JP6442291B2 - Particle dispersion, kit for use in detection of target substance, and target substance detection method - Google Patents

Description

  The present invention relates to a particle dispersion, a kit for use in detecting a target substance, and a method for detecting a target substance.

  An immunoagglutination method using an antigen-antibody reaction is widely used in various fields such as clinical examination and biochemical research. In particular, immunoaggregation methods using carriers typified by latex agglutination methods are the mainstream of immunoagglutination methods because they have better detection sensitivity and the like than methods that detect agglutination without using carriers. .

However, it may be pointed out that the immunoagglutination method has insufficient detection sensitivity as compared with other measurement methods such as chemiluminescence immunoassay.
Therefore, water-soluble polymers such as polyethylene glycol are used as immune aggregation promoters (Patent Documents 1 and 2), but further improvement in detection sensitivity is desired.

Japanese Patent Laid-Open No. 08-278308 JP 2003-294753 A

  Therefore, the problem to be solved by the present invention is to provide a new technique for increasing the sensitivity of detection of a target substance in a specimen by immunoagglutination.

  Therefore, as a result of intensive studies, the present inventors have incorporated a low molecular weight compound having a naphthalene skeleton into a particle dispersion of carrier particles using an aqueous medium as a dispersion medium, and use this to perform detection by immunoaggregation. Thus, it has been found that detection of a target substance in a specimen can be made highly sensitive, and the present invention has been completed.

  That is, the present invention provides the following [1] to [3].

  [1] A particle dispersion of carrier particles using an aqueous medium as a dispersion medium and containing a low molecular compound having a naphthalene skeleton (hereinafter also referred to as “particle dispersion of the present invention”).

  [2] A kit for use in detection of a target substance in a specimen by an immunoagglutination method, comprising the particle dispersion of [1] above (hereinafter also referred to as “kit of the present invention”).

  [3] A method for detecting a target substance in a specimen by immunoagglutination, wherein the carrier particle carrying a substance capable of binding to the target substance is contacted with the specimen in the presence of a low molecular compound having a naphthalene skeleton. A detection method (hereinafter, also referred to as “detection method of the present invention”).

  ADVANTAGE OF THE INVENTION According to this invention, the detection of the target substance in the sample by an immunoagglutination method can be made highly sensitive.

[Particle dispersion]
The particle dispersion of carrier particles using the aqueous medium of the present invention as a dispersion medium contains a low molecular compound having a naphthalene skeleton.

<Low molecular compound having naphthalene skeleton>
The low molecular compound having a naphthalene skeleton is a concept including a non-vinyl low molecular compound having a naphthalene skeleton, a vinyl low molecular compound having a naphthalene skeleton, and an oligomer of the compound. As the low molecular weight compound having a naphthalene skeleton, acetylnaphthalene, naphthaldehyde, ethylnaphthalene, propylnaphthalene, vinylnaphthalene, and naphthyl (meth) acrylate are preferable. Examples of acetylnaphthalene include 1-acetylnaphthalene and 2-acetylnaphthalene. Examples of naphthaldehyde include 1-naphthaldehyde and 2-naphthaldehyde. Examples of ethylnaphthalene include 1-ethylnaphthalene and 2-ethylnaphthalene. Examples of propylnaphthalene include 1-propylnaphthalene and 2-propylnaphthalene. Examples of vinyl naphthalene include 1-vinyl naphthalene and 2-vinyl naphthalene. Examples of (meth) acrylic acid naphthyl include (meth) acrylic acid 1-naphthyl and (meth) acrylic acid 2-naphthyl.
Among these, acetylnaphthalene is particularly preferable as a low molecular compound having a naphthalene skeleton from the viewpoint of detection sensitivity.
In addition, the particle dispersion liquid of the present invention may contain one kind of low molecular compounds having a naphthalene skeleton, or may contain two or more kinds in combination.

  The content of the low molecular compound having a naphthalene skeleton is preferably 0.001 part by mass or more with respect to 100 parts by mass of the carrier particle solid content from the viewpoint of detection sensitivity and the ability of latex particles to aggregate due to the antigen-antibody reaction More preferably 0.005 parts by mass or more, still more preferably 0.01 parts by mass or more, still more preferably 0.05 parts by mass or more, and particularly preferably 0.1 parts by mass or more. From the viewpoint of increasing the sensitivity with a low content, etc., the amount is preferably 10 parts by mass or less, more preferably 5 parts by mass or less, still more preferably 3 parts by mass or less, particularly preferably 1. 5 parts by mass or less. By setting it as such a range, the improvement of detection sensitivity and the improvement of storage stability can be made compatible.

In addition, the content of the low molecular weight compound having a naphthalene skeleton is preferably 0.5 × 10 ⁻¹⁰ mass% or more, more preferably 1 × 10 ⁻¹⁰ mass% or more, and ^still more preferably based on the total amount of the particle dispersion. Is 0.5 × 10 ⁻⁸ mass% or more, more preferably 1 × 10 ⁻⁸ mass% or more, more preferably 1 × 10 ⁻⁷ mass% or more, and further preferably 1 × 10 ⁻⁶ mass% or more. Further, it is preferably 1% by mass or less, more preferably 1 × 10 ⁻¹ % by mass or less, and further preferably 0.5 × 10 ⁻¹ % by mass or less. When the particle dispersion is used as a particle dispersion for producing an agglutinating reagent, 1 × 10 ⁻¹⁰ to 1 × 10 ⁻¹ mass% is preferable, and 1 × 10 ^{−8 to} 0.5 × 10 ⁻¹ mass% is preferable. More preferably, 1 × 10 ^{−6 to} 0.5 × 10 ⁻¹ mass% is particularly preferable. On the other hand, when the particle dispersion is used as an agglutinating reagent, 0.5 × 10 ^{−10 to} 0.5 × 10 ⁻¹ mass% is preferable, and 0.5 × 10 ⁻⁸ to 1 × 10 ⁻² mass% is more preferable. preferable. 0.5 × 10 ^{−6 to} 0.5 × 10 ⁻² mass% is particularly preferable.

The content of the low molecular compound having a naphthalene skeleton can be measured, for example, by the following method. That is, the dispersion liquid component is removed by ultrafiltration of a particle dispersion containing a low molecular compound having a naphthalene skeleton, and then the low molecular compound having a naphthalene skeleton adsorbed on the particles is subjected to thermal analysis gas chromatography / mass spectrometry. By using an apparatus (PyGC / MS) (MS: JMS-600H (manufactured by JEOL Ltd.), GC: Agilent 6890N, Piolizer: PY-2020D Frontier Lab), a low level having a naphthalene skeleton is measured under the following conditions. The content of the molecular compound can be measured.
(Measurement condition)
Thermal decomposition temperature: 600 ° C
Column: SPB-5 (SPELCO)
Carrier gas: He 1mL / min

<Carrier particles>
The carrier particles are not particularly limited as long as they are insoluble carrier particles, but latex particles are preferable from the viewpoint of being suitable for immunolatex aggregation measurement.
Further, as the carrier particles, synthetic polymer type or natural polymer type polymer particles are preferable, and polymer particles derived from a monomer component containing a monomer having an aryl group are more preferable.
Examples of the monomer having an aryl group (hereinafter, also referred to as monomer (a)) include a monomer having a phenyl group and a monomer having a naphthyl group. One type may be used alone, or two or more types may be used in combination. May be. As the monomer having a phenyl group, a styrene monomer is preferable. Specific examples of the monomer having a phenyl group include styrene, aminostyrene, 2-methylstyrene, 4-methylstyrene, 4-chlorostyrene, 4-vinylbenzoic acid, divinylbenzene, vinyltoluene and the like. Moreover, as a monomer which has a naphthyl group, the unsaturated monomer which has a naphthyl group is preferable. Specific examples of the monomer having a naphthyl group include 1-vinylnaphthalene, 2-vinylnaphthalene, 1-naphthyl (meth) acrylate, 2-naphthyl (meth) acrylate, and the like.
The content of the monomer (a) is preferably 70 to 100% by mass, more preferably 90 to 99.8% by mass, and particularly preferably 95 to 99.5% by mass with respect to the total amount of the monomer components.

Moreover, as monomers other than the monomer (a) (hereinafter also referred to as monomer (b)), unsaturated carboxylic acid monomers such as (meth) acrylic acid, itaconic acid, maleic anhydride and crotonic acid, or unsaturated carboxylic anhydrides Acid monomer; unsaturated sulfonic acid such as 2-acrylamido-2-methylpropanesulfonic acid, isoprenesulfonic acid, styrenesulfonic acid, divinylbenzenesulfonic acid, 2-methylstyrenesulfonic acid, 4-methylstyrenesulfonic acid, and salts thereof Monomer or salt thereof; (meth) acrylamide, N-hydroxyethyl (meth) acrylamide, N, N-dimethyl (meth) acrylamide, N-isopropyl (meth) acrylamide, (meth) acryloylmorpholine, N, N-dimethylaminopropyl Acrylamide, N, N- (Meth) acrylamide monomers such as methyl chloride quaternary salt of methylaminopropylacrylamide, diacetone acrylamide, N-vinylacetamide; methoxyethyl (meth) acrylate, methyl (meth) acrylate, ethyl (meth) acrylate, ( Propyl (meth) acrylate, lauryl (meth) acrylate, cyclohexyl (meth) acrylate, isobornyl (meth) acrylate, benzyl (meth) acrylate, glycerol (meth) acrylate, polyethylene glycol (meth) acrylate, 2-hydroxy (Meth) acrylate monomers such as ethyl (meth) acrylate; unsaturated aldehyde monomers such as acrolein, as well as allylamine, N-vinyl-2-pyrrolidone, and the like. It may be used in combination.
Among these, as the monomer (b), an unsaturated carboxylic acid monomer, an unsaturated carboxylic anhydride monomer, an unsaturated sulfonic acid monomer, or a salt thereof is preferable. The salt of the unsaturated sulfonic acid monomer is not particularly limited, and examples thereof include alkali metal salts such as sodium salt, potassium salt and lithium salt; alkaline earth metal salts such as calcium salt; ammonium salt and the like.
The content of the monomer (b) is preferably 0 to 30% by mass, more preferably 0.2 to 10% by mass, and particularly preferably 0.5 to 5% by mass with respect to the total amount of the monomer components.

  The average particle size of the carrier particles is not particularly limited, but is preferably 0.01 to 1 μm, more preferably 0.02 to 0.75 μm, and particularly preferably 0.05 to 0.5 μm from the viewpoint of detection sensitivity and production cost. It is. In the present specification, the average particle diameter means a volume average particle diameter, and is a value measured in the same manner as in the examples.

The carrier particles may be those on which a substance that can bind to the target substance is supported or those that do not support a substance that can bind to the target substance. As the substance capable of binding to the target substance, an antigen or antibody against the target substance is preferable. Examples thereof include antigens such as receptors, enzymes, blood proteins, infectious disease-related antigens, microorganisms, viruses, hormones, and environmental substances (for example, environmental hormones), antibodies against these antigens, and the like. The antibody only needs to have a binding property to a specific antigen, and includes a fragment of the antibody.
Specifically, antibodies for coagulation / fibrinolysis related tests such as anti-antiplasmin antibodies, anti-D dimer antibodies, anti-FDP antibodies, anti-tPA antibodies, anti-thrombin / antithrombin complex antibodies, anti-FPA antibodies or antigens therefor; BFP antibody, anti-CEA antibody, anti-AFP antibody, anti-TSH antibody, anti-ferritin antibody, anti-CA19-9 antibody and other tumor-related test antibodies or antigens thereto; anti-apolipoprotein antibody, anti-β2-microbrovulin antibody, anti-antibody Antibody for serum protein-related test such as α1-microglobulin antibody, anti-immunoglobulin antibody, anti-CRP antibody or the antigen thereto; antibody for endocrine function test such as anti-HCG antibody or the antigen thereto; anti-digoxin antibody, anti-lidocaine antibody, etc. Antibodies for drug analysis or antigens thereto; HBs antigen, HCV antigen, HIV-1 antigen, HIV-2 Antigens for infectious disease-related tests such as antigens, HTLV-1 antigens, mycoplasma antigens, toxoplasma antigens, streptridine O antigens or antibodies thereto; antigens for autoimmune-related tests such as DNA antigens, heat-denatured human IgG, or antibodies thereto Is mentioned. The antibody may be a polyclonal antibody or a monoclonal antibody.
In addition, what is necessary is just to carry | support to the support particle of the substance which can be couple | bonded with a target substance in accordance with conventional methods, such as physical adsorption by hydrophobic-hydrophobic interaction, chemical bonding using a water-soluble carbodiimide type condensing agent, etc.

The carrier particles may be coated with a blocking agent. The blocking agent is not particularly limited as long as it can block a non-specific reaction, and examples thereof include biologically derived water-soluble polymers such as bovine serum albumin and chemically synthesized water-soluble polymers.
The coating of the blocking agent may be performed according to a conventional method such as dispersing the particles in a solution containing the blocking agent, then centrifuging, removing the supernatant, and redispersing the particles again with water or a buffer solution. Good.

  The content of the carrier particles (in terms of solid content) is preferably 0.001% by mass or more, more preferably 0.005% by mass or more, still more preferably 0.01% by mass or more, based on the total amount of the particle dispersion. Preferably it is 0.05% by mass or more, preferably 50% by mass or less, more preferably 40% by mass or less, still more preferably 30% by mass or less, still more preferably 20% by mass or less, and further preferably 15% by mass. It is as follows. And when making a particle dispersion into the particle dispersion for agglutination reagent manufacture, 0.1-20 mass% is preferable, 0.1-15 mass% is more preferable, 0.1-10 mass% is especially preferable. On the other hand, when the particle dispersion is used as an agglutinating reagent, 0.005 to 5% by mass is preferable, and 0.01 to 2% by mass is particularly preferable.

<Aqueous medium>
The aqueous medium may be any medium that can disperse the carrier particles, but preferably includes water. The content of water is preferably 40 to 100% by mass, more preferably 60 to 100% by mass, still more preferably 80 to 100% by mass, and particularly preferably 90 to 100% by mass with respect to the total amount of the aqueous medium.
The aqueous medium may be a buffer solution such as a Good buffer solution, a Tris buffer solution, a phosphate buffer solution, a glycine buffer solution, or an ammonia buffer solution.
The aqueous medium may contain a lower alcohol such as methanol, ethanol, isopropanol in addition to water and a buffer solution.
The content of the aqueous medium is preferably 50 to 99.9% by mass and more preferably 70 to 99.9% by mass with respect to the total amount of the particle dispersion. And when making a particle dispersion into the particle dispersion for aggregating reagent manufacture, 80-99.9 mass% is still more preferable, and 85-99.9 mass% is especially preferable. On the other hand, when the particle dispersion is used as an aggregating reagent, it is more preferably 95 to 99.9% by mass, and particularly preferably 98 to 99.9% by mass.

  Moreover, the particle dispersion liquid of the present invention may contain a surfactant, a blocking agent and the like in addition to the above-described components.

<PH>
The pH at 25 ° C. of the particle dispersion of the present invention is preferably in the range of 3.0 to 12.0, and more preferably in the range of 5.0 to 9.0 when used as an agglutinating reagent. .

Then, as described in Examples below, by incorporating a low molecular weight compound having a naphthalene skeleton in a particle dispersion of carrier particles using an aqueous medium as a dispersion medium, and performing detection by immunoaggregation using this, Sensitivity of the target substance in the sample can be increased.
Therefore, the particle dispersion of the present invention is useful as an agglutinating reagent or as a material for producing the agglutinating reagent (particle dispersion for producing an agglutinating reagent). The agglutinating reagent means a reagent that is used for detection of a target substance in a specimen by immunoagglutination and that quantifies the amount of the target substance based on the amount of change in turbidity when the particles aggregate.
When the particle dispersion liquid of the present invention is used as a particle dispersion liquid for producing an agglutinating reagent, the carrier particles that do not carry a substance that can bind to the target substance are dispersed as carrier particles in consideration of storage stability. Is preferred. On the other hand, when the particle dispersion liquid of the present invention is used as an agglutinating reagent, carrier particles carrying substances capable of binding to the target substance are dispersed as carrier particles in consideration of the reactivity with the target substance. Those are preferred.
The particle dispersion of the present invention is particularly useful as an agglutination reagent for immunoagglutination turbidimetry and a material for producing the agglutination reagent.

〔kit〕
A kit for use in detection of a target substance in a specimen by the immunoaggregation method of the present invention comprises the above particle dispersion. When provided in the kit, the particle dispersion is preferably a dispersion in which carrier particles carrying a substance capable of binding to the target substance are dispersed as carrier particles in consideration of reactivity with the target substance.

The kit of the present invention may include a reagent (also referred to as a first reagent) containing at least one selected from a blocking agent and an immune aggregation promoter in addition to the particle dispersion (also referred to as a second reagent). Good. Examples of the blocking agent and immune aggregation promoter include biologically derived water-soluble polymers such as bovine serum albumin and chemically synthesized water-soluble polymers. The first reagent may contain an aqueous medium similar to the above.
Moreover, the kit of this invention may be equipped with positive control, negative control, serum dilution liquid, etc. other than the said 1st reagent and 2nd reagent. The medium for positive control and negative control may be a solvent other than serum and physiological saline containing no target substance. Examples of the solvent include the above aqueous medium.

The kit of the present invention can be used in a method for detecting a target substance in the same manner as a kit for use in detecting a target substance in a specimen by a general immunoagglutination method. In addition, the concentration of the target substance can be measured according to a conventional method.
The kit of the present invention is particularly useful as a kit for immunoagglutination turbidimetry.

[Detection method]
The method for detecting a target substance in a specimen by the immunoaggregation method of the present invention is a step of bringing a carrier particle carrying a substance capable of binding to the target substance into contact with the specimen in the presence of a low molecular compound having a naphthalene skeleton. It is characterized by including. This detection method may be carried out in the same manner as a conventional detection method, for example, by mixing carrier particles and a specimen, except that it is performed in the presence of a low molecular compound having a naphthalene skeleton. In addition, examples of the low molecular weight compound having a naphthalene skeleton include the same compounds as those contained in the particle dispersion, and the amount used thereof is the content with respect to 100 parts by mass of carrier particle solid content in the particle dispersion. A similar range is preferred.

  The specimen only needs to contain the target substance or may contain the target substance. Specifically, various biological liquid samples such as serum, plasma, urine, saliva, feces and food Examples include pulverized specimens. Further, it may be a sample diluent obtained by diluting a sample with a pH buffer solution, protein, amino acid or the like.

The contact between the carrier particles and the specimen is preferably performed in the pH range of 4-10. The contact temperature is usually in the range of 25 to 45 ° C., and the contact time is usually 1 to 20 minutes.
In addition, the contact between the carrier particles and the specimen is performed in the presence of a low molecular compound having a naphthalene skeleton, but it is preferable to use a solvent together with the low molecular compound having a naphthalene skeleton. Examples of the solvent include those exemplified as an aqueous medium, lower alcohols such as ethanol, methanol, and isopropanol; ketones such as acetone, methyl ethyl ketone, and methyl isobutyl ketone; acetate esters such as ethyl acetate and butyl acetate; amides such as dimethylformamide; dimethyl Examples thereof include sulfoxides such as sulfoxide, and one of these may be used alone or two or more of them may be combined.

  The target substance in the specimen is detected by optically detecting the agglutination reaction that occurs as a result of contact between the carrier particles and the specimen, and the concentration of the target substance can also be measured. The optical detection of the agglutination reaction may be performed according to a conventional method using an optical instrument that can detect scattered light intensity, transmitted light intensity, absorbance, and the like. The detection method of the present invention is particularly useful for detection by immunoagglutination turbidimetry.

  EXAMPLES Hereinafter, although an Example is given and this invention is demonstrated in detail, this invention is not limited to these Examples. In addition, the average particle diameter (volume average particle diameter) of the particles was measured with a nanotrack particle size distribution analyzer UPA-EX150 (manufactured by Nikkiso Co., Ltd.).

[Preparation Example 1 Preparation of Latex Particles]
In a 1 L four-necked flask equipped with a condenser, a temperature controller, and a stirrer, 100 parts by mass of water, 0.1 part by mass of sodium dodecylbenzenesulfonate, 18 parts by mass of styrene, and 2 parts by mass of methacrylic acid are placed. While stirring at 200 revolutions per minute, 1 part by weight of potassium persulfate dissolved in 50 parts by weight of water was added. After reacting for 2 hours after the temperature reaches 75 ° C., a monomer comprising 75 parts by mass of styrene, 5 parts by mass of methacrylic acid, 0.6 part by mass of sodium dodecylbenzenesulfonate, and 200 parts by mass of water The emulsion was added dropwise over 5 hours and polymerization was continued for 4 hours to obtain an aqueous dispersion of polymer particles. Thereafter, the aqueous dispersion of the polymer particles was purified, and the solid content of the particles was adjusted to 10% by mass. This particle aqueous dispersion is referred to as seed particle aqueous dispersion (A). The average particle size of the seed particles contained in the seed particle aqueous dispersion (A) was 0.15 μm.
100 parts by mass of the above seed particle aqueous dispersion (A) (10 parts by mass as particle solids and 90 parts by mass as water) are put into a four-necked flask similar to the above and heated to 75 ° C. while stirring at 200 rpm. The mixture was charged with 1.5 parts by mass of potassium persulfate dissolved in 70 parts by mass of water, 89 parts by mass of styrene, 1 part by mass of methacrylic acid, 2 parts by mass of sodium dodecylbenzenesulfonate, and 400 parts by mass of water. Part of the monomer emulsion was added dropwise over 5 hours and then polymerized for 3 hours.
The particles obtained above are referred to as latex particles (B). The average particle size of the latex particles (B) was about 0.3 μm.

[Preparation Example 2 Preparation of First Reagent]
A 10 mM HEPES buffer (pH 7.5) containing 0.1% by mass of bovine serum albumin (BSA) was prepared. This is referred to as “first reagent”.

[Example 1 Preparation of second reagent (1)]
The latex particles (B) obtained in Preparation Example 1 were purified by centrifugation and resuspension, and added to water so that the particle solid content concentration was 10% by mass. 1 mL of this particle-containing solution and 9 mL of HEPES buffer (0.05 M, pH 7.5) were mixed (particle solid content concentration = 1% by mass), and 10 μL of an ethanol solution containing 1% by mass of 2-acetylnaphthalene was added thereto ( 2-acetylnaphthalene was added in an amount of 0.1 parts by mass with respect to 100 parts by mass of the particle solid content). This dispersion is designated as dispersion (C) for producing a second reagent (aggregation reagent).
In the dispersion (C) for producing the second reagent (aggregation reagent), 1-ethyl-3- (3-dimethylaminopropyl) carbodiimide hydrochloride (manufactured by Dojin Chemical Co., Ltd.) was added at a final concentration of 0.05% by mass. It added so that it might become. Furthermore, 0.5 mL of a 10 mg / mL aqueous solution of anti-CRP (C-reactive protein) antibody (rabbit) was added to this particle dispersion and stirred at room temperature for 3 hours to bind the antibody to the particle surface.
Thereafter, 0.5 mL of a 1% by weight aqueous solution of BSA was added to the particle dispersion, and the mixture was rotated and stirred at room temperature for 10 hours, and then centrifuged at 15,000 rpm for 15 minutes to collect particles as a precipitate.
The particles were then resuspended in 10 mL of HEPES buffer (0.05 M, pH 7.5) and dispersed with ultrasound for 10 minutes. This suspension was centrifuged again at 15,000 rpm for 15 minutes to collect the particles as a precipitate, and then the above resuspension and dispersion operations were repeated once more. To this, 190 mL of 10 mM HEPES buffer solution (pH 7.5) containing 0.02% by mass of BSA was added to adjust the particle solid content to 0.05% by mass, and then dispersed ultrasonically for 10 minutes, 0.8 μm I passed the disc filter.
By the above operation, a dispersion of antibody-bound latex particles coated with BSA as a blocking agent was obtained. This is designated as “second reagent (1)”. The content of 2-acetylnaphthalene contained in the second reagent (1) was about 0.1 parts by mass with respect to 100 parts by mass of the particle solid content.

[Example 2 Preparation of second reagent (2)]
In Example 1, instead of adding 10 μL of an ethanol solution containing 1% by mass of 2-acetylnaphthalene, 5 μL of an ethanol solution containing 10% by mass of 2-acetylnaphthalene (0.5% of 2-acetylnaphthalene in terms of 100 parts by mass of particle solid content) was added. (Mass part) A series of operations were performed in the same manner except that the second reagent (2) was obtained. The content of 2-acetylnaphthalene contained in the second reagent (2) was about 0.5 parts by mass with respect to 100 parts by mass of the particle solid content.

[Example 3 Preparation of second reagent (3)]
In Example 1, instead of adding 10 μL of an ethanol solution containing 1% by mass of 2-acetylnaphthalene, 10 μL of an ethanol solution containing 10% by mass of 2-acetylnaphthalene (2-acetylnaphthalene 1.0 in terms of 100 parts by mass of particle solid content) was added. (Mass part) A series of operations were performed in the same manner except that the second reagent (3) was obtained. The content of 2-acetylnaphthalene contained in the second reagent (3) was about 1.0 part by mass with respect to 100 parts by mass of the particle solid content.

[Example 4 Preparation of Second Reagent (4)]
In Example 1, instead of adding 10 μL of an ethanol solution containing 1% by mass of 2-acetylnaphthalene, 20 μL of an ethanol solution containing 10% by mass of 2-acetylnaphthalene (in contrast to 100 parts by mass of particle solid content, 2.0-acetylnaphthalene 2.0 A second reagent (4) was obtained in the same manner as described above except that (mass parts) was added. The content of 2-acetylnaphthalene contained in the second reagent (4) was about 2.0 parts by mass with respect to 100 parts by mass of the particle solid content.

[Example 5: Preparation of second reagent (5)]
In Example 1, instead of adding 10 μL of an ethanol solution containing 1% by mass of 2-acetylnaphthalene, 80 μL of an ethanol solution containing 10% by mass of 2-acetylnaphthalene (compared to 100 parts by mass of particle solid content of 8.0% 2-acetylnaphthalene). (Mass part) A series of operations were performed in the same manner except that the second reagent (5) was obtained. The content of 2-acetylnaphthalene contained in the second reagent (5) was about 8.0 parts by mass with respect to 100 parts by mass of the particle solid content.

[Example 6: Preparation of second reagent (6)]
In Example 1, a series of operations was performed in the same manner except that the ethanol solution containing 1% by mass of 2-acetylnaphthalene was replaced with an ethanol solution containing 1% by mass of 1-acetylnaphthalene, to obtain a second reagent (6). The content of 1-acetylnaphthalene contained in the second reagent (6) was about 0.1 parts by mass with respect to 100 parts by mass of the particle solid content.

[Example 7: Preparation of second reagent (7)]
In Example 2, a series of operations was performed in the same manner except that the ethanol solution containing 10% by mass of 2-acetylnaphthalene was replaced with the ethanol solution containing 10% by mass of 1-acetylnaphthalene, to obtain a second reagent (7). The content of 1-acetylnaphthalene contained in the second reagent (7) was about 0.5 parts by mass with respect to 100 parts by mass of the particle solid content.

[Example 8: Preparation of second reagent (8)]
In Example 3, a series of operations was performed in the same manner except that the ethanol solution containing 10% by mass of 2-acetylnaphthalene was replaced with an ethanol solution containing 10% by mass of 1-acetylnaphthalene, to obtain a second reagent (8). The content of 1-acetylnaphthalene contained in the second reagent (8) was about 1.0 part by mass with respect to 100 parts by mass of the particle solid content.

[Comparative Example 1 Preparation of Second Reagent (9)]
In Example 1, instead of adding 10 μL of an ethanol solution containing 1% by mass of 2-acetylnaphthalene, a series of operations were performed in the same manner except that 20 μL of ethanol was added to obtain a second reagent (9).

[Comparative Example 2 Preparation of Second Reagent (10)]
In Example 1, instead of adding 10 μL of an ethanol solution containing 1% by mass of 2-acetylnaphthalene, 40 μL of an aqueous solution containing 5% by mass of polyethylene glycol (molecular weight: 20000) (as compared with 100 parts by mass of particle solid content of polyethylene glycol 2.0) (Mass part) A series of operations were performed in the same manner except that the second reagent (10) was obtained.

[Test Example 1 Evaluation of Latex Aggregation Reaction]
In the measurement of immune latex agglutination, Hitachi 7180 type automatic analyzer was used. The measurement wavelength was 570 nm and the measurement temperature was 37 ° C. After adding 3 μL of physiological saline (CRP antigen concentration 0 mg / dL) or CRP antigen physiological saline dilution (CRP antigen concentration: 0.01 mg / dL or 0.04 mg / dL) to the measurement cell, 150 μL of the obtained first reagent was added to the cell and stirred uniformly, and then held for 5 minutes.
Thereafter, 150 μL of the second reagent (1) obtained in Example 1 was added to the cell and stirred uniformly, and then the difference between the absorbance after 50 seconds and the absorbance after 200 seconds was measured.
Further, the absorbance difference was measured in the same manner as described above except that the second reagent (1) obtained in Example 1 was changed to the second reagents (2) to (10).
The test results are shown in Table 1.

[Test Example 2 Storage Stability Test]
The second reagents (1) to (8) obtained in Examples 1 to 8 were stored at 25 ° C. for one week from the preparation of the second reagent, and the presence or absence of particle precipitation was visually confirmed. It was evaluated with. The test results are shown in Table 1.
(Evaluation criteria)
○: Precipitation of particles could not be confirmed ×: Precipitation of particles was confirmed

  From the results of Test Example 1 shown in Table 1, it was found that the low molecular weight compound having a naphthalene skeleton can greatly accelerate the latex agglutination reaction and increase the detection sensitivity.

Claims (11)

A particle dispersion liquid of carrier particles containing an aqueous medium as a dispersion medium and containing acetylnaphthalene .   The particle dispersion according to claim 1, wherein the aqueous medium contains water. The content of acetyl naphthalene, with respect to the support particles to 100 parts by weight of solid content, is 0.01 to 10 parts by weight, the particle dispersion according to claim 1 or 2. The content of acetyl naphthalene, with respect to the support particles to 100 parts by weight of solid content, 0.01 to 5 parts by weight, the particle dispersion liquid according to any one of claims 1-3. Wherein the carrier particles are latex particles, particle dispersion liquid according to any one of claims 1-4. Wherein the carrier particles are polymer particles derived from a monomer component containing a monomer having an aryl group, particle dispersion liquid according to any one of claims 1-5. The particle dispersion according to any one of claims 1 to 6 , wherein the carrier particles are carrier particles on which a substance capable of binding to a target substance is supported. The particle dispersion liquid according to claim 7 , wherein the substance capable of binding to the target substance is an antigen or an antibody against the target substance. The particle dispersion according to claim 7 or 8 , which is for use as an agglutinating reagent. A kit for use in detection of a target substance in a specimen by immunoagglutination, comprising the particle dispersion according to any one of claims 1 to 9 . A method for detecting a target substance in a specimen by immunoagglutination, comprising a step of contacting a carrier particle carrying a substance capable of binding to the target substance and the specimen in the presence of acetylnaphthalene Detection method.