JP5147039B2 - Resin particles and method for producing the same - Google Patents

Description

  The present invention relates to resin particles, and more particularly to identifiable resin particles that can be suitably used for detection of biologically relevant substances such as DNA, RNA, and proteins.

Conventionally, particles (or beads) used for such a purpose are, as seen in, for example, Patent Document 1, a polymer fine particle in which nanoparticles are dyed with a fluorescent dye whose concentration has been changed in advance and this is bound to the surface. It is disclosed. The polymer microparticles can be identified by flow cytometry or the like. Patent Document 2 discloses particles in which fluorescent polymeric core particles are coated with a polymer layer containing a metal oxide.
However, the fluorescent dyes used in these particles are weak against sunlight as well as UV light used for excitation, and have the disadvantage that the substance collapses when exposed to light and the fluorescence intensity becomes weak. . Therefore, care must be taken not only during measurement but also during fading.

On the other hand, Patent Document 3, Patent Document 4, and Patent Document 5 disclose a technique using a semiconductor nanocrystal having a thickness of 1 to 10 nm as a fluorescent material. Semiconductor nanocrystals can be tuned to emit characteristic spectra by changing their composition and size, or size distribution. In addition, semiconductor nanocrystals have a sharp fluorescence spectrum, are easily identified for identification, and have a strong fluorescence intensity due to the quantum effect due to the nanosize. Furthermore, it is stable even during long-term measurement and storage.
Note that in the case of performing an immunological test, nucleic acid detection, or the like by binding a biological substance to particles, it is necessary to separate the bound substance from the released substance without being bound. However, these inventions do not show a composition for performing a specific separation.
In this regard, Non-Patent Document 1 discloses particles in which a semiconductor nanocrystal called Qdot (Quantum Dot) is coated on the surface of a magnetic bead (size: about 10 μm).

Japanese Patent No. 3468750 Japanese Patent No. 2558918 Special table 2003-523718 gazette Special Table 2003-53734 JP 2002-311027 A Sumisho Bioscience Co., Ltd. Materials: “Mosaic System” for Gene Expression Analysis (December 2004)

However, since the particles disclosed in Non-Patent Document 1 are semiconductor nanocrystals arranged on the particle surface by coating, the semiconductor nanocrystals are detected in a reaction process for detecting DNA, RNA, protein, etc. Due to dropping off or dissolution, problems such as failure to obtain predetermined fluorescence intensity and fluorescence spectrum may occur. There is a method of further coating the surface in order to prevent dropping or dissolution, but the manufacturing process increases and the cost increases.
Further, even in the multicoloring technique using two or more kinds of semiconductor nanocrystals having different sizes, it is difficult to provide a sufficient concentration only with the particle surface, and thus multicoloring is naturally limited.
An object of this invention is to provide the resin particle which prevents deterioration of fluorescence intensity, and is advantageous to multi-coloring, and the manufacturing method of this resin particle in view of said problem.

According to the present invention, the above problem has been solved by the means described in the following means [1] and [21]. It is described below together with [2] to [20], [22] and [23] which are preferred embodiments.
[1] Substantially spherical resin particles having a structure including nanocrystals that emit light by excitation energy,
[2] Substantially spherical resin particles having a structure including a nanocrystal that emits light by excitation energy and a magnetic material,
[3] The resin particles according to [1] or [2], wherein the average particle diameter of the resin particles is 1 to 100 μm,
[4] The resin particles according to any one of [1] to [3], wherein the density of the resin particles is 1 to 1.5 g / ml,
[5] The resin particle according to any one of [1] to [4], wherein the nanocrystal is a compound semiconductor composed of 1 to 10 nm.
[6] The resin particle according to any one of [1] to [5], wherein the emission wavelength of the nanocrystal is 350 nm to 900 nm.
[7] The resin particles according to any one of [1] to [6], wherein the nanocrystal is a core / shell type and the surface of the shell is protected with an organic group.
[8] The resin particles according to any one of [1] to [7], including two or more kinds of nanocrystals having different emission wavelengths,
[9] The resin particles according to any one of [1] to [8], wherein at least a part thereof transmits excitation energy,
[10] The resin particles according to any one of [1] to [9], wherein at least a part transmits the emission wavelength.
[11] The resin particle according to any one of [1] to [10], wherein a peak of an emission spectrum emitted from the resin by excitation energy is different from a peak of the emission spectrum of the nanocrystal.
[12] The resin particles according to any one of [1] to [11], wherein the excitation energy is ultraviolet light,
[13] The resin particles according to any one of [1] to [12], wherein the resin is a thermoplastic resin,
[14] The structure according to any one of [1] to [13], wherein the surface has at least one functional group selected from the group consisting of a carboxyl group, a sulfo group, a glycidyl group, a hydroxyl group, an amino group, and a thiol group. Resin particles,
[15] The resin particles according to [13], wherein the thermoplastic resin is a polyolefin copolymer having a carboxyl group in the side chain,
[16] The resin particles according to any one of [2] to [15], wherein the magnetic material is a ferromagnetic material,
[17] The resin particles according to any one of [2] to [16], wherein the magnetic material is a metal.
[18] The resin particles according to any one of [2] to [17], wherein the magnetic material includes at least one selected from the group consisting of iron, nickel, and cobalt,
[19] The resin particles according to any one of [2] to [18], wherein the magnetic substance has a size of 0.1 to 10 μm,
[20] The resin particles according to any one of [1] to [19] manufactured using a melt dispersion method,
[21] The method for producing resin particles according to any one of [1] to [19] using a melt dispersion method,
[22] The thermoplastic resin 1, the thermoplastic resin 2 incompatible with the thermoplastic resin 1, the nanocrystal or the magnetic material, or the nanocrystal and the magnetic particle are melt-kneaded at a temperature lower than the melting point of the nanocrystal and the magnetic particle. [1] thru | or including the process and the process of melt | dissolving in the developing solvent of the thermoplastic resin 2 below the melting temperature of the thermoplastic resin 1, and isolate | separating a substantially spherical resin particle from the solution of the thermoplastic resin 2 characterized by the above-mentioned. [19] The method for producing resin particles according to any one of the above,
[23] A step of producing a nanocrystal-dispersed thermoplastic resin (A) by melting and kneading the thermoplastic resin 1 and nanocrystals below the melting point of the nanocrystals, and (A) and the magnetic particles being below the melting points of the nanocrystals and magnetic particles. A step of producing a thermoplastic resin (B) including nanocrystals and magnetic particles by melt-kneading at a temperature of, a resin (B) and a thermoplastic resin 2 incompatible with the resin (B) below the melting point of the nanocrystals and magnetic material And a step of dissolving the substantially spherical resin particles from the solution of the thermoplastic resin 2 by dissolving in a developing solvent of the resin 2 at a temperature equal to or lower than the melting temperature of the thermoplastic resin 1. [2] to [19] The method for producing resin particles according to any one of [1] to [19].

  According to the resin particles of the present invention and the method for producing the same, it is possible to provide resin particles that are prevented from being deteriorated in fluorescence intensity, can be easily multicolored and can be identified.

This will be described in detail below.
(Structure of resin particles)
The resin particle of the present invention is a substantially spherical resin particle having a structure in which nanocrystals are included in the resin particle, and strongly emits light by excitation energy by various energy rays such as ultraviolet rays, electron beams, X-rays, and gamma rays. Is. Here, “substantially spherical” means that 95% or more of those having a shape factor defined by the ratio of the longest diameter to the shortest diameter of the projected sectional shape of the resin particles is 0.8 or more.
The resin particles of the present invention have a structure in which nanocrystals are included, and preferably have a dispersed structure, more preferably the nanocrystals are dispersed throughout the resin particles, and uniformly throughout the resin particles. More preferably, it is dispersed. The “nanocrystal” refers to a crystal having a crystal size in the nano order.
Further, for the purpose of identifying two or more kinds of resin particles, two or more kinds of nanocrystals having different concentrations and two or more kinds of nanocrystals having different emission wavelengths can be included.

The resin particles of the present invention preferably contain a magnetic material in addition to the above-mentioned nanocrystals because of the ease of handling in the detection processing step for DNA, protein and the like.
The magnetic material is preferably in the form of particles dispersed in resin particles. The size of the magnetic particles (the size of the magnetic material) is preferably 0.1 to 50 μm, more preferably 0.1 to 20 μm, and even more preferably 0.1 to 10 μm. It is preferable that the size of the magnetic particles is within the above range because the production is easy. Moreover, the size of 1/2 to 1/20 is preferable compared with a resin particle. Further, the number of magnetic particles contained in the resin particles is preferably about 1 to 10, more preferably about 1 to 3.
Here, as a magnetic body, any shape can be used, and spherical, rod-like, or tabular particles can be exemplified. The size of the magnetic particles means an average particle size, and the particle size means a diameter when a sphere equivalent to the volume of the magnetic particles is considered.

The size of the resin particles of the present invention is preferably 1 to 100 μm. When the size of the resin particles is 1 μm or more, handling is easy, and particularly when a magnetic material is contained, the operation by an external magnetic field is favorable, which is preferable. Moreover, it is preferable that the resin particle size is 100 μm or less because a favorable capture specific surface area can be obtained with respect to biologically related substances such as DNA and protein.
Here, the size of the resin particles means an average volume particle diameter, and the average volume particle diameter can be measured by a laser diffraction particle size distribution meter or the like.

The density of the resin particles of the present invention is preferably 1 to 1.5 g / ml. It is preferable for the density of the resin particles to be in the above-mentioned range since good floating properties can be obtained in an aqueous solution.
In particular, when the resin particles contain a magnetic material, the greater the content of the magnetic material, the more sensitive the operation to an external magnetic field, but at the same time the density increases, and when various treatments are performed in an aqueous solution, the resin particles There is a tendency for the floating nature of the to be impaired.
From the viewpoint of the floatability of the resin particles, the density is preferably 1 to 1.2 g / ml, and more preferably 1 to 1.05 g / ml.

It is preferable that at least a part of the resin particles of the present invention transmit excitation energy. In particular, when a magnetic material is contained, the excitation energy tends to be shielded. Therefore, it is preferable to reduce the content of the magnetic material and improve the permeability of the excitation energy.
It is preferable that at least a part of the resin particles of the present invention transmit the emission wavelength. In particular, when a magnetic material is contained, there is a tendency to shield the emission wavelength. Therefore, it is preferable to reduce the content of the magnetic material and improve the transmittance of the emission wavelength.

In the present invention, it is preferable that a functional group is provided on the surface of the resin particle from the viewpoint of capturing a biological substance such as DNA. Examples of the functional group include a carboxyl group, a sulfo group (—SO ₃ H), a glycidyl group, a hydroxyl group, an amino group, and a thiol group. In particular, a carboxyl group is preferably used.

(Nanocrystal)
The nanocrystal is preferably a semiconductor nanocrystal, and more preferably a compound semiconductor nanocrystal. Here, the compound semiconductor means a semiconductor made of a plurality of elements. In the present invention, it is preferable that the nanocrystal has a crystal size of 1 to 10 nm at which a quantum effect is manifested in that the emission wavelength is in the visible region. In the present invention, the nanocrystal is particularly preferably a compound semiconductor having a crystal size of 1 to 10 nm.

  In the present invention, the nanocrystal is preferably a nanocrystal that emits light in the range of 350 nm to 900 nm. It is preferable for the emission wavelength to be within the above-mentioned range since it can be detected by a commonly used measuring apparatus.

  Also, the smaller the variation in the size of one kind of crystal, the smaller the half width of the peak at the emission wavelength corresponding to that size, which is advantageous for the identification of many kinds of resin particles. That is, since the detection efficiency is improved when the number of colors is increased, it is preferable that the variation in the size of one type of crystal is small.

Semiconductor nanocrystals that emit light in the visible region include, but are not limited to, CdS, CdSe, CdTe, ZnSe, ZnTe, GaP, and GaAs.
In addition, examples of semiconductor crystals that emit light in the near infrared include InP, InAs, InSb, PbS, and PbSe, but are not limited thereto.
Examples of semiconductor nanocrystals that emit light from blue to near ultraviolet include, but are not limited to, ZnS and GaN.

The nanocrystal is preferably a core / shell type for the purpose of improving the excitation energy resistance, and the shell material is preferably larger in optical band gap than the core material for the purpose of improving the extraction efficiency of emitted light.
Specifically, a nanocrystal that emits energy in the visible (for example, CdS, CdSe, ZnSe, ZnTe, GaP, GaAs) or near infrared (for example, InP, InAs, InSb, PbS, PbSe) is used as the core material. Examples of the shell material include ZnS, GaN, and magnesium chalcogenides such as MgS, MgSe, and MgTe that emit energy in the ultraviolet region.
In the present invention, CdSe / ZnS or the like is preferably used.
Furthermore, the surface of the shell is preferably protected with an organic group for the purpose of uniform dispersion in the resin, and is particularly protected with a hydrophobic group such as trioctyl phosphate (TOPO) or trioctyl phosphine (TOP). Are preferably used.

  The content of the nanocrystals in the resin particles can be appropriately selected according to the application, and is not particularly limited. When resin particles are used for detection of biological substances, the content of nanocrystals is preferably 1 to 2,000 ppm, preferably 10 to 1,000 ppm, based on the weight of the entire resin particles of the present invention. Is more preferable, and 50 to 700 ppm is even more preferable. Depending on the detector used, the nanocrystal content is preferably within the above range, but it is preferable to use a semiconductor detector or the like because it is more than detection sensitivity and the output is often proportional to the content.

(Resin material)
According to the present invention, any resin material can be used without particular limitation as the resin material used for the substantially spherical resin particles, but the peak of the emission spectrum emitted from the resin by the excitation energy is the peak of the emission spectrum of the nanocrystal. It is preferable to use a different resin. Further, a resin that does not emit light in the visible region (wavelength 350 nm to 900 nm) by excitation energy is more preferable.

  As the excitation energy, ultraviolet rays are preferably used for convenience of use. As the ultraviolet rays, ultraviolet rays emitted from a low-pressure mercury lamp, a high-pressure mercury lamp, an ultraviolet laser, or the like are preferably used, and a low-pressure mercury lamp that is easy to handle is particularly preferably used.

  Moreover, it is preferable that it is a thermoplastic resin from a viewpoint of the manufacturing method (melt dispersion method) mentioned later. Examples of the thermoplastic resin include polyolefins such as various nylons, polyethylene, and polypropylene, polyolefin copolymers, ethylene methacrylic acid copolymers, polycarbonate, polystyrene, and PDMS. In particular, from the viewpoint of imparting a surface functional group, a polyolefin copolymer having a carboxyl group in the side chain is preferable, and for example, an ethylene methacrylic acid copolymer is preferably used.

(Magnetic material)
The magnetic material of the present invention is preferably a ferromagnetic material that can be easily operated with a magnet. Here, the ferromagnetic material means a magnetic material having spontaneous magnetization. Among the ferromagnetic materials, a material having a large spontaneous magnetization is preferable, a metal (including an intermetallic compound) is preferable, and iron, an alloy including iron, nickel, cobalt, and the like can be exemplified. Furthermore, magnetic oxides such as ferrite, particularly soft ferrite with little residual magnetization is preferred. Examples of the soft ferrite include NiMn ferrite and MnZn ferrite.

The magnetic substance is preferably added in an amount of 1 to 50% by weight, more preferably 5 to 20% by weight, based on the resin particles of the present invention.
It is preferable for the amount of magnetic substance added to be in the above-mentioned range since good resin particle density and permeability can be obtained.

(Melt dispersion method)
The resin particles of the present invention can be produced by a method for producing composite microspheres developed by one of the inventors (Japanese Patent Laid-Open No. 61-174229). In this method, the thermoplastic resin 1 constituting the target substantially spherical particles and the thermoplastic resin 2 that is not compatible with the thermoplastic resin 1 are melt-kneaded, and the spheroidized thermoplastic resin 1 is cooled to a melting temperature or lower to obtain the thermoplastic resin 2. Only the thermoplastic resin 1 dispersed and spheroidized in the developing solvent is separated and recovered from the solution of the thermoplastic resin 2.
The thermoplastic resin 2 of the present invention forms a continuous phase for dispersing the thermoplastic resin composition 1 in fine particles and is not compatible with the thermoplastic resin 1. “Not compatible” means that the composition does not have a solubility of 1% by weight or more at the heating temperature. The thermoplastic resin 2 is preferably not compatible with the thermoplastic resin 1 and is preferably a poor solvent. Here, the poor solvent refers to a solvent that reduces the solubility of the thermoplastic resin 1 when added to a solution containing the thermoplastic resin 1 at a predetermined temperature. In the present invention, the thermoplastic resin 2 may be a mixture of two or more thermoplastic resins, and is preferably a poor solvent for the thermoplastic resin 1 over a range of room temperature to heating temperature. In the present invention, the thermoplastic resin 2 is preferably used in a capacity of 0.5 to 5 times that of the thermoplastic resin 1.

  Examples of the thermoplastic resin 2 include polyalkylene oxides such as polyethylene oxide, polyethylene glycol, polyvinyl alcohol and derivatives thereof (acetalized products, etc.), polybutene, wax, natural rubber, and synthetic rubber such as polybutadiene and styrene / butadiene copolymer rubber. Petroleum resin and the like can be exemplified, and these can be used alone or in combination. Polyalkylene oxides having different degrees of polymerization are commercially available, and it is also preferable to combine these components as appropriate.

  When the melt dispersion method is applied to the production of the resin particles of the present invention, the nanocrystals are prepared so as to have a desired composition with respect to the thermoplastic resin 1, and the nanocrystals are similarly melt-dispersed below the melting point of the nanocrystals. Resin particles containing can be easily obtained. Further, when a magnetic substance is contained in addition to the nanocrystal, the nanocrystal is similarly prepared by charging the thermoplastic resin 1 so as to have a desired composition and melt-kneading the nanocrystal and the magnetic substance at a melting temperature or lower. And magnetic substance-containing resin particles are obtained. Furthermore, by classifying as necessary, resin particles having a uniform particle diameter can be obtained.

When the resin particles of the present invention are produced by the melt dispersion method, resin particles having uniform quality can be produced by the following improved method.
That is, first, the thermoplastic resin 1 and nanocrystals are melt-kneaded below the melting point of the nanocrystals to produce a thermoplastic resin (A) containing nanocrystals, and then (A) and the magnetic particles are combined with the nanocrystals and magnetic particles. A thermoplastic resin (B) containing nanocrystals and magnetic particles is produced by melt-kneading at a temperature below the melting point of the particles. Finally, the resin (B) and the thermoplastic resin 2 that is not compatible with the resin (B) are melt-kneaded at a temperature below the melting point of the nanocrystals and the magnetic material, and used as a developing solvent for the thermoplastic resin 2 below the melting temperature of the thermoplastic resin 1. By dissolving, the substantially spherical resin particles can be separated and recovered from the solution of the thermoplastic resin 2.

  The resin particles of the present invention can be used for various applications, and are preferably used for detection of biologically relevant substances such as DNA, RNA, and protein. Specifically, the surface of the resin particle is modified with a biological substance such as DNA, RNA, protein, etc., and the biological substance reacting with the modified substance can be detected.

[Example 1]
(Nanocrystal)
As the nanocrystal, a CdSe / ZnS core / shell type Evidot (manufactured by Evident Technologies) particle size of 2.4 nm was prepared. The surface of the nanocrystal is protected with TOPO (Tri-n-octylphosphine oxide) and dispersed in a toluene solvent. The emission color of this nanocrystal by excitation with ultraviolet light (excitation wavelength: 400 nm or less) was green (emission wavelength: 520 nm).
(Magnetic material)
As the magnetic material, commercially available iron particles (melting point: 1,500 ° C.) having a particle diameter of 2 μm produced from carbonyl iron were prepared.

(Dispersion of nanocrystals)
Ethylene methacrylic acid copolymer (N1860 manufactured by Mitsui DuPont Polychemical Co., Ltd .; ethylene: methacrylic acid = 82: 18, melting point 90, so that nanocrystals of green color dispersed in the toluene solvent have a concentration of 200 ppm. ° C) and melt-kneaded at 170 ° C to produce a nanocrystal-containing thermoplastic resin (A).

(Dispersion of magnetic material)
The iron particles were mixed with the thermoplastic resin (A) so as to be 10% by weight, and melt-kneaded at 170 ° C. to prepare a thermoplastic resin (B) containing nanocrystals and a magnetic material.

(Production of substantially spherical resin particles)
2 times the amount of polyethylene glycol as thermoplastic resin 2 that is not compatible with the thermoplastic resin (B) is added and melt-kneaded in a twin-screw extruder at 170 ° C., and then extruded into water to dissolve the polyethylene glycol. Then, approximately spherical resin particles containing nanocrystals and iron particles were centrifuged from the solution. Here, the solubility of the ethylene methacrylic acid copolymer in polyethylene glycol was 1% by weight or less, and there was no compatibility.
As a result of classifying the obtained resin particles using a JIS standard stainless steel sieve, substantially spherical resin particles having an average particle diameter of 20 μm to 25 μm were obtained.
In the obtained resin particles, 97% of the particles had a shape factor of 0.8 or more expressed by the ratio of the longest diameter to the shortest diameter of the projected cross-sectional area shape. In addition, the shape factor was measured with a micrograph for 100 resin particles.

[Example 2]
In Example 1, a nanocrystal (CdSe / ZnS, excitation wavelength: 400 nm or less, emission wavelength: 620 nm) having a particle size of 5.0 nm and a red emission color was prepared. Substantially spherical resin particles having an average particle diameter of red light emission of 20 μm to 25 μm were prepared in the same manner except that 200 ppm of this was added.
In the obtained resin particles, 98% of the particles had a shape factor of 0.8 or more expressed by the ratio of the longest diameter to the shortest diameter of the projected cross-sectional area shape.

[Example 3]
In Example 2, substantially spherical resin particles having an average particle diameter of 20 μm to 25 μm were prepared in the same manner except that 100 ppm of red light emitting nanoparticles were added.
In the obtained resin particles, 98% of the particles had a shape factor of 0.8 or more expressed by the ratio of the longest diameter to the shortest diameter of the projected cross-sectional area shape.

[Example 4]
In Example 1, instead of 200 ppm of green luminescent nanocrystals, the average particle size was the same as in Example 1 except that 100 ppm of green luminescent nanocrystals used in Example 1 and 100 ppm of red luminescent nanocrystals used in Example 2 were added. Produced substantially spherical resin particles having a particle diameter of 20 μm to 25 μm.
In the obtained resin particles, 97% of the particles had a shape factor of 0.8 or more expressed by the ratio of the longest diameter to the shortest diameter of the projected cross-sectional area shape.

(Evaluation of resin particles)
(1) Density of resin particles:
Measurement was performed using a glass pycnometer and isopropyl alcohol as a replacement liquid.

(2) Magnetization:
An aqueous dispersion of resin particles is prepared so as to have a particle concentration of about 10% by weight, and this is shaken and dispersed in a polystyrene container with a lid having a diameter of 15 mm. The state of being sucked into the tube wall was observed, and the case where it was sucked into the tube wall within a few seconds was considered good.

(3) Floating evaluation:
An aqueous dispersion of resin particles obtained to a particle concentration of about 10% by weight was prepared, and after shaking and dispersing in a 15 mm diameter lidded polystyrene container, no sedimentation was observed for more than 1 minute. The case was considered good.

(4) Fluorescence microscope observation:
About 5 μl of the dispersion prepared in (3) was dropped on a slide glass, and the surface was covered with a cover glass. An optical filter that cuts visible light was placed using a low-pressure mercury lamp as the light source, and the appearance of light emission when the resin particles were irradiated with ultraviolet rays was observed. The light emission color and the light emission uniformity in the particles were evaluated, and the case where light emission without spots was observed as a whole was regarded as uniform light emission.
Thereby, the inclusion property of a nanocrystal can be evaluated. Further, it can be evaluated that a part of the resin particles transmits excitation energy and emission wavelength.

(5) Luminous intensity of resin particles:
The Olympus microscope digital camera DP10 was installed in the fluorescence microscope of (4), and the resin particles of Example 2 and Example 3 were photographed under constant photographing conditions. Using the image analysis software “Image-Pro PLUS” manufactured by Nippon Roper, the average luminance in the resin particles was measured, and the relative value of Example 2 with respect to Example 3 was obtained.

(6) Measurement of emission peak wavelength:
A fluorescence spectrophotometer F7000 manufactured by Hitachi High-Technology Corporation was used for measurement of the emission peak wavelength of the resin particles. The excitation wavelength was fixed at 365 nm and the spectral wavelength was scanned from 400 to 900 nm.
(7) Evaluation of labeling:
The resin particles were labeled according to the following procedure. A digital cooled CCD camera “Cool SNAP” manufactured by Nippon Roper and an optical filter suitable for Alexa Fluor 647 (manufactured by Molecular PROBES) with an excitation wavelength of 650 nm and an emission wavelength of 665 nm are installed in the microscope of (4), and labeling is not performed. The case where discrimination was possible under a microscope compared with the light emission luminance of the original resin particles was considered good. Labeling was performed by the following method.
1) 50 μl was taken from a dispersed aqueous solution of resin particles (10 mg / ml), and the solvent was replaced with 125 μl of 100 mM MES, pH 6.0.
2) 100 μl of a 1.25 × 10 ⁻² mg / ml solution of BSA with Biotin (Arista Biologicals, Inc.) was added to 1).
3) A 26 mM MES solution of EDAC (N- (3-dimethylaminopropyl) -N′-ethylcarbodiimide hydrochloride: Sigma-Aldrich) was prepared, and 25 μl was added to 2). Thereafter, the mixture was stirred for 1 hour with a microtube rotator (manufactured by ASONE). As a result, BSA with Biotin was bound to the carboxyl group on the resin particle surface.
4) After stirring, the solvent was replaced with 250 μl of PBS (−).
5) 50 μl of the solution of 4) was collected, and 1 μl of 50 μg / μl solution of Alexa Fluor 647 (manufactured by Molecular PROBES) with streptavidin was added and stirred. After 5 minutes, the solvent was replaced with 250 μl of PBS (−).

  The evaluation results are shown in Table 1. In addition, light emission from the used ethylene methacrylic acid copolymer was not recognized in the measurement range of 400 nm to 900 nm.

Claims (22)

It is a substantially spherical resin particle having a structure including nanocrystals that emit light by excitation energy ,
The resin is a thermoplastic resin;
The thermoplastic resin is a polyolefin copolymer having a carboxyl group in the side chain.
Resin particles . It is a substantially spherical resin particle having a structure including nanocrystals and magnetic materials that emit light by excitation energy ,
The resin is a thermoplastic resin;
The thermoplastic resin is a polyolefin copolymer having a carboxyl group in the side chain.
Resin particles .   The resin particles according to claim 1 or 2, wherein the resin particles have an average particle diameter of 1 to 100 µm.   The resin particles according to any one of claims 1 to 3, wherein the density of the resin particles is 1 to 1.5 g / ml.   The resin particle according to claim 1, wherein the nanocrystal is a compound semiconductor composed of 1 to 10 nm.   The resin particle according to any one of claims 1 to 5, wherein the emission wavelength of the nanocrystal is 350 nm to 900 nm.   The resin particle according to any one of claims 1 to 6, wherein the nanocrystal is of a core / shell type and the surface of the shell is protected with an organic group.   The resin particle according to any one of claims 1 to 7, comprising two or more kinds of nanocrystals having different emission wavelengths.   The resin particle according to any one of claims 1 to 8, wherein at least a part thereof transmits excitation energy.   The resin particle according to any one of claims 1 to 9, wherein at least a part thereof transmits an emission wavelength.   The resin particle according to any one of claims 1 to 10, wherein a peak of an emission spectrum emitted from the resin by excitation energy is different from a peak of an emission spectrum of the nanocrystal.   The resin particle according to any one of claims 1 to 11, wherein the excitation energy is ultraviolet light. The resin particle according to any one of claims 1 to 12, which has at least one functional group selected from the group consisting of a carboxyl group, a sulfo group, a glycidyl group, a hydroxyl group, an amino group, and a thiol group on the surface. Resin particles according to the magnetic material of any one claims 2 to 13 of a ferromagnetic material. Resin particles according to the magnetic material of any one claims 2 to 14 which is a metal. The resin particle according to any one of claims 2 to 15 , wherein the magnetic body contains at least one selected from the group consisting of iron, nickel, and cobalt. The resin particle according to any one of claims 2 to 16, wherein the magnetic substance has a size of 0.1 to 10 µm.   The resin particle according to any one of claims 1 to 17, wherein the polyolefin copolymer having a carboxyl group in a side chain is an ethylene methacrylic acid copolymer. The resin particle according to any one of claims 1 to 18, which is produced by using a melt dispersion method. The method for producing resin particles according to any one of claims 1 to 18, wherein a melt dispersion method is used. Melt-kneading the thermoplastic resin 1, the thermoplastic resin 2 incompatible with the thermoplastic resin 1, nanocrystals or magnetic bodies, or nanocrystals and magnetic particles at a temperature below the melting point of the nanocrystals and magnetic particles; and any claims 1 to 18, characterized in that it comprises a step of separating the resin particles of substantially spherical shape was dissolved in a developing solvent of thermoplastic resin 2 in the following melting temperature of the thermoplastic resin 1 from a solution of a thermoplastic resin 2 The manufacturing method of the resin particle as described in one. A step of producing a nanocrystal-dispersed thermoplastic resin (A) by melt-kneading the thermoplastic resin 1 and nanocrystals below the melting point of the nanocrystals;
(A) and magnetic particles are melt-kneaded at a temperature below the melting point of the nanocrystals and magnetic particles to produce a thermoplastic resin (B) including the nanocrystals and magnetic particles,
A step of melt-kneading the resin (B) and the thermoplastic resin 2 incompatible with the resin at a temperature lower than the melting point of the nanocrystal and the magnetic material, and dissolving in the developing solvent of the resin 2 at a temperature lower than the melting temperature of the thermoplastic resin 1 method for producing resinous particles according to the resin particles of substantially spherical to any one of claims 2 to 18, characterized in that it comprises a step of separating from a solution of a thermoplastic resin 2 Te.