JPS60152954A - Measurement of biological sample - Google Patents

Abstract

PURPOSE:To enable a highly accurate measurement of a biological sample by separating a carrier from a vapor phase through a centrifugal force to measure optical characteristic based on material in the vapor phase. CONSTITUTION:A plurality of reaction vessels 2 having a trapping section each are set on a reaction table 1 and a plurality of sample cups 4 containing a standard substance and a sample to be inspected are set on a sample table 3 separately. A sample distribution arm 5 discharges a sample liquid at the fixed position 6 on the table 3 into a vessel 2 at the fixed position 7 on the table 1. The table 1 is fed rotatively by steps according to intervals between the vessels 2 and the table 3 is turned based on input information on analysis items. As each vessel 2 reaches the specified position 7, arms 8 and 9 move to discharge a reagent of a cold insulation storage 10 into the vessel 2 to mix the sample with the reagent to react. Then, the table 1 is turned at a high speed to separate a fluorescent labeled substance bonded to a solid phase from an isolated fluorescent labeled substance and then, the vessel 2 is irradiated with a laser light to indicate the results of analysis on a printer 14 and a CRT15 through an A/D converter 40. These operations are controlled with a microcomputer 11 based on analysis conditions previously inputted.

Description

DETAILED DESCRIPTION OF THE INVENTION [Field of Application of the Invention] The present invention relates to a method for measuring a biological sample, and particularly to a method for optically measuring a target substance in a biological sample.

[Background of the invention]

In recent years, there has been a strong desire to quantify trace substances in the medical field. Conventionally, various methods have been devised to measure substances in biological fluids using antigen-antibody reactions using immunological techniques. For example, methods that directly measure sedimentation and agglutination, such as capillary sedimentation, immunoturbidimetry, immunoturbidimetry, latex agglutination, and 5RID; laws are being developed. However, these methods have advantages and disadvantages. Among measurement methods with excellent quantitative performance, the measurement method using a homogeneous immune reaction system with good operability has insufficient measurement sensitivity, while the measurement method using a heterogeneous immune reaction system has high sensitivity but is complicated to operate and results in poor results. Because it takes several hours to obtain the desired results, it is inconvenient in regular axillary examinations.

Among these, the so-called B/F fractionation method, which separates the labeled antigen-antibody complex (Bound straddling B) and the excess labeled antigen remaining in a free state (pree or F), The phase method is relatively easy to operate. This solid phase method is
In this method, the activity of B is measured by performing a reaction using a large-particle carrier that precipitates in a liquid phase, and then washing to remove free labeled antigen. However, the solid-phase method has extremely low reaction efficiency and requires several hours to obtain measurement results. Furthermore, because it was difficult to maintain a constant level of cleaning, sufficient measurement accuracy could not be obtained.

[Purpose of the invention]

An object of the present invention is to provide a method for measuring a biological sample that can shorten operation time and improve measurement accuracy.

[Summary of the invention]

In the present invention, a target substance or a receptor having a specific binding property to the target substance is supported on a small-sized insoluble carrier, and a sample containing one target substance and a labeled target substance or label are attached to the supported target substance or receptor. Add the receptor and/or receptor and a solvent or dispersion medium to react while the carrier is suspended, then centrifugal force is applied to separate the carrier from the liquid phase, leaving the carrier in the reaction vessel. It is characterized by measuring a target substance contained in a sample by measuring optical properties based on the substance inside.

[Embodiments of the invention]

In a preferred embodiment of the present invention, an antibody or a fish antigen is supported on a fine particle insoluble carrier suspended in a liquid phase;
An antigen, an antibody, a fluorescently labeled antigen, or a fluorescently labeled antibody is added to this to perform an antigen-antibody reaction.Then, the reaction mixture is separated into a solid phase and a liquid phase by means such as centrifugation, and the reaction mixture is separated into a solid phase and a liquid phase by means such as centrifugation. By irradiating the liquid phase region with excitation light while avoiding the solid phase region, the antigen-antibody reaction is optically and quantitatively measured, thereby quantifying the target antigen or antibody.

According to a preferred embodiment of the present invention, trace components that could only be measured practically by radioisotope labeling immunoassay can be quantitatively measured much more quickly and safely than that. Moreover, in a preferred embodiment, although the reagent composition is a solid phase method using carrier particles, it is possible to provide a measurement method that does not require washing the solid phase and is suitable for complete automation of measurement.

As the carrier for suspension, those having an average particle size of 0.01 μm to 1 fin are suitable for the purpose, and it is particularly preferable to use inert carrier particles with an average particle size of 0.1 μm to 10 μm as the solid phase. When such a carrier is reacted with a test sample, the reaction efficiency becomes very high and the measurement proceeds rapidly. Furthermore, by adding a fluorescent labeling substance to the reagent system and allowing it to react for a certain period of time, the reaction mixture can be centrifuged or other means to easily separate the solid-phase bound fluorescent labeling substance and the free fluorescent labeling substance without introducing a washing operation. There is a certain relationship between the fluorescence intensity of the liquid phase (free) and the concentration of the target component in the test sample. With such a method, 2L can quantify target components without washing operations even though it is a solid-phase reaction system using carrier particles, and it is also possible to obtain a sensitivity measurement comparable to that of conventional heterogeneous reaction systems. can.

In order to accurately and quickly quantify a target substance in a sample using a solid-phase method, a receptor or a labeled target substance supported on an inert carrier such as latex particles should be used at as high a concentration as possible. It goes without saying that it is desirable to bring the specimen into contact with a specimen containing a reactive target substance.

The inventors made insoluble carriers with various diameters support a target substance, such as a receptor (anti-human IgO antibody) for human immunoglobulin G (human IgG), while keeping the weight percent of the carrier in the final reaction solution constant. After preparing the reagent and considering the measurement sensitivity and reaction time to be achieved with the reagent, we found that if a carrier with an average particle size of IIal+ or less was used, the target substance (H)IgG) could be quantified within 1 hour of reaction time. It was found that it is possible to detect Furthermore, it has been found that when fine particles having an average particle size of 10 μm or less are used, the target substance (IgG) can be quantitatively detected within a reaction time of 15 minutes.

In automating the measurement of a target substance, it is preferable to use a carrier with an average particle size of 10 μm or less, since the allowable reaction time from the viewpoint of sample processing speed is 1 hour, preferably 15 minutes or less. Furthermore, polystyrene latex particles (specific gravity 1.
When the reagent was prepared using 05) as a carrier, the mixed reaction solution of the reagent and the test sample was collected at 2000 rpm, which is the maximum rotational speed of the centrifugal capacity that can be loaded into the automatic analyzer.
In order to separate the carrier reaction product and the reaction liquid phase within 1 hour of centrifugation at
It turns out that it is necessary to be greater than or equal to m.

According to the above results, the average particle size of the inert carrier particles is 1
It has been found that the following is preferable, and a carrier having an average particle size of 0.1 μm to 10 μm is suitable.

Examples of the inert carrier particles include fine particles of an organic polymer substance that is substantially insoluble in the liquid medium used when measuring biological fluids and has the above-mentioned average particle size, such as organic polymer latex such as polystyrene; Individually dispersed bacteria or cells such as staphylococci, streptococci, or eg silica.

Inorganic oxides such as silica-alumina and alumina, other mineral powders, metals, etc. are used.

Various physical quantities can be considered depending on the type of labeling substance used as a reagent. For example, when the labeling substance is a fluorescent substance, the fluorescence intensity is measured. Since the fluorescence intensity (■) is expressed as the sum of the fluorescence polarization degree (1) of the vertical component and the fluorescence polarization degree (2)/of the parallel component, similar results can be obtained even if one measurement or (2)/ is measured. In addition, in order to eliminate measurement errors due to fluorescent contaminants (proteins, etc.) in the test sample, it is necessary to support the fluorescent labeling substance (target substance or receptor) in the reagent on an insoluble carrier that is larger than the contaminant. Use the method of

Since contaminants are small molecules compared to insoluble carriers, rotational Brownian motion occurs actively in solution and the degree of fluorescence polarization is small, whereas fluorescent labeling substances supported on insoluble carriers are large molecules. The rotational Brownian motion of fluorescent molecules is suppressed and the degree of polarization of fluorescence increases. That is, the above h)
In an IgG measurement experiment, when the polarizing filter on the excitation light side was fixed vertically and the polarizing filter on the fluorescent side was set parallel to it, the fluorescence polarization intensity I/ was measured, and compared to when measuring the total fluorescence intensity. It was found that the influence of contaminant fluorescent substances was less. At this time, an anti-IgG antibody against the target substance IgG was supported on polystyrene latex with a diameter of 10 μm and a specific gravity of 1.05.
When the gG antibody was supported on polystyrene latex with a diameter of 0.1 μm and a specific gravity of 1.05 and reacted with the target substance IgG, the anti-I
gG antibody-supported latex reaction complex and free F'I'
The I'C-labeled anti-IgG antibody-supported latex could be separated, and excitation light was irradiated avoiding the site of the anti-1gG antibody-supported latex reaction complex, and the results were measured.

The present invention is not limited to the measurement of fluorescence intensity or fluorescence polarization degree as described above. If the labeling substance is an enzyme, the enzyme activity may be measured, and if the labeling substance is a phosphorescent substance, the intensity of phosphorescence may be measured, and other substances may also be used.

Hereinafter, details of embodiments of the present invention will be described.

Example 1: (1) Preparation of anti-α-fetoprotein antibody sensitized latex reagent (anti-AFP latex reagent) An anti-human AFP antibody with an average particle size of 0 was added to a glycine buffer (2~/dt) 10-
, 55 μm polystyrene latex (manufactured by Block Chemical Co., Ltd., solid content concentration 10% by weight) was added, and
After stirring for an hour, centrifugation (12,000 rll) is performed for 40 minutes while cooling at 2-4C. The anti-AP'P antibody sensitized (supported) latex particles in which the precipitate is dispersed are suspended in a bovine serum albumin solution (0.2% by weight) to prepare an anti-AEP latex reagent whose concentration of the sensitized latex particles is 1% by weight. Prepare.

0) F of anti-α-7 protein antibody globulin fraction
0.5 M carbonate-bicarbonate buffer (
pH 9,0) was added, and p I T C (fluo
rescoinlsothiocyanate 31
0.5M carbonate-bicarbonate buffer containing m&(1)H9,O
)0.5- and react at 4C for 4 hours. After the reaction was completed, it was fractionated using a 9 ephadex O-25 column, the absorbance at wavelengths of 280 nm and 495 nm was measured, and a fraction with matching peaks was used as a PITC-labeled antibody.

9 5mM phosphate buffer (pH 7,2) was used for elution from the ephadex Q-25 column.

(3) Measurement method FIG. 1 shows a plan view of the analyzer of this embodiment.

FIG. 2 shows a side view of a table on which a reaction vessel with a trap is placed and a photometric system. The reaction vessel is a molded product of methacrylic resin, and the main body is similar to a normal square cuvette, but a trap 2 is provided on one side.

In FIGS. 1 and 2, 60 reaction vessels 2 (only 12 are shown for convenience of illustration) are installed around the circumference of the reaction table 10, with the trap portions 21 facing toward the outer periphery. Forty sample cups 4 containing standard substances and test samples are placed on the circumference of the sample table 30. Sample dispensing arm 5
works in conjunction with a dispensing syringe (not shown) to aspirate the sample liquid located at a fixed position 6 on the sample table 3 into the probe 13 and discharge it into the reaction container located at a fixed position 7 on the reaction table 1. . Reaction table 1 is reaction container 1
It is rotated by step feed with the interval between each piece being 1 pitch,
The sample table 3 is rotated based on the input information of the analysis request item. When the reaction table 1 is fed step by step and the reaction container 2 in which the sample is discharged and accommodated reaches the predetermined position 7, the arms 8 and 9 each having a reagent dispensing probe interlocked with a dispensing syringe are moved and moved to the cold storage. A certain amount of the reagent corresponding to the analysis item in 10 is taken out and discharged into a reaction container in a fixed position. The sample and reagent are mixed and the reaction proceeds. Each of these operations is controlled by the microphone α computer 11 based on analysis conditions input in advance. The analysis results are displayed on the printer 14 and CRT 15.

When the reaction proceeds for a certain period of time in the reaction vessel 2, the reaction table 1 rotates at high speed in order to separate the fluorescent labeling substance (B) bound to the solid phase from the free fluorescent labeling substance (F). The reaction table 1 on which the reaction container 2 is mounted is fixed to the shaft of a motor 24, and in addition to the step feed operation of the reaction container 2, it can perform high-speed rotation operation up to 20,000 rpm.

A standard AFP solution having the α-fetoprotein (AFP) concentration shown in Table 1 was placed in sample table 3, and anti-AFP latex reagent and FI'rC-textured anti-AFP antibody were placed in cold storage 1.
The analysis was started by setting 0Vc. The analysis conditions are sample 5.
μt1 anti-AFP latex reagent 300 μt% FITC-labeled anti-AFP antibody 100 μt. That is, sample 5μ
After separating t, add 300μ of anti-AFP antibody latex reagent.
was added and reacted at 37C for 10 minutes. To this reaction mixture, 100μ of FITC-labeled anti-ARP antibody was added and dispensed, and further reacted at 37tll'' for 10 minutes.

After this, the reaction container was centrifuged at 1 oooor- for 5 minutes. After the centrifugation was completed, the reaction table was moved in steps at a pitch of 3 seconds to measure the fluorescence intensity at a wavelength of 5251 m. The results are shown in Table 1.

Table 1 The reaction vessel 2 is irradiated with an argon laser (wavelength 488 nm) from the laser light source 31,
oressence was measured. That is, the light source side shutter 32 and the photodetector 35 side shutter 33 are controlled in opposite phases, and the shutter 32 is opened and the shutter 33 is closed during excitation. Further, the shutter 32 is closed during measurement,
Shutter 33 is opened. Each time each reaction vessel is positioned at a photometric position, the shutters 32, 33 are opened and closed. Filters 37 and 38 are buffer filters for cutting stray light.

The measurement was performed 20 times at intervals of 10 milliseconds (m(5)) and the average value was calculated. 39 is an amplifier, and 40 is an A/D converter.

A standard curve is created by plotting the measurement results in Table 1 on a graph with the concentration of the standard APP solution as the horizontal axis and the fluorescence intensity as M1''@. The digital computer 11 reads the AFP concentration corresponding to the fluorescence intensity measured using the standard curve obtained as above.The results of the measurement of the subject sample are shown in Table 2. For comparison, Table 2 also shows the measurement results by the conventional radioisotope labeling method (RIA method).

Table 2 From the above results, it can be seen that the AFP concentration obtained by the method of this example is in extremely good agreement with the measurement results by the RIA method, which is known as the most accurate quantitative method among conventional methods. In the embodiments of the present invention, the fluorescent labeling substance is F.
Although ITC was used, the labeling substance is not limited thereto.

Example 2; (1) Preparation of anti-centamycin antibody-sensitized immunobeads 20
- Immunobeads (Bi.

- To 2001 ng of commercially available anti-gentamicin antibody 151Q was added 10 mM phosphate buffer (1) H6,2)4- containing 22 tidimethylformamide, and then 401111 iJ of 1-ethl-3-(3-
dimethyla-minopropyl) c
art) Add odiimide and react at room temperature for 8 hours. After the reaction was completed, the immunobeads were mixed with 10 mM phosphate buffer (pH 6,2), SO% Meptol, 0.
11 gelatin-containing 0.06 M barbital buffer (p
Hs, a), and finally 0.06M barbital buffer containing 0.1 predominant gelatin (1) H8,
6) is used for measurement as a suspension of 1rv/- of immunobeads.

(2) Preparation of FITC-labeled gettamycin Dissolve 4 ml of gentamicin in 0.5-methanol and add 0.1 N NaO
After adjusting the pH to 9.0 with H, a dioxane solution containing 20 my of FITC was added, and after reacting at room temperature for 4 hours, the reaction product was separated and purified by thin layer chromatography to obtain FITC.
Obtain TC-labeled gentamicin.

(3) Measurement method: Add 10 μt of sample, 100 μt of anti-gentamicin antibody-sensitized immunopeads, and 60 μt of FI'l'C-labeled antibody.
The reaction was carried out at 7C for 20 minutes. After this, the reaction container was
After centrifugation at 00 rpm for 5 minutes, the fluorescence intensity was measured in the same manner as in Example 1.

(4) Measurement Results The calibration curve for gentamicin obtained by the method in (3) is shown in FIG. Furthermore, a correlation diagram with the RIA method is shown in FIG.

Example 3: (1) Preparation of reagent Same as Example 1 except that anti-IgG antibody was used instead of anti-human AFP antibody and latex particles with an average particle size of 10 μm were used.
An anti-IgG antibody sensitized latex reagent was prepared in the same manner as in (1). In addition, a commercially available FITC-labeled anti-IgG antibody was used instead of the anti-Human AFP antibody, and the average particle size was 0.
A FITC-labeled anti-IgG antibody-sensitized latex reagent was prepared in the same manner as in Example 1 (1) except that 1 μm latex particles were used.

(2) Measurement method The analysis conditions were as follows: sample 10 μt1 anti-IgG antibody latex reagent 300 μt% FITC-labeled anti-IgG antibody sensitized latex reagent 50 μt. That is, after 10 μt of the sample was taken, 300 μt of anti-IgG antibody latex reagent was added and reacted at 37 C for 10 minutes. FIT this reaction mixture
50 μt of C-labeled anti-IgG antibody-sensitized latex reagent was added and dispensed, and the mixture was further reacted with 37 U for 10 minutes. Thereafter, the reaction container was centrifuged at 1000 rIIl for 10 minutes, and after the centrifugation, the reaction table was stepped at a 3 second pitch to measure the degree of fluorescence polarization.

If the degree of reaction is to be measured by the degree of fluorescence polarization, filters 37 and 38 in FIG. 2 may be replaced with polarizing filters. The polarizing filter 9 can be switched between being parallel to the polarizing filter 8 and being perpendicular to it. The fluorescence intensity obtained when the polarizing filter 8 on the excitation light side was fixed vertically and the polarizing filter 9 on the fluorescent side was set parallel to it was measured.

(3) Measurement results A calibration curve for IgG measurement is shown in FIG.

Next, some embodiments based on the present invention will be listed.

(1) A target substance is reacted with a fluorescently labeled receptor that has specific binding properties to the target substance. The reaction mixture and the average particle size are preferably from 0.1 μm to 10 μm.
After reacting the immobilized target substance in which the target substance is supported on μm-sized inert carrier particles, the reaction vessel is subjected to centrifugation or other operations to cause the fluorescently labeled receptor that has reacted with the immobilized target substance to react with the target substance. The fluorescently labeled receptors are isolated. The amount of fluorescence derived from the fluorescently labeled receptor that has reacted with the target substance is measured by irradiating the reaction vessel with a light flux that excites the labeled fluorescent substance while avoiding the site of the in-phase target substance, and the amount of fluorescence derived from the fluorescently labeled receptor that has reacted with the target substance is measured. Quantify the target substance.

(2) A solid-phase receptor and a target in which receptor-1 having a specific binding property to a target substance is supported on an inert carrier particle having a cold particle size of 0.01 μm to IIO + preferably 0.1 μm to 10 μm. make substances react. Reaction mixture and receptor-1
After reacting with a fluorescently labeled receptor that does not have specific binding properties to the target substance but does have specific binding properties to the target substance, the reaction vessel is subjected to centrifugation or other operations to detect the fluorescent label that has reacted with the target substance. Separate the receptor and free fluorescently labeled receptor. By irradiating the reaction vessel with a light flux that excites the labeled fluorescent substance while avoiding the site of the immobilized complex, the amount of fluorescence derived from the free fluorescently labeled acceptor is measured, and based on this amount of fluorescence, the target is detected. Quantify substances.

(3) The average particle diameter is preferably from 0.01 μm to 0.01 μm.
The target substance is reacted with a solid-phase receptor in which receptor-1 that specifically binds to the target substance is supported on inert carrier particles of 1 μm to 10 μm.

Receptor-2 that does not have specific binding properties to the reaction mixture and the solid-phase receptor but has specific binding properties to the target substance, and the receptor-2
After reacting with fluorescently labeled receptor-3 that has specific binding properties, the reaction vessel is subjected to centrifugation or other operations to separate the fluorescently labeled receptor-3 that has reacted with the immobilized complex and the free fluorescently labeled receptor. Separate 3. The amount of fluorescence originating from the free fluorescently labeled receptor is measured by irradiating the reaction vessel with a light flux that excites the labeled fluorescent substance while avoiding the site of the immobilized complex.
The target substance is quantified based on this amount of fluorescence.

(4) A solid-phase receptor in which a receptor having specific binding property to a target substance is supported on inert carrier particles with an average particle diameter of 0.01 μm to IIal, preferably 0.1 μm to 10 μm, a target substance, and fluorescence After the labeled target substance is reacted, the reaction vessel is subjected to centrifugation or other operations to separate the fluorescently labeled target substance that has reacted with the solid-phase receptor from the free fluorescently labeled target substance. The amount of fluorescence derived from the free fluorescent labeling substance is measured by irradiating the reaction vessel with a light flux that excites the labeled fluorescent substance while avoiding the site of the immobilized complex, and the target substance is quantified based on the amount of fluorescence. .

(5) React the fluorescently labeled target substance with the target substance and the receptor-1 that has specific binding properties to the target substance. The reaction mixture and the average particle size are preferably from 0.01 μm to 041 μm.
After reacting a solid-phase receptor carrying receptor-2 having a specific binding property to receptor-1 with inert carrier particles of 10 μm in size, the reaction vessel is subjected to centrifugation or other operations,
It reacts with the immobilized complex and separates the nine fluorescently labeled target substances from the free fluorescently labeled target substances. The amount of fluorescence derived from the free fluorescently labeled target substance is measured by irradiating the reaction vessel with a light flux that excites the labeled fluorescent substance while avoiding the site of the immobilized complex, and the target substance is detected based on the amount of fluorescence. Quantify.

(6) Fluorescent substrate-labeled target substance (non-fluorescent) that generates a fluorescent substance only after undergoing enzyme action with the target substance and an average particle size of 0.01 μm to 1 m, preferably 0.1 μm to 10 μm.
41) A solid-phase receptor carrying a receptor having a different binding property to a target substance is reacted with a μm-sized inert carrier particle.

In a reaction system in which a reaction mixture is reacted with an enzyme that acts on a fluorescent substrate to produce a fluorescent substance, the reaction vessel is subjected to centrifugation or other operations to separate the fluorescent substrate ma that has reacted with the immobilized receptor, the target substance, and the free Separate fluorescent substrate-labeled target substances. The amount of fluorescence derived from the free fluorescent substrate-labeled target substance is measured by irradiating the reaction vessel with a light flux that excites the substance that has become fluorescent due to the action of the enzyme, avoiding the site of the phase-forming complex. The target substance is quantified based on the amount of fluorescence.

(7) Fluorescently labeled target substance (non-fluorescent) that generates a fluorescent substance only after receiving enzyme action with the target substance and an average particle size of 0
．． 0.1μm to 1m+preferably 0.1μm to 10μ
A solid-phase receptor carrying a receptor having specific binding property to the target substance is reacted with the inert carrier particles of m. In a reaction system in which a reaction mixture is reacted with an enzyme that has the property of acting on a fluorescent substance (non-fluorescent) to produce a fluorescent substance,
The reaction vessel is subjected to centrifugation or other operations to separate the fluorescent label R target substance that has reacted with the immobilized complex from the fluorescent substance that has not reacted with the immobilized complex but has been subjected to the action of the enzyme.

The amount of fluorescence is measured by irradiating the reaction container with a light flux that excites the fluorescent substance while avoiding the site of the immobilized complex, and the target substance is quantified based on the amount of fluorescence.

(8) a sample supply mechanism that supplies a sample containing a target substance, a sampling mechanism that transfers these samples to a reaction container, a solid-phase receptor, a fluorescently labeled receptor, a solid-phase target substance, a receptor, A reagent supply mechanism that transfers a fluorescently labeled target substance, an enzyme solution, etc. as a reagent to a reactor, and a mechanism that allows the target substance and reagent to react in the reaction vessel for a certain period of time, and then centrifuges the reaction vessel containing the reaction mixture at 500 to 20,000 rfl. a reaction container centrifugation mechanism that can be applied to a reaction container; a light source for irradiating a liquid phase of the reaction mixture in the reaction container with a light flux that excites a labeled fluorescent substance; and a detection means for detecting the intensity of fluorescence emitted from the liquid phase. A target substance is measured using an automatic analyzer characterized by having a part for calculating the concentration of a target substance in a sample based on the amount of fluorescence, and a means for printing the result of the calculation.

(9) A sample supply mechanism that supplies a sample containing a target substance, a sampling mechanism that transfers these samples to a reaction container, and a magnetic solid phase receptor or a magnetic solid phase target substance and a receptor. , a reagent supply mechanism that transfers a fluorescently labeled receptor, a fluorescently labeled target substance, an enzyme solution, etc. as reagents to a reaction vessel, and a part of the reaction vessel containing a reaction mixture after reacting the target substance and reagent for a certain period of time. A reaction container holding mechanism that can magnetically separate the reaction mixture into a liquid phase and a solid phase, a light source for irradiating the liquid phase with a light flux that excites the labeled fluorescent substance, and fluorescence emitted from the liquid phase. The target substance is measured using an automatic analyzer that has a detection means for checking the intensity and a function to calculate the concentration of the target substance in the sample based on the amount of fluorescence.

〔Effect of the invention〕

According to the present invention, since the reaction can proceed while the carrier is in suspension, the reaction efficiency can be increased and the operation time can be shortened. Furthermore, since no cleaning operation is required, measurement accuracy can be improved.

[Brief explanation of the drawing]

FIG. 1 is a plan view showing a main part of an embodiment of the present invention, FIG. 2 is a vertical cross-sectional view of the vicinity of the reaction table of the embodiment of FIG.
The figure shows an example of the calibration curve for gentamicin, FIG. 4 shows the correlation between the measurement results of water droplets and the RIA method, and FIG. 5 shows an example of the calibration curve for immunoglobulin O. DESCRIPTION OF SYMBOLS 1... Reaction table, 2... Reaction container, 3... Sample table, 31... Laser light source, 32.33.
・・Shake ゛゛nfumyshishi(, u3/+nRIASiDirect(1)I/Dedication)

Claims (1)

[Claims] 1. Suspending insoluble carrier particles in a biological sample containing a target substance, causing the carrier particles to carry the target substance, and causing the target substance to undergo an antigen-antibody reaction in a reaction vessel. , separating the carrier particles from the liquid phase by centrifugation;
and a method for measuring a biological sample, comprising measuring the optical properties of the liquid phase while the carrier particles are present in the reaction vessel. 2. The method for measuring a biological sample according to claim 1, wherein the particle size of the carrier particles is within the range of 0.01 μm to one fin. 3. The method for measuring a biological sample according to claim 1, wherein the optical property is one of fluorescence intensity and fluorescence polarization degree. 4. Suspending insoluble carrier particles in a biological sample containing the target substance, adding a labeled target substance to the biological sample,
A biological method comprising causing the target substance and the labeled target substance to undergo an antigen-antibody reaction in a reaction container, separating the carrier particles from a liquid phase by centrifugation, and measuring optical properties of the liquid phase. How to measure the sample. 5. In a method for quantifying a target substance using an antigen-antibody reaction, after reacting a sample containing a target substance with a labeled target substance and a first receptor having specific binding property to the target substance,
□By adding a second receptor having specific binding properties to the receptor and causing a reaction, and then applying centrifugal force,
A method for measuring a biological sample in which a target substance contained in a sample is measured by separating a first receptor-reactive labeling compound that has reacted with a second receptor and a free labeling substance and measuring the physical quantity of the latter label. . 6. Suspending insoluble carrier particles in a biological sample containing the target substance, adding a receptor to the sample and allowing the receptor to be supported on the carrier particles, adding a labeled receptor, and separating the carrier particles by centrifugation. A method for measuring a biological sample, the method comprising: separating a liquid from a liquid phase; and measuring optical properties of the liquid phase. 7. A solid-phase receptor, in which inert carrier particles with an average particle size of 0.1 μm to 10 μm carry a receptor that has specific binding properties with the target substance, and the target substance are placed in a reaction vessel. After reacting the reaction mixture with the fluorescently labeled receptor, the reaction vessel is centrifugally rotated to separate the fluorescently labeled receptor that has reacted with the inphase complex and the free fluorescently labeled receptor,
Measurement of a biological sample, characterized in that the reaction vessel is irradiated with a light beam that excites the labeled fluorescent substance, the amount of fluorescence derived from the free fluorescently labeled receptor is measured, and the labeled substance is quantified based on the amount of fluorescence. Method. 8. The method for measuring a biological sample according to claim 7, wherein the carrier particles are a fine powder of an organic polymer substance or a fine powder of an inorganic substance that is substantially insoluble in an aqueous liquid medium. 9. The method for measuring a biological sample according to claim 8, wherein the organic polymer substance fine powder is a synthetic resin fine powder, bacteria, or cell membrane fragment. 10. The method for measuring a biological sample according to claim 8, wherein the organic polymer substance fine powder is polystyrene latex particles or styrene/pudadiene latex particles. 11. The method for measuring a biological sample according to claim 10, wherein the organic polymer substance fine powder is a magnetic latex particle. 12. The method for measuring a biological sample according to claim 8, wherein the inorganic substance fine powder is any one of metal, inorganic oxide, and mineral fine powder. 13. The method for measuring a biological sample according to claim 8, wherein the inorganic substance fine powder is any one of silica, alumina, and silica-alumina fine powder.