WO2020196786A1 - Magnetic particles and test agent - Google Patents

Magnetic particles and test agent Download PDF

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Publication number
WO2020196786A1
WO2020196786A1 PCT/JP2020/013814 JP2020013814W WO2020196786A1 WO 2020196786 A1 WO2020196786 A1 WO 2020196786A1 JP 2020013814 W JP2020013814 W JP 2020013814W WO 2020196786 A1 WO2020196786 A1 WO 2020196786A1
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WO
WIPO (PCT)
Prior art keywords
magnetic
particles
substance
resin particles
layer
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Application number
PCT/JP2020/013814
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French (fr)
Japanese (ja)
Inventor
秀平 大日方
脇屋　武司
祐也 稲葉
Original Assignee
積水化学工業株式会社
積水メディカル株式会社
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Application filed by 積水化学工業株式会社, 積水メディカル株式会社 filed Critical 積水化学工業株式会社
Priority to CN202080023454.4A priority Critical patent/CN113614530A/en
Priority to JP2021509614A priority patent/JPWO2020196786A1/ja
Publication of WO2020196786A1 publication Critical patent/WO2020196786A1/en

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    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/543—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/543—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals
    • G01N33/551—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals the carrier being inorganic
    • G01N33/553—Metal or metal coated

Definitions

  • the present invention relates to magnetic particles containing a magnetic material.
  • the present invention also relates to a test agent using the above magnetic particles.
  • Magnetic particles are used to measure the concentration of target substances in samples in fields such as pharmaceutical research and development and clinical tests.
  • immunoassays such as chemiluminescence immunoassay (CLIA method)
  • magnetic particles having an antibody or antigen on the surface are widely used.
  • these magnetic particles are magnetized by a magnet or the like after being bound to an antigen or an antibody which is a target substance.
  • magnetic particles magnetic particles having a magnetic substance inside the resin particles (see, for example, Patent Documents 1 and 2) and magnetic particles having a magnetic layer formed on the outer surface of the resin particles (for example, Patent Documents). 3) is used.
  • the conventional magnetic particles may have low magnetic collection.
  • the magnetic material may oxidize and the magnetic collection may decrease over time.
  • conventional magnetic particles may have low dispersibility.
  • the number of magnetic particles that cannot be completely collected by the magnet increases, and the measurement accuracy and measurement sensitivity deteriorate when measuring the concentration of the target substance such as an antibody or antigen.
  • the magnetic particles may not be sufficiently redispersed after magnetic separation, and the measurement accuracy, measurement sensitivity and measurement reproducibility may decrease.
  • An object of the present invention is to provide magnetic particles capable of enhancing magnetic collection and dispersibility, and maintaining high magnetic collection. Further, a limited object of the present invention is to provide magnetic particles capable of maintaining high magnetic collection for a long period of time. Another object of the present invention is to provide a test agent using the above magnetic particles.
  • magnetic particles used for specifically interacting with a target substance the magnetic inclusion resin particles containing the first magnetic substance inside, and the magnetic inclusion resin particles.
  • a magnetism having a magnetic layer arranged on the outer surface of the magnetic layer and containing a second magnetic substance, and a substance supported on the outer surface side of the magnetic layer and specifically interacting with the target substance. Particles are provided.
  • the first magnetic material is a metal or a metal oxide
  • the second magnetic material is a metal or a metal oxide
  • a shell layer is further provided on the outer surface of the magnetic layer, the material of the shell layer containing an inorganic oxide or an organic polymer, and the shell layer. , The substance is bound.
  • the material of the shell layer is an inorganic oxide containing the inorganic oxide, wherein the inorganic oxide has a silicon atom, a germanium atom, a titanium atom or a zirconium atom. is there.
  • the substance is an antigen or antibody.
  • the substance is avidin or streptavidin.
  • the total content of the first magnetic substance and the second magnetic substance in 100% by volume of the magnetic particles is 10% by weight or more and 95% by weight or less. ..
  • the content of the first magnetic substance is 10% by weight in a total of 100% by weight of the contents of the first magnetic substance and the second magnetic substance. More than 90% by weight or less.
  • the content of the first magnetic substance in 100% by volume of the region from the outer surface to the center of the magnetic inclusion resin particles up to 1/3 of the thickness is 0.8 or more and 4.0 or less. is there.
  • the magnetic particles are used as a test agent.
  • a test agent containing the above-mentioned magnetic particles is provided.
  • the magnetic particles according to the present invention are used to specifically interact with a target substance.
  • the magnetic particles according to the present invention include magnetic encapsulating resin particles containing a first magnetic substance inside, and a magnetic layer arranged on the outer surface of the magnetic encapsulating resin particles and containing a second magnetic substance. It includes a substance that is supported on the outer surface side of the magnetic layer and that specifically interacts with the target substance. Since the magnetic particles according to the present invention have the above-mentioned structure, the magnetic collection and dispersibility can be enhanced, and the magnetic collection can be maintained high.
  • FIG. 1 is a cross-sectional view schematically showing magnetic particles according to the first embodiment of the present invention.
  • FIG. 2 is a cross-sectional view schematically showing magnetic particles according to a second embodiment of the present invention.
  • FIG. 3 is a cross-sectional view schematically showing the magnetic particles according to the third embodiment of the present invention.
  • FIG. 4 is a cross-sectional view schematically showing the magnetic particles according to the fourth embodiment of the present invention.
  • FIG. 5 is a schematic diagram for explaining each region for obtaining the content of the first magnetic substance in the magnetic inclusion resin particles.
  • the magnetic particles according to the present invention are used to specifically interact with a target substance.
  • the magnetic particles according to the present invention are magnetic particles that can specifically interact with a target substance.
  • the magnetic particles according to the present invention include magnetic encapsulating resin particles containing a first magnetic substance inside, and a magnetic layer arranged on the outer surface of the magnetic encapsulating resin particles and containing a second magnetic substance. It includes a substance that is supported on the outer surface side of the magnetic layer and that specifically interacts with the target substance.
  • the magnetic particles according to the present invention have the above-mentioned structure, the magnetic collection and dispersibility can be enhanced, and the magnetic collection can be maintained high. In the present invention, the magnetism collection can be maintained high for a long period of time. In the magnetic particles according to the present invention, in the measurement of the target substance using the magnetic particles, the amount of binding to the target substance per unit weight of the magnetic particles can be increased, and therefore the measurement sensitivity can be increased.
  • Conventional magnetic particles may have low magnetic collection or dispersibility.
  • the conventional magnetic particles having a magnetic substance inside the resin particles it is difficult to sufficiently increase the content of the magnetic substance because the space inside the resin particles is limited. Therefore, it is difficult to improve the magnetic collection with the conventional magnetic particles.
  • the conventional magnetic particles having a magnetic layer on the outer surface of the resin particles when the content of the magnetic substance contained in the magnetic layer is increased, the magnetic collection can be increased to some extent, but the particles of the magnetic particles. This is not preferable because the diameter becomes large and the amount of the target substance bonded per unit area of the magnetic particles decreases. Further, when the magnetic layer on the outer surface of the resin particles is thickened, the repulsive force between the magnetic particles is lowered, and the dispersibility may be lowered.
  • the smaller the particle diameter the lower the content of the magnetic substance.
  • the magnetic collection may decrease, and as a result, the measurement sensitivity may decrease.
  • the magnetic substance is oxidized and the magnetic collection property is lowered with time. There is.
  • the content of the magnetic substance can be increased and the magnetic collection can be enhanced while keeping the particle diameter of the magnetic particles small.
  • the magnetic particles according to the present invention can enhance the dispersibility.
  • high magnetic collection can be maintained even if the magnetic material deteriorates to some extent.
  • the magnetic particles according to the present invention even if the specific magnetic layer containing the second magnetic material is deteriorated to some extent, the magnetic particles contain the first magnetic material, so that the magnetic collection property can be maintained high.
  • the measurement accuracy, measurement sensitivity and measurement reproducibility can be improved in the measurement of the target substance using the magnetic particles.
  • the average particle size of the magnetic particles is preferably 0.5 ⁇ m or more, more preferably 1.0 ⁇ m or more, preferably 10 ⁇ m or less, more preferably 7 ⁇ m or less, still more preferably 5 ⁇ m or less, particularly preferably 4 ⁇ m or less, most preferably. It is preferably 3.5 ⁇ m or less.
  • the average particle size of the magnetic particles is at least the above lower limit, the magnetism collection can be further enhanced.
  • the average particle size of the magnetic particles is not more than the upper limit, the content of the substance that specifically interacts with the target substance per unit weight can be increased, and the binding amount of the target substance can be increased. it can.
  • the average particle size of the magnetic particles is a number average particle size.
  • the particle size of the magnetic particles is determined, for example, by observing 50 arbitrary magnetic particles with an electron microscope or an optical microscope and calculating the average value of the particle sizes of each magnetic particle. It is preferable to prepare a sample obtained by drying the magnetic particles and observe the obtained sample with an electron microscope or an optical microscope.
  • the coefficient of variation (CV value) of the average particle size of the magnetic particles is preferably 10% or less, more preferably 5% or less.
  • the coefficient of variation of the average particle size of the magnetic particles is not more than the above upper limit, the measurement accuracy can be further improved in the measurement of the target substance using the magnetic particles.
  • the coefficient of variation (CV value) can be measured as follows.
  • CV value (%) ( ⁇ / Dn) ⁇ 100 ⁇ : Standard deviation of the particle size of the magnetic particles Dn: Average value of the particle size of the magnetic particles
  • FIG. 1 is a cross-sectional view schematically showing magnetic particles according to the first embodiment of the present invention.
  • the magnetic particle 1 shown in FIG. 1 is used to specifically interact with the target substance.
  • the magnetic particles 1 include magnetic inclusion resin particles 2, a magnetic layer 3, a shell layer 4, and a substance 5 that specifically interacts with a target substance.
  • the substance 5 is, for example, a physiologically active substance such as avidin, streptavidin, an antigen and an antibody.
  • the magnetic inclusion resin particles 2 have resin particles 21 and a first magnetic material 22.
  • the magnetic inclusion resin particles 2 contain a first magnetic substance 22 inside.
  • the resin particles 21 contain a first magnetic material 22 inside.
  • the first magnetic material 22 is dispersed inside the magnetic inclusion resin particles 2.
  • the first magnetic material 22 is dispersed inside the resin particles 21.
  • the first magnetic material 22 is uniformly distributed inside the magnetic inclusion resin particles 2.
  • the first magnetic material does not have to be uniformly distributed inside the magnetic inclusion resin particles.
  • the first magnetic material may be arranged from the center of the magnetic inclusion resin particles to the outer surface so that the content of the first magnetic material increases.
  • the magnetic layer 3 is arranged on the outer surface of the magnetic inclusion resin particles 2.
  • the magnetic layer 3 is arranged on the outer surface of the resin particles 21 in the magnetic inclusion resin particles 2.
  • the magnetic layer 3 contains a second magnetic material.
  • the shell layer 4 is arranged on the outer surface of the magnetic layer 3.
  • the substance 5 is supported on the outer surface side of the magnetic layer 3.
  • the substance 5 is supported on the outer surface of the shell layer 4.
  • the shell layer 4 and the substance 5 are bonded to each other.
  • the substance 5 is present on the surface of the magnetic particles 1.
  • the magnetic layer 3 is a single magnetic layer.
  • the magnetic layer 3 covers the entire outer surface of the magnetic inclusion resin particles 2.
  • the magnetic layer may cover the entire outer surface of the magnetic inclusion resin particles, or the magnetic layer may cover a part of the surface of the magnetic inclusion resin particles.
  • the magnetic layer may be a single magnetic layer or a multi-layered magnetic layer composed of two or more layers.
  • the shell layer 4 is a single-layer shell layer.
  • the shell layer 4 covers the entire outer surface of the magnetic layer 3.
  • the shell layer may cover the entire outer surface of the magnetic layer, or the shell layer may cover a part of the surface of the magnetic layer.
  • FIG. 2 is a cross-sectional view schematically showing the magnetic particles according to the second embodiment of the present invention.
  • the magnetic particles 1A shown in FIG. 2 are used to specifically interact with the target substance.
  • the magnetic particles 1A include magnetic inclusion resin particles 2A, a magnetic layer 3A, and a substance 5A that specifically interacts with a target substance.
  • the substance 5A is, for example, avidin, streptavidin, an antigen, an antibody, or the like.
  • the magnetic particles 1A do not have the shell layer.
  • the magnetic inclusion resin particles 2A have resin particles 21A and a first magnetic body 22A.
  • the distribution state of the first magnetic substance is different between the magnetic encapsulating resin particles 2 shown in FIG. 1 and the magnetic encapsulating resin particles 2A shown in FIG.
  • the first magnetic substance 22A is not uniformly distributed inside the magnetic inclusion resin particles 2A. From the center of the magnetic inclusion resin particles 2A to the outer surface, the first magnetic body 22A is arranged so that the content of the first magnetic body 22A increases.
  • the first magnetic material may be uniformly distributed inside the magnetic inclusion resin particles.
  • the magnetic layer 3A is arranged on the outer surface of the magnetic inclusion resin particles 2A.
  • the magnetic layer 3A contains a second magnetic material.
  • the substance 5A is supported on the outer surface of the magnetic layer 3A.
  • the substance 5A is present on the surface of the magnetic particles 1A.
  • the magnetic layer 3A is a single magnetic layer.
  • the magnetic layer 3A covers the entire outer surface of the magnetic inclusion resin particles 2A.
  • the magnetic layer may be a single magnetic layer or a multi-layered magnetic layer composed of two or more layers.
  • FIG. 3 is a cross-sectional view schematically showing the magnetic particles according to the third embodiment of the present invention.
  • the magnetic particles 1B shown in FIG. 3 are used to specifically interact with the target substance.
  • the magnetic particles 1B include magnetic inclusion resin particles 2B, a magnetic layer 3B, a shell layer 4B, and a substance 5B that specifically interacts with a target substance.
  • the substance 5B is, for example, a bioactive substance such as avidin, streptavidin, an antigen and an antibody.
  • the magnetic inclusion resin particles 2B have resin particles 211B, a first magnetic body 22B, and a resin layer 212B.
  • the magnetic inclusion resin particles 2B contain a first magnetic substance 22B inside.
  • the first magnetic substance 22B is contained in a layered manner.
  • the magnetic inclusion resin particles 2B have a magnetic layer containing the first magnetic body 22B inside.
  • the resin particles 211B do not contain the first magnetic material 22B.
  • the magnetic inclusion resin particles 2B contains the first magnetic body 22B inside.
  • a magnetic layer containing the first magnetic substance 22B is arranged on the outer surface of the resin particles 211B, and the resin layer 212B is arranged on the outer surface of the magnetic layer.
  • the magnetic layer 3B is arranged on the outer surface of the magnetic inclusion resin particles 2B.
  • the magnetic layer 3B contains a second magnetic material.
  • the shell layer 4B is arranged on the outer surface of the magnetic layer 3B.
  • the substance 5B is supported on the outer surface side of the magnetic layer 3B.
  • the substance 5B is supported on the outer surface of the shell layer 4B.
  • the shell layer 4B and the substance 5B are bonded.
  • the substance 5B exists on the surface of the magnetic particles 1B.
  • the magnetic layer 3B is a single magnetic layer.
  • the magnetic layer 3B covers the entire outer surface of the magnetic inclusion resin particles 2B.
  • the magnetic layer may be a single magnetic layer or a multi-layered magnetic layer composed of two or more layers.
  • the shell layer 4B is a single-layer shell layer.
  • the shell layer 4B covers the entire outer surface of the magnetic layer 3B.
  • FIG. 4 is a cross-sectional view schematically showing the magnetic particles according to the fourth embodiment of the present invention.
  • the magnetic particles 1C shown in FIG. 4 are used to specifically interact with the target substance.
  • the magnetic particles 1C include magnetic inclusion resin particles 2C, a magnetic layer 3C, a shell layer 4C, and a substance 5C that specifically interacts with a target substance.
  • the substance 5C is, for example, a bioactive substance such as avidin, streptavidin, an antigen and an antibody.
  • the magnetic inclusion resin particles 2C have resin particles 211C, first magnetic materials 221C and 222C, and a resin layer 212C.
  • the magnetic inclusion resin particles 2C contain the first magnetic substances 221C and 222C inside.
  • the first magnetic body 221C is dispersed inside the magnetic inclusion resin particles 2C.
  • the first magnetic material 221C is dispersed inside the resin particles 211C.
  • the first magnetic body 221C is dispersed inside the resin particles 211C.
  • the first magnetic substance 222C is contained in a layered manner.
  • the magnetic inclusion resin particles 2C have a magnetic layer containing the first magnetic substance 222C inside.
  • the magnetic inclusion resin particles 2C contains the first magnetic material 222C inside.
  • a magnetic layer containing the first magnetic substance 222C is arranged on the outer surface of the resin particles 211C, and the resin layer 212C is arranged on the outer surface of the magnetic layer.
  • the magnetic layer 3C is arranged on the outer surface of the magnetic inclusion resin particles 2C.
  • the magnetic layer 3C contains a second magnetic material.
  • the shell layer 4C is arranged on the outer surface of the magnetic layer 3C.
  • the substance 5C is supported on the outer surface side of the magnetic layer 3C.
  • the substance 5C is supported on the outer surface of the shell layer 4C.
  • the shell layer 4C and the substance 5C are bonded.
  • the substance 5C is present on the surface of the magnetic particles 1C.
  • the magnetic layer 3C is a single magnetic layer.
  • the magnetic layer 3C covers the entire outer surface of the magnetic inclusion resin particles 2C.
  • the magnetic layer may be a single magnetic layer or a multi-layered magnetic layer composed of two or more layers.
  • the shell layer 4C is a single-layer shell layer.
  • the shell layer 4C covers the entire outer surface of the magnetic layer 3C.
  • the magnetic inclusion resin particles contain a resin.
  • the magnetic inclusion resin particles include resin particles.
  • the magnetic encapsulating resin particles contain a first magnetic substance inside. It is preferable that the magnetic encapsulating resin particles contain the first magnetic substance inside by having the following constitution (1) or (2). (1) It contains resin particles and a first magnetic substance, and the first magnetic substance is dispersed inside the resin particles. (2) The resin particles, the first magnetic material, and the resin layer are included, and the first magnetic material is contained on the outer surface of the resin particles from the center of the magnetic inclusion resin particles toward the outer surface. The magnetic layer and the resin layer are arranged alternately.
  • the magnetic inclusion resin particles may have only the configuration (1), may have only the configuration (2), or may have the configuration (1) and the configuration (2).
  • the magnetic encapsulating resin particles have the configuration (2), it is preferable that a magnetic layer containing the first magnetic substance is arranged on the outer surface of the resin particles, and the magnetic encapsulating resin particles The outermost layer is preferably a resin layer.
  • the magnetic encapsulating resin particles having the above configuration (1) are, for example, the magnetic encapsulating resin particles shown in FIGS. 1 and 2.
  • the magnetic encapsulating resin particles having the above configuration (2) are, for example, the magnetic encapsulating resin particles shown in FIG.
  • the magnetic encapsulating resin particles having the above-mentioned configuration (1) and the above-mentioned configuration (2) are, for example, the magnetic encapsulating resin particles shown in FIG.
  • the magnetic encapsulating resin particles may have one resin layer and one magnetic layer containing the first magnetic material. Often, it may have two or more resin layers and two or more magnetic layers including the first magnetic material.
  • Resin particles and resin layer examples include polyolefin resins such as polyethylene, polypropylene, polystyrene, polyvinyl chloride, polyvinylidene chloride, polyisobutylene and polybutadiene; acrylic resins such as polymethylmethacrylate and polymethylacrylate; polycarbonate and polyamide.
  • divinylbenzene copolymer and the like examples include a divinylbenzene-styrene copolymer and a divinylbenzene- (meth) acrylic acid ester copolymer.
  • the material of the resin particles and the resin layer only one kind may be used, or two or more kinds may be used in combination. Further, when the magnetic inclusion resin particles have the above configuration (2), the material of the resin particles and the material of the resin layer may be the same or different.
  • the resin particles preferably have a porous structure.
  • the resin particles in the magnetic inclusion resin particles having the above configuration (1) preferably have a porous structure.
  • the resin particles in the magnetic inclusion resin particles having the above configuration (2) may or may not have a porous structure.
  • the BET specific surface area of the resin particles is preferably 20 m 2 / g or more, more preferably 40 m 2 / g or more, further preferably 100 m 2 / g or more, preferably 800 m 2 / g or less, and more preferably 700 m 2 It is / g or less, more preferably 650 m 2 / g or less.
  • the BET specific surface area is not less than the above lower limit and not more than the above upper limit, the content of the first magnetic substance contained in the magnetic inclusion resin particles can be increased, and the magnetic collection can be further enhanced.
  • the average pore diameter of the resin particles is preferably 0.5 nm or more, more preferably 1 nm or more, preferably 30 nm or less, and more preferably 10 nm or less.
  • the average pore diameter is equal to or greater than the above lower limit and equal to or less than the above upper limit, the magnetic substance can be more easily contained inside the resin particles, and the content of the first magnetic substance contained in the magnetic inclusion resin particles can be contained. The amount can be increased, and the magnetic collection can be further enhanced.
  • the BET specific surface area and the average pore diameter of the resin particles can be measured from the adsorption isotherm of nitrogen in accordance with the BJH method.
  • Examples of the measuring device for measuring the BET specific surface area and the average pore diameter of the resin particles include "NOVA4200e" manufactured by Cantachrome Instruments.
  • Resin particles satisfying the preferable ranges such as the BET specific surface area and the average pore diameter can be obtained, for example, by a method for producing resin particles including the following steps.
  • the polymerizable monomer include a monofunctional monomer and a polyfunctional monomer.
  • the organic solvent that does not react with the polymerizable monomer is not particularly limited as long as it is incompatible with a polar solvent such as water, which is a polymerization medium.
  • the organic solvent include cyclohexane, toluene, xylene, ethyl acetate, butyl acetate, allyl acetate, propyl acetate, chloroform, methylcyclohexane, methyl ethyl ketone and the like.
  • the amount of the organic solvent added is preferably 1 part to 80 parts by weight, more preferably 20 parts by weight to 60 parts by weight, based on 100 parts by weight of the polymerizable monomer component.
  • the amount of the organic solvent added is in the above-mentioned preferable range, the BET specific surface area, the above-mentioned average pore diameter, and the like can be controlled in a more suitable range, and it becomes easy to obtain dense pores inside the particles.
  • the average particle size of the resin particles is preferably 0.5 ⁇ m or more, more preferably 1 ⁇ m or more, preferably 10 ⁇ m or less, more preferably 7 ⁇ m or less, still more preferably 5 ⁇ m or less, particularly preferably 4 ⁇ m or less, and most preferably 3. It is 5 ⁇ m or less.
  • the average particle size of the resin particles is at least the above lower limit, the magnetism collection can be further enhanced.
  • the average particle size of the resin particles is not more than the above upper limit, the sedimentation of the magnetic particles can be effectively suppressed, the dispersibility can be further improved, and the particle size of the magnetic particles can be reduced.
  • the target substance can be effectively interacted with each other.
  • the average particle size of the resin particles is a number average particle size.
  • the average particle size of the resin particles is obtained by observing 50 arbitrary resin particles with an electron microscope or an optical microscope and calculating the average value of the particle size of each resin particle. It is preferable to prepare a sample obtained by drying the magnetic particles or the resin particles, and observe the obtained sample with an electron microscope or an optical microscope.
  • the first magnetic material is preferably a metal or a metal oxide, and more preferably a ferromagnetic material or a paramagnetic material.
  • the first magnetic material examples include iron, cobalt, nickel, ruthenium, lanthanoids, ferrite and the like.
  • the ferrite in chromite to mug ( ⁇ Fe 2 O 3) and MFe compounds represented by 2 O 4 (MFe 2 O 4 , M is, Co, Ni, Mn, Zn , Mg, Cu, Fe, Li 0 .5 Fe 0.5 etc.) and the like.
  • the ferrite is preferably ferric tetroxide (Fe 3 O 4 ).
  • the first magnetic material may be an alloy. Examples of the alloy include nickel-cobalt alloy, cobalt-tungsten alloy, iron-platinum alloy, iron-cobalt alloy and the like.
  • the said metal may be a metal ion.
  • the first magnetic material only one kind may be used, or two or more kinds may be used in combination.
  • the first magnetic material is preferably cobalt or ferrite, and more preferably cobalt or triiron tetroxide. It is preferable, and cobalt is more preferable.
  • the content of the first magnetic substance in 100% by volume of the magnetic particles is preferably 10% by volume or more, more preferably 20% by volume or more, preferably 80% by volume or less, and more preferably 70% by volume or less. ..
  • the content of the first magnetic substance is at least the above lower limit, the magnetism collection can be further enhanced.
  • the content of the first magnetic substance is not more than the above upper limit, the dispersibility can be further enhanced.
  • the content of the first magnetic material is preferably 10% by weight or more, more preferably 20% by weight or more, and preferably 90% by weight in the total of 100% by weight of the first magnetic material and the second magnetic material. % Or less, more preferably 85% by weight or less.
  • the content of the first magnetic substance is not less than the above lower limit and not more than the above upper limit, the magnetic collection can be further enhanced and the high magnetic collection can be maintained for a longer period of time.
  • the content of the first magnetic substance (hereinafter referred to as the content (1)) in 100% by volume of the region (R1) from the outer surface to the center of the magnetic inclusion resin particles to the thickness of 1/3 is described.
  • the content (1) is at least the above lower limit and at least the above upper limit, the magnetism collection can be further enhanced. Further, when the content (1) is not more than the above upper limit, the resin skeleton is well retained and the particle shape can be well maintained.
  • the region (R1) is a region outside the broken line L1 of the magnetic inclusion resin particles 2 in FIG.
  • the content of the first magnetic substance (hereinafter referred to as the content (2)) in 100% by volume of the region (R2) having a thickness of 2/3 from the center of the magnetic inclusion resin particles to the outer surface is described.
  • the region (R2) is a region inside the broken line L1 of the magnetic inclusion resin particles 2 in FIG.
  • the ratio of the content (1) to the content (2) is preferably 0.8 or more, more preferably 1.0 or more, and preferably 4 It is 0.0 or less, more preferably 3.0 or less.
  • the above ratio (content (1) / content (2)) is at least the above lower limit and at least the above upper limit, the magnetism collection and dispersibility can be further enhanced.
  • the absolute value of the difference between the content (1) and the content (2) is preferably 25% by volume or less, and more preferably 10% by volume or less.
  • the first magnetic material is preferably uniformly present inside the magnetic inclusion resin particles, and is preferably uniformly contained inside the magnetic inclusion resin particles.
  • the above-mentioned content (1) and the above-mentioned content (2) can be measured as follows.
  • the above-mentioned content (1) and the above-mentioned content (2) can be calculated.
  • the above-mentioned content (1) and the above-mentioned content (2) are averages calculated by arithmetically averaging the contents (1) and the contents (2) of 20 arbitrarily selected magnetic inclusion resin particles.
  • the content is preferably.
  • the method for producing the magnetic encapsulating resin particles is not particularly limited. For example, by mixing the resin particles having a porous structure and the first magnetic material and introducing the first magnetic material into the resin particles, a magnetic body having the above configuration (1) is provided. Encapsulating resin particles can be obtained. Further, for example, the resin particles having a solid structure and the first magnetic material are mixed, the outer surface of the resin particles is coated with the first magnetic material, and then the outer surface of the first magnetic material is coated. By coating the above with a resin, magnetic inclusion resin particles having the above configuration (2) can be obtained.
  • the magnetic particles according to the present invention include a magnetic layer containing a second magnetic material.
  • the magnetic layer containing the second magnetic material is arranged on the outer surface of the magnetic inclusion resin particles.
  • the first magnetic material and the second magnetic material may be the same or different.
  • the second magnetic material is preferably a metal or a metal oxide, and more preferably a ferromagnetic material or a paramagnetic material.
  • the second magnetic material examples include iron, cobalt, nickel, ruthenium, lanthanoids, ferrite and the like.
  • the ferrite in chromite to mug ( ⁇ Fe 2 O 3) and MFe compounds represented by 2 O 4 (MFe 2 O 4 , M is, Co, Ni, Mn, Zn , Mg, Cu, Fe, Li 0 .5 Fe 0.5 etc.) and the like.
  • the ferrite is preferably ferric tetroxide (Fe 3 O 4 ).
  • the second magnetic material may be an alloy. Examples of the alloy include nickel-cobalt alloy, cobalt-tungsten alloy, iron-platinum alloy, iron-cobalt alloy and the like.
  • the said metal may be a metal ion.
  • the second magnetic material only one kind may be used, or two or more kinds may be used in combination.
  • the second magnetic material is preferably ferrite, and more preferably triiron tetroxide.
  • the content of the second magnetic material is preferably 10% by weight or more, more preferably 30% by weight or more, preferably 90% by weight, based on 100% by weight of the total of the first magnetic material and the second magnetic material. % Or less, more preferably 70% by weight or less.
  • the content of the second magnetic substance is not less than the above lower limit and not more than the above upper limit, the magnetic collection can be further enhanced and the high magnetic collection can be maintained for a longer period of time.
  • the content of the second magnetic substance in 100% by volume of the magnetic particles is preferably 4% by volume or more, more preferably 8% by volume or more, preferably 40% by volume or less, and more preferably 25% by volume or less. ..
  • the content of the second magnetic substance is at least the above lower limit, the magnetism collection can be further enhanced.
  • the content of the second magnetic substance is not more than the above upper limit, the dispersibility can be further enhanced.
  • the total content of the first magnetic substance and the second magnetic substance in 100% by weight of the magnetic particles is preferably 10% by weight or more, more preferably 15% by weight or more, and preferably 95% by weight or less. , More preferably 80% by weight or less.
  • the total content is at least the above lower limit, the magnetism collection can be further enhanced.
  • the dispersibility can be further enhanced.
  • the total content of the first magnetic substance and the second magnetic substance in 100% by volume of the magnetic particles is preferably 20% by volume or more, more preferably 30% by volume or more, and preferably 80% by volume or less. , More preferably 70% by volume or less.
  • the total content is at least the above lower limit, the magnetism collection can be further enhanced.
  • the dispersibility can be further enhanced.
  • the surface area covered by the magnetic layer containing the second magnetic substance is preferably 70% or more, more preferably 80% or more, still more preferably. It is 95% or more, most preferably 100%.
  • the surface area is at least the above lower limit, the magnetic collection can be further enhanced, the high magnetic collection can be maintained for a longer period of time, and the dispersibility can be further enhanced.
  • the thickness of the magnetic layer is preferably 20 nm or more, more preferably 50 nm or more, preferably 1000 nm or less, and more preferably 200 nm or less.
  • the thickness of the magnetic layer is the thickness of the entire magnetic layer when the magnetic layer is multi-layered.
  • the thickness of the magnetic layer is at least the above lower limit, the magnetic collection can be further enhanced, and the high magnetic collection can be maintained for a longer period of time.
  • the thickness of the magnetic layer is not more than the above upper limit, the sedimentation of the magnetic particles can be effectively suppressed, the dispersibility can be further improved, and the particle size of the magnetic particles can be reduced, so that the magnetic particles can be reduced.
  • the amount of binding of the target substance per unit weight of the above can be increased.
  • the thickness of the magnetic layer can be measured by observing the cross section of the magnetic particles, for example, using a transmission electron microscope (TEM). Regarding the thickness of the magnetic layer, it is preferable to calculate the average value of five thicknesses of any magnetic layer as the thickness of the magnetic layer of one magnetic particle, and the average value of the thickness of the entire magnetic layer is one magnetic. It is more preferable to calculate as the thickness of the magnetic layer of the particles.
  • the thickness of the magnetic layer is preferably obtained by calculating the average value of the thickness of the magnetic layer of each magnetic particle for 10 arbitrary magnetic particles.
  • the magnetic layer may be a continuous layer, or may be a layer formed by particles in which the second magnetic material is aggregated in the form of particles. Further, the magnetic layer may be entirely covered on the outer surface of the resin particles, or may be partially covered. Further, the magnetic layer may have a sea-island structure. From the viewpoint of effectively suppressing the decrease in magnetic collection, the magnetic layer is preferably a continuous layer.
  • the continuous layer has a structure having few seams or no seams, unlike a shape in which innumerable seams (for example, 1000 or more per magnetic particle) exist in the magnetic layer such as an aggregate of fine particles. Point to. From the viewpoint of further enhancing the magnetic collection, the magnetic layer is preferably a layer formed by particles in which the second magnetic material is aggregated in the form of particles.
  • the average particle size of the particles is preferably 1 nm or more, more preferably 2 nm or more, preferably 2 nm or more. Is 50 nm or less, more preferably 20 nm or less.
  • the average particle size of the granules is at least the above lower limit, the magnetism collection can be further enhanced.
  • the second magnetic substance can be satisfactorily arranged on the outer surface of the magnetic inclusion resin particles, and the magnetic layer can be satisfactorily formed.
  • the method for forming the magnetic layer is not particularly limited. For example, by mixing the magnetic inclusion resin particles and the particles of the second magnetic substance, a magnetic layer can be formed on the outer surface of the magnetic inclusion resin particles.
  • the magnetic particles according to the present invention preferably include a shell layer.
  • the material of the shell layer contains an inorganic oxide or an organic polymer.
  • the shell layer is an inorganic oxide shell layer containing the inorganic oxide as the material of the shell layer, or an organic polymer shell layer containing the organic polymer as the material of the shell layer.
  • the shell layer is preferably arranged on the outer surface of the magnetic layer containing the second magnetic material.
  • the magnetic particles include a shell layer, it is possible to strongly prevent the elution of impurities from the magnetic inclusion resin particles and the like, the elution of the magnetic material, and the elution of impurities from the magnetic layer. Therefore, when the magnetic particles include the shell layer, they can be suitably used as a test agent.
  • the shell layer may or may not contain a magnetic material. It is more preferable that the shell layer does not contain a magnetic material.
  • the shell layer is more preferably a non-magnetic layer containing no magnetic material.
  • the material of the shell layer preferably contains the above-mentioned inorganic oxide, and more preferably an inorganic oxide shell layer containing the inorganic oxide.
  • the inorganic oxide means a compound having at least a metal element or a metalloid element and an oxygen atom.
  • the inorganic oxide is not particularly limited. Only one kind of the above-mentioned inorganic oxide may be used, or two or more kinds may be used.
  • the inorganic oxide is preferably an inorganic oxide having a silicon atom, a germanium atom, a titanium atom or a zirconium atom. Further, the inorganic oxide preferably has a functional group capable of reacting with the outer surface of the magnetic layer.
  • the method for reacting the magnetic surface with the non-magnetic layer is not particularly limited, and examples thereof include covalent bonds and coordination bonds.
  • the inorganic oxide examples include an alkoxysilane such as tetraethyl orthosilicate and a silane compound typified by a hydrolyzate thereof, and an alkoxygermanium such as germanium tetraethoxydo and a germanium compound typified by the hydrolyzate thereof.
  • an alkoxysilane such as tetraethyl orthosilicate and a silane compound typified by a hydrolyzate thereof
  • an alkoxygermanium such as germanium tetraethoxydo and a germanium compound typified by the hydrolyzate thereof.
  • Titanium compounds typified by alkoxytitanium such as titanium tetraethoxydo and its hydrolyzate
  • zirconium compounds such as zirconium tetrabutoxide and zirconium compounds typified by its hydrolyzate.
  • the inorganic oxide is preferably a compound having a small specific gravity, and among the above examples, the above silane compound is most preferable.
  • silane compound examples include tetraethyl orthosilicate; a vinyl group-containing silane compound such as vinyltrimethoxysilane, vinyltriethoxysilane, and 7-octenyltrimethoxysilane; 2- (3,4-epoxycyclohexyl) ethyltrimethoxy.
  • Silane 3-glycidoxypropylmethyldimethoxysilane, 3-glycidoxypropyltrimethoxysilane, 3-glycidoxypropylmethyldiethoxysilane, 3-glycidoxypropyltriethoxysilane, 8-glycidoxyoctyltri Epoxy group-containing silane compounds such as methoxysilane; styryl group-containing silane compounds such as p-styryltrimethoxysilane; 3-methacryloxypropylmethyldimethoxysilane, 3-methacryloxypropyltrimethoxysilane, 3-methacryloxypropylmethyldiethoxy Methacrylic group-containing silane compounds such as silane, 3-methacryloxypropyltriethoxysilane, 8-methacryloxyoctyltrimethoxysilane; acrylic group-containing silane compounds such as 3-acryloxypropyltrimethoxysilane; N-2- (aminoethyl)
  • an inorganic oxide shell layer having a silicon atom can be formed on the outer surface of the magnetic layer.
  • the content of the inorganic oxide in 100% by weight of the inorganic oxide shell layer is preferably 70% by weight or more, more preferably 80% by weight or more.
  • Organic polymer is not particularly limited, but is preferably a vinyl polymer.
  • vinyl-based monomer used as the material for the vinyl-based polymer examples include styrene monomers such as styrene, ⁇ -methylstyrene, chlorostyrene, and divinylbenzene; vinyl ether compounds such as methylvinyl ether, ethylvinyl ether, and propylvinyl ether; vinyl acetate, Acid vinyl ester compounds such as vinyl butyrate, vinyl laurate, vinyl stearate; halogen-containing monomers such as vinyl chloride and vinyl fluoride; (meth) acrylic acid; methyl (meth) acrylate, ethyl (meth) acrylate, propyl (Meta) acrylate, butyl (meth) acrylate, 2-ethylhexyl (meth) acrylate, lauryl (meth) acrylate, cetyl (meth) acrylate, stearyl (meth) acrylate, cyclohexyl (meth)
  • Halogen-containing (meth) acrylate compound such as trifluoromethyl (meth) acrylate and pentafluoroethyl (meth) acrylate; vinyl trimethoxysilane, vinyl triethoxysilane, dimethoxymethyl vinyl cysilane, dimethoxyethyl vinyl silane, diethoxy Methylvinylsilane, diethoxyethylvinylsilane, ethylmethyldivinylsilane, methylvinyldimethoxysilane, ethylvinyldimethoxysilane, methylvinyldiethoxysilane, ethylvinyldiethoxysilane, p-styryltrimethoxysilane, 3-methacryloxypropylmethyldimethoxysilane , 3-methacryloxypropyltrimethoxysilane, 3-methacryloxypropylmethyldiethoxysilane, 3-meth
  • the vinyl-based polymer may be a homopolymer obtained by polymerizing one kind of vinyl-based monomer, or may be a copolymer obtained by polymerizing two or more kinds of vinyl-based monomers.
  • a copolymerization monomer such as a vinyl monomer as a main raw material and, if necessary, a polymerization initiator, an emulsifier, a dispersant, a surfactant, an electrolyte, a cross-linking agent, and a molecular weight as auxiliary raw materials.
  • An organic polymer shell layer can be formed on the outer surface of the magnetic layer by adding a modifier or the like and polymerizing in a liquid.
  • the content of the organic polymer in 100% by weight of the organic polymer shell layer is preferably 70% by weight or more, more preferably 80% by weight or more.
  • the shell layer has a carboxyl group, a hydroxyl group, an epoxy group, an amino group, a tosyl group, a thiol group, a triethylammonium group, a dimethylamino group and a sulfonic acid before binding to the substance that specifically interacts with the target substance. It is preferable to have a functional group such as a group.
  • the substance that specifically interacts with the target substance can be well supported on the outer surface of the shell layer, and the substance can be supported on the surface of the magnetic particles. It can be arranged well.
  • the shell layer may have a linker portion on the outer surface.
  • the functional group which is a bonding point with the substance that specifically interacts with the target substance, is arranged at a position farther from the outermost surface of the shell layer.
  • the functional group and the substance can come into contact with each other at a position where there are less steric obstacles. Therefore, the substance is easily bonded, and the substance can be satisfactorily arranged on the surface of the magnetic particles.
  • the linker portion can be covalently bonded to a target substance such as a carboxyl group, a hydroxyl group, an epoxy group, an amino group, a tosyl group, or a thiol group at the terminal before binding to the above substance that specifically interacts with the target substance. It is preferable to have a functional group. By reacting the functional group with the functional group of the substance that specifically interacts with the target substance, the shell layer and the substance can be chemically bonded.
  • the epoxy group may be an epoxy group derived from a glycidyl group-containing monomer.
  • the hydroxyl group may be a hydroxyl group generated by ring-opening of the epoxy group.
  • the material of the linker portion is preferably an epoxy compound having a plurality of epoxy groups at the ends.
  • the epoxy compound having a plurality of epoxy groups at the end is preferably polyethylene glycol diglycidyl ether, polypropylene glycol diglycidyl ether, neopentyl glycol diglycidyl ether, or trimethylol propanepolyglycidyl ether.
  • the epoxy compound having a plurality of epoxy groups at the end is more preferably polyethylene glycol diglycidyl ether.
  • the shell layer and the substance that specifically interacts with the target substance are preferably bonded, and more preferably chemically bonded.
  • the surface area covered by the shell layer is preferably 95% or more, more preferably 99% or more, and most preferably 100%. is there.
  • the surface area is equal to or greater than the above lower limit, the content of the substance that specifically interacts with the target substance can be increased, and as a result, the measurement accuracy and measurement in the measurement of the target substance using magnetic particles The sensitivity can be increased.
  • the thickness of the shell layer is preferably 20 nm or more, more preferably 40 nm or more, preferably 500 nm or less, and more preferably 300 nm or less.
  • the thickness of the shell layer is not less than the above lower limit and not more than the above upper limit, the sedimentation of the magnetic particles can be effectively suppressed, the dispersibility can be further improved, and the particle diameter of the magnetic particles can be reduced. Therefore, the target substance can be effectively bound.
  • the thickness of the shell layer can be measured by observing the cross section of the magnetic particles, for example, using a transmission electron microscope (TEM). Regarding the thickness of the shell layer, it is preferable to calculate the average value of the thickness of any shell layer at five points as the thickness of the shell layer of one magnetic particle, and the average value of the thickness of the entire shell layer is one magnetism. It is more preferable to calculate as the thickness of the shell layer of particles.
  • the thickness of the shell layer is preferably obtained by calculating the average value of the thickness of the shell layer of each magnetic particle for 10 arbitrary magnetic particles.
  • Substances that specifically interact with the target substance On the surface of the magnetic particles according to the present invention, there is a substance that specifically interacts with the target substance.
  • the substance include sugar chains, peptide chains, proteins, antigens, nucleotide chains and the like.
  • the substance can be appropriately changed depending on the type of the target substance. Only one type of the above substance may be used, or two or more types may be used in combination.
  • Examples of the interaction between the substance and the target substance include an antigen-antibody reaction and an interaction between an enzyme and a substrate.
  • the interaction between the substance and the target substance may be a non-covalent interaction between the substance and the target substance, or may be a covalent bond between the substance and the target substance.
  • Examples of the non-covalent interaction include hydrophobic interaction, electrostatic interaction, van der Waals force, hydrogen bond, coordination bond, ionic bond and the like.
  • the substance that specifically interacts with the target substance is preferably a substance capable of non-covalent interaction with the target substance.
  • the combination of the above-mentioned substance that specifically interacts with the above-mentioned target substance and the target substance includes a combination of an antibody and an antigen, a combination of a sugar chain and a protein such as a lectin, and a combination of a protein such as an enzyme and an inhibitor.
  • Examples thereof include a combination of a peptide and a protein, a combination of a nucleotide chain and a nucleotide chain, and a combination of a nucleotide chain and a protein.
  • the substance that specifically interacts with the target substance is preferably a protein, more preferably avidin or streptavidin.
  • the substance that specifically interacts with the target substance is preferably an antigen or an antibody.
  • the above antibody may be a polyclonal antibody or a monoclonal antibody.
  • the antibody may be treated with a proteolytic enzyme such as papain and pepsin, or may be a Fab and F (ab') 2 fragment or the like.
  • the method of arranging the substance that specifically interacts with the target substance is not particularly limited.
  • the substance can be arranged on the surface of the magnetic particles by mixing the substance with the magnetic particles before the substance is supported.
  • the target substance is a substance that specifically interacts with the above-mentioned substance.
  • target substance examples include proteins, nucleic acids, hormones, cancer markers, respiratory-related markers, heart disease markers, drugs and the like.
  • lipid proteins such as high specific gravity lipoprotein (HDL), low specific gravity lipoprotein (LDL), and ultralow specific gravity lipoprotein; alkaline phosphatase, amylase, acidic phosphatase, ⁇ -glutamyltransferase ( ⁇ -GTP), lipase.
  • HDL high specific gravity lipoprotein
  • LDL low specific gravity lipoprotein
  • ⁇ -GTP ultralow specific gravity lipoprotein
  • CK Cleatin kinase
  • LDH lactic acid dehydrogenase
  • GAT glutamate oxaloacetate transaminase
  • GPT glutamate pyruvate transaminase
  • PK protein kinase
  • Antibodies such as streptidine O antibody, anti-human hepatitis B virus surface antigen antibody (HBs antigen), anti-human hepatitis C virus antibody, anti-rheumatic factor; albumin, hemoglobin, myoglobin, transferase, protein A, C reactive protein ( CRP) and
  • nucleic acid examples include DNA and RNA.
  • hormone examples include thyroid stimulating hormone (TSH), thyroid hormone (FT3, FT4, T3, T4), parathyroid hormone (PTH), and human chorionic gonadotropin (hCG) estradiol (E2).
  • TSH thyroid stimulating hormone
  • FT3, FT4, T3, T4 thyroid hormone
  • PTH parathyroid hormone
  • hCG human chorionic gonadotropin estradiol
  • cancer marker examples include ⁇ -fetoprotein (AFP), PIVKA-II, carcinoembryonic antigen (CEA), CA19-9, and prostate-specific antigen (PSA).
  • AFP ⁇ -fetoprotein
  • PIVKA-II carcinoembryonic antigen
  • CA19-9 carcinoembryonic antigen
  • PSA prostate-specific antigen
  • Examples of the respiratory-related marker include KL-6 and the like.
  • Examples of the above-mentioned heart disease marker include troponin T (TnT), human brain natriuretic peptide precursor N-terminal fragment (NT-proBNP), and the like.
  • TnT troponin T
  • NT-proBNP human brain natriuretic peptide precursor N-terminal fragment
  • Examples of the above-mentioned drugs include antiepileptic drugs, antibiotics, theophylline and the like.
  • the magnetic particles are used for radioimmunoassay (RIA), enzyme immunoassay (EIA), fluorescence immunoassay (FIA), chemiluminescence immunoassay (ECLIA), chemiluminescence immunoassay (CLIA and CLEIA), and absorbance measurement. , And in the measurement of surface plasmon resonance and the like, it is preferably used.
  • the magnetic particles are preferably used in the measurement of the sandwich method and the competitive method.
  • the magnetic particles are suitably used for measuring the concentration of the target substance in a sample.
  • the magnetic particles are preferably used as a test agent.
  • the magnetic particles are preferably magnetic particles that can be used as a test agent.
  • the concentration of the target substance can be measured using the magnetic particles as follows, for example.
  • a liquid containing the above magnetic particles (for example, a test agent described later) and a sample containing the target substance are mixed to obtain a mixed liquid.
  • the obtained mixed solution is heated or the like to obtain a reaction solution in which the substance that specifically interacts with the target substance in the magnetic particles and the target substance in the sample are bound to each other (first reaction step).
  • a magnetic force is applied to the reaction solution by a magnet or the like to collect magnetic particles (magnetic collection step).
  • a washing solution is added and mixed (washing step).
  • the magnetic collection step and the cleaning step may be repeated a plurality of times.
  • the target substance and the labeling substance are reacted to measure the concentration of the target substance (second reaction step).
  • labeling substance examples include alkaline phosphatase, ⁇ -galactosidase, peroxidase, microperoxidase, glucose oxidase, glucose-6-phosphate dehydrogenase, malic acid dehydrogenase, which are preferably used in enzyme immunoassay (EIA).
  • Enzymes such as luciferase, tyrosinase, acidic phosphatase; radioactive isotopes such as 99mTc, 131I, 125I, 14C, 3H, 32P preferably used in radioimmunosassay (RIA); suitable in fluorescence immunoassay (FIA), for example.
  • Fluorescent substances such as fluorescein, dancil, fluorescamine, coumarin, naphthylamine and derivatives thereof, and fluorescent substances such as green fluorescent protein (GFP) used in, for example, luciferin, isolminol, luminol, bis (2,4,6-trifluorophenyl).
  • GFP green fluorescent protein
  • Luminescent substances such as oxalate, such as phenol, naphthol, anthracene and derivatives thereof that have ultraviolet absorption, such as 4-amino-2,2,6,6-tetramethylpiperidine-1-oxyl, 3 -Amino-2,2,5,5-tetramethylpyrrolidin-1-oxyl, 2,6-di-t-butyl- ⁇ - (3,5-di-t-butyl-4-oxo-2,5-)
  • spin labeling agents such as compounds having an oxyl group such as cyclohexadien-1-iriden) -p-trioxyl.
  • the labeling substance is preferably an enzyme or a fluorescent substance, more preferably alkaline phosphatase, peroxidase or glucose oxidase, and even more preferably peroxidase.
  • Test drug The above-mentioned test agent contains the above-mentioned magnetic particles.
  • the above test agent preferably contains a buffer solution.
  • the buffer solution is preferably a buffer solution having a buffering ability at pH 5.0 or more and 9.0 or less.
  • the buffer solution include phosphate buffer solution, glycine buffer solution, veronal buffer solution, Tris buffer solution, borate buffer solution, citrate buffer solution, and Good buffer solution.
  • test agent may contain a sensitizer, a polymer compound such as a protein, an amino acid, and other components such as a surfactant.
  • the reaction between the target substance and the compound capable of binding to the target substance can be efficiently promoted, and the measurement accuracy can be improved.
  • the sensitizer include alkylated polysaccharide compounds such as methyl cellulose and ethyl cellulose, pullulan, polyvinylpyrrolidone and the like.
  • protein examples include albumin (bovine serum albumin, egg albumin, etc.), casein, gelatin, and the like.
  • the content of the magnetic particles in 100% by weight of the test agent is preferably 0.5% by weight or more, more preferably 2% by weight or more, preferably 10% by weight or less, and more preferably 5% by weight or less.
  • the measurement accuracy of the target substance can be further improved.
  • Example 1 Preparation of magnetic encapsulating resin particles: Polystyrene particles having an average particle diameter of 0.69 ⁇ m were prepared as seed particles. A mixed solution was prepared by mixing 3.9 parts by weight of the polystyrene particles, 500 parts by weight of ion-exchanged water, and 120 parts by weight of a 5.0% by weight polyvinyl alcohol aqueous solution. After the above mixed solution was dispersed by ultrasonic waves, it was placed in a separable flask and stirred uniformly.
  • the emulsion was added to the mixed solution in the separable flask in several portions and stirred for 12 hours to allow the seed particles to absorb the monomer to obtain a suspension containing the seed particles in which the monomer was swollen. ..
  • Magnetic layer (magnetic layer containing a second magnetic material): 1 part by weight of magnetic inclusion resin particles and 4 parts by weight of magnetic fluid EMG707 (aqueous dispersion of triiron tetroxide nanoparticles with an average particle diameter of about 10 nm, manufactured by Fellow Tech) (however, the content of magnetic material is about about 17% by weight) was stirred at 250 rpm for 10 minutes. The obtained particle dispersion is filtered and washed with water to form a magnetic layer containing triiron tetroxide as a second magnetic substance on the outer surface of the magnetic inclusion resin particles to form a magnetic layer. The magnetic encapsulating resin particles having were obtained.
  • EMG707 aqueous dispersion of triiron tetroxide nanoparticles with an average particle diameter of about 10 nm, manufactured by Fellow Tech
  • the obtained particle dispersion is filtered and washed with water to form a magnetic layer containing triiron tetroxide as a second magnetic substance on the outer surface of the magnetic inclusion
  • Arrangement of substances that specifically interact with the target substance 0.5 mL of an aqueous dispersion of magnetic encapsulating resin particles having a magnetic layer was added to a test tube, and the mixture was washed 3 times with a PBS solution. After removing the dispersion medium, 0.5 mL of a PBS solution (0.75 mg / mL) containing a sialylated sugar chain antigen KL-6 (hereinafter abbreviated as KL-6) antibody was added, and the mixture was stirred at 25 ° C. overnight. Then, 1.5 mL of 1.0 wt% BSA solution was added, and the mixture was stirred at 25 ° C. for 4 hours.
  • KL-6 sialylated sugar chain antigen KL-6
  • the average particle size, BET specific surface area and average pore size of the obtained resin particles, and the contents and contents (1) of the first magnetic substance and the second magnetic substance in the obtained magnetic particles are shown. Table 1 shows the ratio to the amount (2), the absolute value of the difference between the content (1) and the content (2), and the average particle size. These were obtained by the method described in the evaluation items described later.
  • Example 2 Magnetic particles were prepared in the same manner as in Example 1 except that resin particles were prepared using 80 parts by weight of toluene instead of 150 parts by weight of toluene and the composition of the magnetic particles was changed as shown in Table 1. did.
  • Example 3 Magnetic particles were prepared in the same manner as in Example 1 except that resin particles were prepared using 30 parts by weight of toluene instead of 150 parts by weight of toluene and the composition of the magnetic particles was changed as shown in Table 1. did.
  • Example 4 In the same manner as in Example 1, magnetic inclusion resin particles having a magnetic layer were obtained.
  • Shell layer formation To 1.0 part by weight of the magnetic encapsulating resin particles having the obtained magnetic layer, 400 parts by weight of ethanol and 20 parts by weight of a 28% aqueous ammonia solution (manufactured by Nacalai Tesque) were added. Then, 5.0 parts by weight of tetraethyl orthosilicate and 15 parts by weight of 8-glycidoxyoctyltrimethoxysilane were added, and the mixture was stirred for 1 hour. The obtained dispersion was filtered and then washed with water. In this way, magnetic inclusion particles having an inorganic oxide shell layer having a functional group on the surface were obtained.
  • Example 5 Magnetic particles were obtained in the same manner as in Example 4 except that the configurations of the resin particles and the magnetic particles were changed as shown in Table 1.
  • Comparative Example 1 No toluene was used, 300 parts by weight of divinylbenzene was used instead of 150 parts by weight of divinylbenzene, and 4.0 parts by weight of benzoyl peroxide was used instead of 2.0 parts by weight of benzoyl peroxide.
  • Resin particles were obtained in the same manner as in Example 1 except for the above. Further, in Comparative Example 1, the first magnetic material was not used. Using the obtained resin particles, a magnetic layer was formed on the outer surface of the resin particles by a second magnetic material in the same manner as in Example 1.
  • Shell layer formation Using the obtained resin particles having a magnetic layer, a shell layer was formed on the outer surface of the magnetic layer in the same manner as in Example 4.
  • Comparative Example 2 Resin particles were obtained in the same manner as in Comparative Example 1. Next, acetone was added to an oil-based magnetic fluid (organic solvent dispersion of triiron tetroxide nanoparticles with an average particle diameter of about 10 nm, "EXP series” manufactured by Fellow Tech) to precipitate and precipitate the particles, and then dried. By doing so, ferrite-based magnetic fine particles (average primary particle diameter: 10 nm) having a hydrophobically treated surface were obtained.
  • oil-based magnetic fluid organic solvent dispersion of triiron tetroxide nanoparticles with an average particle diameter of about 10 nm, "EXP series” manufactured by Fellow Tech
  • Shell layer formation 10 parts by weight of the obtained particles having a magnetic layer and 300 parts by weight of a 0.5% aqueous solution of a nonionic emulsifier (“Emulgen 150” manufactured by Kao Corporation) as a dispersant were put into a 1 L separable flask and stirred.
  • a nonionic emulsifier (“Emulgen 150” manufactured by Kao Corporation) as a dispersant
  • 24 parts by weight of cyclohexyl methacrylate and 60 parts by weight of 2-methacryloyloxyethyl succinic acid were used as monomers, and di (3,5,5-trimethylhexanoyl) peroxide (NOF Corporation "Perloyl”) was used as an initiator.
  • 355 1.0 part by weight was added, and the mixture was stirred at 80 ° C.
  • the organic polymer is a copolymer of cyclohexyl methacrylate and 2-methacryloyloxyethyl succinic acid.
  • Comparative Example 3 Magnetic particles were obtained in the same manner as in Comparative Example 1 except that the configurations of the resin particles and the magnetic particles were changed as shown in Table 1. The magnetic particles obtained in Comparative Example 3 do not have the first magnetic material.
  • Comparative Example 4 Magnetic particles were produced in the same manner as in Example 1 except that the configurations of the resin particles and the magnetic particles were changed as shown in Table 1. The magnetic particles obtained in Comparative Example 4 do not have a magnetic layer.
  • Comparative Example 5 Magnetic particles were obtained in the same manner as in Comparative Example 4, except that the configurations of the resin particles and the magnetic particles were changed as shown in Table 1.
  • a shell layer organic polymer shell layer
  • PGMA polyglycidyl methacrylate
  • the measurement was performed to obtain the distribution results of the contents of the first magnetic substance and the second magnetic substance inside the magnetic particles. From the obtained results, the contents of the first magnetic substance and the second magnetic substance were calculated. The contents of the first magnetic material and the second magnetic material were calculated by arithmetically averaging the contents of the first magnetic material and the second magnetic material of 20 arbitrarily selected magnetic particles.
  • the content of the first magnetic substance was defined as the content (1) in 100% by volume of the region (R1) having a thickness of 1/3 from the outer surface to the inside of the magnetic particles.
  • the content of the first magnetic substance was defined as the content (2) in 100% by volume of the region (R2) having a thickness of 2/3 from the center of the magnetic particles to the outside.
  • An embedded resin body for magnetic particle inspection was prepared by adding and dispersing it to "Technobit 4000" manufactured by Kulzer so that the content of magnetic particles was 30% by weight.
  • a cross section of the magnetic particles was cut out using an ion milling device (“IM4000” manufactured by Hitachi High-Technologies Corporation) so as to pass near the center of the magnetic particles dispersed in the embedded resin body for inspection.
  • IM4000 manufactured by Hitachi High-Technologies Corporation
  • JEM-2010FEF electric field radiation type transmission electron microscope
  • EDS energy dispersive X-ray analyzer
  • the content was measured, and the distribution result of the content of the first magnetic substance in the thickness direction of the magnetic inclusion resin particles was obtained. From the obtained results, the above-mentioned content (1) and the above-mentioned content (2) were calculated.
  • the content (1) and the content (2) were calculated by arithmetically averaging the content (1) and the content (2) of 20 arbitrarily selected magnetic particles.
  • Magnetic Collection Rate As a sample solution, a solution in which magnetic particles whose absorbance at a wavelength of 550 nm was adjusted to 0.9 to 1.1 was dispersed in water was prepared. Sample solution 1. In a quartz cell installed in a spectrophotometer (“U-3900H” manufactured by Hitachi, Ltd.) with a magnet (2800G, W10mm ⁇ D10mm ⁇ H1mm) applied via a spacer (W10mm ⁇ D10mm ⁇ H4mm). 3 mL was added, and the absorbance at a wavelength of 550 nm was measured 5 to 125 seconds after the sample solution was added. The absorbance attenuation rate in 120 seconds was calculated by the following formula and used as the magnetic collection rate.
  • Magnetic collection rate (%) [ ⁇ (absorbance after 5 seconds)-(absorbance after 125 seconds) ⁇ / (absorbance after 5 seconds)] x 100
  • Dispersion rate As a sample solution, a solution in which magnetic particles whose absorbance at a wavelength of 550 nm was adjusted to 0.9 to 1.1 was dispersed in water was prepared. 1.3 mL of the sample solution was put into a quartz cell installed in a spectrophotometer (“U-3900H” manufactured by Hitachi, Ltd.), and the absorbance at a wavelength of 550 nm was measured. Next, a magnet (28000 G, W40 mm ⁇ D40 mm ⁇ H10 mm) was used to collect magnetism until the absorbance of the supernatant became zero. Then, the magnetic particles were dispersed by vortex at 2000 rpm for 5 seconds, and the absorbance at a wavelength of 550 nm was measured. From the absorbance before magnetic collection and the absorbance after magnetic collection and dispersion, the rate of change in absorbance was calculated by the following formula and used as the dispersion rate.
  • Dispersion rate (%) ⁇ (absorbance after magnetic collection and dispersion) / (absorbance before magnetic collection) ⁇ x 100
  • Dispersion rate is 95% or more ⁇ : Dispersion rate is 90% or more and less than 95% ⁇ : Dispersion rate is 85% or more and less than 90% ⁇ : Dispersion rate is less than 85%
  • Magnetic collection rate (after leaving for 1 month) The obtained magnetic particles were left at 25 ° C. for 1 month. After being left to stand, the magnetic collection coefficient was determined by the same method as in (5) magnetic collection rate.
  • ruthenium complex-labeled anti-KL-6 antibody (secondary antibody): To a polypropylene tube, 0.5 mL of a PBS-1 solution of anti-KL-6 antibody (anti-KL-6 antibody concentration 2.0 mg / mL) was added, and then 13 ⁇ L of Ru-NHS (10 mg / mL) was added. After vibrating and stirring at 25 ° C., purification was performed using a Sephadex G25 column to obtain a ruthenium complex-labeled anti-KL-6 antibody.
  • the amount of luminescence was measured as follows using an ECLIA automatic analyzer (“Picormi III” manufactured by Sekisui Medical Co., Ltd.) based on the electrochemical luminescence immunoassay method.
  • the reaction solution buffer solution containing normal rabbit serum
  • 20 ⁇ L of a solution containing 5000 U / mL KL-6 (antigen-containing solution) was added, and then 25 ⁇ L of magnetic particles were added.
  • 350 ⁇ L of Picormi BF washing solution (10 mM Tris buffer) was added, and the magnetic particles were washed three times while being trapped with a magnet.

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Abstract

Provided are magnetic particles improved in magnetism and dispersibility, and capable of maintaining a high level of magnetism.　Magnetic particles according to the present invention are used to be specifically interact with a target substance. The magnetic particles each comprise: a magnetic material-containing resin particle containing a first magnetic material; a magnetic layer disposed on the outer surface of the magnetic material-containing resin particle and containing a second magnetic material; and a substance that is supported on the outer surface side of the magnetic layer and that specifically interacts with the target substance.

Description

磁性粒子及び検査薬Magnetic particles and test agents
　本発明は、磁性体を含む磁性粒子に関する。また、本発明は、上記磁性粒子を用いた検査薬に関する。 The present invention relates to magnetic particles containing a magnetic material. The present invention also relates to a test agent using the above magnetic particles.
　医薬品の研究開発及び臨床検査等の分野において、検体中の目的物質の濃度等を測定するために磁性粒子が用いられている。例えば、化学発光免疫測定法（ＣＬＩＡ法）等のイムノアッセイでは、表面に抗体又は抗原等を有する磁性粒子が広く用いられている。この磁性粒子は、一般に、目的物質である抗原又は抗体等と結合した後、磁石等によって集磁される。 Magnetic particles are used to measure the concentration of target substances in samples in fields such as pharmaceutical research and development and clinical tests. For example, in immunoassays such as chemiluminescence immunoassay (CLIA method), magnetic particles having an antibody or antigen on the surface are widely used. Generally, these magnetic particles are magnetized by a magnet or the like after being bound to an antigen or an antibody which is a target substance.
　従来、磁性粒子として、樹脂粒子の内部に磁性体を有する磁性粒子（例えば、特許文献１，２参照）、及び樹脂粒子の外表面上に形成された磁性層を有する磁性粒子（例えば、特許文献３参照）が用いられている。 Conventionally, as magnetic particles, magnetic particles having a magnetic substance inside the resin particles (see, for example, Patent Documents 1 and 2) and magnetic particles having a magnetic layer formed on the outer surface of the resin particles (for example, Patent Documents). 3) is used.
ＷＯ２００４／０５６８９５Ａ１WO2004 / 056895A1 特開昭５９－５００６９１号公報JP-A-59-500691 特開平０７－０６３７６１号公報Japanese Unexamined Patent Publication No. 07-063761
　樹脂粒子の内部に磁性体を有する従来の磁性粒子や、樹脂粒子の外表面上に磁性層を有する従来の磁性粒子では、該磁性粒子中の磁性体の含有量を十分に多くすることが困難である。そのため、従来の磁性粒子では、集磁性が低いことがある。 With conventional magnetic particles having a magnetic substance inside the resin particles and conventional magnetic particles having a magnetic layer on the outer surface of the resin particles, it is difficult to sufficiently increase the content of the magnetic substance in the magnetic particles. Is. Therefore, the conventional magnetic particles may have low magnetic collection.
　また、従来の磁性粒子では、磁性体が酸化して、経時的に集磁性が低下することがある。 Also, with conventional magnetic particles, the magnetic material may oxidize and the magnetic collection may decrease over time.
　また、従来の磁性粒子では、分散性が低いことがある。 In addition, conventional magnetic particles may have low dispersibility.
　集磁性の低い磁性粒子を用いた場合には、磁石で集磁しきれない磁性粒子が多くなり、抗体や抗原等の目的物質の濃度等を測定する際に、測定精度及び測定感度が低下することがある。また、分散性の低い磁性粒子を用いた場合には、磁気分離後に磁性粒子が十分に再分散せず、測定精度、測定感度及び測定再現性が低下することがある。 When magnetic particles with low magnetic collection are used, the number of magnetic particles that cannot be completely collected by the magnet increases, and the measurement accuracy and measurement sensitivity deteriorate when measuring the concentration of the target substance such as an antibody or antigen. Sometimes. Further, when magnetic particles having low dispersibility are used, the magnetic particles may not be sufficiently redispersed after magnetic separation, and the measurement accuracy, measurement sensitivity and measurement reproducibility may decrease.
　本発明の目的は、集磁性及び分散性を高めることができ、かつ集磁性を高く維持することができる磁性粒子を提供することである。また、本発明の限定的な目的は、長期間集磁性を高く維持することができる磁性粒子を提供することである。また、本発明の目的は、上記磁性粒子を用いた検査薬を提供することである。 An object of the present invention is to provide magnetic particles capable of enhancing magnetic collection and dispersibility, and maintaining high magnetic collection. Further, a limited object of the present invention is to provide magnetic particles capable of maintaining high magnetic collection for a long period of time. Another object of the present invention is to provide a test agent using the above magnetic particles.
　本発明の広い局面によれば、目的物質と特異的に相互作用させるために用いられる磁性粒子であって、内部に第１の磁性体を含む磁性体内包樹脂粒子と、前記磁性体内包樹脂粒子の外表面上に配置され、かつ第２の磁性体を含む磁性層と、前記磁性層の外表面側に担持されており、かつ前記目的物質と特異的に相互作用する物質とを備える、磁性粒子が提供される。 According to a broad aspect of the present invention, magnetic particles used for specifically interacting with a target substance, the magnetic inclusion resin particles containing the first magnetic substance inside, and the magnetic inclusion resin particles. A magnetism having a magnetic layer arranged on the outer surface of the magnetic layer and containing a second magnetic substance, and a substance supported on the outer surface side of the magnetic layer and specifically interacting with the target substance. Particles are provided.
　本発明に係る磁性粒子のある特定の局面では、前記第１の磁性体が、金属又は金属酸化物であり、前記第２の磁性体が、金属又は金属酸化物である。 In a specific aspect of the magnetic particles according to the present invention, the first magnetic material is a metal or a metal oxide, and the second magnetic material is a metal or a metal oxide.
　本発明に係る磁性粒子のある特定の局面では、前記磁性層の外表面上に配置されたシェル層を更に備え、前記シェル層の材料が、無機酸化物又は有機ポリマーを含み、前記シェル層と、前記物質とが結合している。 In certain aspects of the magnetic particles according to the present invention, a shell layer is further provided on the outer surface of the magnetic layer, the material of the shell layer containing an inorganic oxide or an organic polymer, and the shell layer. , The substance is bound.
　本発明に係る磁性粒子のある特定の局面では、前記シェル層の材料が、前記無機酸化物を含み、前記無機酸化物が、ケイ素原子、ゲルマニウム原子、チタン原子又はジルコニウム原子を有する無機酸化物である。 In certain aspects of the magnetic particles according to the present invention, the material of the shell layer is an inorganic oxide containing the inorganic oxide, wherein the inorganic oxide has a silicon atom, a germanium atom, a titanium atom or a zirconium atom. is there.
　本発明に係る磁性粒子のある特定の局面では、前記物質が、抗原又は抗体である。 In certain aspects of the magnetic particles according to the present invention, the substance is an antigen or antibody.
　本発明に係る磁性粒子のある特定の局面では、前記物質が、アビジン又はストレプトアビジンである。 In certain aspects of the magnetic particles according to the present invention, the substance is avidin or streptavidin.
　本発明に係る磁性粒子のある特定の局面では、磁性粒子１００体積％中、前記第１の磁性体と前記第２の磁性体と合計の含有量が、１０重量％以上９５重量％以下である。 In a specific aspect of the magnetic particles according to the present invention, the total content of the first magnetic substance and the second magnetic substance in 100% by volume of the magnetic particles is 10% by weight or more and 95% by weight or less. ..
　本発明に係る磁性粒子のある特定の局面では、前記第１の磁性体と前記第２の磁性体の含有量の合計１００重量％中、前記第１の磁性体の含有量が、１０重量％以上９０重量％以下である。 In a specific aspect of the magnetic particles according to the present invention, the content of the first magnetic substance is 10% by weight in a total of 100% by weight of the contents of the first magnetic substance and the second magnetic substance. More than 90% by weight or less.
　本発明に係る磁性粒子のある特定の局面では、前記磁性体内包樹脂粒子の外表面から中心に向かって厚み１／３までの領域の１００体積％中における前記第１の磁性体の含有量の、前記磁性体内包樹脂粒子の中心から外表面に向かって厚み２／３までの領域の１００体積％中における前記第１の磁性体の含有量に対する比が、０．８以上４．０以下である。 In a specific aspect of the magnetic particles according to the present invention, the content of the first magnetic substance in 100% by volume of the region from the outer surface to the center of the magnetic inclusion resin particles up to 1/3 of the thickness. The ratio of the content of the first magnetic substance to the content of the first magnetic substance in 100% by volume of the region from the center of the magnetic inclusion resin particles to the outer surface to the thickness of 2/3 is 0.8 or more and 4.0 or less. is there.
　本発明に係る磁性粒子のある特定の局面では、前記磁性粒子は、検査薬として用いられる。 In a specific aspect of the magnetic particles according to the present invention, the magnetic particles are used as a test agent.
　本発明の広い局面によれば、上述した磁性粒子を含む、検査薬が提供される。 According to a broad aspect of the present invention, a test agent containing the above-mentioned magnetic particles is provided.
　本発明に係る磁性粒子は、目的物質と特異的に相互作用させるために用いられる。本発明に係る磁性粒子は、内部に第１の磁性体を含む磁性体内包樹脂粒子と、上記磁性体内包樹脂粒子の外表面上に配置され、かつ第２の磁性体を含む磁性層と、上記磁性層の外表面側に担持されており、かつ上記目的物質と特異的に相互作用する物質とを備える。本発明に係る磁性粒子では、上記の構成が備えられているので、集磁性及び分散性を高めることができ、かつ集磁性を高く維持することができる。 The magnetic particles according to the present invention are used to specifically interact with a target substance. The magnetic particles according to the present invention include magnetic encapsulating resin particles containing a first magnetic substance inside, and a magnetic layer arranged on the outer surface of the magnetic encapsulating resin particles and containing a second magnetic substance. It includes a substance that is supported on the outer surface side of the magnetic layer and that specifically interacts with the target substance. Since the magnetic particles according to the present invention have the above-mentioned structure, the magnetic collection and dispersibility can be enhanced, and the magnetic collection can be maintained high.
図１は、本発明の第１の実施形態に係る磁性粒子を模式的に示す断面図である。FIG. 1 is a cross-sectional view schematically showing magnetic particles according to the first embodiment of the present invention. 図２は、本発明の第２の実施形態に係る磁性粒子を模式的に示す断面図である。FIG. 2 is a cross-sectional view schematically showing magnetic particles according to a second embodiment of the present invention. 図３は、本発明の第３の実施形態に係る磁性粒子を模式的に示す断面図である。FIG. 3 is a cross-sectional view schematically showing the magnetic particles according to the third embodiment of the present invention. 図４は、本発明の第４の実施形態に係る磁性粒子を模式的に示す断面図である。FIG. 4 is a cross-sectional view schematically showing the magnetic particles according to the fourth embodiment of the present invention. 図５は、磁性体内包樹脂粒子において、第１の磁性体の含有量を求める各領域を説明するための模式図である。FIG. 5 is a schematic diagram for explaining each region for obtaining the content of the first magnetic substance in the magnetic inclusion resin particles.
　以下、本発明の詳細を説明する。 The details of the present invention will be described below.
　（磁性粒子）
　本発明に係る磁性粒子は、目的物質と特異的に相互作用させるために用いられる。本発明に係る磁性粒子は、目的物質と特異的に相互作用させることができる磁性粒子である。本発明に係る磁性粒子は、内部に第１の磁性体を含む磁性体内包樹脂粒子と、上記磁性体内包樹脂粒子の外表面上に配置され、かつ第２の磁性体を含む磁性層と、上記磁性層の外表面側に担持されており、かつ上記目的物質と特異的に相互作用する物質とを備える。 (Magnetic particles)
The magnetic particles according to the present invention are used to specifically interact with a target substance. The magnetic particles according to the present invention are magnetic particles that can specifically interact with a target substance. The magnetic particles according to the present invention include magnetic encapsulating resin particles containing a first magnetic substance inside, and a magnetic layer arranged on the outer surface of the magnetic encapsulating resin particles and containing a second magnetic substance. It includes a substance that is supported on the outer surface side of the magnetic layer and that specifically interacts with the target substance.
　本発明に係る磁性粒子では、上記の構成が備えられているので、集磁性及び分散性を高めることができ、かつ集磁性を高く維持することができる。本発明では、長期間集磁性を高く維持することができる。本発明に係る磁性粒子では、該磁性粒子を用いた目的物質の測定において、磁性粒子の単位重量あたりの目的物質との結合量を多くすることができ、従って、測定感度を高めることができる。 Since the magnetic particles according to the present invention have the above-mentioned structure, the magnetic collection and dispersibility can be enhanced, and the magnetic collection can be maintained high. In the present invention, the magnetism collection can be maintained high for a long period of time. In the magnetic particles according to the present invention, in the measurement of the target substance using the magnetic particles, the amount of binding to the target substance per unit weight of the magnetic particles can be increased, and therefore the measurement sensitivity can be increased.
　従来の磁性粒子では、集磁性又は分散性が低いことがある。樹脂粒子の内部に磁性体を有する従来の磁性粒子では、樹脂粒子内部の空間が限られているため、磁性体の含有量を十分に多くすることが困難である。そのため、従来の磁性粒子では、集磁性を高めることが困難である。また、樹脂粒子の外表面上に磁性層を有する従来の磁性粒子において、磁性層に含まれる磁性体の含有量を多くした場合には、集磁性をある程度高めることができるものの、磁性粒子の粒子径が大きくなり、磁性粒子の単位面積あたりの目的物質の結合量が低下するため好ましくない。また、樹脂粒子の外表面上の磁性層を厚くすると、磁性粒子間での反発力が低下し、分散性が低下することがある。さらに、樹脂粒子の内部に磁性体を有する従来の磁性粒子及び樹脂粒子の外表面上に磁性層を有する従来の磁性粒子の双方において、粒子径が小さくなるほど磁性体の含有量が減少するため、集磁性が低下し、その結果、測定感度が低下することがある。 Conventional magnetic particles may have low magnetic collection or dispersibility. In the conventional magnetic particles having a magnetic substance inside the resin particles, it is difficult to sufficiently increase the content of the magnetic substance because the space inside the resin particles is limited. Therefore, it is difficult to improve the magnetic collection with the conventional magnetic particles. Further, in the conventional magnetic particles having a magnetic layer on the outer surface of the resin particles, when the content of the magnetic substance contained in the magnetic layer is increased, the magnetic collection can be increased to some extent, but the particles of the magnetic particles. This is not preferable because the diameter becomes large and the amount of the target substance bonded per unit area of the magnetic particles decreases. Further, when the magnetic layer on the outer surface of the resin particles is thickened, the repulsive force between the magnetic particles is lowered, and the dispersibility may be lowered. Further, in both the conventional magnetic particles having a magnetic substance inside the resin particles and the conventional magnetic particles having a magnetic layer on the outer surface of the resin particles, the smaller the particle diameter, the lower the content of the magnetic substance. The magnetic collection may decrease, and as a result, the measurement sensitivity may decrease.
　さらに、樹脂粒子の内部に磁性体を有する従来の磁性粒子や、樹脂粒子の外表面上に磁性層を有する従来の磁性粒子では、磁性体が酸化して、経時的に集磁性が低下することがある。 Further, in the conventional magnetic particles having a magnetic substance inside the resin particles and the conventional magnetic particles having a magnetic layer on the outer surface of the resin particles, the magnetic substance is oxidized and the magnetic collection property is lowered with time. There is.
　これに対して、本発明に係る磁性粒子では、磁性粒子の粒子径を小さく維持したままで、磁性体の含有量を多くすることができ、集磁性を高めることができる。その結果、磁性粒子の単位重量あたりの表面積を大きくしつつ集磁性を高くすることができるため、測定感度を高めることができる。また、本発明に係る磁性粒子では、分散性を高めることができる。さらに、本発明に係る磁性粒子では、磁性体がある程度劣化したとしても、集磁性を高く維持することができる。例えば、本発明に係る磁性粒子では、上記第２の磁性体を含む特定の磁性層がある程度劣化したとしても、上記第１の磁性体を含むので、集磁性を高く維持することができる。本発明に係る磁性粒子では、該磁性粒子を用いた目的物質の測定において、測定精度、測定感度及び測定再現性を高めることができる。 On the other hand, in the magnetic particles according to the present invention, the content of the magnetic substance can be increased and the magnetic collection can be enhanced while keeping the particle diameter of the magnetic particles small. As a result, it is possible to increase the magnetic collection while increasing the surface area per unit weight of the magnetic particles, so that the measurement sensitivity can be increased. Further, the magnetic particles according to the present invention can enhance the dispersibility. Further, in the magnetic particles according to the present invention, high magnetic collection can be maintained even if the magnetic material deteriorates to some extent. For example, in the magnetic particles according to the present invention, even if the specific magnetic layer containing the second magnetic material is deteriorated to some extent, the magnetic particles contain the first magnetic material, so that the magnetic collection property can be maintained high. With the magnetic particles according to the present invention, the measurement accuracy, measurement sensitivity and measurement reproducibility can be improved in the measurement of the target substance using the magnetic particles.
　上記磁性粒子の平均粒子径は、好ましくは０．５μｍ以上、より好ましくは１．０μｍ以上であり、好ましくは１０μｍ以下、より好ましくは７μｍ以下、更に好ましくは５μｍ以下、特に好ましくは４μｍ以下、最も好ましくは３．５μｍ以下である。上記磁性粒子の平均粒子径が上記下限以上であると、集磁性をより一層高めることができる。上記磁性粒子の平均粒子径が上記上限以下であると、単位重量あたりの上記目的物質と特異的に相互作用する物質の含有量を多くすることができ、目的物質の結合量を多くすることができる。 The average particle size of the magnetic particles is preferably 0.5 μm or more, more preferably 1.0 μm or more, preferably 10 μm or less, more preferably 7 μm or less, still more preferably 5 μm or less, particularly preferably 4 μm or less, most preferably. It is preferably 3.5 μm or less. When the average particle size of the magnetic particles is at least the above lower limit, the magnetism collection can be further enhanced. When the average particle size of the magnetic particles is not more than the upper limit, the content of the substance that specifically interacts with the target substance per unit weight can be increased, and the binding amount of the target substance can be increased. it can.
　上記磁性粒子の平均粒子径は、数平均粒子径である。上記磁性粒子の粒子径は、例えば、任意の磁性粒子５０個を電子顕微鏡又は光学顕微鏡にて観察し、各磁性粒子の粒子径の平均値を算出することにより求められる。上記磁性粒子を乾燥させた試料を作製し、得られた試料を電子顕微鏡又は光学顕微鏡にて観察することが好ましい。 The average particle size of the magnetic particles is a number average particle size. The particle size of the magnetic particles is determined, for example, by observing 50 arbitrary magnetic particles with an electron microscope or an optical microscope and calculating the average value of the particle sizes of each magnetic particle. It is preferable to prepare a sample obtained by drying the magnetic particles and observe the obtained sample with an electron microscope or an optical microscope.
　上記磁性粒子の平均粒子径の変動係数（ＣＶ値）は、好ましくは１０％以下、より好ましくは５％以下である。上記磁性粒子の平均粒子径の変動係数が上記上限以下であると、磁性粒子を用いた目的物質の測定において、測定精度をより一層高めることができる。 The coefficient of variation (CV value) of the average particle size of the magnetic particles is preferably 10% or less, more preferably 5% or less. When the coefficient of variation of the average particle size of the magnetic particles is not more than the above upper limit, the measurement accuracy can be further improved in the measurement of the target substance using the magnetic particles.
　上記変動係数（ＣＶ値）は、以下のようにして測定できる。 The coefficient of variation (CV value) can be measured as follows.
　ＣＶ値（％）＝（ρ／Ｄｎ）×１００
　ρ：磁性粒子の粒子径の標準偏差
　Ｄｎ：磁性粒子の粒子径の平均値 CV value (%) = (ρ / Dn) × 100
ρ: Standard deviation of the particle size of the magnetic particles Dn: Average value of the particle size of the magnetic particles
　以下、図面を参照しつつ、本発明を具体的に説明する。 Hereinafter, the present invention will be specifically described with reference to the drawings.
　図１は、本発明の第１の実施形態に係る磁性粒子を模式的に示す断面図である。 FIG. 1 is a cross-sectional view schematically showing magnetic particles according to the first embodiment of the present invention.
　図１に示す磁性粒子１は、目的物質と特異的に相互作用させるために用いられる。磁性粒子１は、磁性体内包樹脂粒子２と、磁性層３と、シェル層４と、目的物質と特異的に相互作用する物質５とを備える。物質５は、例えば、アビジン、ストレプトアビジン、抗原及び抗体等の生理活性物質等である。 The magnetic particle 1 shown in FIG. 1 is used to specifically interact with the target substance. The magnetic particles 1 include magnetic inclusion resin particles 2, a magnetic layer 3, a shell layer 4, and a substance 5 that specifically interacts with a target substance. The substance 5 is, for example, a physiologically active substance such as avidin, streptavidin, an antigen and an antibody.
　磁性体内包樹脂粒子２は、樹脂粒子２１と、第１の磁性体２２とを有する。磁性体内包樹脂粒子２は、内部に第１の磁性体２２を含む。磁性体内包樹脂粒子２において、樹脂粒子２１は、内部に第１の磁性体２２を含む。第１の磁性体２２が、磁性体内包樹脂粒子２の内部に分散している。磁性体内包樹脂粒子２において、第１の磁性体２２が、樹脂粒子２１の内部に分散している。 The magnetic inclusion resin particles 2 have resin particles 21 and a first magnetic material 22. The magnetic inclusion resin particles 2 contain a first magnetic substance 22 inside. In the magnetic inclusion resin particles 2, the resin particles 21 contain a first magnetic material 22 inside. The first magnetic material 22 is dispersed inside the magnetic inclusion resin particles 2. In the magnetic inclusion resin particles 2, the first magnetic material 22 is dispersed inside the resin particles 21.
　第１の磁性体２２が、磁性体内包樹脂粒子２の内部に均一に分布している。上記第１の磁性体は、磁性体内包樹脂粒子の内部に均一に分布していなくてもよい。例えば、磁性体内包樹脂粒子の中心から外表面にかけて、第１の磁性体の含有量が多くなるように第１の磁性体が配置されていてもよい。 The first magnetic material 22 is uniformly distributed inside the magnetic inclusion resin particles 2. The first magnetic material does not have to be uniformly distributed inside the magnetic inclusion resin particles. For example, the first magnetic material may be arranged from the center of the magnetic inclusion resin particles to the outer surface so that the content of the first magnetic material increases.
　磁性層３は、磁性体内包樹脂粒子２の外表面上に配置されている。磁性層３は、磁性体内包樹脂粒子２における樹脂粒子２１の外表面上に配置されている。磁性層３は、第２の磁性体を含む。シェル層４は、磁性層３の外表面上に配置されている。磁性層３の外表面側に、物質５が担持されている。シェル層４の外表面上に、物質５が担持されている。シェル層４と、物質５とが結合している。物質５は、磁性粒子１の表面に存在する。 The magnetic layer 3 is arranged on the outer surface of the magnetic inclusion resin particles 2. The magnetic layer 3 is arranged on the outer surface of the resin particles 21 in the magnetic inclusion resin particles 2. The magnetic layer 3 contains a second magnetic material. The shell layer 4 is arranged on the outer surface of the magnetic layer 3. The substance 5 is supported on the outer surface side of the magnetic layer 3. The substance 5 is supported on the outer surface of the shell layer 4. The shell layer 4 and the substance 5 are bonded to each other. The substance 5 is present on the surface of the magnetic particles 1.
　磁性層３は、単層の磁性層である。磁性層３は、磁性体内包樹脂粒子２の外表面の全体を覆っている。上記磁性粒子では、上記磁性層が上記磁性体内包樹脂粒子の外表面の全体を覆っていてもよく、上記磁性層が上記磁性体内包樹脂粒子の表面の一部を覆っていてもよい。上記磁性粒子では、上記磁性層は、単層の磁性層であってもよく、２層以上の層から構成される多層の磁性層であってもよい。 The magnetic layer 3 is a single magnetic layer. The magnetic layer 3 covers the entire outer surface of the magnetic inclusion resin particles 2. In the magnetic particles, the magnetic layer may cover the entire outer surface of the magnetic inclusion resin particles, or the magnetic layer may cover a part of the surface of the magnetic inclusion resin particles. In the magnetic particles, the magnetic layer may be a single magnetic layer or a multi-layered magnetic layer composed of two or more layers.
　シェル層４は、単層のシェル層である。シェル層４は、磁性層３の外表面の全体を覆っている。上記磁性粒子では、上記シェル層が上記磁性層の外表面の全体を覆っていてもよく、上記シェル層が上記磁性層の表面の一部を覆っていてもよい。 The shell layer 4 is a single-layer shell layer. The shell layer 4 covers the entire outer surface of the magnetic layer 3. In the magnetic particles, the shell layer may cover the entire outer surface of the magnetic layer, or the shell layer may cover a part of the surface of the magnetic layer.
　図２は、本発明の第２の実施形態に係る磁性粒子を模式的に示す断面図である。 FIG. 2 is a cross-sectional view schematically showing the magnetic particles according to the second embodiment of the present invention.
　図２に示す磁性粒子１Ａは、目的物質と特異的に相互作用させるために用いられる。磁性粒子１Ａは、磁性体内包樹脂粒子２Ａと、磁性層３Ａと、目的物質と特異的に相互作用する物質５Ａとを備える。物質５Ａは、例えば、アビジン、ストレプトアビジン、抗原及び抗体等である。磁性粒子１Ａは、上記シェル層を備えない。 The magnetic particles 1A shown in FIG. 2 are used to specifically interact with the target substance. The magnetic particles 1A include magnetic inclusion resin particles 2A, a magnetic layer 3A, and a substance 5A that specifically interacts with a target substance. The substance 5A is, for example, avidin, streptavidin, an antigen, an antibody, or the like. The magnetic particles 1A do not have the shell layer.
　磁性体内包樹脂粒子２Ａは、樹脂粒子２１Ａと、第１の磁性体２２Ａとを有する。図１に示す磁性体内包樹脂粒子２と、図２に示す磁性体内包樹脂粒子２Ａとでは、第１の磁性体の分布状態が異なっている。 The magnetic inclusion resin particles 2A have resin particles 21A and a first magnetic body 22A. The distribution state of the first magnetic substance is different between the magnetic encapsulating resin particles 2 shown in FIG. 1 and the magnetic encapsulating resin particles 2A shown in FIG.
　磁性体内包樹脂粒子２Ａでは、第１の磁性体２２Ａが、磁性体内包樹脂粒子２Ａの内部に均一に分布していない。磁性体内包樹脂粒子２Ａの中心から外表面にかけて、第１の磁性体２２Ａの含有量が多くなるように第１の磁性体２２Ａが配置されている。なお、上記第１の磁性体は、磁性体内包樹脂粒子の内部に均一に分布していてもよい。 In the magnetic inclusion resin particles 2A, the first magnetic substance 22A is not uniformly distributed inside the magnetic inclusion resin particles 2A. From the center of the magnetic inclusion resin particles 2A to the outer surface, the first magnetic body 22A is arranged so that the content of the first magnetic body 22A increases. The first magnetic material may be uniformly distributed inside the magnetic inclusion resin particles.
　磁性層３Ａは、磁性体内包樹脂粒子２Ａの外表面上に配置されている。磁性層３Ａは、第２の磁性体を含む。磁性層３Ａの外表面上に、物質５Ａが担持されている。物質５Ａは、磁性粒子１Ａの表面に存在する。 The magnetic layer 3A is arranged on the outer surface of the magnetic inclusion resin particles 2A. The magnetic layer 3A contains a second magnetic material. The substance 5A is supported on the outer surface of the magnetic layer 3A. The substance 5A is present on the surface of the magnetic particles 1A.
　磁性層３Ａは、単層の磁性層である。磁性層３Ａは、磁性体内包樹脂粒子２Ａの外表面の全体を覆っている。上記磁性粒子では、上記磁性層は、単層の磁性層であってもよく、２層以上の層から構成される多層の磁性層であってもよい。 The magnetic layer 3A is a single magnetic layer. The magnetic layer 3A covers the entire outer surface of the magnetic inclusion resin particles 2A. In the magnetic particles, the magnetic layer may be a single magnetic layer or a multi-layered magnetic layer composed of two or more layers.
　図３は、本発明の第３の実施形態に係る磁性粒子を模式的に示す断面図である。 FIG. 3 is a cross-sectional view schematically showing the magnetic particles according to the third embodiment of the present invention.
　図３に示す磁性粒子１Ｂは、目的物質と特異的に相互作用させるために用いられる。磁性粒子１Ｂは、磁性体内包樹脂粒子２Ｂと、磁性層３Ｂと、シェル層４Ｂと、目的物質と特異的に相互作用する物質５Ｂとを備える。物質５Ｂは、例えば、アビジン、ストレプトアビジン、抗原及び抗体等の生理活性物質等である。 The magnetic particles 1B shown in FIG. 3 are used to specifically interact with the target substance. The magnetic particles 1B include magnetic inclusion resin particles 2B, a magnetic layer 3B, a shell layer 4B, and a substance 5B that specifically interacts with a target substance. The substance 5B is, for example, a bioactive substance such as avidin, streptavidin, an antigen and an antibody.
　磁性体内包樹脂粒子２Ｂは、樹脂粒子２１１Ｂと、第１の磁性体２２Ｂと、樹脂層２１２Ｂとを有する。磁性体内包樹脂粒子２Ｂは、内部に第１の磁性体２２Ｂを含む。磁性体内包樹脂粒子２Ｂにおいて、第１の磁性体２２Ｂが層状に含まれている。磁性体内包樹脂粒子２Ｂは、内部に第１の磁性体２２Ｂを含む磁性層を有する。樹脂粒子２１１Ｂは、第１の磁性体２２Ｂを含まない。樹脂粒子２１１Ｂと樹脂層２１２Ｂとの間に第１の磁性体２２Ｂが配置されていることによって、磁性体内包樹脂粒子２Ｂが、内部に第１の磁性体２２Ｂを含む。磁性体内包樹脂粒子２Ｂにおいて、樹脂粒子２１１Ｂの外表面上に第１の磁性体２２Ｂを含む磁性層が配置されており、該磁性層の外表面上に樹脂層２１２Ｂが配置されている。 The magnetic inclusion resin particles 2B have resin particles 211B, a first magnetic body 22B, and a resin layer 212B. The magnetic inclusion resin particles 2B contain a first magnetic substance 22B inside. In the magnetic inclusion resin particles 2B, the first magnetic substance 22B is contained in a layered manner. The magnetic inclusion resin particles 2B have a magnetic layer containing the first magnetic body 22B inside. The resin particles 211B do not contain the first magnetic material 22B. By arranging the first magnetic body 22B between the resin particles 211B and the resin layer 212B, the magnetic inclusion resin particles 2B contains the first magnetic body 22B inside. In the magnetic inclusion resin particles 2B, a magnetic layer containing the first magnetic substance 22B is arranged on the outer surface of the resin particles 211B, and the resin layer 212B is arranged on the outer surface of the magnetic layer.
　磁性層３Ｂは、磁性体内包樹脂粒子２Ｂの外表面上に配置されている。磁性層３Ｂは、第２の磁性体を含む。シェル層４Ｂは、磁性層３Ｂの外表面上に配置されている。磁性層３Ｂの外表面側に、物質５Ｂが担持されている。シェル層４Ｂの外表面上に、物質５Ｂが担持されている。シェル層４Ｂと、物質５Ｂとが結合している。物質５Ｂは、磁性粒子１Ｂの表面に存在する。 The magnetic layer 3B is arranged on the outer surface of the magnetic inclusion resin particles 2B. The magnetic layer 3B contains a second magnetic material. The shell layer 4B is arranged on the outer surface of the magnetic layer 3B. The substance 5B is supported on the outer surface side of the magnetic layer 3B. The substance 5B is supported on the outer surface of the shell layer 4B. The shell layer 4B and the substance 5B are bonded. The substance 5B exists on the surface of the magnetic particles 1B.
　磁性層３Ｂは、単層の磁性層である。磁性層３Ｂは、磁性体内包樹脂粒子２Ｂの外表面の全体を覆っている。上記磁性粒子では、上記磁性層は、単層の磁性層であってもよく、２層以上の層から構成される多層の磁性層であってもよい。 The magnetic layer 3B is a single magnetic layer. The magnetic layer 3B covers the entire outer surface of the magnetic inclusion resin particles 2B. In the magnetic particles, the magnetic layer may be a single magnetic layer or a multi-layered magnetic layer composed of two or more layers.
　シェル層４Ｂは、単層のシェル層である。シェル層４Ｂは、磁性層３Ｂの外表面の全体を覆っている。 The shell layer 4B is a single-layer shell layer. The shell layer 4B covers the entire outer surface of the magnetic layer 3B.
　図４は、本発明の第４の実施形態に係る磁性粒子を模式的に示す断面図である。 FIG. 4 is a cross-sectional view schematically showing the magnetic particles according to the fourth embodiment of the present invention.
　図４に示す磁性粒子１Ｃは、目的物質と特異的に相互作用させるために用いられる。磁性粒子１Ｃは、磁性体内包樹脂粒子２Ｃと、磁性層３Ｃと、シェル層４Ｃと、目的物質と特異的に相互作用する物質５Ｃとを備える。物質５Ｃは、例えば、アビジン、ストレプトアビジン、抗原及び抗体等の生理活性物質等である。 The magnetic particles 1C shown in FIG. 4 are used to specifically interact with the target substance. The magnetic particles 1C include magnetic inclusion resin particles 2C, a magnetic layer 3C, a shell layer 4C, and a substance 5C that specifically interacts with a target substance. The substance 5C is, for example, a bioactive substance such as avidin, streptavidin, an antigen and an antibody.
　磁性体内包樹脂粒子２Ｃは、樹脂粒子２１１Ｃと、第１の磁性体２２１Ｃ，２２２Ｃと、樹脂層２１２Ｃとを有する。磁性体内包樹脂粒子２Ｃは、内部に第１の磁性体２２１Ｃ，２２２Ｃを含む。第１の磁性体２２１Ｃが、磁性体内包樹脂粒子２Ｃの内部に分散している。第１の磁性体２２１Ｃが、樹脂粒子２１１Ｃの内部に分散している。磁性体内包樹脂粒子２Ｃにおいて、第１の磁性体２２１Ｃが、樹脂粒子２１１Ｃの内部に分散している。磁性体内包樹脂粒子２Ｃにおいて、第１の磁性体２２２Ｃが層状に含まれている。磁性体内包樹脂粒子２Ｃは、内部に第１の磁性体２２２Ｃを含む磁性層を有する。樹脂粒子２１１Ｃと樹脂層２１２Ｃとの間に第１の磁性体２２２Ｃが配置されていることによって、磁性体内包樹脂粒子２Ｃが、内部に第１の磁性体２２２Ｃを含む。磁性体内包樹脂粒子２Ｃにおいて、樹脂粒子２１１Ｃの外表面上に第１の磁性体２２２Ｃを含む磁性層が配置されており、該磁性層の外表面上に樹脂層２１２Ｃが配置されている。 The magnetic inclusion resin particles 2C have resin particles 211C, first magnetic materials 221C and 222C, and a resin layer 212C. The magnetic inclusion resin particles 2C contain the first magnetic substances 221C and 222C inside. The first magnetic body 221C is dispersed inside the magnetic inclusion resin particles 2C. The first magnetic material 221C is dispersed inside the resin particles 211C. In the magnetic inclusion resin particles 2C, the first magnetic body 221C is dispersed inside the resin particles 211C. In the magnetic inclusion resin particles 2C, the first magnetic substance 222C is contained in a layered manner. The magnetic inclusion resin particles 2C have a magnetic layer containing the first magnetic substance 222C inside. By arranging the first magnetic material 222C between the resin particles 211C and the resin layer 212C, the magnetic inclusion resin particles 2C contains the first magnetic material 222C inside. In the magnetic inclusion resin particles 2C, a magnetic layer containing the first magnetic substance 222C is arranged on the outer surface of the resin particles 211C, and the resin layer 212C is arranged on the outer surface of the magnetic layer.
　磁性層３Ｃは、磁性体内包樹脂粒子２Ｃの外表面上に配置されている。磁性層３Ｃは、第２の磁性体を含む。シェル層４Ｃは、磁性層３Ｃの外表面上に配置されている。磁性層３Ｃの外表面側に、物質５Ｃが担持されている。シェル層４Ｃの外表面上に、物質５Ｃが担持されている。シェル層４Ｃと、物質５Ｃとが結合している。物質５Ｃは、磁性粒子１Ｃの表面に存在する。 The magnetic layer 3C is arranged on the outer surface of the magnetic inclusion resin particles 2C. The magnetic layer 3C contains a second magnetic material. The shell layer 4C is arranged on the outer surface of the magnetic layer 3C. The substance 5C is supported on the outer surface side of the magnetic layer 3C. The substance 5C is supported on the outer surface of the shell layer 4C. The shell layer 4C and the substance 5C are bonded. The substance 5C is present on the surface of the magnetic particles 1C.
　磁性層３Ｃは、単層の磁性層である。磁性層３Ｃは、磁性体内包樹脂粒子２Ｃの外表面の全体を覆っている。上記磁性粒子では、上記磁性層は、単層の磁性層であってもよく、２層以上の層から構成される多層の磁性層であってもよい。 The magnetic layer 3C is a single magnetic layer. The magnetic layer 3C covers the entire outer surface of the magnetic inclusion resin particles 2C. In the magnetic particles, the magnetic layer may be a single magnetic layer or a multi-layered magnetic layer composed of two or more layers.
　シェル層４Ｃは、単層のシェル層である。シェル層４Ｃは、磁性層３Ｃの外表面の全体を覆っている。 The shell layer 4C is a single-layer shell layer. The shell layer 4C covers the entire outer surface of the magnetic layer 3C.
　以下、磁性粒子の他の詳細について説明する。 The other details of the magnetic particles will be described below.
　（磁性体内包樹脂粒子）
　上記磁性体内包樹脂粒子は、樹脂を含む。上記磁性体内包樹脂粒子は、樹脂粒子を含む。上記磁性体内包樹脂粒子は、内部に第１の磁性体を含む。上記磁性体内包樹脂粒子は、以下の（１）又は（２）の構成を備えることによって、内部に第１の磁性体を含むことが好ましい。（１）樹脂粒子と第１の磁性体を含み、該樹脂粒子の内部に該第１の磁性体が分散している。（２）樹脂粒子と、第１の磁性体と、樹脂層とを含み、該樹脂粒子の外表面上において、磁性体内包樹脂粒子の中心から外表面に向かって、第１の磁性体を含む磁性層と、樹脂層とが交互に配置されている。上記磁性体内包樹脂粒子は、構成（１）のみを備えていてもよく、構成（２）のみを備えていてもよく、構成（１）と構成（２）とを備えていてもよい。なお、上記磁性体内包樹脂粒子が構成（２）を備える場合には、樹脂粒子の外表面上に第１の磁性体を含む磁性層が配置されていることが好ましく、磁性体内包樹脂粒子の最外層は樹脂層であることが好ましい。上記構成（１）を備える磁性体内包樹脂粒子は、例えば、図１，２に示す磁性体内包樹脂粒子である。上記構成（２）を備える磁性体内包樹脂粒子は、例えば、図３に示す磁性体内包樹脂粒子である。上記構成（１）と上記構成（２）とを備える磁性体内包樹脂粒子は、例えば、図４に示す磁性体内包樹脂粒子である。 (Magnetic encapsulating resin particles)
The magnetic inclusion resin particles contain a resin. The magnetic inclusion resin particles include resin particles. The magnetic encapsulating resin particles contain a first magnetic substance inside. It is preferable that the magnetic encapsulating resin particles contain the first magnetic substance inside by having the following constitution (1) or (2). (1) It contains resin particles and a first magnetic substance, and the first magnetic substance is dispersed inside the resin particles. (2) The resin particles, the first magnetic material, and the resin layer are included, and the first magnetic material is contained on the outer surface of the resin particles from the center of the magnetic inclusion resin particles toward the outer surface. The magnetic layer and the resin layer are arranged alternately. The magnetic inclusion resin particles may have only the configuration (1), may have only the configuration (2), or may have the configuration (1) and the configuration (2). When the magnetic encapsulating resin particles have the configuration (2), it is preferable that a magnetic layer containing the first magnetic substance is arranged on the outer surface of the resin particles, and the magnetic encapsulating resin particles The outermost layer is preferably a resin layer. The magnetic encapsulating resin particles having the above configuration (1) are, for example, the magnetic encapsulating resin particles shown in FIGS. 1 and 2. The magnetic encapsulating resin particles having the above configuration (2) are, for example, the magnetic encapsulating resin particles shown in FIG. The magnetic encapsulating resin particles having the above-mentioned configuration (1) and the above-mentioned configuration (2) are, for example, the magnetic encapsulating resin particles shown in FIG.
　上記磁性体内包樹脂粒子が上記構成（２）を備える場合に、該磁性体内包樹脂粒子は、１層の樹脂層と、第１の磁性体を含む１層の磁性層を有していてもよく、２層以上の樹脂層と、第１の磁性体を含む２層以上の磁性層を有していてもよい。 When the magnetic encapsulating resin particles have the above configuration (2), the magnetic encapsulating resin particles may have one resin layer and one magnetic layer containing the first magnetic material. Often, it may have two or more resin layers and two or more magnetic layers including the first magnetic material.
　＜樹脂粒子及び樹脂層＞
　上記樹脂粒子及び上記樹脂層の材料としては、ポリエチレン、ポリプロピレン、ポリスチレン、ポリ塩化ビニル、ポリ塩化ビニリデン、ポリイソブチレン、ポリブタジエン等のポリオレフィン樹脂；ポリメチルメタクリレート及びポリメチルアクリレート等のアクリル樹脂；ポリカーボネート、ポリアミド、フェノールホルムアルデヒド樹脂、メラミンホルムアルデヒド樹脂、ベンゾグアナミンホルムアルデヒド樹脂、尿素ホルムアルデヒド樹脂、フェノール樹脂、メラミン樹脂、ベンゾグアナミン樹脂、尿素樹脂、エポキシ樹脂、不飽和ポリエステル樹脂、飽和ポリエステル樹脂、ポリエチレンテレフタレート、ポリスルホン、ポリフェニレンオキサイド、ポリアセタール、ポリイミド、ポリアミドイミド、ポリエーテルエーテルケトン、ポリエーテルスルホン、ジビニルベンゼン重合体、並びにジビニルベンゼン共重合体等が挙げられる。上記ジビニルベンゼン共重合体等としては、ジビニルベンゼン－スチレン共重合体及びジビニルベンゼン－（メタ）アクリル酸エステル共重合体等が挙げられる。上記樹脂粒子及び上記樹脂層の材料は、１種のみが用いられてもよく、２種以上が併用されてもよい。また、上記磁性体内包樹脂粒子が上記構成（２）を備える場合、上記樹脂粒子の材料と上記樹脂層の材料とは同一であってもよく、異なっていてもよい。 <Resin particles and resin layer>
Examples of the resin particles and the material of the resin layer include polyolefin resins such as polyethylene, polypropylene, polystyrene, polyvinyl chloride, polyvinylidene chloride, polyisobutylene and polybutadiene; acrylic resins such as polymethylmethacrylate and polymethylacrylate; polycarbonate and polyamide. , Phenol formaldehyde resin, melamine formaldehyde resin, benzoguanamine formaldehyde resin, urea formaldehyde resin, phenol resin, melamine resin, benzoguanamine resin, urea resin, epoxy resin, unsaturated polyester resin, saturated polyester resin, polyethylene terephthalate, polysulfone, polyphenylene oxide, polyacetal , Polygonide, polyamideimide, polyether ether ketone, polyether sulfone, divinylbenzene polymer, divinylbenzene copolymer and the like. Examples of the divinylbenzene copolymer and the like include a divinylbenzene-styrene copolymer and a divinylbenzene- (meth) acrylic acid ester copolymer. As the material of the resin particles and the resin layer, only one kind may be used, or two or more kinds may be used in combination. Further, when the magnetic inclusion resin particles have the above configuration (2), the material of the resin particles and the material of the resin layer may be the same or different.
　上記磁性体内包樹脂粒子を良好に得る観点からは、上記樹脂粒子は、多孔質構造を有することが好ましい。上記構成（１）を備える磁性体内包樹脂粒子における樹脂粒子は、多孔質構造を有することが好ましい。上記構成（２）を備える磁性体内包樹脂粒子における樹脂粒子は、多孔質構造を有していてもよく、多孔質構造を有していなくてもよい。 From the viewpoint of obtaining the magnetic encapsulating resin particles satisfactorily, the resin particles preferably have a porous structure. The resin particles in the magnetic inclusion resin particles having the above configuration (1) preferably have a porous structure. The resin particles in the magnetic inclusion resin particles having the above configuration (2) may or may not have a porous structure.
　上記樹脂粒子のＢＥＴ比表面積は、好ましくは２０ｍ２／ｇ以上、より好ましくは４０ｍ２／ｇ以上、更に好ましくは１００ｍ２／ｇ以上であり、好ましくは８００ｍ２／ｇ以下、より好ましくは７００ｍ２／ｇ以下、更に好ましくは６５０ｍ２／ｇ以下である。上記ＢＥＴ比表面積が上記下限以上及び上記上限以下であると、磁性体内包樹脂粒子に内包される第１の磁性体の含有量を多くすることができ、集磁性をより一層高めることができる。 The BET specific surface area of the resin particles is preferably 20 m 2 / g or more, more preferably 40 m 2 / g or more, further preferably 100 m 2 / g or more, preferably 800 m 2 / g or less, and more preferably 700 m 2 It is / g or less, more preferably 650 m 2 / g or less. When the BET specific surface area is not less than the above lower limit and not more than the above upper limit, the content of the first magnetic substance contained in the magnetic inclusion resin particles can be increased, and the magnetic collection can be further enhanced.
　上記樹脂粒子の平均細孔径は、好ましくは０．５ｎｍ以上、より好ましくは１ｎｍ以上であり、好ましくは３０ｎｍ以下、より好ましくは１０ｎｍ以下である。上記平均細孔径が上記下限以上及び上記上限以下であると、樹脂粒子の内部に磁性体をより一層容易に含有させることができ、磁性体内包樹脂粒子に内包される第１の磁性体の含有量を多くすることができ、集磁性をより一層高めることができる。 The average pore diameter of the resin particles is preferably 0.5 nm or more, more preferably 1 nm or more, preferably 30 nm or less, and more preferably 10 nm or less. When the average pore diameter is equal to or greater than the above lower limit and equal to or less than the above upper limit, the magnetic substance can be more easily contained inside the resin particles, and the content of the first magnetic substance contained in the magnetic inclusion resin particles can be contained. The amount can be increased, and the magnetic collection can be further enhanced.
　上記樹脂粒子のＢＥＴ比表面積、及び平均細孔径は、ＢＪＨ法に準拠して、窒素の吸着等温線から測定することができる。上記樹脂粒子のＢＥＴ比表面積、及び平均細孔径を測定するための測定装置としては、カンタクローム・インスツルメンツ社製「ＮＯＶＡ４２００ｅ」等が挙げられる。 The BET specific surface area and the average pore diameter of the resin particles can be measured from the adsorption isotherm of nitrogen in accordance with the BJH method. Examples of the measuring device for measuring the BET specific surface area and the average pore diameter of the resin particles include "NOVA4200e" manufactured by Cantachrome Instruments.
　上記ＢＥＴ比表面積及び上記平均細孔径等の好ましい範囲を満足する樹脂粒子は、例えば、下記の工程を備える樹脂粒子の製造方法により得ることができる。重合性モノマーと、上記重合性モノマーとは反応しない有機溶剤とを混合し、重合性モノマー溶液を調整する工程。上記重合性モノマー溶液と、アニオン性分散安定剤とを極性溶媒に添加して乳化させて乳化液を得る工程。上記乳化液を数回に分けて添加し、種粒子にモノマーを吸収させて、モノマーが膨潤した種粒子を含む懸濁液を得る工程。上記重合性モノマーを重合させて樹脂粒子を得る工程。上記重合性モノマーとしては、単官能性モノマー、及び多官能性モノマー等が挙げられる。上記重合性モノマーとは反応しない有機溶剤は、重合系の媒体である水等の極性溶媒と相溶しないものであれば、特に限定されない。上記有機溶剤としては、シクロヘキサン、トルエン、キシレン、酢酸エチル、酢酸ブチル、酢酸アリル、酢酸プロピル、クロロホルム、メチルシクロヘキサン、メチルエチルケトン等が挙げられる。上記有機溶剤の添加量は、上記重合性モノマー成分１００重量部に対して、１重量部～８０重量部であることが好ましく、２０重量部～６０重量部であることがより好ましい。上記有機溶剤の添加量が上記の好ましい範囲であると、ＢＥＴ比表面積及び上記平均細孔径等をより一層好適な範囲に制御することができ、粒子内部で緻密な細孔が得られやすくなる。 Resin particles satisfying the preferable ranges such as the BET specific surface area and the average pore diameter can be obtained, for example, by a method for producing resin particles including the following steps. A step of preparing a polymerizable monomer solution by mixing a polymerizable monomer with an organic solvent that does not react with the polymerizable monomer. A step of adding the above-mentioned polymerizable monomer solution and an anionic dispersion stabilizer to a polar solvent and emulsifying them to obtain an emulsion. A step of adding the emulsion in several portions to allow the seed particles to absorb the monomer to obtain a suspension containing the seed particles in which the monomer is swollen. A step of polymerizing the above-mentioned polymerizable monomer to obtain resin particles. Examples of the polymerizable monomer include a monofunctional monomer and a polyfunctional monomer. The organic solvent that does not react with the polymerizable monomer is not particularly limited as long as it is incompatible with a polar solvent such as water, which is a polymerization medium. Examples of the organic solvent include cyclohexane, toluene, xylene, ethyl acetate, butyl acetate, allyl acetate, propyl acetate, chloroform, methylcyclohexane, methyl ethyl ketone and the like. The amount of the organic solvent added is preferably 1 part to 80 parts by weight, more preferably 20 parts by weight to 60 parts by weight, based on 100 parts by weight of the polymerizable monomer component. When the amount of the organic solvent added is in the above-mentioned preferable range, the BET specific surface area, the above-mentioned average pore diameter, and the like can be controlled in a more suitable range, and it becomes easy to obtain dense pores inside the particles.
　上記樹脂粒子の平均粒子径は、好ましくは０．５μｍ以上、より好ましくは１μｍ以上、好ましくは１０μｍ以下、より好ましくは７μｍ以下、更に好ましくは５μｍ以下、特に好ましくは４μｍ以下、最も好ましくは３．５μｍ以下である。上記樹脂粒子の平均粒子径が上記下限以上であると、集磁性をより一層高めることができる。上記樹脂粒子の平均粒子径が上記上限以下であると、磁性粒子の沈降を効果的に抑え、分散性をより一層高めることができ、また、磁性粒子の粒子径を小さくすることができるので、目的物質を効果的に相互作用させることができる。 The average particle size of the resin particles is preferably 0.5 μm or more, more preferably 1 μm or more, preferably 10 μm or less, more preferably 7 μm or less, still more preferably 5 μm or less, particularly preferably 4 μm or less, and most preferably 3. It is 5 μm or less. When the average particle size of the resin particles is at least the above lower limit, the magnetism collection can be further enhanced. When the average particle size of the resin particles is not more than the above upper limit, the sedimentation of the magnetic particles can be effectively suppressed, the dispersibility can be further improved, and the particle size of the magnetic particles can be reduced. The target substance can be effectively interacted with each other.
　上記樹脂粒子の平均粒子径は、数平均粒子径である。上記樹脂粒子の平均粒子径は、任意の樹脂粒子５０個を電子顕微鏡又は光学顕微鏡にて観察し、各樹脂粒子の粒子径の平均値を算出することにより求められる。上記磁性粒子又は上記樹脂粒子を乾燥させた試料を作製し、得られた試料を電子顕微鏡又は光学顕微鏡にて観察することが好ましい。 The average particle size of the resin particles is a number average particle size. The average particle size of the resin particles is obtained by observing 50 arbitrary resin particles with an electron microscope or an optical microscope and calculating the average value of the particle size of each resin particle. It is preferable to prepare a sample obtained by drying the magnetic particles or the resin particles, and observe the obtained sample with an electron microscope or an optical microscope.
　＜第１の磁性体＞
　上記第１の磁性体は、金属又は金属酸化物であることが好ましく、強磁性体又は常磁性体であることがより好ましい。 <First magnetic material>
The first magnetic material is preferably a metal or a metal oxide, and more preferably a ferromagnetic material or a paramagnetic material.
　上記第１の磁性体としては、鉄、コバルト、ニッケル、ルテニウム、ランタノイド、フェライト等が挙げられる。上記フェライトとしては、マグへマイト（γＦｅ２Ｏ３）及びＭＦｅ２Ｏ４で表される化合物（ＭＦｅ２Ｏ４中、Ｍは、Ｃｏ、Ｎｉ、Ｍｎ、Ｚｎ、Ｍｇ、Ｃｕ、Ｆｅ、Ｌｉ０．５Ｆｅ０．５等）等が挙げられる。上記フェライトは、四酸化三鉄（Ｆｅ３Ｏ４）であることが好ましい。上記第１の磁性体は、合金であってもよい。上記合金としては、ニッケル－コバルト合金、コバルト－タングステン合金、鉄－白金合金、及び鉄－コバルト合金等が挙げられる。また、上記金属は、金属イオンであってもよい。上記第１の磁性体は、１種のみが用いられてもよく、２種以上が併用されてもよい。 Examples of the first magnetic material include iron, cobalt, nickel, ruthenium, lanthanoids, ferrite and the like. As the ferrite, in chromite to mug (γFe 2 O 3) and MFe compounds represented by 2 O 4 (MFe 2 O 4 , M is, Co, Ni, Mn, Zn , Mg, Cu, Fe, Li 0 .5 Fe 0.5 etc.) and the like. The ferrite is preferably ferric tetroxide (Fe 3 O 4 ). The first magnetic material may be an alloy. Examples of the alloy include nickel-cobalt alloy, cobalt-tungsten alloy, iron-platinum alloy, iron-cobalt alloy and the like. Moreover, the said metal may be a metal ion. As the first magnetic material, only one kind may be used, or two or more kinds may be used in combination.
　集磁性をより一層高め、かつ高い集磁性をより一層長期間維持する観点からは、上記第１の磁性体は、コバルト又はフェライトであることが好ましく、コバルト又は四酸化三鉄であることがより好ましく、コバルトであることが更に好ましい。 From the viewpoint of further enhancing the magnetic collection and maintaining a high magnetic collection for a longer period of time, the first magnetic material is preferably cobalt or ferrite, and more preferably cobalt or triiron tetroxide. It is preferable, and cobalt is more preferable.
　上記磁性粒子１００体積％中、上記第１の磁性体の含有量は、好ましくは１０体積％以上、より好ましくは２０体積％以上、好ましくは８０体積％以下、より好ましくは７０体積％以下である。上記第１の磁性体の含有量が上記下限以上であると、集磁性をより一層高めることができる。上記第１の磁性体の含有量が上記上限以下であると、分散性をより一層高めることができる。 The content of the first magnetic substance in 100% by volume of the magnetic particles is preferably 10% by volume or more, more preferably 20% by volume or more, preferably 80% by volume or less, and more preferably 70% by volume or less. .. When the content of the first magnetic substance is at least the above lower limit, the magnetism collection can be further enhanced. When the content of the first magnetic substance is not more than the above upper limit, the dispersibility can be further enhanced.
　上記第１の磁性体と上記第２の磁性体の合計１００重量％中、上記第１の磁性体の含有量は、好ましくは１０重量％以上、より好ましくは２０重量％以上、好ましくは９０重量％以下、より好ましくは８５重量％以下である。上記第１の磁性体の含有量が上記下限以上及び上記上限以下であると、集磁性をより一層高め、かつ高い集磁性をより一層長期間維持することができる。 The content of the first magnetic material is preferably 10% by weight or more, more preferably 20% by weight or more, and preferably 90% by weight in the total of 100% by weight of the first magnetic material and the second magnetic material. % Or less, more preferably 85% by weight or less. When the content of the first magnetic substance is not less than the above lower limit and not more than the above upper limit, the magnetic collection can be further enhanced and the high magnetic collection can be maintained for a longer period of time.
　上記磁性体内包樹脂粒子の外表面から中心に向かって厚み１／３までの領域（Ｒ１）の１００体積％中、上記第１の磁性体の含有量（以下、含有量（１）と記載することがある）は、好ましくは２０体積％以上、より好ましくは３０体積％以上であり、好ましくは７０体積％以下、より好ましくは６０体積％以下である。上記含有量（１）が上記下限以上及び上記上限以下であると、集磁性をより一層高めることができる。また、上記含有量（１）が上記上限以下であると、樹脂骨格が良好に保持され、粒子形状を良好に保持することができる。上記領域（Ｒ１）は、図５において、磁性体内包樹脂粒子２の破線Ｌ１よりも外側の領域である。 The content of the first magnetic substance (hereinafter referred to as the content (1)) in 100% by volume of the region (R1) from the outer surface to the center of the magnetic inclusion resin particles to the thickness of 1/3 is described. ) Is preferably 20% by volume or more, more preferably 30% by volume or more, preferably 70% by volume or less, and more preferably 60% by volume or less. When the content (1) is at least the above lower limit and at least the above upper limit, the magnetism collection can be further enhanced. Further, when the content (1) is not more than the above upper limit, the resin skeleton is well retained and the particle shape can be well maintained. The region (R1) is a region outside the broken line L1 of the magnetic inclusion resin particles 2 in FIG.
　上記磁性体内包樹脂粒子の中心から外表面に向かって厚み２／３までの領域（Ｒ２）の１００体積％中、上記第１の磁性体の含有量（以下、含有量（２）と記載することがある）は、好ましくは１５体積％以上、より好ましくは３０体積％以上であり、好ましくは７０体積％以下、より好ましくは６０体積％以下である。上記含有量（２）が上記下限以上及び上記上限以下であると、集磁性をより一層高めることができる。上記領域（Ｒ２）は、図５において、磁性体内包樹脂粒子２の破線Ｌ１よりも内側の領域である。 The content of the first magnetic substance (hereinafter referred to as the content (2)) in 100% by volume of the region (R2) having a thickness of 2/3 from the center of the magnetic inclusion resin particles to the outer surface is described. ) Is preferably 15% by volume or more, more preferably 30% by volume or more, preferably 70% by volume or less, and more preferably 60% by volume or less. When the content (2) is at least the above lower limit and at least the above upper limit, the magnetism collection can be further enhanced. The region (R2) is a region inside the broken line L1 of the magnetic inclusion resin particles 2 in FIG.
　上記含有量（１）の上記含有量（２）に対する比（含有量（１）／含有量（２））は、好ましくは０．８以上、より好ましくは１．０以上であり、好ましくは４．０以下、より好ましくは３．０以下である。上記比（含有量（１）／含有量（２））が上記下限以上及び上記上限以下であると、集磁性及び分散性をより一層高めることができる。 The ratio of the content (1) to the content (2) (content (1) / content (2)) is preferably 0.8 or more, more preferably 1.0 or more, and preferably 4 It is 0.0 or less, more preferably 3.0 or less. When the above ratio (content (1) / content (2)) is at least the above lower limit and at least the above upper limit, the magnetism collection and dispersibility can be further enhanced.
　上記含有量（１）と上記含有量（２）との差の絶対値は、２５体積％以下であることが好ましく、１０体積％以下であることがより好ましい。上記第１の磁性体は、上記磁性体内包樹脂粒子の内部に均一に存在していることが好ましく、上記磁性体内包樹脂粒子の内部に均一に含まれていることが好ましい。 The absolute value of the difference between the content (1) and the content (2) is preferably 25% by volume or less, and more preferably 10% by volume or less. The first magnetic material is preferably uniformly present inside the magnetic inclusion resin particles, and is preferably uniformly contained inside the magnetic inclusion resin particles.
　上記含有量（１）及び上記含有量（２）は、以下のようにして測定できる。 The above-mentioned content (1) and the above-mentioned content (2) can be measured as follows.
　磁性粒子の含有量が３０重量％となるように、Ｋｕｌｚｅｒ社製「テクノビット４０００」に添加し、分散させて、磁性粒子検査用埋め込み樹脂体を作製する。検査用埋め込み樹脂体中に分散した磁性粒子の中心付近を通るようにイオンミリング装置（日立ハイテクノロジーズ社製「ＩＭ４０００」）を用いて、磁性粒子の断面を切り出す。そして、電界放射型透過電子顕微鏡（日本電子社製「ＪＥＭ－２０１０ＦＥＦ」）を用いて、エネルギー分散型Ｘ線分析装置（ＥＤＳ）により、磁性体内包樹脂粒子の厚み方向における第１の磁性体の含有量を測定することで、磁性体内包樹脂粒子の厚み方向における第１の磁性体の含有量の分布結果が得られる。この結果から、上記含有量（１）及び上記含有量（２）を算出することができる。上記含有量（１）及び上記含有量（２）は、任意に選択された２０個の磁性体内包樹脂粒子の含有量（１）及び含有量（２）を算術平均することにより算出された平均含有量であることが好ましい。 Add to "Technobit 4000" manufactured by Kulzer so that the content of magnetic particles is 30% by weight, and disperse to prepare an embedded resin body for magnetic particle inspection. A cross section of the magnetic particles is cut out using an ion milling device (“IM4000” manufactured by Hitachi High-Technologies Corporation) so as to pass near the center of the magnetic particles dispersed in the embedded resin body for inspection. Then, using an electric field radiation type transmission electron microscope (“JEM-2010FEF” manufactured by JEOL Ltd.), an energy dispersive X-ray analyzer (EDS) is used to obtain the first magnetic material in the thickness direction of the magnetic inclusion resin particles. By measuring the content, the distribution result of the content of the first magnetic substance in the thickness direction of the magnetic inclusion resin particles can be obtained. From this result, the above-mentioned content (1) and the above-mentioned content (2) can be calculated. The above-mentioned content (1) and the above-mentioned content (2) are averages calculated by arithmetically averaging the contents (1) and the contents (2) of 20 arbitrarily selected magnetic inclusion resin particles. The content is preferably.
　上記磁性体内包樹脂粒子の製造方法は特に限定されない。例えば、多孔質構造を有する上記樹脂粒子と、上記第１の磁性体とを混合し、該樹脂粒子の内部に該第１の磁性体を導入することで、上記構成（１）を備える磁性体内包樹脂粒子を得ることができる。また、例えば、中実構造を有する上記樹脂粒子と上記第１の磁性体を混合し、該樹脂粒子の外表面に該第１の磁性体を被覆し、次いで該第１の磁性体の外表面を樹脂で被覆することにより、上記構成（２）を備える磁性体内包樹脂粒子を得ることができる。 The method for producing the magnetic encapsulating resin particles is not particularly limited. For example, by mixing the resin particles having a porous structure and the first magnetic material and introducing the first magnetic material into the resin particles, a magnetic body having the above configuration (1) is provided. Encapsulating resin particles can be obtained. Further, for example, the resin particles having a solid structure and the first magnetic material are mixed, the outer surface of the resin particles is coated with the first magnetic material, and then the outer surface of the first magnetic material is coated. By coating the above with a resin, magnetic inclusion resin particles having the above configuration (2) can be obtained.
　（磁性層）
　本発明に係る磁性粒子は、第２の磁性体を含む磁性層を備える。上記第２の磁性体を含む磁性層は、上記磁性体内包樹脂粒子の外表面上に配置される。上記第１の磁性体と上記第２の磁性体とは、同一であってもよく、異なっていてもよい。 (Magnetic layer)
The magnetic particles according to the present invention include a magnetic layer containing a second magnetic material. The magnetic layer containing the second magnetic material is arranged on the outer surface of the magnetic inclusion resin particles. The first magnetic material and the second magnetic material may be the same or different.
　上記第２の磁性体は、金属又は金属酸化物であることが好ましく、強磁性体又は常磁性体であることがより好ましい。 The second magnetic material is preferably a metal or a metal oxide, and more preferably a ferromagnetic material or a paramagnetic material.
　上記第２の磁性体としては、鉄、コバルト、ニッケル、ルテニウム、ランタノイド及びフェライト等が挙げられる。上記フェライトとしては、マグへマイト（γＦｅ２Ｏ３）及びＭＦｅ２Ｏ４で表される化合物（ＭＦｅ２Ｏ４中、Ｍは、Ｃｏ、Ｎｉ、Ｍｎ、Ｚｎ、Ｍｇ、Ｃｕ、Ｆｅ、Ｌｉ０．５Ｆｅ０．５等）等が挙げられる。上記フェライトは、四酸化三鉄（Ｆｅ３Ｏ４）であることが好ましい。上記第２の磁性体は、合金であってもよい。上記合金としては、ニッケル－コバルト合金、コバルト－タングステン合金、鉄－白金合金、及び鉄－コバルト合金等が挙げられる。また、上記金属は、金属イオンであってもよい。上記第２の磁性体は、１種のみが用いられてもよく、２種以上が併用されてもよい。 Examples of the second magnetic material include iron, cobalt, nickel, ruthenium, lanthanoids, ferrite and the like. As the ferrite, in chromite to mug (γFe 2 O 3) and MFe compounds represented by 2 O 4 (MFe 2 O 4 , M is, Co, Ni, Mn, Zn , Mg, Cu, Fe, Li 0 .5 Fe 0.5 etc.) and the like. The ferrite is preferably ferric tetroxide (Fe 3 O 4 ). The second magnetic material may be an alloy. Examples of the alloy include nickel-cobalt alloy, cobalt-tungsten alloy, iron-platinum alloy, iron-cobalt alloy and the like. Moreover, the said metal may be a metal ion. As the second magnetic material, only one kind may be used, or two or more kinds may be used in combination.
　集磁性をより一層高め、かつ高い集磁性をより一層長期間維持する観点からは、上記第２の磁性体は、フェライトであることが好ましく、四酸化三鉄であることがより好ましい。 From the viewpoint of further enhancing the magnetic collection and maintaining a high magnetic collection for a longer period of time, the second magnetic material is preferably ferrite, and more preferably triiron tetroxide.
　上記第１の磁性体と上記第２の磁性体の合計１００重量％中、上記第２の磁性体の含有量は、好ましくは１０重量％以上、より好ましくは３０重量％以上、好ましくは９０重量％以下、より好ましくは７０重量％以下である。上記第２の磁性体の含有量が上記下限以上及び上記上限以下であると、集磁性をより一層高め、かつ高い集磁性をより一層長期間維持することができる。 The content of the second magnetic material is preferably 10% by weight or more, more preferably 30% by weight or more, preferably 90% by weight, based on 100% by weight of the total of the first magnetic material and the second magnetic material. % Or less, more preferably 70% by weight or less. When the content of the second magnetic substance is not less than the above lower limit and not more than the above upper limit, the magnetic collection can be further enhanced and the high magnetic collection can be maintained for a longer period of time.
　上記磁性粒子１００体積％中、上記第２の磁性体の含有量は、好ましくは４体積％以上、より好ましくは８体積％以上、好ましくは４０体積％以下、より好ましくは２５体積％以下である。上記第２の磁性体の含有量が上記下限以上であると、集磁性をより一層高めることができる。上記第２の磁性体の含有量が上記上限以下であると、分散性をより一層高めることができる。 The content of the second magnetic substance in 100% by volume of the magnetic particles is preferably 4% by volume or more, more preferably 8% by volume or more, preferably 40% by volume or less, and more preferably 25% by volume or less. .. When the content of the second magnetic substance is at least the above lower limit, the magnetism collection can be further enhanced. When the content of the second magnetic substance is not more than the above upper limit, the dispersibility can be further enhanced.
　上記磁性粒子１００重量％中、上記第１の磁性体と上記第２の磁性体との合計の含有量は、好ましくは１０重量％以上、より好ましくは１５重量％以上、好ましくは９５重量％以下、より好ましくは８０重量％以下である。上記合計の含有量が上記下限以上であると、集磁性をより一層高めることができる。上記合計の含有量が上記上限以下であると、分散性をより一層高めることができる。 The total content of the first magnetic substance and the second magnetic substance in 100% by weight of the magnetic particles is preferably 10% by weight or more, more preferably 15% by weight or more, and preferably 95% by weight or less. , More preferably 80% by weight or less. When the total content is at least the above lower limit, the magnetism collection can be further enhanced. When the total content is not more than the above upper limit, the dispersibility can be further enhanced.
　上記磁性粒子１００体積％中、上記第１の磁性体と上記第２の磁性体との合計の含有量は、好ましくは２０体積％以上、より好ましくは３０体積％以上、好ましくは８０体積％以下、より好ましくは７０体積％以下である。上記合計の含有量が上記下限以上であると、集磁性をより一層高めることができる。上記合計の含有量が上記上限以下であると、分散性をより一層高めることができる。 The total content of the first magnetic substance and the second magnetic substance in 100% by volume of the magnetic particles is preferably 20% by volume or more, more preferably 30% by volume or more, and preferably 80% by volume or less. , More preferably 70% by volume or less. When the total content is at least the above lower limit, the magnetism collection can be further enhanced. When the total content is not more than the above upper limit, the dispersibility can be further enhanced.
　上記磁性体内包樹脂粒子の外表面の全表面積１００％中、上記第２の磁性体を含む磁性層により覆われている表面積は、好ましくは７０％以上、より好ましくは８０％以上、更に好ましくは９５％以上、最も好ましくは１００％である。上記表面積が上記下限以上であると、集磁性をより一層高めることができ、かつ高い集磁性をより一層長期間維持することができ、さらに分散性をより一層高めることができる。 Of the total surface area of the outer surface of the magnetic inclusion resin particles, the surface area covered by the magnetic layer containing the second magnetic substance is preferably 70% or more, more preferably 80% or more, still more preferably. It is 95% or more, most preferably 100%. When the surface area is at least the above lower limit, the magnetic collection can be further enhanced, the high magnetic collection can be maintained for a longer period of time, and the dispersibility can be further enhanced.
　上記磁性層の厚みは、好ましくは２０ｎｍ以上、より好ましくは５０ｎｍ以上であり、好ましくは１０００ｎｍ以下、より好ましくは２００ｎｍ以下である。上記磁性層の厚みは、磁性層が多層である場合には磁性層全体の厚みである。上記磁性層の厚みが上記下限以上であると、集磁性をより一層高めることができ、かつ高い集磁性をより一層長期間維持することができる。上記磁性層の厚みが上記上限以下であると、磁性粒子の沈降を効果的に抑え、分散性をより一層高めることができ、また、磁性粒子の粒子径を小さくすることができるので、磁性粒子の単位重量当たりの目的物質の結合量を多くすることができる。 The thickness of the magnetic layer is preferably 20 nm or more, more preferably 50 nm or more, preferably 1000 nm or less, and more preferably 200 nm or less. The thickness of the magnetic layer is the thickness of the entire magnetic layer when the magnetic layer is multi-layered. When the thickness of the magnetic layer is at least the above lower limit, the magnetic collection can be further enhanced, and the high magnetic collection can be maintained for a longer period of time. When the thickness of the magnetic layer is not more than the above upper limit, the sedimentation of the magnetic particles can be effectively suppressed, the dispersibility can be further improved, and the particle size of the magnetic particles can be reduced, so that the magnetic particles can be reduced. The amount of binding of the target substance per unit weight of the above can be increased.
　上記磁性層の厚みは、例えば透過型電子顕微鏡（ＴＥＭ）を用いて、磁性粒子の断面を観察することにより測定できる。上記磁性層の厚みについては、任意の磁性層の厚み５箇所の平均値を１個の磁性粒子の磁性層の厚みとして算出することが好ましく、磁性層全体の厚みの平均値を１個の磁性粒子の磁性層の厚みとして算出することがより好ましい。上記磁性層の厚みは、任意の磁性粒子１０個について、各磁性粒子の磁性層の厚みの平均値を算出することにより求めることが好ましい。 The thickness of the magnetic layer can be measured by observing the cross section of the magnetic particles, for example, using a transmission electron microscope (TEM). Regarding the thickness of the magnetic layer, it is preferable to calculate the average value of five thicknesses of any magnetic layer as the thickness of the magnetic layer of one magnetic particle, and the average value of the thickness of the entire magnetic layer is one magnetic. It is more preferable to calculate as the thickness of the magnetic layer of the particles. The thickness of the magnetic layer is preferably obtained by calculating the average value of the thickness of the magnetic layer of each magnetic particle for 10 arbitrary magnetic particles.
　上記磁性層は、連続層であってもよく、上記第２の磁性体が粒子状に集合した粒状物によって形成された層であってもよい。また、上記磁性層は、樹脂粒子の外表面上の全部を被覆していてもよく、一部を被覆していてもよい。また、上記磁性層は、海島構造を有していてもよい。集磁性の低下を効果的に抑える観点からは、上記磁性層は、連続層であることが好ましい。上記連続層は、例えば、微粒子の凝集体のように磁性層に無数の継ぎ目（例えば磁性粒子１個あたり１０００個以上）が存在する形状とは異なり、継ぎ目が少ない或いは継ぎ目が無い構造を有する層を指す。集磁性をより一層高める観点からは、上記磁性層は、第２の磁性体が粒子状に集合した粒状物によって形成された層であることが好ましい。 The magnetic layer may be a continuous layer, or may be a layer formed by particles in which the second magnetic material is aggregated in the form of particles. Further, the magnetic layer may be entirely covered on the outer surface of the resin particles, or may be partially covered. Further, the magnetic layer may have a sea-island structure. From the viewpoint of effectively suppressing the decrease in magnetic collection, the magnetic layer is preferably a continuous layer. The continuous layer has a structure having few seams or no seams, unlike a shape in which innumerable seams (for example, 1000 or more per magnetic particle) exist in the magnetic layer such as an aggregate of fine particles. Point to. From the viewpoint of further enhancing the magnetic collection, the magnetic layer is preferably a layer formed by particles in which the second magnetic material is aggregated in the form of particles.
　上記磁性層が、上記第２の磁性体が粒子状に集合した粒状物によって形成された層である場合に、該粒状物の平均粒径は、好ましくは１ｎｍ以上、より好ましくは２ｎｍ以上、好ましくは５０ｎｍ以下、より好ましくは２０ｎｍ以下である。上記粒状物の平均粒径が上記下限以上であると、集磁性をより一層高めることができる。上記粒状物の平均粒径が上記上限以下であると、上記磁性体内包樹脂粒子の外表面に第２の磁性体を良好に配置することができ、磁性層を良好に形成させることができる。 When the magnetic layer is a layer formed by particles in which the second magnetic material is aggregated in the form of particles, the average particle size of the particles is preferably 1 nm or more, more preferably 2 nm or more, preferably 2 nm or more. Is 50 nm or less, more preferably 20 nm or less. When the average particle size of the granules is at least the above lower limit, the magnetism collection can be further enhanced. When the average particle size of the granules is not more than the upper limit, the second magnetic substance can be satisfactorily arranged on the outer surface of the magnetic inclusion resin particles, and the magnetic layer can be satisfactorily formed.
　上記磁性層の形成方法は、特に限定されない。例えば上記磁性体内包樹脂粒子と上記第２の磁性体の粒状物とを混合することにより、上記磁性体内包樹脂粒子の外表面上に磁性層を形成させることができる。 The method for forming the magnetic layer is not particularly limited. For example, by mixing the magnetic inclusion resin particles and the particles of the second magnetic substance, a magnetic layer can be formed on the outer surface of the magnetic inclusion resin particles.
　（シェル層）
　本発明に係る磁性粒子は、シェル層を備えることが好ましい。上記シェル層の材料は、無機酸化物又は有機ポリマーを含む。上記シェル層は、該シェル層の材料として上記無機酸化物を含む無機酸化物シェル層であるか、又は該シェル層の材料として上記有機ポリマーを含む有機ポリマーシェル層である。上記シェル層は、上記第２の磁性体を含む上記磁性層の外表面上に配置されることが好ましい。上記磁性粒子が上記シェル層を備えることにより、上記目的物質と特異的に相互作用する物質を磁性粒子の表面に良好に配置することができ、また、磁性粒子の分散性をより一層高めることができる。また、上記磁性粒子がシェル層を備えることにより、磁性体内包樹脂粒子等からの不純物の溶出、磁性体の溶出、及び磁性層からの不純物の溶出を強固に防ぐことができる。そのため、上記磁性粒子が上記シェル層を備える場合には、検査薬として好適に用いることができる。 (Shell layer)
The magnetic particles according to the present invention preferably include a shell layer. The material of the shell layer contains an inorganic oxide or an organic polymer. The shell layer is an inorganic oxide shell layer containing the inorganic oxide as the material of the shell layer, or an organic polymer shell layer containing the organic polymer as the material of the shell layer. The shell layer is preferably arranged on the outer surface of the magnetic layer containing the second magnetic material. When the magnetic particles are provided with the shell layer, a substance that specifically interacts with the target substance can be satisfactorily arranged on the surface of the magnetic particles, and the dispersibility of the magnetic particles can be further enhanced. it can. Further, when the magnetic particles include a shell layer, it is possible to strongly prevent the elution of impurities from the magnetic inclusion resin particles and the like, the elution of the magnetic material, and the elution of impurities from the magnetic layer. Therefore, when the magnetic particles include the shell layer, they can be suitably used as a test agent.
　上記シェル層は、磁性体を含んでもよく、磁性体を含まなくてもよい。上記シェル層は、磁性体を含まないことがより好ましい。上記シェル層は、磁性体を含まない非磁性層であることがより好ましい。 The shell layer may or may not contain a magnetic material. It is more preferable that the shell layer does not contain a magnetic material. The shell layer is more preferably a non-magnetic layer containing no magnetic material.
　磁性粒子の分散性をより一層高める観点からは、上記シェル層の材料は、上記無機酸化物を含むことが好ましく、無機酸化物を含む無機酸化物シェル層であることがより好ましい。 From the viewpoint of further enhancing the dispersibility of the magnetic particles, the material of the shell layer preferably contains the above-mentioned inorganic oxide, and more preferably an inorganic oxide shell layer containing the inorganic oxide.
　＜無機酸化物＞
　上記無機酸化物とは、金属元素又は半金属元素と、酸素原子とを少なくとも有する化合物を意味する。上記無機酸化物としては、特に限定されない。上記無機酸化物は、１種のみが用いられてもよく、２種以上が用いられてもよい。 <Inorganic oxide>
The inorganic oxide means a compound having at least a metal element or a metalloid element and an oxygen atom. The inorganic oxide is not particularly limited. Only one kind of the above-mentioned inorganic oxide may be used, or two or more kinds may be used.
　上記無機酸化物は、ケイ素原子、ゲルマニウム原子、チタン原子又はジルコニウム原子を有する無機酸化物であることが好ましい。また、上記無機酸化物は、上記磁性層の外表面と反応可能な官能基を有することが好ましい。磁性表面と非磁性層を反応させる方法は特に限定されないが、例えば共有結合や配位結合が挙げられる。 The inorganic oxide is preferably an inorganic oxide having a silicon atom, a germanium atom, a titanium atom or a zirconium atom. Further, the inorganic oxide preferably has a functional group capable of reacting with the outer surface of the magnetic layer. The method for reacting the magnetic surface with the non-magnetic layer is not particularly limited, and examples thereof include covalent bonds and coordination bonds.
　上記無機酸化物としては、具体的には、オルトケイ酸テトラエチル等のアルコキシシラン及びその加水分解物に代表されるシラン化合物、ゲルマニウムテトラエトキシド等のアルコキシゲルマニウム及びその加水分解物に代表されるゲルマニウム化合物、チタンテトラエトキシド等のアルコキシチタン及びその加水分解物に代表されるチタン化合物、ジルコニウムテトラブトキシド等のアルコキシジルコニウム及びその加水分解物に代表されるジルコニウム化合物等が挙げられる。 Specific examples of the inorganic oxide include an alkoxysilane such as tetraethyl orthosilicate and a silane compound typified by a hydrolyzate thereof, and an alkoxygermanium such as germanium tetraethoxydo and a germanium compound typified by the hydrolyzate thereof. , Titanium compounds typified by alkoxytitanium such as titanium tetraethoxydo and its hydrolyzate, zirconium compounds such as zirconium tetrabutoxide and zirconium compounds typified by its hydrolyzate.
　磁性粒子の分散性を高く維持する観点からは、上記無機酸化物は、比重の小さい化合物であることが好ましく、上記の例示の中では、上記シラン化合物であることが最も好ましい。 From the viewpoint of maintaining high dispersibility of the magnetic particles, the inorganic oxide is preferably a compound having a small specific gravity, and among the above examples, the above silane compound is most preferable.
　上記シラン化合物としては、例えば、オルトケイ酸テトラエチル；ビニルトリメトキシシラン、ビニルトリエトキシシラン、７－オクテニルトリメトキシシラン等のビニル基含有シラン化合物；２－（３，４－エポキシシクロヘキシル）エチルトリメトキシシラン、３－グリシドキシプロピルメチルジメトキシシラン、３－グリシドキシプロピルトリメトキシシラン、３－グリシドキシプロピルメチルジエトキシシラン、３－グリシドキシプロピルトリエトキシシラン、８－グリシドキシオクチルトリメトキシシラン等のエポキシ基含有シラン化合物；ｐ－スチリルトリメトキシシラン等のスチリル基含有シラン化合物；３－メタクリロキシプロピルメチルジメトキシシラン、３－メタクリロキシプロピルトリメトキシシラン、３－メタクリロキシプロピルメチルジエトキシシラン、３－メタクリロキシプロピルトリエトキシシラン、８－メタクリロキシオクチルトリメトキシシラン等のメタクリル基含有シラン化合物；３－アクリロキシプロピルトリメトキシシラン等のアクリル基含有シラン化合物；Ｎ－２－（アミノエチル）－３－アミノプロピルメチルジメトキシシラン、Ｎ－２－（アミノエチル）－３－アミノプロピルトリメトキシシラン、３－アミノプロピルトリメトキシシラン、３－アミノプロピルトリエトキシシラン、３－トリエトキシシリル－Ｎ－（１，３－ジメチル－ブチリデン）プロピルアミン、Ｎ－フェニル－３－アミノプロピルトリメトキシシラン、Ｎ－（ビニルベンジル）－２－アミノエチル－３－アミノプロピルトリメトキシシラン塩酸塩、Ｎ－２－（アミノエチル）－８－アミノオクチルトリメトキシシラン等のアミノ基含有シラン化合物；トリス－（トリメトキシシリルプロピル）イソシアヌレート等のイソシアヌレート基含有シラン化合物；３－ウレイドプロピルトリアルコキシシラン等のウレイド基含有シラン化合物；３－メルカプトプロピルメチルジメトキシシラン、３－メルカプトプロピルトリメトキシシラン等のメルカプト基含有シラン化合物；３－イソシアネートプロピルトリエトキシシラン等のイソシアネート基含有シラン化合物；３－トリメトキシシリルプロピルコハク酸無水物等のカルボン酸無水物含有シラン化合物；３－トリメトキシシリルプロピルコハク酸無水物の加水分解物等のカルボン酸含有シラン化合物等が挙げられる。 Examples of the silane compound include tetraethyl orthosilicate; a vinyl group-containing silane compound such as vinyltrimethoxysilane, vinyltriethoxysilane, and 7-octenyltrimethoxysilane; 2- (3,4-epoxycyclohexyl) ethyltrimethoxy. Silane, 3-glycidoxypropylmethyldimethoxysilane, 3-glycidoxypropyltrimethoxysilane, 3-glycidoxypropylmethyldiethoxysilane, 3-glycidoxypropyltriethoxysilane, 8-glycidoxyoctyltri Epoxy group-containing silane compounds such as methoxysilane; styryl group-containing silane compounds such as p-styryltrimethoxysilane; 3-methacryloxypropylmethyldimethoxysilane, 3-methacryloxypropyltrimethoxysilane, 3-methacryloxypropylmethyldiethoxy Methacrylic group-containing silane compounds such as silane, 3-methacryloxypropyltriethoxysilane, 8-methacryloxyoctyltrimethoxysilane; acrylic group-containing silane compounds such as 3-acryloxypropyltrimethoxysilane; N-2- (aminoethyl) ) -3-Aminopropylmethyldimethoxysilane, N-2- (aminoethyl) -3-aminopropyltrimethoxysilane, 3-aminopropyltrimethoxysilane, 3-aminopropyltriethoxysilane, 3-triethoxysilyl-N -(1,3-Dimethyl-butylidene) propylamine, N-phenyl-3-aminopropyltrimethoxysilane, N- (vinylbenzyl) -2-aminoethyl-3-aminopropyltrimethoxysilane hydrochloride, N-2 -Amino group-containing silane compound such as (aminoethyl) -8-aminooctyltrimethoxysilane; isocyanurate group-containing silane compound such as tris- (trimethoxysilylpropyl) isocyanurate; ureido such as 3-ureidopropyltrialkoxysilane Group-containing silane compound; mercapto group-containing silane compound such as 3-mercaptopropylmethyldimethoxysilane and 3-mercaptopropyltrimethoxysilane; isocyanate group-containing silane compound such as 3-isocyanoxidetriethoxysilane; 3-trimethoxysilylpropylkohaku Examples thereof include carboxylic acid anhydride-containing silane compounds such as acid anhydride; and carboxylic acid-containing silane compounds such as a hydrolyzate of 3-trimethoxysilylpropyl succinic acid anhydride.
　例えば、磁性層の表面を種々の官能基を有するシラン化合物で処理することにより、上記磁性層の外表面上にケイ素原子を有する無機酸化物シェル層を形成させることができる。 For example, by treating the surface of the magnetic layer with a silane compound having various functional groups, an inorganic oxide shell layer having a silicon atom can be formed on the outer surface of the magnetic layer.
　上記無機酸化物シェル層１００重量％中、上記無機酸化物の含有量は、好ましくは７０重量％以上、より好ましくは８０重量％以上である。 The content of the inorganic oxide in 100% by weight of the inorganic oxide shell layer is preferably 70% by weight or more, more preferably 80% by weight or more.
　＜有機ポリマー＞
　上記有機ポリマーは、特に限定されないが、ビニル系ポリマーであることが好ましい。 <Organic polymer>
The organic polymer is not particularly limited, but is preferably a vinyl polymer.
　上記ビニル系ポリマーの材料に用いられるビニル系モノマーとしては、スチレン、α－メチルスチレン、クロルスチレン、ジビニルベンゼン等のスチレン単量体；メチルビニルエーテル、エチルビニルエーテル、プロピルビニルエーテル等のビニルエーテル化合物；酢酸ビニル、酪酸ビニル、ラウリン酸ビニル、ステアリン酸ビニル等の酸ビニルエステル化合物；塩化ビニル、フッ化ビニル等のハロゲン含有単量体；（メタ）アクリル酸；メチル（メタ）アクリレート、エチル（メタ）アクリレート、プロピル（メタ）アクリレート、ブチル（メタ）アクリレート、２－エチルヘキシル（メタ）アクリレート、ラウリル（メタ）アクリレート、セチル（メタ）アクリレート、ステアリル（メタ）アクリレート、シクロヘキシル（メタ）アクリレート、イソボルニル（メタ）アクリレート、エチレングリコール（メタ）アクリレート等のアルキル（メタ）アクリレート化合物；２－ヒドロキシエチル（メタ）アクリレート、グリセロール（メタ）アクリレート、ポリオキシエチレン（メタ）アクリレート、グリシジル（メタ）アクリレート等の酸素原子含有（メタ）アクリレート化合物；トリフルオロメチル（メタ）アクリレート、ペンタフルオロエチル（メタ）アクリレート等のハロゲン含有（メタ）アクリレート化合物；ビニルトリメトキシシラン、ビニルトリエトキシシラン、ジメトキシメチルビニルシシラン、ジメトキシエチルビニルシラン、ジエトキシメチルビニルシラン、ジエトキシエチルビニルシラン、エチルメチルジビニルシラン、メチルビニルジメトキシシラン、エチルビニルジメトキシシラン、メチルビニルジエトキシシラン、エチルビニルジエトキシシラン、ｐ－スチリルトリメトキシシラン、３－メタクリロキシプロピルメチルジメトキシシラン、３－メタクリロキシプロピルトリメトキシシラン、３－メタクリロキシプロピルメチルジエトキシシラン、３－メタクリロキシプロピルトリエトキシシラン、３－アクリロキシプロピルトリメトキシシラン等の重合性二重結合含有シランアルコキシド等が挙げられる。 Examples of the vinyl-based monomer used as the material for the vinyl-based polymer include styrene monomers such as styrene, α-methylstyrene, chlorostyrene, and divinylbenzene; vinyl ether compounds such as methylvinyl ether, ethylvinyl ether, and propylvinyl ether; vinyl acetate, Acid vinyl ester compounds such as vinyl butyrate, vinyl laurate, vinyl stearate; halogen-containing monomers such as vinyl chloride and vinyl fluoride; (meth) acrylic acid; methyl (meth) acrylate, ethyl (meth) acrylate, propyl (Meta) acrylate, butyl (meth) acrylate, 2-ethylhexyl (meth) acrylate, lauryl (meth) acrylate, cetyl (meth) acrylate, stearyl (meth) acrylate, cyclohexyl (meth) acrylate, isobornyl (meth) acrylate, ethylene Alkyl (meth) acrylate compounds such as glycol (meth) acrylate; oxygen atom-containing (meth) such as 2-hydroxyethyl (meth) acrylate, glycerol (meth) acrylate, polyoxyethylene (meth) acrylate, and glycidyl (meth) acrylate. Acrylate compound; Halogen-containing (meth) acrylate compound such as trifluoromethyl (meth) acrylate and pentafluoroethyl (meth) acrylate; vinyl trimethoxysilane, vinyl triethoxysilane, dimethoxymethyl vinyl cysilane, dimethoxyethyl vinyl silane, diethoxy Methylvinylsilane, diethoxyethylvinylsilane, ethylmethyldivinylsilane, methylvinyldimethoxysilane, ethylvinyldimethoxysilane, methylvinyldiethoxysilane, ethylvinyldiethoxysilane, p-styryltrimethoxysilane, 3-methacryloxypropylmethyldimethoxysilane , 3-methacryloxypropyltrimethoxysilane, 3-methacryloxypropylmethyldiethoxysilane, 3-methacryloxypropyltriethoxysilane, 3-acryloxypropyltrimethoxysilane, and other polymerizable double bond-containing silane alkoxides. Be done.
　上記ビニル系ポリマーは１種のビニル系モノマーが重合した単独重合体であってもよく、２種以上のビニル系モノマーが重合した共重合体であってもよい。 The vinyl-based polymer may be a homopolymer obtained by polymerizing one kind of vinyl-based monomer, or may be a copolymer obtained by polymerizing two or more kinds of vinyl-based monomers.
　例えば、母粒子の存在下で、主原料としてのビニル系モノマー等の共重合モノマーと、必要に応じて副原料である重合開始剤、乳化剤、分散剤、界面活性剤、電解質、架橋剤、分子量調節剤等を添加し、液体中で重合を行うことにより、上記磁性層の外表面上に有機ポリマーシェル層を形成させることができる。 For example, in the presence of mother particles, a copolymerization monomer such as a vinyl monomer as a main raw material and, if necessary, a polymerization initiator, an emulsifier, a dispersant, a surfactant, an electrolyte, a cross-linking agent, and a molecular weight as auxiliary raw materials. An organic polymer shell layer can be formed on the outer surface of the magnetic layer by adding a modifier or the like and polymerizing in a liquid.
　上記有機ポリマーシェル層１００重量％中、上記有機ポリマーの含有量は、好ましくは７０重量％以上、より好ましくは８０重量％以上である。 The content of the organic polymer in 100% by weight of the organic polymer shell layer is preferably 70% by weight or more, more preferably 80% by weight or more.
　上記シェル層は、上記目的物質と特異的に相互作用する上記物質と結合する前において、カルボキシル基、水酸基、エポキシ基、アミノ基、トシル基、チオール基、トリエチルアンモニウム基、ジメチルアミノ基及びスルホン酸基等の官能基を有することが好ましい。上記シェル層が上記官能基を有する場合には、上記目的物質と特異的に相互作用する上記物質を上記シェル層の外表面上に良好に担持させることができ、該物質を磁性粒子の表面に良好に配置することができる。 The shell layer has a carboxyl group, a hydroxyl group, an epoxy group, an amino group, a tosyl group, a thiol group, a triethylammonium group, a dimethylamino group and a sulfonic acid before binding to the substance that specifically interacts with the target substance. It is preferable to have a functional group such as a group. When the shell layer has the functional group, the substance that specifically interacts with the target substance can be well supported on the outer surface of the shell layer, and the substance can be supported on the surface of the magnetic particles. It can be arranged well.
　上記シェル層は、外表面上にリンカー部を有していてもよい。上記シェル層が上記リンカー部を有することにより、上記目的物質と特異的に相互作用する上記物質との結合点である上記官能基を、上記シェル層の最表面からより離れた位置に配置することができ、立体的な障害がより少ない位置で上記官能基と上記物質とが接触可能になる。このため、上記物質が結合しやすくなり、上記物質を磁性粒子の表面に良好に配置することができる。 The shell layer may have a linker portion on the outer surface. When the shell layer has the linker portion, the functional group, which is a bonding point with the substance that specifically interacts with the target substance, is arranged at a position farther from the outermost surface of the shell layer. The functional group and the substance can come into contact with each other at a position where there are less steric obstacles. Therefore, the substance is easily bonded, and the substance can be satisfactorily arranged on the surface of the magnetic particles.
　上記リンカー部は、上記目的物質と特異的に相互作用する上記物質と結合する前において、末端に、カルボキシル基、水酸基、エポキシ基、アミノ基、トシル基、チオール基等の目的物質と共有結合可能な官能基を有していることが好ましい。上記官能基と、上記目的物質と特異的に相互作用する上記物質の官能基とを反応させることにより、上記シェル層と、該物質とを化学結合させることができる。上記エポキシ基は、グリシジル基含有モノマー由来のエポキシ基であってもよい。上記水酸基は、上記エポキシ基の開環により生じる水酸基であってもよい。 The linker portion can be covalently bonded to a target substance such as a carboxyl group, a hydroxyl group, an epoxy group, an amino group, a tosyl group, or a thiol group at the terminal before binding to the above substance that specifically interacts with the target substance. It is preferable to have a functional group. By reacting the functional group with the functional group of the substance that specifically interacts with the target substance, the shell layer and the substance can be chemically bonded. The epoxy group may be an epoxy group derived from a glycidyl group-containing monomer. The hydroxyl group may be a hydroxyl group generated by ring-opening of the epoxy group.
　上記リンカー部の材料は、複数のエポキシ基を末端に有するエポキシ化合物であることが好ましい。上記複数のエポキシ基を末端に有するエポキシ化合物は、ポリエチレングリコールジグリシジルエーテル、ポリプロピレングリコールジグリシジルエーテル、ネオペンチルグリコールジグリシジルエーテル、又はトリメチロールプロパンポリグリシジルエーテルであることが好ましい。上記複数のエポキシ基を末端に有するエポキシ化合物は、ポリエチレングリコールジグリシジルエーテルであることがより好ましい。 The material of the linker portion is preferably an epoxy compound having a plurality of epoxy groups at the ends. The epoxy compound having a plurality of epoxy groups at the end is preferably polyethylene glycol diglycidyl ether, polypropylene glycol diglycidyl ether, neopentyl glycol diglycidyl ether, or trimethylol propanepolyglycidyl ether. The epoxy compound having a plurality of epoxy groups at the end is more preferably polyethylene glycol diglycidyl ether.
　上記シェル層と、上記目的物質と特異的に相互作用する上記物質とは、結合していることが好ましく、化学結合していることがより好ましい。 The shell layer and the substance that specifically interacts with the target substance are preferably bonded, and more preferably chemically bonded.
　上記第２の磁性体を含む磁性層の外表面の全表面積１００％中、上記シェル層により覆われている表面積は、好ましくは９５％以上、より好ましくは９９％以上、最も好ましくは１００％である。上記表面積が上記下限以上であると、上記目的物質と特異的に相互作用する上記物質の含有量を多くすることができ、その結果、磁性粒子を用いた目的物質の測定において、測定精度及び測定感度を高めることができる。 Of the total surface area of the outer surface of the magnetic layer containing the second magnetic material, the surface area covered by the shell layer is preferably 95% or more, more preferably 99% or more, and most preferably 100%. is there. When the surface area is equal to or greater than the above lower limit, the content of the substance that specifically interacts with the target substance can be increased, and as a result, the measurement accuracy and measurement in the measurement of the target substance using magnetic particles The sensitivity can be increased.
　上記シェル層の厚みは、好ましくは２０ｎｍ以上、より好ましくは４０ｎｍ以上であり、好ましくは５００ｎｍ以下、より好ましくは３００ｎｍ以下である。上記シェル層の厚みが上記下限以上及び上記上限以下であると、磁性粒子の沈降を効果的に抑え、分散性をより一層高めることができ、また、磁性粒子の粒子径を小さくすることができるので、目的物質を効果的に結合させることができる。 The thickness of the shell layer is preferably 20 nm or more, more preferably 40 nm or more, preferably 500 nm or less, and more preferably 300 nm or less. When the thickness of the shell layer is not less than the above lower limit and not more than the above upper limit, the sedimentation of the magnetic particles can be effectively suppressed, the dispersibility can be further improved, and the particle diameter of the magnetic particles can be reduced. Therefore, the target substance can be effectively bound.
　上記シェル層の厚みは、例えば透過型電子顕微鏡（ＴＥＭ）を用いて、磁性粒子の断面を観察することにより測定できる。上記シェル層の厚みについては、任意のシェル層の厚み５箇所の平均値を１個の磁性粒子のシェル層の厚みとして算出することが好ましく、シェル層全体の厚みの平均値を１個の磁性粒子のシェル層の厚みとして算出することがより好ましい。上記シェル層の厚みは、任意の磁性粒子１０個について、各磁性粒子のシェル層の厚みの平均値を算出することにより求めることが好ましい。 The thickness of the shell layer can be measured by observing the cross section of the magnetic particles, for example, using a transmission electron microscope (TEM). Regarding the thickness of the shell layer, it is preferable to calculate the average value of the thickness of any shell layer at five points as the thickness of the shell layer of one magnetic particle, and the average value of the thickness of the entire shell layer is one magnetism. It is more preferable to calculate as the thickness of the shell layer of particles. The thickness of the shell layer is preferably obtained by calculating the average value of the thickness of the shell layer of each magnetic particle for 10 arbitrary magnetic particles.
　（目的物質と特異的に相互作用する物質）
　本発明に係る磁性粒子の表面において、目的物質と特異的に相互作用する物質が存在する。上記物質としては、糖鎖、ペプチド鎖、タンパク質、抗原、及びヌクレオチド鎖等が挙げられる。上記物質は、該目的物質の種類によって適宜変更可能である。上記物質は、１種のみが用いられてもよく、２種以上が併用されてもよい。 (Substances that specifically interact with the target substance)
On the surface of the magnetic particles according to the present invention, there is a substance that specifically interacts with the target substance. Examples of the substance include sugar chains, peptide chains, proteins, antigens, nucleotide chains and the like. The substance can be appropriately changed depending on the type of the target substance. Only one type of the above substance may be used, or two or more types may be used in combination.
　上記物質と上記目的物質との相互作用としては、例えば、抗原抗体反応及び酵素と基質の相互作用等が挙げられる。上記物質と上記目的物質との相互作用は、上記物質と上記目的物質との非共有結合的な相互作用であってもよく、上記物質と上記目的物質との共有結合であってもよい。上記非共有結合的な相互作用としては、疎水性相互作用、静電的相互作用、ファンデルワールス力、水素結合、配位結合及びイオン結合等が挙げられる。 Examples of the interaction between the substance and the target substance include an antigen-antibody reaction and an interaction between an enzyme and a substrate. The interaction between the substance and the target substance may be a non-covalent interaction between the substance and the target substance, or may be a covalent bond between the substance and the target substance. Examples of the non-covalent interaction include hydrophobic interaction, electrostatic interaction, van der Waals force, hydrogen bond, coordination bond, ionic bond and the like.
　上記目的物質と特異的に相互作用する物質は、上記目的物質と非共有結合的な相互作用が可能な物質であること好ましい。 The substance that specifically interacts with the target substance is preferably a substance capable of non-covalent interaction with the target substance.
　上記目的物質と特異的に相互作用する上記物質と、該目的物質の組み合わせとしては、抗体と抗原との組み合わせ、糖鎖とレクチン等のタンパク質との組み合わせ、酵素等のタンパク質とインヒビターとの組み合わせ、ペプチドとタンパク質との組み合わせ、ヌクレオチド鎖とヌクレオチド鎖との組み合わせ、ヌクレオチド鎖とタンパク質との組み合わせ等が挙げられる。 The combination of the above-mentioned substance that specifically interacts with the above-mentioned target substance and the target substance includes a combination of an antibody and an antigen, a combination of a sugar chain and a protein such as a lectin, and a combination of a protein such as an enzyme and an inhibitor. Examples thereof include a combination of a peptide and a protein, a combination of a nucleotide chain and a nucleotide chain, and a combination of a nucleotide chain and a protein.
　上記目的物質と特異的に相互作用する上記物質は、タンパク質であることが好ましく、アビジン又はストレプトアビジンであることがより好ましい。また、上記目的物質と特異的に相互作用する上記物質は、抗原又は抗体であることが好ましい。 The substance that specifically interacts with the target substance is preferably a protein, more preferably avidin or streptavidin. In addition, the substance that specifically interacts with the target substance is preferably an antigen or an antibody.
　上記抗体は、ポリクローナル抗体であってもよく、モノクロナール抗体であってもよい。上記抗体は、パパイン及びペプシン等のタンパク質分解酵素により処理されていてもよく、Ｆａｂ及びＦ（ａｂ’）２フラグメント等であってもよい。 The above antibody may be a polyclonal antibody or a monoclonal antibody. The antibody may be treated with a proteolytic enzyme such as papain and pepsin, or may be a Fab and F (ab') 2 fragment or the like.
　上記目的物質と特異的に相互作用する上記物質の配置方法は、特に限定されない。例えば、上記物質と、該物質が担持される前の磁性粒子とを混合することにより、該物質を該磁性粒子の表面に配置することができる。 The method of arranging the substance that specifically interacts with the target substance is not particularly limited. For example, the substance can be arranged on the surface of the magnetic particles by mixing the substance with the magnetic particles before the substance is supported.
　（目的物質）
　上記目的物質とは、上述した物質と特異的に相互作用する物質である。 (Target substance)
The target substance is a substance that specifically interacts with the above-mentioned substance.
　上記目的物質としては、タンパク質、核酸、ホルモン、癌マーカー、呼吸器関連マーカー、心疾患マーカー及び薬物等挙げられる。 Examples of the target substance include proteins, nucleic acids, hormones, cancer markers, respiratory-related markers, heart disease markers, drugs and the like.
　上記タンパク質としては、高比重リポタンパク質（ＨＤＬ）、低比重リポタンパク質（ＬＤＬ）、超低比重リポタンパク質等の脂質タンパク質；アルカリホスファターゼ、アミラーゼ、酸性ホスファターゼ、γ－グルタミルトランスフェラーゼ（γ－ＧＴＰ）、リパーゼ、クレアチンキナーゼ（ＣＫ）、乳酸脱水素酵素（ＬＤＨ）、グルタミン酸オキザロ酢酸トランスアミナーゼ（ＧＯＴ）、グルタミン酸ピルビン酸トランスアミナーゼ（ＧＰＴ）、レニン、プロテインキナーゼ（ＰＫ）、チロシンキナーゼ等の酵素；ＩｇＧ、ＩｇＭ、ＩｇＡ、ＩｇＤ、ＩｇＥ等の免疫グロブリン（又はこれらのＦｃ部、Ｆａｂ部及びＦ（ａｂ）２部等の断片）；例えばフィブリノーゲン、フィブリン分解産物（ＦＤＰ）、プロトロンビン、トロンビン等の血液凝固関連因子；抗ストレプトリジンＯ抗体、抗ヒトＢ型肝炎ウイルス表面抗原抗体（ＨＢｓ抗原）、抗ヒトＣ型肝炎ウイルス抗体、抗リュウマチ因子等の抗体；アルブミン、ヘモグロビン、ミオグロビン、トランスフェリン、プロテインＡ、Ｃ反応性蛋白質（ＣＲＰ）等が挙げられる。 Examples of the above proteins include lipid proteins such as high specific gravity lipoprotein (HDL), low specific gravity lipoprotein (LDL), and ultralow specific gravity lipoprotein; alkaline phosphatase, amylase, acidic phosphatase, γ-glutamyltransferase (γ-GTP), lipase. , Cleatin kinase (CK), lactic acid dehydrogenase (LDH), glutamate oxaloacetate transaminase (GOT), glutamate pyruvate transaminase (GPT), renin, protein kinase (PK), tyrosine kinase and other enzymes; IgG, IgM, IgA , IgD, IgE and other immunoglobulins (or fragments of these Fc, Fab and F (ab) 2 parts, etc.); for example, blood coagulation-related factors such as fibrinogen, fibrin degradation products (FDP), prothrombin, thrombin; Antibodies such as streptidine O antibody, anti-human hepatitis B virus surface antigen antibody (HBs antigen), anti-human hepatitis C virus antibody, anti-rheumatic factor; albumin, hemoglobin, myoglobin, transferase, protein A, C reactive protein ( CRP) and the like.
　上記核酸としては、ＤＮＡ及びＲＮＡ等が挙げられる。 Examples of the nucleic acid include DNA and RNA.
　上記ホルモンとしては、甲状腺刺激ホルモン（ＴＳＨ）、甲状腺ホルモン（ＦＴ３、ＦＴ４、Ｔ３、Ｔ４）、副甲状腺ホルモン（ＰＴＨ）、及びヒト絨毛性ゴナドトロピン（ｈＣＧ）エストラジオール（Ｅ２）等が挙げられる。 Examples of the hormone include thyroid stimulating hormone (TSH), thyroid hormone (FT3, FT4, T3, T4), parathyroid hormone (PTH), and human chorionic gonadotropin (hCG) estradiol (E2).
　上記癌マーカーとしては、α－フェトプロテイン（ＡＦＰ）、ＰＩＶＫＡ－ＩＩ、癌胎児性抗原（ＣＥＡ）、ＣＡ１９－９、及び前立腺特異抗原（ＰＳＡ）等が挙げられる。 Examples of the cancer marker include α-fetoprotein (AFP), PIVKA-II, carcinoembryonic antigen (CEA), CA19-9, and prostate-specific antigen (PSA).
　上記呼吸器関連マーカーとしては、ＫＬ－６等が挙げられる。 Examples of the respiratory-related marker include KL-6 and the like.
　上記心疾患マーカーとしては、トロポニンＴ（ＴｎＴ）、及びヒト脳性ナトリウム利尿ペプチド前駆体Ｎ端フラグメント（ＮＴ－ｐｒｏＢＮＰ）等が挙げられる。 Examples of the above-mentioned heart disease marker include troponin T (TnT), human brain natriuretic peptide precursor N-terminal fragment (NT-proBNP), and the like.
　上記薬物としては、抗てんかん薬、抗生物質、及びテオフィリン等が挙げられる。 Examples of the above-mentioned drugs include antiepileptic drugs, antibiotics, theophylline and the like.
　（磁性粒子の他の詳細）
　上記磁性粒子は、放射免疫測定法（ＲＩＡ）、酵素免疫測定法（ＥＩＡ）、蛍光免疫測定法（ＦＩＡ）、電気化学発光法（ＥＣＬＩＡ）、化学発光免疫測定法（ＣＬＩＡ及びＣＬＥＩＡ）、吸光度測定、並びに表面プラズモン共鳴等の測定において、好適に用いられる。上記磁性粒子は、サンドイッチ法及び競合法の測定において、好適に用いられる。 (Other details of magnetic particles)
The magnetic particles are used for radioimmunoassay (RIA), enzyme immunoassay (EIA), fluorescence immunoassay (FIA), chemiluminescence immunoassay (ECLIA), chemiluminescence immunoassay (CLIA and CLEIA), and absorbance measurement. , And in the measurement of surface plasmon resonance and the like, it is preferably used. The magnetic particles are preferably used in the measurement of the sandwich method and the competitive method.
　上記磁性粒子は、検体中の上記目的物質の濃度を測定にするために好適に用いられる。 The magnetic particles are suitably used for measuring the concentration of the target substance in a sample.
　上記磁性粒子は、検査薬として好適に用いられる。上記磁性粒子は、検査薬として用いることができる磁性粒子であることが好ましい。 The magnetic particles are preferably used as a test agent. The magnetic particles are preferably magnetic particles that can be used as a test agent.
　上記磁性粒子を用いた目的物質の濃度測定は、例えば、以下のようにして行うことができる。 The concentration of the target substance can be measured using the magnetic particles as follows, for example.
　上記磁性粒子を含む液（例えば、後述の検査薬）と、目的物質を含む検体とを混合し、混合液を得る。得られた混合液を加温等して、磁性粒子における上記目的物質と特異的に相互作用する上記物質と、検体中の目的物質を結合させた反応液を得る（第１の反応工程）。次いで、磁石等により、反応液に磁力を加え、磁性粒子を集める（集磁工程）。未反応の検体を除去したのち、洗浄液を添加し、混合する（洗浄工程）。なお、集磁工程及び洗浄工程は、複数回繰り返されてもよい。次いで、洗浄液を除去した後、該目的物質と標識物質とを反応させて、目的物質の濃度を測定する（第２の反応工程）。 A liquid containing the above magnetic particles (for example, a test agent described later) and a sample containing the target substance are mixed to obtain a mixed liquid. The obtained mixed solution is heated or the like to obtain a reaction solution in which the substance that specifically interacts with the target substance in the magnetic particles and the target substance in the sample are bound to each other (first reaction step). Next, a magnetic force is applied to the reaction solution by a magnet or the like to collect magnetic particles (magnetic collection step). After removing the unreacted sample, a washing solution is added and mixed (washing step). The magnetic collection step and the cleaning step may be repeated a plurality of times. Next, after removing the cleaning liquid, the target substance and the labeling substance are reacted to measure the concentration of the target substance (second reaction step).
　上記標識物質としては、例えば酵素免疫測定法（ＥＩＡ）において好適に用いられるアルカリホスファターゼ、β－ガラクトシダーゼ、ペルオキシダーゼ、マイクロペルオキシダーゼ、グルコースオキシダーゼ、グルコース－６－リン酸脱水素酵素、リンゴ酸脱水素酵素、ルシフェラーゼ、チロシナーゼ、酸性ホスファターゼ等の酵素；例えば放射免疫測定法（ＲＩＡ）において好適に用いられる９９ｍＴｃ、１３１Ｉ、１２５Ｉ、１４Ｃ、３Ｈ、３２Ｐ等の放射性同位元素；例えば蛍光免疫測定法（ＦＩＡ）において好適に用いられるフルオレセイン、ダンシル、フルオレスカミン、クマリン、ナフチルアミン及びこれらの誘導体、グリーン蛍光タンパク質（ＧＦＰ）等の蛍光性物質、例えばルシフェリン、イソルミノール、ルミノール、ビス（２，４，６－トリフロロフェニル）オキザレート等の発光性物質、例えばフェノール、ナフトール、アントラセン及びこれらの誘導体等の紫外部に吸収を有する物質、例えば４－アミノ－２，２，６，６－テトラメチルピペリジン－１－オキシル、３－アミノ－２，２，５，５－テトラメチルピロリジン－１－オキシル、２，６－ジ－ｔ－ブチル－α－（３，５－ジ－ｔ－ブチル－４－オキソ－２，５－シクロヘキサジエン－１－イリデン）－ｐ－トリオキシル等のオキシル基を有する化合物等のスピンラベル化剤としての性質を有する化合物等が挙げられる。測定感度を高める観点からは、上記標識物質は、酵素又は蛍光性物質であることが好ましく、アルカリホスファターゼ、ペルオキシダーゼ又はグルコースオキシダーゼであることがより好ましく、ペルオキシダーゼであることが更に好ましい。 Examples of the labeling substance include alkaline phosphatase, β-galactosidase, peroxidase, microperoxidase, glucose oxidase, glucose-6-phosphate dehydrogenase, malic acid dehydrogenase, which are preferably used in enzyme immunoassay (EIA). Enzymes such as luciferase, tyrosinase, acidic phosphatase; radioactive isotopes such as 99mTc, 131I, 125I, 14C, 3H, 32P preferably used in radioimmunosassay (RIA); suitable in fluorescence immunoassay (FIA), for example. Fluorescent substances such as fluorescein, dancil, fluorescamine, coumarin, naphthylamine and derivatives thereof, and fluorescent substances such as green fluorescent protein (GFP) used in, for example, luciferin, isolminol, luminol, bis (2,4,6-trifluorophenyl). ) Luminescent substances such as oxalate, such as phenol, naphthol, anthracene and derivatives thereof that have ultraviolet absorption, such as 4-amino-2,2,6,6-tetramethylpiperidine-1-oxyl, 3 -Amino-2,2,5,5-tetramethylpyrrolidin-1-oxyl, 2,6-di-t-butyl-α- (3,5-di-t-butyl-4-oxo-2,5-) Examples thereof include compounds having properties as spin labeling agents such as compounds having an oxyl group such as cyclohexadien-1-iriden) -p-trioxyl. From the viewpoint of increasing the measurement sensitivity, the labeling substance is preferably an enzyme or a fluorescent substance, more preferably alkaline phosphatase, peroxidase or glucose oxidase, and even more preferably peroxidase.
　（検査薬）
　上記検査薬は、上述した磁性粒子を含む。 (Test drug)
The above-mentioned test agent contains the above-mentioned magnetic particles.
　上記検査薬は、緩衝液を含むことが好ましい。 The above test agent preferably contains a buffer solution.
　上記緩衝液は、ｐＨ５．０以上９．０以下において緩衝能を有する緩衝液であることが好ましい。上記緩衝液としては、リン酸緩衝液、グリシン緩衝液、ベロナール緩衝液、トリス緩衝液、ホウ酸緩衝液、クエン酸緩衝液、及びグッド緩衝液等が挙げられる。 The buffer solution is preferably a buffer solution having a buffering ability at pH 5.0 or more and 9.0 or less. Examples of the buffer solution include phosphate buffer solution, glycine buffer solution, veronal buffer solution, Tris buffer solution, borate buffer solution, citrate buffer solution, and Good buffer solution.
　上記検査薬は、増感剤、タンパク質等の高分子化合物、アミノ酸、及び界面活性剤等の他の成分を含んでいてもよい。 The above-mentioned test agent may contain a sensitizer, a polymer compound such as a protein, an amino acid, and other components such as a surfactant.
　上記検査薬が上記増感剤を含むことにより、目的物質と該目的物質と結合可能である化合物との反応を効率的に進行させることができ、また、測定精度を高めることができる。上記増感剤としては、メチルセルロース及びエチルセルロース等のアルキル化多糖化合物、プルラン、並びにポリビニルピロリドン等が挙げられる。 When the test agent contains the sensitizer, the reaction between the target substance and the compound capable of binding to the target substance can be efficiently promoted, and the measurement accuracy can be improved. Examples of the sensitizer include alkylated polysaccharide compounds such as methyl cellulose and ethyl cellulose, pullulan, polyvinylpyrrolidone and the like.
　上記タンパク質としては、アルブミン（牛血清アルブミン及び卵性アルブミン等）、カゼイン、及びゼラチン等が挙げられる。 Examples of the protein include albumin (bovine serum albumin, egg albumin, etc.), casein, gelatin, and the like.
　上記検査薬１００重量％中、上記磁性粒子の含有量は、好ましくは０．５重量％以上、より好ましくは２重量％以上、好ましくは１０重量％以下、より好ましくは５重量％以下である。上記磁性粒子の含有量が上記下限以上及び上記上限以下であると、目的物質の測定精度をより一層高めることができる。 The content of the magnetic particles in 100% by weight of the test agent is preferably 0.5% by weight or more, more preferably 2% by weight or more, preferably 10% by weight or less, and more preferably 5% by weight or less. When the content of the magnetic particles is not less than the above lower limit and not more than the above upper limit, the measurement accuracy of the target substance can be further improved.
　以下、実施例及び比較例を挙げて、本発明を具体的に説明する。本発明は、以下の実施例のみに限定されない。 Hereinafter, the present invention will be specifically described with reference to Examples and Comparative Examples. The present invention is not limited to the following examples.
　（実施例１）
　磁性体内包樹脂粒子の作製：
　種粒子として平均粒子径０．６９μｍのポリスチレン粒子を用意した。上記ポリスチレン粒子３．９重量部と、イオン交換水５００重量部と、５．０重量％ポリビニルアルコール水溶液１２０重量部とを混合し、混合液を調製した。上記混合液を超音波により分散させた後、セパラブルフラスコに入れて、均一に撹拌した。 (Example 1)
Preparation of magnetic encapsulating resin particles:
Polystyrene particles having an average particle diameter of 0.69 μm were prepared as seed particles. A mixed solution was prepared by mixing 3.9 parts by weight of the polystyrene particles, 500 parts by weight of ion-exchanged water, and 120 parts by weight of a 5.0% by weight polyvinyl alcohol aqueous solution. After the above mixed solution was dispersed by ultrasonic waves, it was placed in a separable flask and stirred uniformly.
　次に、ジビニルベンゼン（モノマー成分）１５０重量部と、過酸化ベンゾイル（日油社製「ナイパーＢＷ」）２．０重量部とを混合した。さらに、ラウリル硫酸トリエタノールアミン９．０重量部と、トルエン（溶媒）１５０重量部と、イオン交換水１１００重量部とを添加し、乳化液を調製した。 Next, 150 parts by weight of divinylbenzene (monomer component) and 2.0 parts by weight of benzoyl peroxide (“NOF BW” manufactured by NOF Corporation) were mixed. Further, 9.0 parts by weight of triethanolamine lauryl sulfate, 150 parts by weight of toluene (solvent), and 1100 parts by weight of ion-exchanged water were added to prepare an emulsion.
　セパラブルフラスコ中の上記混合液に、上記乳化液を数回に分けて添加し、１２時間撹拌し、種粒子にモノマーを吸収させて、モノマーが膨潤した種粒子を含む懸濁液を得た。 The emulsion was added to the mixed solution in the separable flask in several portions and stirred for 12 hours to allow the seed particles to absorb the monomer to obtain a suspension containing the seed particles in which the monomer was swollen. ..
　その後、５．０重量％ポリビニルアルコール水溶液４９０重量部を添加し、加熱を開始して８５℃で９時間反応させ、樹脂粒子を得た。 Then, 490 parts by weight of a 5.0% by weight polyvinyl alcohol aqueous solution was added, heating was started, and the mixture was reacted at 85 ° C. for 9 hours to obtain resin particles.
　撹拌子を入れた３００ｍＬ容のビーカーに、得られた樹脂粒子１重量部、６０％硫酸５０重量部を秤量した後、撹拌し、９０℃で２４時間反応させ、スルホン基が導入された樹脂粒子を得た。 1 part by weight of the obtained resin particles and 50 parts by weight of 60% sulfuric acid were weighed in a 300 mL beaker containing a stirrer, and then stirred and reacted at 90 ° C. for 24 hours to introduce the resin particles into which a sulfone group was introduced. Got
　その後、撹拌子を入れた２００ｍＬビーカーに、スルホン基が導入された樹脂粒子１．０重量部、硫酸コバルト・７水和物５．０重量部、及び蒸留水２５ｍＬを秤量した後、室温下で２４時間撹拌した。その後、上記粒子及び蒸留水２０ｍＬ、ジメチルアミンボラン５．０重量部を秤量し、超音波照射下、６０℃で１時間反応させ、第１の磁性体としてコバルトを含む磁性体内包樹脂粒子を得た。 Then, 1.0 part by weight of the resin particles into which the sulfone group was introduced, 5.0 parts by weight of cobalt sulfate heptahydrate, and 25 mL of distilled water were weighed in a 200 mL beaker containing a stirrer, and then at room temperature. The mixture was stirred for 24 hours. Then, the particles, 20 mL of distilled water, and 5.0 parts by weight of dimethylamine borane were weighed and reacted at 60 ° C. for 1 hour under ultrasonic irradiation to obtain magnetic encapsulating resin particles containing cobalt as the first magnetic material. It was.
　磁性層（第２の磁性体を含む磁性層）の形成：
　磁性体内包樹脂粒子１重量部と、磁性流体ＥＭＧ７０７（平均粒子径約１０ｎｍの四酸化三鉄ナノ粒子の水分散液、フェローテック社製）４重量部（但し、そのうち磁性体の含有量は約１７重量％）とを２５０ｒｐｍで１０間撹拌した。得られた粒子分散液を、水を用いてろ過及び洗浄し、磁性体内包樹脂粒子の外表面上に第２の磁性体としての四酸化三鉄を含む磁性層を形成させて、磁性層を有する磁性体内包樹脂粒子を得た。 Formation of magnetic layer (magnetic layer containing a second magnetic material):
1 part by weight of magnetic inclusion resin particles and 4 parts by weight of magnetic fluid EMG707 (aqueous dispersion of triiron tetroxide nanoparticles with an average particle diameter of about 10 nm, manufactured by Fellow Tech) (however, the content of magnetic material is about about 17% by weight) was stirred at 250 rpm for 10 minutes. The obtained particle dispersion is filtered and washed with water to form a magnetic layer containing triiron tetroxide as a second magnetic substance on the outer surface of the magnetic inclusion resin particles to form a magnetic layer. The magnetic encapsulating resin particles having were obtained.
　目的物質と特異的に相互作用する物質の配置：
　磁性層を有する磁性体内包樹脂粒子の水分散液０．５ｍＬを試験管に加え、ＰＢＳ溶液を用いて３回洗浄した。分散媒を除去した後、シアル化糖鎖抗原ＫＬ－６（以下、ＫＬ－６と略記）抗体を含むＰＢＳ溶液（０．７５ｍｇ／ｍＬ）０．５ｍＬを加え、２５℃にて一夜撹拌した。その後、１．０重量％ＢＳＡ溶液を１．５ｍＬ添加し、２５℃にて４時間撹拌した。続いて、１．０重量％ＢＳＡ溶液１．５ｍＬを用いて分散と磁石を用いた試験管壁面への集磁を３回繰り返し、抗ＫＬ－６抗体が感作された磁性粒子分散液を調製した。 Arrangement of substances that specifically interact with the target substance:
0.5 mL of an aqueous dispersion of magnetic encapsulating resin particles having a magnetic layer was added to a test tube, and the mixture was washed 3 times with a PBS solution. After removing the dispersion medium, 0.5 mL of a PBS solution (0.75 mg / mL) containing a sialylated sugar chain antigen KL-6 (hereinafter abbreviated as KL-6) antibody was added, and the mixture was stirred at 25 ° C. overnight. Then, 1.5 mL of 1.0 wt% BSA solution was added, and the mixture was stirred at 25 ° C. for 4 hours. Subsequently, dispersion using 1.5 mL of 1.0 wt% BSA solution and magnetic collection on the test tube wall surface using a magnet were repeated three times to prepare a magnetic particle dispersion sensitized with an anti-KL-6 antibody. did.
　なお、得られた樹脂粒子における平均粒子径、ＢＥＴ比表面積及び平均細孔径、並びに、得られた磁性粒子における第１の磁性体、第２の磁性体の含有量、含有量（１）の含有量（２）に対する比、及び含有量（１）と含有量（２）との差の絶対値、平均粒子径を表１に示す。なお、これらは、後述の評価の項目に記載の方法で求めた。 The average particle size, BET specific surface area and average pore size of the obtained resin particles, and the contents and contents (1) of the first magnetic substance and the second magnetic substance in the obtained magnetic particles. Table 1 shows the ratio to the amount (2), the absolute value of the difference between the content (1) and the content (2), and the average particle size. These were obtained by the method described in the evaluation items described later.
　（実施例２）
　トルエン１５０重量部に変えて、トルエン８０重量部用いて樹脂粒子を作製したこと、磁性粒子の構成を表１に示すように変更したこと以外は、実施例１と同様して、磁性粒子を作製した。 (Example 2)
Magnetic particles were prepared in the same manner as in Example 1 except that resin particles were prepared using 80 parts by weight of toluene instead of 150 parts by weight of toluene and the composition of the magnetic particles was changed as shown in Table 1. did.
　（実施例３）
　トルエン１５０重量部に変えて、トルエン３０重量部用いて樹脂粒子を作製したこと、磁性粒子の構成を表１に示すように変更したこと以外は、実施例１と同様して、磁性粒子を作製した。 (Example 3)
Magnetic particles were prepared in the same manner as in Example 1 except that resin particles were prepared using 30 parts by weight of toluene instead of 150 parts by weight of toluene and the composition of the magnetic particles was changed as shown in Table 1. did.
　（実施例４）
　実施例１と同様にして、磁性層を有する磁性体内包樹脂粒子を得た。 (Example 4)
In the same manner as in Example 1, magnetic inclusion resin particles having a magnetic layer were obtained.
　シェル層の形成：
　得られた磁性層を有する磁性体内包樹脂粒子１．０重量部にエタノール４００重量部及び２８％アンモニア水溶液（ナカライテスク社製）２０重量部を加えた。次いで、オルトケイ酸テトラエチル５．０重量部及び８－グリシドキシオクチルトリメトキシシラン１５重量部を加え、１時間撹拌した。得られた分散液をろ過後、水で洗浄した。このようにして、表面に官能基を有する無機酸化物シェル層を有する磁性体内包粒子を得た。 Shell layer formation:
To 1.0 part by weight of the magnetic encapsulating resin particles having the obtained magnetic layer, 400 parts by weight of ethanol and 20 parts by weight of a 28% aqueous ammonia solution (manufactured by Nacalai Tesque) were added. Then, 5.0 parts by weight of tetraethyl orthosilicate and 15 parts by weight of 8-glycidoxyoctyltrimethoxysilane were added, and the mixture was stirred for 1 hour. The obtained dispersion was filtered and then washed with water. In this way, magnetic inclusion particles having an inorganic oxide shell layer having a functional group on the surface were obtained.
　目的物質と特異的に相互作用する物質の配置：
　実施例１と同様にして、磁性粒子の表面に抗ＫＬ－６抗体が存在する磁性粒子を得た。 Arrangement of substances that specifically interact with the target substance:
In the same manner as in Example 1, magnetic particles having an anti-KL-6 antibody on the surface of the magnetic particles were obtained.
　（実施例５，６）
　樹脂粒子及び磁性粒子の構成を表１に示すように変更したこと以外は、実施例４と同様にして磁性粒子を得た。 (Examples 5 and 6)
Magnetic particles were obtained in the same manner as in Example 4 except that the configurations of the resin particles and the magnetic particles were changed as shown in Table 1.
　（比較例１）
　トルエンを用いなかったこと、ジビニルベンゼン１５０重量部に変えて、ジビニルベンゼン３００重量部を用いたこと、過酸化ベンゾイル２．０重量部に変えて、過酸化ベンゾイル４．０重量部を用いたこと以外は、実施例１と同様にして樹脂粒子を得た。また、比較例１では、第１の磁性体を用いなかった。得られた樹脂粒子を用いて、実施例１と同様にして、樹脂粒子の外表面上に第２の磁性体により磁性層を形成した。 (Comparative Example 1)
No toluene was used, 300 parts by weight of divinylbenzene was used instead of 150 parts by weight of divinylbenzene, and 4.0 parts by weight of benzoyl peroxide was used instead of 2.0 parts by weight of benzoyl peroxide. Resin particles were obtained in the same manner as in Example 1 except for the above. Further, in Comparative Example 1, the first magnetic material was not used. Using the obtained resin particles, a magnetic layer was formed on the outer surface of the resin particles by a second magnetic material in the same manner as in Example 1.
　シェル層の形成：
　得られた磁性層を有する樹脂粒子を用いて、実施例４と同様にして、該磁性層の外表面上にシェル層を形成した。 Shell layer formation:
Using the obtained resin particles having a magnetic layer, a shell layer was formed on the outer surface of the magnetic layer in the same manner as in Example 4.
　目的物質と特異的に相互作用する物質の配置：
　得られたシェル層及び磁性層を有する樹脂粒子を用いて、実施例１と同様にして、抗ＫＬ－６抗体を配置した。このようにして、第１の磁性体を有さない磁性粒子を得た。 Arrangement of substances that specifically interact with the target substance:
Using the obtained resin particles having a shell layer and a magnetic layer, an anti-KL-6 antibody was placed in the same manner as in Example 1. In this way, magnetic particles having no first magnetic substance were obtained.
　（比較例２）
　比較例１と同様にして、樹脂粒子を得た。次に、油性磁性流体（平均粒子径約１０ｎｍの四酸化三鉄ナノ粒子の有機溶剤分散液、フェローテック社製「ＥＸＰシリーズ」）にアセトンを加えて粒子を析出沈殿させた後、これを乾燥することにより、疎水化処理された表面を有するフェライト系の磁性体微粒子（平均一次粒子径：１０ｎｍ）を得た。 (Comparative Example 2)
Resin particles were obtained in the same manner as in Comparative Example 1. Next, acetone was added to an oil-based magnetic fluid (organic solvent dispersion of triiron tetroxide nanoparticles with an average particle diameter of about 10 nm, "EXP series" manufactured by Fellow Tech) to precipitate and precipitate the particles, and then dried. By doing so, ferrite-based magnetic fine particles (average primary particle diameter: 10 nm) having a hydrophobically treated surface were obtained.
　次いで、上記樹脂粒子１．０重量部と上記疎水化された表面を有するフェライト系の磁性体微粒子１．０重量部とをミキサーでよく混合した。得られた混合物を、ハイブリダイゼーションシステムＮＨＳ－０型（奈良機械製作所社製）を用いて、羽根（撹拌翼）の周速度１００ｍ／秒（１６２００ｒｐｍ）で５分間処理し、樹脂粒子の外表面上に第２の磁性体により磁性層を形成した。 Next, 1.0 part by weight of the resin particles and 1.0 part by weight of the ferritic magnetic fine particles having a hydrophobic surface were well mixed with a mixer. The obtained mixture was treated with a hybridization system NHS-0 (manufactured by Nara Machinery Co., Ltd.) at a peripheral speed of 100 m / sec (16200 rpm) for a blade (stirring blade) for 5 minutes on the outer surface of the resin particles. A magnetic layer was formed from the second magnetic material.
　シェル層の形成：
　得られた磁性層を有する粒子１０重量部と、分散剤としてノニオン性乳化剤（花王社製「エマルゲン１５０」）の０．５％水溶液３００重量部とを１Ｌセパラブルフラスコに投入し、撹拌した。これに、モノマーとして、シクロヘキシルメタクリレート２４重量部及び２－メタクリロイロキシエチルコハク酸６０重量部と、開始剤として、ジ（３，５，５－トリメチルヘキサノイル）パーオキサイド（日油社製「パーロイル３５５」）１．０重量部とを添加し、８０℃、２００ｒｐｍで８時間撹拌した。このようにして、該磁性層の外表面上にカルボキシル基を有する有機ポリマーシェル層を形成した。上記有機ポリマーは、シクロヘキシルメタクリレートと２－メタクリロイロキシエチルコハク酸との共重合体である。 Shell layer formation:
10 parts by weight of the obtained particles having a magnetic layer and 300 parts by weight of a 0.5% aqueous solution of a nonionic emulsifier (“Emulgen 150” manufactured by Kao Corporation) as a dispersant were put into a 1 L separable flask and stirred. To this, 24 parts by weight of cyclohexyl methacrylate and 60 parts by weight of 2-methacryloyloxyethyl succinic acid were used as monomers, and di (3,5,5-trimethylhexanoyl) peroxide (NOF Corporation "Perloyl") was used as an initiator. 355 ”) 1.0 part by weight was added, and the mixture was stirred at 80 ° C. and 200 rpm for 8 hours. In this way, an organic polymer shell layer having a carboxyl group was formed on the outer surface of the magnetic layer. The organic polymer is a copolymer of cyclohexyl methacrylate and 2-methacryloyloxyethyl succinic acid.
　目的物質と特異的に相互作用する物質の配置：
　得られたシェル層及び磁性層を有する樹脂粒子を用いて、実施例１と同様にして、抗ＫＬ－６抗体を配置した。このようにして、第１の磁性体を有さない磁性粒子を得た。 Arrangement of substances that specifically interact with the target substance:
Using the obtained resin particles having a shell layer and a magnetic layer, an anti-KL-6 antibody was placed in the same manner as in Example 1. In this way, magnetic particles having no first magnetic substance were obtained.
　（比較例３）
　樹脂粒子及び磁性粒子の構成を表１に示すように変更したこと以外は、比較例１と同様にして磁性粒子を得た。比較例３で得られた磁性粒子は、第１の磁性体を有さない。 (Comparative Example 3)
Magnetic particles were obtained in the same manner as in Comparative Example 1 except that the configurations of the resin particles and the magnetic particles were changed as shown in Table 1. The magnetic particles obtained in Comparative Example 3 do not have the first magnetic material.
　（比較例４）
　樹脂粒子及び磁性粒子の構成を表１に示すように変更したこと以外は、実施例１と同様にして、磁性粒子を作製した。比較例４で得られた磁性粒子は、磁性層を有さない。 (Comparative Example 4)
Magnetic particles were produced in the same manner as in Example 1 except that the configurations of the resin particles and the magnetic particles were changed as shown in Table 1. The magnetic particles obtained in Comparative Example 4 do not have a magnetic layer.
　（比較例５）
　樹脂粒子及び磁性粒子の構成を表１に示すように変更したこと以外は、比較例４と同様にして磁性粒子を得た。なお、比較例５では、シェル層として、ポリグリシジルメタクリレート（ＰＧＭＡ）を含むシェル層（有機ポリマーシェル層）を形成した。比較例５で得られた磁性粒子は、磁性層を有さない。 (Comparative Example 5)
Magnetic particles were obtained in the same manner as in Comparative Example 4, except that the configurations of the resin particles and the magnetic particles were changed as shown in Table 1. In Comparative Example 5, a shell layer (organic polymer shell layer) containing polyglycidyl methacrylate (PGMA) was formed as the shell layer. The magnetic particles obtained in Comparative Example 5 do not have a magnetic layer.
　（評価）
　（１）樹脂粒子及び磁性粒子の平均粒子径
　得られた樹脂粒子及び磁性粒子について、走査型電子顕微鏡（日立ハイテクノロジーズ社製「Ｒｅｇｕｌｕｓ８２２０」）を用いて、樹脂粒子及び磁性粒子の平均粒子径を算出した。具体的には、得られた磁性粒子を乾燥させた試料を作製し、得られた試料を走査型電子顕微鏡にて観察し、任意の５０個の磁性粒子、及び磁性粒子における樹脂粒子の粒子径を測定し、平均値を算出することにより求めた。 (Evaluation)
(1) Average Particle Diameter of Resin Particles and Magnetic Particles For the obtained resin particles and magnetic particles, the average particle diameter of the resin particles and magnetic particles was determined using a scanning electron microscope (“Regulus 8220” manufactured by Hitachi High-Technologies Corporation). Calculated. Specifically, a sample obtained by drying the obtained magnetic particles is prepared, the obtained sample is observed with a scanning electron microscope, and the particle diameters of any 50 magnetic particles and the resin particles in the magnetic particles are observed. Was measured and the average value was calculated.
　（２）樹脂粒子のＢＥＴ比表面積
　得られた樹脂粒子について、カンタクローム・インスツルメンツ社製「ＮＯＶＡ４２００ｅ」を用いて、窒素の吸着等温線を測定した。測定結果から、ＢＥＴ法に準拠して、樹脂粒子の比表面積を算出した。 (2) BET Specific Surface Area of Resin Particles With respect to the obtained resin particles, the adsorption isotherm of nitrogen was measured using "NOVA4200e" manufactured by Cantachrome Instruments. From the measurement results, the specific surface area of the resin particles was calculated according to the BET method.
　（３）樹脂粒子の平均細孔径
　得られた樹脂粒子について、カンタクローム・インスツルメンツ「ＮＯＶＡ４２００ｅ」を用いて、窒素の吸着等温線を測定した。測定結果から、ＢＪＨ法に準拠して、樹脂粒子の平均細孔径を算出した。 (3) Average Pore Diameter of Resin Particles With respect to the obtained resin particles, the adsorption isotherm of nitrogen was measured using Cantachrome Instruments "NOVA4200e". From the measurement results, the average pore size of the resin particles was calculated according to the BJH method.
　（４）第１の磁性体及び第２の磁性体の含有量
　磁性粒子の含有量が３０重量％となるように、Ｋｕｌｚｅｒ社製「テクノビット４０００」に添加し、分散させて、磁性粒子検査用埋め込み樹脂体を作製した。検査用埋め込み樹脂体中に分散した磁性粒子の中心付近を通るようにイオンミリング装置（日立ハイテクノロジーズ社製「ＩＭ４０００」）を用いて、磁性粒子の断面を切り出した。そして、電界放射型透過電子顕微鏡（日本電子社製「ＪＥＭ－２０１０ＦＥＦ」）を用いて、エネルギー分散型Ｘ線分析装置（ＥＤＳ）により、第１の磁性体及び第２の磁性体の含有量を測定し、磁性粒子の内部における第１の磁性体及び第２の磁性体の含有量の分布結果を得た。得られた結果から、第１の磁性体及び第２の磁性体の含有量を算出した。第１の磁性体及び第２の磁性体の含有量は、任意に選択された２０個の磁性粒子の第１の磁性体及び第２の磁性体の含有量を算術平均することにより算出した。 (4) Content of 1st magnetic substance and 2nd magnetic substance Add to "Technobit 4000" manufactured by Kulzer so that the content of magnetic particles is 30% by weight, disperse and inspect magnetic particles. An embedded resin body for use was prepared. A cross section of the magnetic particles was cut out using an ion milling device (“IM4000” manufactured by Hitachi High-Technologies Corporation) so as to pass near the center of the magnetic particles dispersed in the embedded resin body for inspection. Then, using an electric field radiation transmission electron microscope (“JEM-2010FEF” manufactured by JEOL Ltd.), the contents of the first magnetic substance and the second magnetic substance are measured by an energy dispersive X-ray analyzer (EDS). The measurement was performed to obtain the distribution results of the contents of the first magnetic substance and the second magnetic substance inside the magnetic particles. From the obtained results, the contents of the first magnetic substance and the second magnetic substance were calculated. The contents of the first magnetic material and the second magnetic material were calculated by arithmetically averaging the contents of the first magnetic material and the second magnetic material of 20 arbitrarily selected magnetic particles.
　また、磁性粒子の外表面から内側に向かって厚み１／３までの領域（Ｒ１）の１００体積％中、第１の磁性体の含有量を含有量（１）とした。磁性粒子の中心から外側に向かって厚み２／３までの領域（Ｒ２）の１００体積％中、第１の磁性体の含有量を含有量（２）とした。 Further, the content of the first magnetic substance was defined as the content (1) in 100% by volume of the region (R1) having a thickness of 1/3 from the outer surface to the inside of the magnetic particles. The content of the first magnetic substance was defined as the content (2) in 100% by volume of the region (R2) having a thickness of 2/3 from the center of the magnetic particles to the outside.
　磁性粒子の含有量が３０重量％となるように、Ｋｕｌｚｅｒ社製「テクノビット４０００」に添加し、分散させて、磁性粒子検査用埋め込み樹脂体を作製した。検査用埋め込み樹脂体中に分散した磁性粒子の中心付近を通るようにイオンミリング装置（日立ハイテクノロジーズ社製「ＩＭ４０００」）を用いて、磁性粒子の断面を切り出した。そして、電界放射型透過電子顕微鏡（日本電子社製「ＪＥＭ－２０１０ＦＥＦ」）を用いて、エネルギー分散型Ｘ線分析装置（ＥＤＳ）により、磁性体内包樹脂粒子の厚み方向における第１の磁性体の含有量を測定し、磁性体内包樹脂粒子の厚み方向における第１の磁性体の含有量の分布結果を得た。得られた結果から、上記含有量（１）及び上記含有量（２）を算出した。上記含有量（１）及び上記含有量（２）は、任意に選択された２０個の磁性粒子の含有量（１）及び含有量（２）を算術平均することにより算出した。 An embedded resin body for magnetic particle inspection was prepared by adding and dispersing it to "Technobit 4000" manufactured by Kulzer so that the content of magnetic particles was 30% by weight. A cross section of the magnetic particles was cut out using an ion milling device (“IM4000” manufactured by Hitachi High-Technologies Corporation) so as to pass near the center of the magnetic particles dispersed in the embedded resin body for inspection. Then, using an electric field radiation type transmission electron microscope (“JEM-2010FEF” manufactured by JEOL Ltd.), an energy dispersive X-ray analyzer (EDS) is used to obtain the first magnetic material in the thickness direction of the magnetic inclusion resin particles. The content was measured, and the distribution result of the content of the first magnetic substance in the thickness direction of the magnetic inclusion resin particles was obtained. From the obtained results, the above-mentioned content (1) and the above-mentioned content (2) were calculated. The content (1) and the content (2) were calculated by arithmetically averaging the content (1) and the content (2) of 20 arbitrarily selected magnetic particles.
　また、得られた結果から、上記含有量（１）の上記含有量（２）に対する比（含有量（１）／含有量（２））、及び上記含有量（１）の上記含有量（２）との差の絶対値を算出した。 Further, from the obtained results, the ratio of the content (1) to the content (2) (content (1) / content (2)) and the content (2) of the content (1). ) And the absolute value of the difference was calculated.
　（５）集磁率
　試料溶液として、波長５５０ｎｍにおける吸光度が０．９～１．１に調整された磁性粒子が水に分散した液を用意した。スペーサー（Ｗ１０ｍｍ×Ｄ１０ｍｍ×Ｈ４ｍｍ）を介し磁石（２８００Ｇ、Ｗ１０ｍｍ×Ｄ１０ｍｍ×Ｈ１ｍｍ）をあてた状態で分光光度計（日立製作所社製「Ｕ－３９００Ｈ」）に設置した石英セルに、試料溶液１．３ｍＬを投入し、試料溶液投入後５秒後～１２５秒後の波長５５０ｎｍにおける吸光度を測定した。この１２０秒間での吸光度減衰率を以下の式により算出し、集磁率とした。 (5) Magnetic Collection Rate As a sample solution, a solution in which magnetic particles whose absorbance at a wavelength of 550 nm was adjusted to 0.9 to 1.1 was dispersed in water was prepared. Sample solution 1. In a quartz cell installed in a spectrophotometer (“U-3900H” manufactured by Hitachi, Ltd.) with a magnet (2800G, W10mm × D10mm × H1mm) applied via a spacer (W10mm × D10mm × H4mm). 3 mL was added, and the absorbance at a wavelength of 550 nm was measured 5 to 125 seconds after the sample solution was added. The absorbance attenuation rate in 120 seconds was calculated by the following formula and used as the magnetic collection rate.
　集磁率（％）＝［｛（５秒後の吸光度）－（１２５秒後の吸光度）｝／（５秒後の吸光度）］×１００ Magnetic collection rate (%) = [{(absorbance after 5 seconds)-(absorbance after 125 seconds)} / (absorbance after 5 seconds)] x 100
　［集磁率の判定基準］
　○○：集磁率が８０％以上
　○：集磁率が６０％以上８０％未満
　△：集磁率が４０％以上６０％未満
　×：集磁率が４０％未満 [Criteria for magnetic collection rate]
○○: Magnetic collection rate of 80% or more ○: Magnetic collection rate of 60% or more and less than 80% Δ: Magnetic collection rate of 40% or more and less than 60% ×: Magnetic collection rate of less than 40%
　（６）分散率
　試料溶液として、波長５５０ｎｍにおける吸光度が０．９～１．１に調整された磁性粒子が水に分散した液を用意した。分光光度計（日立製作所社製「Ｕ－３９００Ｈ」）に設置した石英セルに、試料溶液１．３ｍＬを投入し、波長５５０ｎｍにおける吸光度を測定した。次いで、磁石（２８０００Ｇ、Ｗ４０ｍｍ×Ｄ４０ｍｍ×Ｈ１０ｍｍ）を用いて、上澄みの吸光度が０になるまで集磁した。その後、ボルテックスにより２０００ｒｐｍ、５秒間、磁性粒子を分散させて、波長５５０ｎｍにおける吸光度を測定した。集磁前の吸光度と、集磁及び分散後の吸光度とから、吸光度の変化率を以下の式により算出し、分散率とした。 (6) Dispersion rate As a sample solution, a solution in which magnetic particles whose absorbance at a wavelength of 550 nm was adjusted to 0.9 to 1.1 was dispersed in water was prepared. 1.3 mL of the sample solution was put into a quartz cell installed in a spectrophotometer (“U-3900H” manufactured by Hitachi, Ltd.), and the absorbance at a wavelength of 550 nm was measured. Next, a magnet (28000 G, W40 mm × D40 mm × H10 mm) was used to collect magnetism until the absorbance of the supernatant became zero. Then, the magnetic particles were dispersed by vortex at 2000 rpm for 5 seconds, and the absorbance at a wavelength of 550 nm was measured. From the absorbance before magnetic collection and the absorbance after magnetic collection and dispersion, the rate of change in absorbance was calculated by the following formula and used as the dispersion rate.
　分散率（％）＝｛（集磁及び分散後の吸光度）／（集磁前の吸光度）｝×１００ Dispersion rate (%) = {(absorbance after magnetic collection and dispersion) / (absorbance before magnetic collection)} x 100
　［分散率の判定基準］
　○○：分散率が９５％以上
　○：分散率が９０％以上９５％未満
　△：分散率が８５％以上９０％未満
　×：分散率が８５％未満 [Criteria for determining the dispersion rate]
○ ○: Dispersion rate is 95% or more ○: Dispersion rate is 90% or more and less than 95% Δ: Dispersion rate is 85% or more and less than 90% ×: Dispersion rate is less than 85%
　（７）集磁率（１カ月放置後）
　得られた磁性粒子を２５℃で１カ月放置した。放置後、（５）集磁率と同様の方法で集磁率を求めた。 (7) Magnetic collection rate (after leaving for 1 month)
The obtained magnetic particles were left at 25 ° C. for 1 month. After being left to stand, the magnetic collection coefficient was determined by the same method as in (5) magnetic collection rate.
　［集磁率（１カ月放置後）の判定基準］
　○○：集磁率が８０％以上
　○：集磁率が６０％以上８０％未満
　△：集磁率が４０％以上６０％未満
　×：集磁率が４０％未満 [Criteria for magnetic collection rate (after leaving for 1 month)]
○○: Magnetic collection rate of 80% or more ○: Magnetic collection rate of 60% or more and less than 80% Δ: Magnetic collection rate of 40% or more and less than 60% ×: Magnetic collection rate of less than 40%
　（８）免疫測定（磁性粒子の単位重量当たりの発光量）
　ＫＬ－６を含まない溶液（抗原未含有溶液）と、５０００Ｕ／ｍＬのＫＬ－６を含む溶液（抗原含有溶液）を用意した。抗原未含有溶液を用いた免疫測定における発光量と、抗原含有溶液を用いた免疫測定における発光量との差を求めることにより、磁性粒子と目的物質（ＫＬ－６）との結合能を評価した。具体的には、以下の手順で評価を行った。 (8) Immunometric measurement (amount of light emitted per unit weight of magnetic particles)
A solution containing no KL-6 (antigen-free solution) and a solution containing 5000 U / mL KL-6 (antigen-containing solution) were prepared. The binding ability between the magnetic particles and the target substance (KL-6) was evaluated by determining the difference between the amount of light emitted in the immunoassay using the antigen-free solution and the amount of light emitted in the immunoassay using the antigen-containing solution. .. Specifically, the evaluation was performed according to the following procedure.
　ルテニウム錯体標識抗ＫＬ－６抗体（二次抗体）の作製：
　ポリプロピレンチューブに抗ＫＬ－６抗体のＰＢＳ－１溶液（抗ＫＬ－６抗体濃度２．０ｍｇ／ｍＬ）０．５ｍＬを加え、次いでＲｕ－ＮＨＳ（１０ｍｇ／ｍＬ）を１３μＬ加えた。２５℃にて振動撹拌した後、Ｓｅｐｈａｄｅｘ　Ｇ２５カラムを用いて精製し、ルテニウム錯体標識抗ＫＬ－６抗体を得た。 Preparation of ruthenium complex-labeled anti-KL-6 antibody (secondary antibody):
To a polypropylene tube, 0.5 mL of a PBS-1 solution of anti-KL-6 antibody (anti-KL-6 antibody concentration 2.0 mg / mL) was added, and then 13 μL of Ru-NHS (10 mg / mL) was added. After vibrating and stirring at 25 ° C., purification was performed using a Sephadex G25 column to obtain a ruthenium complex-labeled anti-KL-6 antibody.
　免疫測定：
　電気化学発光免疫測定法を測定原理としたＥＣＬＩＡ自動分析装置（積水メディカル社製「ピコルミＩＩＩ」）を用いて、以下のようにして発光量の測定を行った。反応溶液（正常ウサギ血清を含む緩衝液）２００μＬに５０００Ｕ／ｍＬのＫＬ－６を含む溶液（抗原含有溶液）２０μＬを加えたのち、磁性粒子２５μＬを加えた。３０℃で９分間反応させたのち、ピコルミＢＦ洗浄液（１０ｍＭトリス緩衝液）を３５０μＬ加え、磁性粒子を磁石でトラップしながら３回洗浄した。次いで、１．０μｇ／ｍＬのルテニウム錯体標識抗ＫＬ－６抗体を含むルテニウム標識抗体含有液２００μＬを加え、３０℃で９分間反応させたのち、ピコルミＢＦ洗浄液（１０ｍＭトリス緩衝液）を３５０μＬ加え、磁性粒子を磁石でトラップしながら３回洗浄した。次いで、０．１Ｍトリプロピルアミンを含むピコルミ発光電解液３００μＬを加え、電極表面に送液し、磁性粒子に結合したルテニウム錯体の発光量を測定した。抗原未含有溶液を用いた免疫測定における発光量と、抗原含有溶液を用いた免疫測定における発光量との差Ｘを求めた。 Immune measurement:
The amount of luminescence was measured as follows using an ECLIA automatic analyzer (“Picormi III” manufactured by Sekisui Medical Co., Ltd.) based on the electrochemical luminescence immunoassay method. To 200 μL of the reaction solution (buffer solution containing normal rabbit serum), 20 μL of a solution containing 5000 U / mL KL-6 (antigen-containing solution) was added, and then 25 μL of magnetic particles were added. After reacting at 30 ° C. for 9 minutes, 350 μL of Picormi BF washing solution (10 mM Tris buffer) was added, and the magnetic particles were washed three times while being trapped with a magnet. Next, 200 μL of a ruthenium-labeled antibody-containing solution containing 1.0 μg / mL ruthenium complex-labeled anti-KL-6 antibody was added, and the mixture was reacted at 30 ° C. for 9 minutes, and then 350 μL of Picormi BF washing solution (10 mM Tris buffer) was added. The magnetic particles were washed three times while being trapped with a magnet. Next, 300 μL of a picormi luminescent electrolytic solution containing 0.1 M tripropylamine was added and sent to the electrode surface, and the amount of luminescence of the ruthenium complex bonded to the magnetic particles was measured. The difference X between the amount of luminescence in the immunoassay using the antigen-free solution and the amount of luminescence in the immunoassay using the antigen-containing solution was determined.
　［磁性粒子の単位重量当たりの発光量の判定基準］
　○○：差Ｘが１５００００以上
　○：差Ｘが１２５０００以上１５００００未満
　△：差Ｘが１０００００以上１２５０００未満
　×：差Ｘが１０００００未満 [Criteria for determining the amount of light emitted per unit weight of magnetic particles]
○ ○: Difference X is 150,000 or more ○: Difference X is 125,000 or more and less than 150,000 Δ: Difference X is 100,000 or more and less than 125,000 ×: Difference X is less than 100,000
　磁性粒子の構成及び結果を下記の表１，２に示す。 The composition and results of the magnetic particles are shown in Tables 1 and 2 below.
Figure JPOXMLDOC01-appb-T000001
Figure JPOXMLDOC01-appb-T000001
Figure JPOXMLDOC01-appb-T000002
Figure JPOXMLDOC01-appb-T000002
　１，１Ａ，１Ｂ，１Ｃ…磁性粒子
　２，２Ａ，２Ｂ，２Ｃ…磁性体内包樹脂粒子
　３，３Ａ，３Ｂ，３Ｃ…磁性層
　４，４Ｂ，４Ｃ…シェル層
　５，５Ａ，５Ｂ，５Ｃ…物質
　２１，２１Ａ，２１１Ｂ，２１１Ｃ…樹脂粒子
　２２，２２Ａ，２２Ｂ，２２１Ｃ，２２２Ｃ…第１の磁性体
　２１２Ｂ，２１２Ｃ…樹脂層 1,1A, 1B, 1C ... Magnetic particles 2,2A, 2B, 2C ... Magnetic encapsulating resin particles 3,3A, 3B, 3C ... Magnetic layer 4,4B, 4C ... Shell layer 5,5A, 5B, 5C ... Material 21,21A, 211B, 211C ... Resin particles 22,22A, 22B, 221C, 222C ... First magnetic material 212B, 212C ... Resin layer

Claims (11)

  1. 　目的物質と特異的に相互作用させるために用いられる磁性粒子であって、
    　内部に第１の磁性体を含む磁性体内包樹脂粒子と、
    　前記磁性体内包樹脂粒子の外表面上に配置され、かつ第２の磁性体を含む磁性層と、
    　前記磁性層の外表面側に担持されており、かつ前記目的物質と特異的に相互作用する物質とを備える、磁性粒子。     Magnetic particles used to specifically interact with a target substance.
    Magnetic encapsulating resin particles containing the first magnetic material inside,
    A magnetic layer arranged on the outer surface of the magnetic inclusion resin particles and containing a second magnetic substance,
    A magnetic particle that is supported on the outer surface side of the magnetic layer and includes a substance that specifically interacts with the target substance.
  2. 　前記第１の磁性体が、金属又は金属酸化物であり、
    　前記第２の磁性体が、金属又は金属酸化物である、請求項１に記載の磁性粒子。     The first magnetic material is a metal or a metal oxide.
    The magnetic particles according to claim 1, wherein the second magnetic substance is a metal or a metal oxide.
  3. 　前記磁性層の外表面上に配置されたシェル層を更に備え、
    　前記シェル層の材料が、無機酸化物又は有機ポリマーを含み、
    　前記シェル層と、前記物質とが結合している、請求項１又は２に記載の磁性粒子。     A shell layer arranged on the outer surface of the magnetic layer is further provided.
    The material of the shell layer contains an inorganic oxide or an organic polymer and contains.
    The magnetic particles according to claim 1 or 2, wherein the shell layer and the substance are bonded to each other.
  4. 　前記シェル層の材料が、前記無機酸化物を含み、
    　前記無機酸化物が、ケイ素原子、ゲルマニウム原子、チタン原子又はジルコニウム原子を有する無機酸化物である、請求項３に記載の磁性粒子。     The material of the shell layer contains the inorganic oxide and
    The magnetic particles according to claim 3, wherein the inorganic oxide is an inorganic oxide having a silicon atom, a germanium atom, a titanium atom or a zirconium atom.
  5. 　前記物質が、抗原又は抗体である、請求項１～４のいずれか１項に記載の磁性粒子。 The magnetic particle according to any one of claims 1 to 4, wherein the substance is an antigen or an antibody.
  6. 　前記物質が、アビジン又はストレプトアビジンである、請求項１～４のいずれか１項に記載の磁性粒子。 The magnetic particle according to any one of claims 1 to 4, wherein the substance is avidin or streptavidin.
  7. 　磁性粒子１００重量％中、前記第１の磁性体と前記第２の磁性体と合計の含有量が、１０重量％以上９５重量％以下である、請求項１～６のいずれか１項に記載の磁性粒子。 The method according to any one of claims 1 to 6, wherein the total content of the first magnetic substance and the second magnetic substance in 100% by weight of the magnetic particles is 10% by weight or more and 95% by weight or less. Magnetic particles.
  8. 　前記第１の磁性体と前記第２の磁性体の含有量の合計１００重量％中、前記第１の磁性体の含有量が、１０重量％以上９０重量％以下である、請求項１～７のいずれか１項に記載の磁性粒子。 Claims 1 to 7 in which the content of the first magnetic substance is 10% by weight or more and 90% by weight or less in a total of 100% by weight of the contents of the first magnetic substance and the second magnetic substance. The magnetic particle according to any one of the above items.
  9. 　前記磁性体内包樹脂粒子の外表面から中心に向かって厚み１／３までの領域の１００体積％中における前記第１の磁性体の含有量の、前記磁性体内包樹脂粒子の中心から外表面に向かって厚み２／３までの領域の１００体積％中における前記第１の磁性体の含有量に対する比が、０．８以上４．０以下である、請求項１～８のいずれか１項に記載の磁性粒子。 From the center to the outer surface of the magnetic inclusion resin particles, the content of the first magnetic substance in 100% by volume of the region from the outer surface to the center of the magnetic inclusion resin particles to the thickness 1/3. According to any one of claims 1 to 8, the ratio to the content of the first magnetic substance in 100% by volume of the region up to 2/3 of the thickness is 0.8 or more and 4.0 or less. The magnetic particles described.
  10. 　検査薬として用いられる、請求項１～９のいずれか１項に記載の磁性粒子。 The magnetic particle according to any one of claims 1 to 9, which is used as a test agent.
  11. 　請求項１～９のいずれか１項に記載の磁性粒子を含む、検査薬。 A test agent containing the magnetic particles according to any one of claims 1 to 9.
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