, it is sufficient to cancel the magnetic field action by the magnetic force generation member 7 (to eliminate the magnetic field). In this case, the magnetic interaction (magnetic force or magnetic attractive force) is eliminated. Since the support 11 disengages from the surface force of the piezoelectric vibrator 6, the reactant on the piezoelectric vibrator 6 is released, and the surface of the piezoelectric vibrator 6 can be easily reset.

In addition, as shown in FIGS. 4 (a) and 4 (b), in the embodiment in which the support 11 is composed of the magnetic particles 1 la and the measurement object analogue 1 lc, the support 11 is supported on the piezoelectric vibrator 6. The supported body 11 reacts competitively with the measurement object 10 with respect to the binder 12 or the label body 13. In this case, on the piezoelectric vibrator 6, a measurement object-like substance binder or a complex having a label strength is generated.

In this embodiment, the measurement object analogue 11c reacts competitively with the measurement object 10, so if the reaction is accelerated with the measurement object analogue 11c, the corresponding amount in the sample is increased. This means that the number of measurement objects 10 is small, and the opposite reaction tendency means that there are many measurement objects 10 in the sample. Therefore, it is possible to indirectly increase the measurement sensitivity of the measurement object 10 by accurately measuring the amount of reaction with the measurement object analog 11c.

Furthermore, when the reaction at the detection site 5 is completed, the magnetic field action by the magnetic force generating member 7 is solved in the same manner as in FIG. 3 in order to remove the reaction product generated on the piezoelectric vibrator 6 after the reaction is completed. It can be removed.

Further, as shown in FIG. 5, a carrier 11 (for example, magnetic particles 1 la + trapper ib) carried on the piezoelectric vibrator 6 includes a label 13 (measurement object analogue 13c). ) And the measurement object 10 in a competitive manner, a composite body having a carrier measurement object or a measurement object analog force is generated on the piezoelectric vibrator 6.

[0050] <Measurement method using measurement instrument>

Next, a measurement method for measuring the measurement object 10 using the measurement instrument 1 described above will be described.

First, using the measuring instrument 1 in which the carrier 11 is a magnetic particle 11a and a trapper l ib. When measuring the measurement object 10 in the sample, as shown in Figs. 2 (a) to (c) and Fig. 3 and Figs. 8 (a) and (b), the sample addition site 4 and the piezoelectric vibration A reaction vessel 2 having a detection site 5 with a child 6 and a carrier 11 formed by binding a trapper l ib that specifically binds to a measurement object or a measurement object analog and a magnetic particle 11a. Is a method for measuring a measurement object 10 in a sample by using a reaction vessel 2 in which a measurement object 10 and a support 11 in a sample are reacted in a reaction vessel 2 and a complex including the support measurement object A sample reaction step for generating a magnetic material, a magnetic material 11a of the carrier 11 is made to act on a magnetic force, and a magnetic material 11a is magnetically supported on the piezoelectric vibrator 6 and a composite supported on the piezoelectric vibrator 6 The frequency measurement process for measuring the amount of the body as the amount of change in the frequency of the piezoelectric vibrator 6, and the piezoelectric measured in this frequency measurement process And a previously prepared calibration curve and the frequency of variation of Doko 6 as long as it includes a concentration determination step of determining the concentration of the measuring object 10 in the sample.

In particular, in order to accurately grasp the amount of the complex at the detection site 5, a washing step for the detection site 5 is included between the sample reaction process and the frequency measurement step, and unnecessary components at the detection site 5 are removed. It is preferable.

[0051] As another measurement method using this type of measuring instrument, as shown in FIGS. 5, 6, and 8 (a) and 8 (b), a sample addition site 4 and a piezoelectric vibrator 6 are provided. In the sample, the reaction vessel 2 having the detection site 5 and the support 11 formed by binding the trapper 11b specifically binding to the measurement object or the measurement object analog and the magnetic particle 11a are used. This is a method for measuring the measurement object 10 and carries in the reaction vessel 2 a label 13 formed by binding the measurement object 10 in the sample and the measurement object analog 13c to the insoluble vesicle 13a. The sample reaction step of reacting with the body 11 to generate a complex containing the label body, the magnetic particles 11a of the carrier 11 acting magnetically, and the magnetic particles 11a are magnetically supported on the piezoelectric vibrator 6 Supporting process Measures the amount of composite supported on the piezoelectric vibrator 6 as the amount of change in the frequency of the piezoelectric vibrator 6 A frequency measurement step, and a concentration determination step for determining the concentration of the measurement object 10 in the sample from the amount of change in the frequency of the piezoelectric vibrator 6 measured in the frequency measurement step and a calibration curve prepared in advance. If you do,

[0052] Further, in order to measure the measurement object 10 in the sample using the measurement instrument 1 in the form in which the support 11 is the magnetic particle 11a and the measurement object analogue 11c, FIG. 4 and FIG. 8 (a As shown in (b) , A reaction vessel 2 having a detection part 5 provided with a sample addition part 4 and a piezoelectric vibrator 6, a carrier 11 formed by combining a measurement object analogue 11c and a magnetic particle 11a, and a measurement object 10 or A method for measuring a measurement object 10 in a sample using a binder 12 that specifically binds to a measurement object analog or a labeled body 13 formed by binding the binder 13b and an insoluble vesicle 13a, In reaction container 2, specifically binds to the measurement object 10 or the measurement object analogue in the reaction object 10 and the measurement object analog 1 lc and magnetic particles 1 la. Sample reaction step of reacting the binder 12 or the binder 13b with the insoluble vesicle 13a to form a complex containing the carrier binder or carrier label, and applying a magnetic force to the magnetic particles 11a of the carrier 11; Magnetic particle 11a magnetically piezoelectric vibrator A carrier supporting process for supporting the piezoelectric vibrator 6, a frequency measuring process for measuring the amount of the composite supported on the piezoelectric vibrator 6 as a change in the frequency of the piezoelectric vibrator 6, and this frequency measuring process A concentration determining step for determining the concentration of the measurement object 10 in the sample from the amount of change in the frequency of the piezoelectric vibrator 6 measured in step 1 and a calibration curve prepared in advance may be provided.

In this embodiment as well, it is preferable to include a cleaning step for the detection site 5 between the sample reaction step and the frequency measurement step.

[0053] Furthermore, in this type of measurement method, in a mode in which the binder Z label supply part 8 also serves as the sample addition part 4, the supply step includes the step of supplying the sample and the binder 12 or the label 13 from the sample addition part 4. It may be added at the same time or before and after.

Further, in this type of measurement method, in a mode in which the binder Z labeled body supply portion 8 holds the binder 12 or the labeled body 13 movably in advance in the flow path 3 of the flow cell type reaction container 2, the supply step Any sample may be used as long as the sample is added from the sample addition site 4 and the binder 12 or the label 13 held in advance in the Noinder Z label supply site 8 moves together with the sample.

[0054] In this type of measurement method, when a plurality of items are measured simultaneously, a plurality of different types of carriers 11 in any plurality of detection units 5 are used as magnetic fields generated by the magnetic force generation member 7. It is sufficient that a plurality of measurement objects can be measured simultaneously.

Furthermore, when the detection part 5 is arbitrarily set, any plurality of detection part 5 A plurality of magnetic force generating members 7 may be provided corresponding to the above, and the detection site 5 may be set at an arbitrary location. According to this aspect, it is possible to cause the magnetic fields generated by the plurality of magnetic force generating members 7 to act separately, and to allow only the magnetic field acting region to function as the detection portion 5.

[0055] Further, as an apparatus invention for embodying such a measurement method, as shown in FIG. 8 (b), a sample and a binder 12 or a binder 13b that binds specifically to the measurement object 10 or The above-described measuring instrument 1 that supplies the analyte 13c to the detection site 5 with the labeled body 13 bound to the insoluble endoplasmic reticulum 13a and the piezoelectric vibrator 6 at the detection site 5 of the measurement instrument 1 were generated. Trapper--measurement object or trapper--measurement object--binder or trapper--one measurement object Composite consisting of a label or measurement object analogue Non-one or measurement object analogue The frequency measuring means 15 that measures the amount of the complex as the labeled body force as the amount of change in the vibration frequency of the piezoelectric vibrator 6, and the amount of change in the vibration frequency of the piezoelectric vibrator 6 that is measured by the vibration frequency measuring means 15 and created in advance. Measurement in the sample from the measured calibration curve That a density determining means 16 for determining the concentration of an object 10 and the like.

In such a measuring apparatus, the measuring instrument 1, the frequency measuring means 15, and the concentration determining means 16 may be provided separately, or the measuring instrument 1 is provided with the frequency measuring means 15 and the concentration determining means 16. You may make it provide integrally by incorporating.

Hereinafter, the present invention will be described in more detail based on embodiments shown in the accompanying drawings.

◎ Embodiment 1

FIG. 9 is an explanatory view showing Embodiment 1 of the measuring apparatus to which the present invention is applied. In the figure, the measurement apparatus includes a measurement instrument 20 for measuring a measurement object in a sample, and an analysis processing apparatus 100 that performs analysis processing based on a sensor output from the measurement instrument 20.

[0057] <Outline of measuring instrument>

In the present embodiment, the measuring instrument 20 has a flow cell type reaction vessel 21 in which a measurement object in a sample can move along a flow path 22 as shown in FIG. 9 and FIG. The flow cell type reaction vessel 21 is sealed with a cover 30 so that a sensor substrate 24 of a QCM (Quartz Crystal Microbalance) sensor 23 is the bottom of the vessel and a flow path 22 with a predetermined gap is secured on the sensor substrate 24. Is. In addition, a liquid inflow opening 31 is formed in a part of the cover 30 of the flow cell type reaction vessel 21 located upstream of the flow path 22 and a liquid outflow is provided in a part located downstream of the flow path 22. Opening 32 is established.

In the present embodiment, a sample container 34 is connected to one liquid inflow opening 31 via a tube 33 and functions as a sample addition site A, and the other liquid outflow opening 32 via a tube 35. The syringe pump 36 is connected to function as an absorption site D. In the present embodiment, the label added from the sample addition site A is supplied at the same time as the sample or after the sample is added.

Furthermore, in the present embodiment, the QCM sensor 23 has a quartz resonator 25 mounted on the surface of the sensor substrate 24, and a portion of the flow path 22 corresponding to the surface of the quartz resonator 25 is detected. It is designed to function as C.

Then, a magnet 40 made of, for example, a permanent magnet is installed on the back surface of the sensor substrate 24 at a position where the crystal unit 25 is mounted! The magnet 40 is detachable, and when the magnet 40 is installed, the position is adjusted so that the surface portion of the crystal unit 25 is disposed in the magnetic field application region by the magnet 40.

A carrier 11 (magnetic particle l la + trapper l ib or measurement object analogue 11c) is carried on the surface of the crystal unit 25 by the magnetic field of the magnet 40 (FIG. 1 or FIG. 4). reference).

Furthermore, in the present embodiment, the sensor substrate 24 is placed on a constant temperature block 51 made of, for example, aluminum, which is maintained at a constant temperature by, for example, a Peltier element 52! Consideration is given to prevent changes in the resonance frequency.

[0059] <Overview of analysis processing device>

In the present embodiment, the analysis processing device 100 calculates the concentration of the measurement object in the sample based on the frequency measurement circuit that measures the resonance characteristics of the crystal unit 25 and the information on the frequency measurement circuit force. It should be equipped with a concentration calculation circuit.

In the present embodiment, as the frequency measurement circuit and the concentration calculation circuit, for example, the resonance characteristics of the crystal unit 25 are measured using the network analyzer 101, and the measured resonance characteristic data is stored in a personal computer (PC). Crystal vibration by taking in 102 and calculating An example is a configuration in which the resonance frequency of the element 25 is obtained and the calculation is performed on the resonance frequency force concentration using a calibration curve.

To obtain the resonance frequency from the resonance characteristics of the crystal unit 25, measure the resonance characteristics (frequency-admittance characteristics) near the resonance frequency of the crystal unit 25 using a network analyzer 101, etc. Find the equivalent circuit constants using mathematical means such as the least-squares method to match the admittance characteristics shown by.

As shown in FIG. 11, an equivalent circuit of the crystal unit 25 can be expressed by a parallel connection of a series resonant circuit of an inductance Lx, a capacitance Cx, and a loss resistance Rx and a parallel capacitance Cp such as an electrode capacitance. At this time, the admittance Y of the crystal unit can be obtained by Equation 1 and Equation 2.

Y = J- ω -Cp + 1 / ϋ- ω -Lx + l / (j- ω -Cx) + Rx) (Equation 1)

ω = 2 · π -fo (Equation 2)

Here, J is a complex number (1 1) ^1/2 , ω is an angular frequency, fo is a resonance frequency, π is a circularity, and the resonance frequency fo of the quartz crystal is calculated by the equivalent circuit constant Lx , Formula from Cx

Can be calculated using 3.

fo = lZ (2.7u. (Lx'Cx) ^1/2 ) (Equation 3)

It should be noted that the personal computer 102 preferably has a display device 103 for displaying the concentration calculation result!

[0060] <Details of QCM sensor>

Crystal oscillator

In the present embodiment, the crystal resonator 25 is, for example, an AT-cut crystal resonator having a thickness-shear vibration mode, and is preferably one that oscillates at a fundamental resonance frequency of 5 to 50 MHz. Further, one crystal unit 25 may be used as in the present embodiment, but a plurality of crystal units 25 may be used.

[0061] Sensor substrate

The structure of the sensor substrate 24 may be selected as appropriate, but an example is shown in FIGS. 12 (a) and 12 (b).

In the figure, the sensor substrate 24 is a double-sided printed circuit board made of glass epoxy, for example. Using plate 61, on the top surface, make two land patterns 62 to connect with electrodes 65 and 66 of crystal unit 25 and the vibrating part of crystal unit 25 not to contact printed circuit board 61. A U-shaped pattern 63 for a pedestal is arranged. The crystal unit 25 is arranged such that the electrode connection part of the crystal unit 25 is placed on the electrode connection land pattern 62, and is electrically connected with, for example, conductive silicone resin. In order to prevent the sample solution from seeping between the crystal unit 25 and the sensor substrate 24 and shorting the front electrode 65 and the back electrode 65, the periphery of the crystal unit 25 is, for example, a silicone adhesive 64. It is sealed with. The wiring 67 from the electrode connecting land pattern 62 is drawn to the lower surface of the printed circuit board 61 through the through hole and extends to the connection terminal 68 to avoid short circuit due to the sample solution.

Further, as shown in FIGS. 13 (a) and 13 (b), the front electrode 65 and the back electrode 66 are both provided with connection portions 69a so that they can be connected to the electrode connection land pattern 62 of the printed circuit board 61 on the back surface. It is electrically connected by the extraction electrode 69. The front electrode 65 is routed by the extraction electrode 69 to the connection portion 69a on the back surface via the left side surface of the crystal unit 25.

[0062] Method of fixing a quartz crystal to the surface of a flow cell type reaction vessel

In the present embodiment, the sensor substrate 24 constitutes a part of the flow cell type reaction vessel 21, but the present invention is not limited to this. In order to fix the sensor substrate 24 to the surface of the flow cell type reaction vessel 21, an adhesive is used. You should do it in a scientific way using chemicals! The adhesive used is silicone conductive adhesive, epoxy conductive adhesive, anisotropic conductive film (ACF), anisotropic conductive paste (ACP), etc. for electrical connection, and waterproof and fixing. Silicone adhesives, epoxy adhesives, etc. are used.

[0063] <Method of carrying / removing carrier on quartz crystal>

Method for supporting a carrier on a crystal unit

As shown in FIG. 9, there are no particular limitations on the method of supporting the carrier 11 on the crystal unit 25 as long as it can be supported by applying a magnetic field by an external force magnet 40 in the reaction vessel 21. How to detach the carrier from the crystal unit

As shown in FIG. 9, the carrier 11 carried on the crystal unit 25 is detached from the external force of the reaction vessel 21 and the magnetic field generated by the magnet 40 is released. There is no particular restriction as long as you can leave 11.

As shown in FIG. 9 and FIG. 10, a solution containing a carrier (magnetic particle + trapper or similar object to be measured) is added from the sample addition site A of the measuring instrument 20, and a magnet is placed on the surface of the quartz crystal resonator 25. The magnetic field generated by 40 is applied, the carrier is supported on the crystal unit 25, and cleaning with the cleaning liquid is performed.

After that, the sample and the binder (or labeled body) are added to the sample addition site A of the measuring instrument 20, and the measurement object and the binder (or labeled body) in the sample are the flow cell type of the measuring instrument 20. Move through the flow path 22 of the reaction vessel 21, supply the sample and binder (or label) to the carrier on the crystal unit 25 that is the detection unit C, and if necessary, wash with the above-described cleaning solution. The amount of the complex produced by specifically binding to the carrier is detected as the amount of change in the vibration frequency of the crystal unit 25, and the concentration of the measurement object in the sample is measured via the analysis processing device 100.

[0065] In such a measurement process, the conditions under which the measurement object in the sample reacts with the carrier (for example, a trapper) on the crystal unit 25 or the measurement object in the sample and the label in the reaction solution. Is not particularly limited as long as it allows specific reaction, but the reaction temperature is usually 0 ° C-100 ° C, preferably 10-60 ° C, more preferably 20-40 ° C. Is done. The measurement time is usually 10 seconds to 10 hours, preferably 30 seconds to 5 hours, more preferably 1 minute to 1 hour.

Furthermore, the measurement using a standard substance with a known concentration for preparing a calibration curve may be repeated individually using the same measuring device 20, but using an apparatus configured by combining a plurality of measuring devices 20. Thus, it is preferable to simultaneously perform measurement of the measurement object in the sample and measurement of a standard substance having a known concentration.

In such a measurement process, the states shown in FIG. 2, FIG. 4 and FIG. By these reactions, the measurement object 10 is measured directly or indirectly.

Then, after the reaction is completed, the magnetic field action by the magnet 40 is canceled and the cleaning liquid is allowed to flow, so that the carrier 11 is separated from the surface of the quartz oscillator 25 as shown in FIG. 3, for example. As a result, the reactant on the crystal unit 25 is surely removed. For this reason, in the present embodiment, since the surface of the crystal unit 25 is reset, the measuring instrument 20 can be reused.

[0067] Embodiment 2

FIG. 14 shows Embodiment 2 of the measuring apparatus to which the present invention is applied.

In this figure, the basic configuration of the measuring device is provided with a measuring instrument 20 and an analysis processing device 100 in substantially the same manner as in the first embodiment, but unlike in the first embodiment, it is applied to POCT (point-of-care testing). Considering the correspondence, it is designed to be a small and simple configuration. Components similar to those in the first embodiment are denoted by the same reference numerals as those in the first embodiment, and detailed description thereof is omitted here.

In the present embodiment, the entire apparatus is obtained by incorporating the analysis processing device 100 into the measuring instrument 20, and has two element forces, that is, a measurement unit S1 and a measurement result analysis processing unit S2. The measurement unit S1 includes a QCM sensor 23 (a sensor substrate 24 on which a crystal unit 25 is mounted), a plastic upper cover 71, a lower cover 72, and various membranes 81 to 83.

[0068] The upper cover 71 is formed by hollowing out the channel 22 part, and is placed on the sensor substrate 24 in a detachable and watertight manner, and a gap formed between the two is defined as the channel 22 It is getting used. In this example, as shown in FIG. 14 and FIG. 15, a portion of the flow channel 22 is secured long, and on one side of the flow channel 22, there is a sample holding membrane 81 that functions as the sample addition site A. A label (or binder) holding membrane 82 that functions as a label holding site (or binder holding site) B is sandwiched between the sample holding membrane 81. Furthermore, an opening 75 for adding a sample solution with a pipette or the like is provided in the vicinity of the sample addition site A of the upper cover 71. In this example, the opening 75 also functions as an auxiliary sample container. To do. Further, on the other side of the flow path 22, an absorption membrane 83 is sandwiched on the downstream side of the detection site C, so that it functions as the absorption site D.

Here, the label (binder) holding site B is a site for holding the label (binder) in the flow path 22 of the flow cell type reaction vessel 21 so that the label (binder) can move. In the present embodiment, the material of the label (binder) holding membrane 82 at the label (binder) holding site B may be the same as or different from the material of the sample holding membrane 81 at the sample addition site A. Examples thereof include glass fiber, cellulose, nylon, cross-linked dextran, various chromatographic papers, nitrocellulose and the like, and nitrocellulose is preferable.

Further, as the absorption membrane 83 of the absorption site D, a water-absorbing polymer compound can be used. Examples of the polymer compound include cellulose, glass fiber, cotton, polyurethane and the like.

[0070] Further, a magnet 40 (for example, a rectangular ferrite magnet) is installed at the position where the crystal resonator 25 is mounted on the lower surface of the sensor substrate 24. The surface will be placed.

Furthermore, the mounting method of the sensor substrate 24 and the crystal resonator 25, the electrode drawing method, and the like are substantially the same as those in the first embodiment. The sensor substrate 24 is assembled by bonding or fitting with the upper cover 71 and the lower cover 72 sandwiched therebetween. Further, since the crystal unit 25 is disposed in a space sealed by the upper and lower covers 71 and 72, it is not easily affected by environmental temperature changes or wind.

[0071] Further, the measurement unit S1 and the measurement result analysis processing unit S2 can be mechanically connected and disconnected by a connector 85. At the same time, the wiring from the crystal unit 25 is also connected to the measurement result analysis processing unit S 2 via the connector 85.

In the present embodiment, the measurement result analysis processing unit S2 (100) includes a frequency measurement circuit and a concentration calculation circuit, as in the first embodiment, but the crystal resonator is used as the frequency measurement circuit. A configuration is possible in which 25 is connected to a crystal oscillation circuit 111 to oscillate in the vicinity of the resonance frequency, and the oscillation frequency is measured by the frequency counter 112. In this configuration, a control device 113 such as a personal computer, a microprocessor, or a logic operation circuit can be used as the concentration operation circuit.

In other words, in the present embodiment, unlike the first embodiment, the measurement of the resonance frequency of the crystal unit 25 is performed at a frequency slightly higher than the resonance frequency when the crystal unit 25 is oscillated using the crystal oscillation circuit 111. Since it oscillates, it can be regarded as a resonance frequency. And Then, the resonance frequency is obtained by measuring the oscillation output by the frequency counter 112, and the control device 113 using a CPU or the like uses the calibration curve data to determine the concentration of the measurement object from the resonance frequency. And the result can be displayed on the display device 114.

FIG. 16 shows an example of the crystal oscillation circuit 111, which is a Colpitts type crystal oscillation circuit using a CMOS inverter. In this example, the wiring from the front electrode and the back electrode of the crystal unit 25 is connected to the crystal oscillation circuit 111 through the wiring of the sensor substrate 24. The crystal unit 25 is connected to the ground via the input capacitor 221 and the output capacitor 222 to form a π-type feedback circuit, and this feedback resistor 223 is inserted between the input and output of the amplifier circuit composed of the CMOS inverter 224. The signal is fed back and oscillated. The C MOS inverter 224 is used as an inverting amplifier by connecting the input and output with a feedback resistor 223 of several ΩΩ power MΩ.

[0073] Compared to the first embodiment, the measuring device according to the present embodiment does not require large devices such as a syringe pump 36 or a network analyzer, and is a small-sized device suitable for POCT or the like. A measuring device can be realized.

[0074] ◎ Embodiment 3

The measuring apparatus according to the first and second embodiments is a model using a single crystal unit as the QCM sensor 23. There is no particular problem with these models as long as the frequency drift of the crystal unit itself is negligible compared to the amount of frequency change due to complex coupling, where the frequency drift of the crystal unit itself is relatively small.

Here, the frequency drift means a frequency fluctuation other than a frequency change based on specific binding of the target substance. For example, when the added sample solution comes into contact with the crystal resonator, the resonator itself or the ambient temperature is changed. Change, and the resonance frequency will change, and components other than the target substance contained in the sample solution will bind to the sensor film and cause a frequency change. It is.

In the first and second embodiments, if such a frequency drift exists, the drift component is added to the measurement value, which may cause an error in the measurement value.

In order to reduce the measurement error due to such a drift phenomenon, a method using a reference crystal resonator can be considered. In contrast to the name reference crystal resonator, the crystal resonator that detects the target substance is called the test crystal resonator.

This reference crystal unit is attached to the test crystal unit at the detection site in the flow path, and the change in the resonance frequency of each test crystal unit is measured. By subtracting, it is possible to remove the influence of the drift component and reduce the measurement error.

Next, a model using a reference crystal resonator will be described with reference to the drawings.

FIG. 17 is a diagram showing the configuration of the third embodiment using a reference crystal resonator. The measurement device according to the present embodiment is basically a reference crystal oscillator added to the second embodiment, and further measures the resonance frequency of the added reference crystal resonator and calculates the difference. This is an improvement of the measurement circuit 120 of the result analysis processing unit S2 (analysis processing device 100).

Therefore, in the description of the present embodiment, the description will focus on the parts different from the second embodiment.

In the present embodiment, a reference crystal resonator 252 is installed downstream of the flow of the sample solution with respect to the test crystal resonator 251 provided at the detection site C in the flow path 22. The method of mounting the reference crystal unit 252 can be the same as that of the test crystal unit 251!

The wiring from the reference crystal resonator 252 is connected to the measurement circuit 120 of the measurement instrument analysis processing unit S2 through the connector 85 through the wiring pattern of the sensor substrate 24.

[0076] Further, the magnet 40 for applying a magnetic field is provided separately for the reference crystal resonator 252 and the test crystal resonator 251, but each magnet resonator 251, The magnet shape and installation position may be adjusted so that the magnetic field concentrates on the 252 electrode surfaces.

The positional relationship between the reference crystal unit and the test crystal unit can be either upstream or downstream in the case of series connection with the sample solution flow.

However, if the test crystal unit 251 and the reference crystal unit 252 are placed as close as possible so that they are equally affected by drift, the drift cancellation Preferred in terms of becoming more complete.

FIG. 18 is a diagram showing a configuration of the measurement circuit 120.

Corresponding to the test crystal oscillator 251 and the reference crystal oscillator 252 are connected to the individual crystal oscillator circuits 121 and 122 and frequency counters 123 and 124, respectively. It is configured to continuously measure.

As the crystal oscillation circuits 121 and 122, the Colpitts crystal oscillation circuit used in the second embodiment can be used. The measured resonance frequencies are calculated by calculating the difference in the control device 125 so that the drift component is subtracted. Reference numeral 126 denotes a display device that displays a calculation result by the control device 125.

In this configuration, the difference is calculated after the oscillation frequencies of the two are obtained by the frequency counters 123 and 124, but the difference frequency is created directly from the outputs of the two crystal oscillation circuits 121 and 122 using a logic circuit. Alternatively, a method of measuring the difference frequency with a counter may be used.

FIG. 19 is a diagram showing another configuration of the measurement circuit 120.

In this example, the reference crystal resonator 252 and the test crystal resonator 251 are switched by the switching circuit 130 and are alternately oscillated by one crystal oscillation circuit 131 to measure the frequency.

The switching circuit 130 can be an electromagnetic relay or an electronic relay such as an analog switch.

Switching is performed by a switching signal generated in the control device 133.

The switching timing can be in the range of several 0.1s of force and several tens of seconds. If the switching time is short, the measurement accuracy of the frequency counter 132 will deteriorate. Conversely, if the switching time is long, the resonance frequency on the non-oscillating side Since the state of change disappears, a force of about 1 second per second is usually appropriate. Reference numeral 134 denotes a display device that displays a calculation result by the control device 133.

[0079] The measurement procedure in the present embodiment is exactly the same as in Embodiment 2.

According to the present embodiment, in addition to non-specific adsorption and temperature change, measurement that cancels drift based on an unexpected cause is possible, so that improvement in measurement accuracy can be expected.

In the third embodiment, force using separate crystal resonators for the test crystal resonator 251 and the reference crystal resonator 252, for example, two channels in which two sets of electrodes are provided on one crystal substrate. A quartz crystal can also be used.

In this example, instead of using two separate crystal units, a 2-channel crystal unit is used, and the drift removal capability is superior compared to using two individual crystal units. It is suitable when higher accuracy and detection limit are required.

It is possible to add two samples of one type of measurement object or add two different types of measurement objects one by one and measure simultaneously.

Furthermore, it is possible to easily increase the number of lines to 2 or more, and simultaneous measurement of multiple samples and multiple measurement objects becomes possible.

In addition, it is possible to detect two or more target substances by using two reference type 2-channel crystal resonators. For example, an influenza A virus can be detected with a first reference type 2-channel crystal resonator, and an influenza B virus can be detected with a second reference type 2-channel crystal resonator.

In addition to the 2-channel crystal unit, it can also be applied to crystal units with 3 or more channels.

◎ Embodiment 4

FIG. 20 shows a measuring instrument according to the fourth embodiment.

In this figure, this example is an example of an n-line type measuring instrument (for example, n = 3), and the sample addition site A is made common to each flow path 22 (22-22) of a plurality of lines, and the flow of the line 1 A test crystal unit 25 is located at the detection site of the path 22 (22), and a reference channel is located at the path 22 (22) of the line 2.

1 2

Quartz crystal unit 25, control crystal unit 25 is placed in channel 22 (22) of line n

By increasing the accuracy and detection limit, it is possible to determine whether or not the measurement itself is possible by detecting whether the magnetic particles have normally flowed through the flow path during measurement.

Here, the control crystal unit 25 is a crystal unit on which a sensor film that specifically detects magnetic particles not forming a complex is formed. Can be used as an index to indicate whether the measurement itself has been completed normally, that is, whether measurement is possible.

In addition, if the measuring instrument shown in this figure is used, all n crystal resonators at the detection site are used as test crystal resonators, so that the same measurement object in n samples can be measured simultaneously. And _n measurement objects in the same specimen can be simultaneously measured.

In addition, one of the multiple lines is used as the reference crystal unit 25.

2

It is also possible to measure two different types of measurement objects or one type of measurement object for two samples at the same time.

It is also possible to easily increase the number of lines to 3 or more, and simultaneous measurement of multiple samples and multiple measurement objects becomes possible. Also, by using one of the multiple lines as a reference line, it is possible to cancel the error due to the drift phenomenon and improve the accuracy and detection limit by the method described in Embodiment 3. is there.

Further, in the present embodiment, when several types of measurement objects are measured at the same time, as shown in FIG. 20 (a), crystals are formed at a plurality of locations (P to P) of the reaction vessel 21. The vibrator 25 (25 to 25) and a plurality of carriers (A to 8) are supported by the magnet 40 (40 to 40). At this time, as shown in FIG. 20 (b), by releasing the magnetic field generated by the magnet 40 from the outside of the reaction vessel 21, the respective supporting bodies (A to A) are detached separately from the surface of the crystal unit 25. be able to. The supports (A to A) may be different from each other, but two or more types may be the same. The binder or the label may be different from each other, and two or more kinds of force may be the same.

In this embodiment, a mode in which the sample addition site A is shared is shown, but this is not limited to this. As shown in FIG. 22, the sample addition site for each channel is shown. A may be provided.

In this case, for example, with an n-line type measuring instrument, the test crystal resonators 25, 25 · '· 25 are arranged at the detection site of the flow path 22 of the line 1 and the flow path 22 of the line 1, while the line η By disposing the reference crystal unit 25 in the flow path 22, it is possible to remove the influence of drift components on a plurality of types of measurement objects, for example, and to reduce measurement errors.

Industrial applicability

[0084] According to the present invention, it is possible to provide a measuring instrument effective for measuring a measurement object contained in a sample to be analyzed such as a living body, food, and soil, a measurement method and a measurement apparatus using the measurement instrument. I'll do it.

Claims

The scope of the claims

[1] A carrier formed by binding a trapper that specifically binds to a measurement object or a measurement object analog and a magnetic particle, or a support formed by bonding a measurement object analog and a magnetic particle A measuring instrument for measuring a measurement object in a sample using a body,

A reaction vessel having a sample addition site and a detection site equipped with a piezoelectric vibrator, and a magnetic force generating member for applying a magnetic field to the piezoelectric vibrator,

The measuring instrument, wherein the carrier is magnetically supported on the piezoelectric vibrator by the magnetic force of the magnetic force generating member.

[2] In the measuring instrument according to claim 1,

The magnetic force generating member also has means for releasing the magnetic field applied to the piezoelectric vibrator.

[3] In the measuring instrument according to claim 1 or 2,

The reaction vessel is a binder that specifically binds to the measurement object or the measurement object analog, or a binder that can supply a label formed by binding the binder or the measurement object analog to an insoluble vesicle or A measuring instrument having a label supply part.

[4] In the measuring instrument according to claim 1 or 2,

The reaction container is a flow cell type in which a measurement object and a carrier can move along a flow path, and a sample addition site and a detection site are provided in the flow path of the flow cell type reaction vessel. Measuring instrument to do.

[5] The measuring instrument according to claim 3,

The reaction container is a flow cell type in which a measurement object, a carrier, and a labeling body can move along a flow path, and a sample addition site, a detection site, a binder, or a labeling body are placed in the flow cell of the flow cell type reaction container. A measuring instrument, characterized in that a supply site is provided.

[6] The measuring instrument according to claim 5,

A measuring instrument characterized in that the noinder or labeled body supply site is provided downstream of the flow channel of the flow cell type reaction vessel in the flow channel of the sample.

[7] The measuring instrument according to claim 5, The sample addition site also serves as a binder or label supply site.

[8] In the measuring instrument according to any one of claims 4 to 7,

A measuring instrument comprising an absorption site downstream of a detection site.

[9] In the measuring instrument according to any one of claims 1 to 8,

A measuring instrument using a plurality of types of carriers.

[10] In the measuring instrument according to any one of claims 1 to 9,

A measuring instrument comprising a plurality of piezoelectric vibrators at a detection site.

[11] The measuring instrument according to claim 10,

A measuring instrument comprising a magnetic force generating member for each piezoelectric vibrator.

[12] The measuring instrument according to any one of claims 1 to 11,

A measuring instrument, wherein the piezoelectric vibrator is a quartz crystal vibrator.

[13] In the measuring instrument according to any one of claims 4 to 12,

A flow cell type reaction vessel is provided with a plurality of flow paths.

[14] In the measuring instrument according to any one of claims 5 to 13,

A measuring instrument, wherein a binder or a marker is movably held in advance at a binder or marker supply site.

[15] The measuring instrument according to any one of claims 1 to 14,

A measuring instrument characterized in that the carrier is magnetically carried on the surface of the piezoelectric vibrator in advance.

[16] A measuring kit comprising the measuring instrument according to any one of claims 1 to 13 and a carrier.

[17] A measurement kit comprising the measurement instrument according to any one of claims 1 to 13, a carrier, and a binder or a label.

[18] A reaction vessel having a sample addition site and a detection site equipped with a piezoelectric vibrator, and a carrier formed by binding a trapper that specifically binds to a measurement object or a measurement object analog and magnetic particles A method for measuring a measurement object in a sample using A sample reaction step in which a measurement object in a sample and a carrier are reacted in a reaction container to generate a complex including the carrier measurement object;

A carrier carrying step in which a magnetic force is applied to the magnetic particles of the carrier to magnetically carry the magnetic particles on the piezoelectric vibrator;

A frequency measurement process for measuring the amount of the composite supported on the piezoelectric vibrator as a change in the frequency of the piezoelectric vibrator; and

A measurement method comprising a concentration determination step for determining the concentration of the measurement object in the sample from the amount of change in the vibration frequency of the piezoelectric vibrator measured in the frequency measurement step and a calibration curve prepared in advance. .

[19] The method of claim 18, comprising

Further, in the reaction vessel, the sample and the binder that specifically binds to the measurement object or the measurement object analog or the labeled body formed by binding the binder to the insoluble vesicle is reacted, and the carrier measurement object. A composite comprising a binder, a carrier, and an object to be measured. A measurement method comprising a step of producing a composite comprising a label.

[20] A reaction vessel having a sample addition site and a detection site equipped with a piezoelectric vibrator, and a carrier in which a trapper that specifically binds to a measurement object or a measurement object analogue and magnetic particles are combined. A method for measuring a measurement object in a sample using

A sample reaction in which a labeled body formed by binding a measurement object in a sample and a measurement object analog and an insoluble endoplasmic reticulum in a reaction container reacts with the support body to form a complex containing the support body label body. Process,

[21] a reaction vessel having a detection site with a sample addition site and a piezoelectric vibrator, and a measurement object Using a carrier formed by binding an analog and magnetic particles, and a measurement object or a measurement object or a binder that specifically binds to the analog or a label formed by binding the binder and an insoluble vesicle A method for measuring an object to be measured in a sample,

In a reaction container, a support formed by binding a measurement object and a measurement object analog in the sample to magnetic particles, and a binder or the binder specifically binding to the measurement object or the measurement object analog A sample reaction step for reacting with an insoluble endoplasmic reticulum to produce a complex containing a carrier binder or a carrier label;

[22] In the measurement method according to any one of claims 18 to 21,

A measuring method comprising a cleaning step of cleaning the composite supported on a piezoelectric vibrator.

[23] The measurement method according to any one of claims 18 to 22,

A measuring method comprising performing a carrier supporting step before the sample reaction step.

[24] In the measurement method according to any one of claims 18 to 23,

The reaction container is a flow cell type in which the measurement object and the carrier can move along the flow path, and the sample addition site and the detection site are provided in the flow path of the flow cell type reaction container! Measuring method characterized by

[25] The measurement method according to claim 24,

A measuring method characterized in that a sampler is started by adding a sample to a sample addition site, wherein a noinder or a labeled body is held in a flow path.

[26] The measuring instrument according to any one of claims 1 to 17,

The amount of the composite carried on the piezoelectric vibrator at the detection site of this measuring instrument And a frequency measuring means for measuring the amount of change in the frequency.

[27] The measuring instrument according to any one of claims 1 to 17,

A frequency measuring means for measuring the amount of the composite carried on the piezoelectric vibrator at the detection site of the measuring instrument as a change in the frequency of the piezoelectric vibrator;

A measuring apparatus comprising: a concentration determining means for determining the concentration of the measurement object in the sample from the amount of change in the vibration frequency of the piezoelectric vibrator measured by the frequency measuring means and a calibration curve prepared in advance. .

[28] A carrier formed by binding a trapper that specifically binds to a measurement object or a measurement object analog and a magnetic particle, or a support formed by bonding a measurement object analog and a magnetic particle. A method for regenerating a piezoelectric vibrator in measurement of a measurement object in a sample using a body and a piezoelectric vibrator, comprising: a carrier carried on the piezoelectric vibrator by a magnetic force generating means; and a measurement object A method for regenerating a piezoelectric vibrator, comprising: measuring a composite and then dissociating the carrier by a magnetic field release means.

[29] The playback method according to claim 28,

A reproducing method, wherein the piezoelectric vibrator is a crystal vibrator.