Monomodal narrow-band light emitting microspheres and applications thereof

Abstract

The application belongs to the technical field of luminescence, and discloses a single-peak narrow-band luminescent microsphere and application thereof. The single-peak narrow-band luminescent microsphere comprises organic luminescent molecules, and the diameter of the single-peak narrow-band luminescent microsphere is in the range of nanometer to micrometer. The single-peak narrow-band luminescent microsphere can be used for preparing a homogeneous detection reagent, and the sensitivity of homogeneous detection can be improved. The single-peak narrow-band luminescent microsphere can also be used for preparing an optical probe, a flow detection reagent, a biological marker, an immunochromatographic detection reagent, an immunochromatographic kit and a radionuclide drug.

Description

Technical Field

[0001] This invention belongs to the field of luminescence technology and relates to a single-peak narrowband luminescent microsphere and its applications. Background Technology

[0002] Homogeneous detection is a detection technique that can be performed directly under pure liquid phase conditions without separation and washing steps, and it has attracted much attention in many fields such as basic medical research, immunoassay, biomarker detection, and drug screening. Luminescent materials are key materials in homogeneous detection technology.

[0003] Luminescent materials used in homogeneous detection include quantum dot luminescent materials, rare-earth-doped luminescent materials, and organic luminescent microspheres. Quantum dots, typically only 2-20 nm in size, present numerous challenges for biological applications due to their extremely small size. These challenges include difficulty in effectively encapsulating them into structurally stable and functionally defined carriers, a lack of diversity in surface functional groups, and potential toxicity. Rare-earth luminescent materials often exhibit multiple emission peaks, which are difficult to completely separate, displaying a so-called "multi-peak" characteristic that fails to meet the requirements for high-resolution emission spectra and single-wavelength output. Organic luminescent microspheres, comprising a microsphere carrier and organic luminescent molecules, offer advantages such as simplified operation and enhanced specificity in homogeneous detection. For example, the suspension properties of organic luminescent microspheres allow for uniform dispersion in the liquid phase, facilitating high-throughput sample loading and detection using automated pipetting equipment. Organic luminescent microspheres can also achieve precise identification of target analytes through surface modification with specific biomolecules. However, the sensitivity of existing organic luminescent microspheres for homogeneous detection still needs improvement.

[0004] Therefore, there is an urgent need in this field for an organic light-emitting microsphere that can improve the sensitivity of homogeneous detection. Summary of the Invention

[0005] This invention addresses the aforementioned problems in existing technologies by proposing a single-peak narrow-band luminescent microsphere and its applications. The single-peak narrow-band luminescent microsphere of this invention comprises organic luminescent molecules, and its diameter is in the sub-micrometer to micrometer range. This invention uses a small organic molecule single-peak narrow-band emitter as the luminescent center, covalently bonding or physically encapsulating it within the microsphere to form a spherical microstructure with nanometer to micrometer dimensions, suitable for applications in the biological field, while maintaining its narrow-band and single-peak emission characteristics. The single-peak narrow-band luminescent microsphere of this invention can improve the sensitivity of homogeneous detection and also shows promising application prospects in the preparation of optical probes, flow cytometry reagents, biomarkers, immunochromatographic reagents, immunochromatographic kits, and radiopharmaceuticals.

[0006] Specifically, the present invention provides a single-peak narrowband luminescent microsphere, the single-peak narrowband luminescent microsphere comprising organic luminescent molecules, the diameter of the single-peak narrowband luminescent microsphere being 50 nm to 500 μm, and the half-width at half-maximum (WHM) of the emission spectrum of the single-peak narrowband luminescent microsphere being no greater than 35 nm.

[0007] In one or more embodiments, the peak width at 1% intensity of the highest peak of the emission spectrum of the single-peak narrowband luminescent microsphere is no greater than 150 nm. In one or more embodiments, the peak width at 1% intensity of the highest peak of the emission spectrum of the single-peak narrowband luminescent microsphere is no greater than 130 nm.

[0008] In one or more embodiments, the emission spectrum of the single-peak narrowband luminescent microsphere does not exhibit double or more peaks at an intensity greater than or equal to 5% of the highest peak value.

[0009] In one or more embodiments, the luminescence lifetime of the single-peak narrowband luminescent microsphere is 1 ns to 999 ns.

[0010] In one or more embodiments, the organic light-emitting molecule is a narrow-band organic light-emitting molecule, and the full width at half maximum (FWHM) of the emission spectrum of the narrow-band organic light-emitting molecule is not greater than 30 nm.

[0011] In one or more embodiments, the peak width at 1% intensity of the highest peak of the emission spectrum of the narrow-band organic light-emitting molecule is no greater than 140 nm.

[0012] In one or more embodiments, under external stimulation, the half-width at half maximum (WHM) of the emission spectrum of the single-peak narrowband luminescent microsphere is no greater than 35 nm, and the peak width at 1% intensity of the highest peak is no greater than 150 nm.

[0013] In one or more embodiments, the single-peak narrowband luminescent microspheres further include a component that is responsive to the external stimulus.

[0014] In one or more embodiments, under external stimulation, the half-width at half maximum (WHM) of the emission spectrum of the narrow-band organic light-emitting molecule is no greater than 30 nm, and the peak width at 1% intensity of the highest peak is no greater than 140 nm.

[0015] In one or more embodiments, the external stimulus includes at least one selected from light, chemical reaction, X-ray, ultrasound, radioactive irradiation, thermal stimulation, and mechanical stimulation, and the single-peak narrow-band luminescent microsphere emits light under the external stimulus.

[0016] In one or more embodiments, the external stimulus is illumination, and the single-peak narrow-band luminescent microspheres produce fluorescence under illumination. In one or more embodiments, the external stimulus is illumination, and the single-peak narrow-band luminescent microspheres produce long-afterglow luminescence under illumination. In one or more embodiments, the external stimulus is a chemical reaction, and the single-peak narrow-band luminescent microspheres produce chemiluminescence under the chemical reaction.

[0017] In one or more embodiments, the organic light-emitting molecule contains only a single light group, the light group contains a conjugated π-electron system, and the organic light-emitting molecule has a planar rigid structure.

[0018] In one or more embodiments, the conjugated π-electron system has at least one substituent, and there is a mixed steric hindrance effect among the substituents.

[0019] In one or more embodiments, at least one substituent is present on the conjugated π-electron system, the conformational free energy A of the substituent being ≥1.7 kcal / mol.

[0020] In one or more embodiments, at least one substituent is present on the conjugated π-electron system, and the conformational free energy A of the substituent is ≥4 kcal / mol.

[0021] In one or more embodiments, at least one substituent is present on the conjugated π-electron system, the substituent being selected from halogens, straight-chain alkyl, branched alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, cycloalkylene, heterocycloalkylene, cycloalkenyl, heterocycloalkenyl, alkoxy, alkylsilyl, aryl, alkyl-substituted aryl, alkoxy-substituted aryl, aryl-substituted aryl, fluoroalkyl-substituted aryl, fluoroaryl-substituted aryl, aralkyl, aryloxy, heteroaryl, alkyl-substituted heteroaryl, alkoxy-substituted Heteroaryl, aryl-substituted heteroaryl, fluoroalkyl-substituted heteroaryl, fluoroaryl-substituted heteroaryl, cyano, esters and their derivatives, alkyl ketones, alkyl esters, aryl esters, alkynyl, alkylamine, fluoroalkyl, fluoroaryl, polyalkylene, aniline-substituted fluorenyl, diphenylamino-substituted fluorenyl, triphenylamino-substituted fluorenyl, alkyl-substituted triphenylamino, alkyl-substituted diphenylamino, alkyl-substituted aniline, alkyl-substituted thiophenyl, borate ester, hydroxyl, amino, nitro and trimethylsilylphenoxy.

[0022] In this invention, the aryl group can be selected from phenyl and fluorenyl.

[0023] In this invention, the heteroaryl group can be selected from pyridyl, bipyridyl, tripyridyl, furanyl, thiophenyl, pyrroleyl, pyrazolyl, oxazolyl, thiazolyl, imidazoleyl, pyrazinyl, benzoxazolyl, benzothiadiazolyl, and carbazoleyl.

[0024] In one or more embodiments, the conjugated π-electron system has at least two substituents selected from a first substituent, a second substituent, a third substituent, and a fourth substituent. In one or more embodiments, there is a mixed steric hindrance effect among the substituents.

[0025] In one or more embodiments, the first substituent includes C6-C18 straight-chain alkyl, C6-C18 heteroalkyl, polyalkylene, C6-C18 ester and its derivatives, alkyl ketone, alkyl ester, alkylamine, borate ester, hydroxyl, amino and nitro groups.

[0026] In one or more embodiments, the second substituent includes C4-C8 branched alkyl, aryl, alkyl-substituted aryl, aryl-substituted aryl, fluoroalkyl-substituted aryl, fluoroaryl-substituted aryl, aralkyl, aryloxy, heteroaryl, aniline-substituted fluorenyl, diphenylamino-substituted fluorenyl, triphenylamino-substituted fluorenyl, alkyl-substituted triphenylamino, alkyl-substituted diphenylamino, alkyl-substituted aniline, C6-C18 cycloalkyl, heterocycloalkyl, cycloalkenyl, heterocycloalkenyl, and aryl esters.

[0027] In one or more embodiments, the third substituent includes hydrogen, halogen, C1-C5 straight-chain alkyl, C3 branched alkyl, C3-C5 cycloalkyl, C1-C6 alkoxy, cyano, and C2-C5 alkynyl.

[0028] In one or more embodiments, the fourth substituent includes fluoroalkyl, fluoroaryl, alkyl-substituted heteroaryl, alkoxy-substituted heteroaryl, aryl-substituted heteroaryl, fluoroalkyl-substituted heteroaryl, fluoroaryl-substituted heteroaryl, heterocyclic alkylene, cycloalkylene, alkyl-substituted thiophenyl, alkoxy-substituted aryl, alkylsilyl, and trimethylsilylphenoxy.

[0029] In this invention, the heteroaryl group can be selected from polycyclic pyridyl and fused-ring heteroaryl groups. In this invention, the C6-C18 cycloalkyl group can be adamantyl. In this invention, the alkylsilyl group can be triisopropylsilyl.

[0030] In one or more embodiments, the narrow-band organic light-emitting molecule has the structure shown in Formula I or Formula II: Formula I, Formula II; In Formula I and Formula II, each X is independently selected from fluorine, oxygen, cyano and fluoroalkoxy groups; In formula II, and Each is an independent structural unit, existing or not existing; when When it exists, it may be optionally replaced by one or more R2s; when When present, it may be optionally substituted by one or more R2'; in Formula II, n1 and n2 each independently represent zero or one or more groups; In Formulas I and II, R1, R2, R2′, R3, R3′, R4, and R4′ are independently selected from hydrogen, halogen, straight-chain alkyl, branched alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, cycloalkylene, heterocycloalkylene, cycloalkenyl, heterocycloalkenyl, alkoxy, alkylsilyl, aryl, alkyl-substituted aryl, alkoxy-substituted aryl, aryl-substituted aryl, fluoroalkyl-substituted aryl, fluoroaryl-substituted aryl, aralkyl, aryloxy, heteroaryl, alkyl-substituted heteroaryl, alkoxy-substituted Heteroaryl, aryl-substituted heteroaryl, fluoroalkyl-substituted heteroaryl, fluoroaryl-substituted heteroaryl, cyano, esters and their derivatives, alkyl ketones, alkyl esters, aryl esters, alkynyl, alkylamine, fluoroalkyl, fluoroaryl, polyalkylene, aniline-substituted fluorenyl, diphenylamino-substituted fluorenyl, triphenylamino-substituted fluorenyl, alkyl-substituted triphenylamino, alkyl-substituted diphenylamino, alkyl-substituted aniline, alkyl-substituted thiophenyl, borate ester, hydroxyl, amino, nitro, and trimethylsilylphenoxy.

[0031] In one or more embodiments, in Formula I, R1, R2, R2′, R3, R3′, R4, and R4′ are each independently selected from at least two substituents selected from a first substituent, a second substituent, a third substituent, and a fourth substituent; in Formula II, R1, R2, R2′, R4, and R4′ are each independently selected from at least two substituents selected from a first substituent, a second substituent, a third substituent, and a fourth substituent. The first substituent includes C6-C18 straight-chain alkyl, C6-C18 heteroalkyl, polyalkylene, C6-C18 esters and their derivatives, alkyl ketones, alkyl esters, alkylamines, borate esters, hydroxyl groups, amino groups, and nitro groups; the second substituent includes C4-C8 branched alkyl, aryl, alkyl-substituted aryl, aryl-substituted aryl, fluoroalkyl-substituted aryl, fluoroaryl-substituted aryl, aralkyl, aryloxy, heteroaryl, aniline-substituted fluorenyl, diphenylamino-substituted fluorenyl, triphenylamino-substituted fluorenyl, alkyl-substituted triphenylamino, alkyl-substituted diphenylamino, and alkyl-substituted aniline. The group includes C6-C18 cycloalkyl, heterocycloalkyl, cycloalkenyl, heterocycloalkenyl, and aryl esters; the third substituent includes hydrogen, halogen, C1-C5 straight-chain alkyl, C3 branched alkyl, C3-C5 cycloalkyl, C1-C6 alkoxy, cyano, and C2-C5 alkynyl; the fourth substituent includes fluoroalkyl, fluoroaryl, alkyl-substituted heteroaryl, alkoxy-substituted heteroaryl, aryl-substituted heteroaryl, fluoroalkyl-substituted heteroaryl, fluoroaryl-substituted heteroaryl, heterocyclic alkylene, cycloalkylene, alkyl-substituted thiophenyl, alkoxy-substituted aryl, alkylsilyl, and trimethylsilylphenoxy.

[0032] In one or more embodiments, the narrow-band organic light-emitting molecule has a structure shown in any one of Formula I (a1) to Formula I (a6): Formula I (a1), Equation I (a2), Equation I (a3), Equation I (a4), Formula I (a5), Formula I (a6); In formulas I(a1) to I(a6), n1, n2, n3, and n4 each independently represent zero or one or more groups; in formulas I(a1) to I(a6), R1′, R2, R2′, R3, R3′, R4, R4′, and R′ a 、R′ b 、R′ c and R′ d Each is independently selected from hydrogen, halogen, straight-chain alkyl, branched alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, cycloalkylene, heterocycloalkylene, cycloalkenyl, heterocycloalkenyl, alkoxy, alkylsilyl, aryl, alkyl-substituted aryl, alkoxy-substituted aryl, aryl-substituted aryl, fluoroalkyl-substituted aryl, fluoroaryl-substituted aryl, aralkyl, aryloxy, heteroaryl, alkyl-substituted heteroaryl, alkoxy-substituted heteroaryl, aryl-substituted heteroaryl, fluoroalkyl Alkyl-substituted heteroaryl, fluoroaryl-substituted heteroaryl, cyano, esters and their derivatives, alkyl ketones, alkyl esters, aryl esters, alkynyl, alkylamine, fluoroalkyl, fluoroaryl, polyalkylene, aniline-substituted fluorenyl, diphenylamino-substituted fluorenyl, triphenylamino-substituted fluorenyl, alkyl-substituted triphenylamino, alkyl-substituted diphenylamino, alkyl-substituted aniline, alkyl-substituted thiophenyl, borate ester, hydroxyl, amino, nitro and trimethylsilylphenoxy.

[0033] In one or more embodiments, in Formula I (a1), R1′, R2, R2′, R3, R3′, R4, and R4′ are each independently selected from at least two substituents selected from the first substituent, the second substituent, the third substituent, and the fourth substituent; in Formula I (a2), R2, R2′, R3, R3′, R4, R4′, and R′ are each selected from the first substituent, the second substituent, the third substituent, and the fourth substituent. a 、R′ b Each of the following substituents is independently selected from at least two substituents: a first substituent, a second substituent, a third substituent, and a fourth substituent; in formula I (a3), R2, R2′, R3, R3′, R4, R4′, and R′ a 、R′ b 、R′ cEach of the following substituents is independently selected from at least two substituents: a first substituent, a second substituent, a third substituent, and a fourth substituent; in formula I (a4), R2, R2′, R3, R3′, R4, R4′, and R′ a 、R′ b 、R′ c and R′ d Each of the following substituents is independently selected from at least two substituents: a first substituent, a second substituent, a third substituent, and a fourth substituent; in formula I (a5), R2, R2′, R3, R3′, R4, R4′, and R′ a 、R′ b Each of the following substituents is independently selected from at least two substituents: a first substituent, a second substituent, a third substituent, and a fourth substituent; in formula I (a6), R2, R2′, R3, R3′, R4, R4′, and R′ a 、R′ b 、R′ c Each of the substituents is independently selected from at least two substituents selected from the first substituent, the second substituent, the third substituent, and the fourth substituent; The first substituent includes C6-C18 straight-chain alkyl, C6-C18 heteroalkyl, polyalkylene, C6-C18 esters and their derivatives, alkyl ketones, alkyl esters, alkylamines, borate esters, hydroxyl groups, amino groups, and nitro groups; the second substituent includes C4-C8 branched alkyl, aryl, alkyl-substituted aryl, aryl-substituted aryl, fluoroalkyl-substituted aryl, fluoroaryl-substituted aryl, aralkyl, aryloxy, heteroaryl, aniline-substituted fluorenyl, diphenylamino-substituted fluorenyl, triphenylamino-substituted fluorenyl, alkyl-substituted triphenylamino, alkyl-substituted diphenylamino, and alkyl-substituted aniline. The group includes C6-C18 cycloalkyl, heterocycloalkyl, cycloalkenyl, heterocycloalkenyl, and aryl esters; the third substituent includes hydrogen, halogen, C1-C5 straight-chain alkyl, C3 branched alkyl, C3-C5 cycloalkyl, C1-C6 alkoxy, cyano, and C2-C5 alkynyl; the fourth substituent includes fluoroalkyl, fluoroaryl, alkyl-substituted heteroaryl, alkoxy-substituted heteroaryl, aryl-substituted heteroaryl, fluoroalkyl-substituted heteroaryl, fluoroaryl-substituted heteroaryl, heterocyclic alkylene, cycloalkylene, alkyl-substituted thiophenyl, alkoxy-substituted aryl, alkylsilyl, and trimethylsilylphenoxy.

[0034] In one or more embodiments, in formula I (a1), R1′ is each independently selected from H, phenyl and C1-C4 alkyl-substituted phenyl, and R2, R2′, R3, R3′, R4, R4′ are each independently selected from H and C1-C4 alkyl.

[0035] In one or more embodiments, the narrow-band organic light-emitting molecule has formula I (a 1-1 The structure shown is as follows: Ⅰ(a 1-1 ); Formula I (a) 1-1 In equation (a), n1, n2, n3, and n4 independently represent zero, one, or more groups; 1-1 In this context, R1, R2, R3, R4, R5, R1′, R2′, R3′, R4′, and R5′ are each independently selected from hydrogen, halogen, straight-chain alkyl, branched-chain alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, cycloalkylene, heterocycloalkylene, cycloalkenyl, heterocycloalkenyl, alkoxy, alkylsilyl, aryl, alkyl-substituted aryl, alkoxy-substituted aryl, aryl-substituted aryl, fluoroalkyl-substituted aryl, fluoroaryl-substituted aryl, aralkyl, aryloxy, heteroaryl, alkyl-substituted heteroaryl, alkoxy Alkyl-substituted heteroaryl, aryl-substituted heteroaryl, fluoroalkyl-substituted heteroaryl, fluoroaryl-substituted heteroaryl, cyano, esters and their derivatives, alkyl ketones, alkyl esters, aryl esters, alkynyl, alkylamine, fluoroalkyl, fluoroaryl, polyalkylene, aniline-substituted fluorenyl, diphenylamino-substituted fluorenyl, triphenylamino-substituted fluorenyl, alkyl-substituted triphenylamino, alkyl-substituted diphenylamino, alkyl-substituted aniline, alkyl-substituted thiophenyl, borate ester, hydroxyl, amino, nitro, and trimethylsilylphenoxy.

[0036] In one or more embodiments, formula I (a 1-1 In ), R 1 R 2 R 3 R 4 Each of the substituents is independently selected from at least two substituents selected from the first, second, third, and fourth substituents; the first substituent includes C6-C18 straight-chain alkyl, C6-C18 heteroalkyl, polyalkylene, C6-C18 esters and their derivatives, alkyl ketones, alkyl esters, alkylamines, borate esters, hydroxyl groups, amino groups, and nitro groups; the second substituent includes C4-C8 branched alkyl, aryl, alkyl-substituted aryl, aryl-substituted aryl, fluoroalkyl-substituted aryl, fluoroaryl-substituted aryl, aralkyl, aryloxy, heteroaryl, aniline-substituted fluorenyl, diphenylamino-substituted fluorenyl, triphenylamino-substituted fluorenyl, and alkyl-substituted triphenylene. The group includes amino, alkyl-substituted diphenylamino, alkyl-substituted aniline, C6-C18 cycloalkyl, heterocycloalkyl, cycloalkenyl, heterocycloalkenyl, and aryl esters; the third substituent includes hydrogen, halogen, C1-C5 straight-chain alkyl, C3 branched alkyl, C3-C5 cycloalkyl, C1-C6 alkoxy, cyano, and C2-C5 alkynyl; the fourth substituent includes fluoroalkyl, fluoroaryl, alkyl-substituted heteroaryl, alkoxy-substituted heteroaryl, aryl-substituted heteroaryl, fluoroalkyl-substituted heteroaryl, fluoroaryl-substituted heteroaryl, heterocyclic alkylene, cycloalkylene, alkyl-substituted thiophenyl, alkoxy-substituted aryl, alkylsilyl, and trimethylsilylphenoxy.

[0037] In one or more embodiments, formula I (a 1-1 In ), R 1 R 2 R 3 R 4 Each is independently selected from H and C1-C4 alkyl groups.

[0038] In one or more embodiments, formula I (a 1-1 In ), R 1 R 2 R 3 R 4 Each is independently selected from H and methyl.

[0039] In one or more embodiments, the narrow-band organic light-emitting molecule has one of the following structures: , , , , , , , , , , , , , , , , , , , , .

[0040] In one or more embodiments, the narrow-band organic light-emitting molecule has a structure as shown in any one of Formula III-3 to Formula III-5: Formula III-3; Formula III-4; Formula III-5; In formulas III-3, III-4, and III-5, n1, n2, n3, n4, and n5 independently represent zero, one, or more groups; in formulas III-3, III-4, and III-5, X1 and X2 are nitrogen atoms, and R1, R2, R3, R4, and R5 are independently selected from hydrogen, halogen, straight-chain alkyl, branched-chain alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, cycloalkylene, heterocycloalkylene, cycloalkenyl, heterocycloalkenyl, alkoxy, alkylsilyl, aryl, alkyl-substituted aryl, alkoxy-substituted aryl, aryl-substituted aryl, fluoroalkyl-substituted aryl, fluoroaryl-substituted aryl, aralkyl, aryloxy, heteroaryl, alkyl-substituted heteroaryl, alkoxy-substituted heteroaryl, aryl-substituted heteroaryl, fluoroalkyl-substituted heteroaryl, fluoroaryl-substituted heteroaryl, cyano, esters and their derivatives, alkyl ketones, alkyl esters, aryl esters, alkynyl, alkylamine, etc. Fluoroalkyl, fluoroaryl, polyalkylene, aniline-substituted fluorenyl, diphenylamino-substituted fluorenyl, triphenylamino-substituted fluorenyl, alkyl-substituted triphenylamino, alkyl-substituted diphenylamino, alkyl-substituted aniline, alkyl-substituted thiophenyl, borate ester, hydroxyl, amino, nitro, and trimethylsilylphenoxy; in formulas III-3, III-4, and III-5, R1 and R5 at the positions adjacent to the carbon atom bonded to the nitrogen atom can be linked to form... It can form a nitrogen heterocycle; R2 and R3 at the adjacent positions of the carbon atom bonded to the nitrogen atom can be linked to form a nitrogen heterocycle; R3 and R4 at the adjacent positions of the carbon atom bonded to the nitrogen atom can be linked to form a six-membered heterocycle, wherein the para position of the nitrogen atom in the six-membered heterocycle can be substituted with one of oxygen, nitrogen, hydrogen, carbon, boron, or sulfur, and the para atom of the nitrogen atom in the six-membered heterocycle can be bonded to hydrogen, C1-C8 alkyl, C1-C8 alkoxy, aryl, aralkyl, or heteroaryl.

[0041] In one or more embodiments, in formulas III-3, III-4 and III-5, R1, R2, R3, and R5 are each independently selected from hydrogen, C1-C8 alkyl, C1-C8 alkoxy, aryl, aralkyl, and heteroaryl, and R4 is each independently selected from hydrogen, C1-C8 alkyl, C1-C8 alkoxy, aryl, aralkyl, heteroaryl, borate ester, and halogen.

[0042] In one or more embodiments, in Formulas III-3, III-4, and III-5, R1, R2, R3, R4, and R5 are each independently selected from at least two substituents selected from a first substituent, a second substituent, a third substituent, and a fourth substituent; the first substituent includes C6-C18 straight-chain alkyl, C6-C18 heteroalkyl, polyalkylene, C6-C18 esters and their derivatives, alkyl ketones, alkyl esters, alkylamines, borate esters, hydroxyl groups, amino groups, and nitro groups; the second substituent includes C4-C8 branched alkyl, aryl, alkyl-substituted aryl, aryl-substituted aryl, fluoroalkyl-substituted aryl, fluoroaryl-substituted aryl, aralkyl, aryloxy, heteroaryl, aniline-substituted fluorenyl, and diphenylamine. The substituents include alkyl-substituted fluorenyl, triphenylamino-substituted fluorenyl, alkyl-substituted triphenylamino, alkyl-substituted diphenylamino, alkyl-substituted aniline, C6-C18 cycloalkyl, heterocyclic alkyl, cycloalkenyl, heterocyclic alkenyl, and aryl esters; the third substituent includes hydrogen, halogen, C1-C5 straight-chain alkyl, C3 branched alkyl, C3-C5 cycloalkyl, C1-C6 alkoxy, cyano, and C2-C5 alkynyl; the fourth substituent includes fluoroalkyl, fluoroaryl, alkyl-substituted heteroaryl, alkoxy-substituted heteroaryl, aryl-substituted heteroaryl, fluoroalkyl-substituted heteroaryl, fluoroaryl-substituted heteroaryl, heterocyclic alkylene, cycloalkylene, alkyl-substituted thiophenyl, alkoxy-substituted aryl, alkylsilyl, and trimethylsilylphenoxy.

[0043] In one or more embodiments, in Formula III-5, X1 and X2 are nitrogen atoms, R1, R2, R3 and R5 are each independently selected from H, C1-C4 alkyl and phenyl, and R4 is H.

[0044] In one or more embodiments, the narrow-band organic light-emitting molecule has the structure shown in Formula III-5a, Formula III-5b, Formula III-5c, Formula III-5d, Formula III-5e, or Formula III-5f: Formula III-5a Formula III-5b Formula III-5c Formula III-5d Formula III-5e Formula III-5f; In Formula III-5a, Formula III-5b, Formula III-5c, Formula III-5d, Formula III-5e and Formula III-5f, n1, n2, n3, n4 and n5 each independently represent zero or one or more groups; In Equations III-5a, III-5b, III-5c, III-5d, III-5e, and III-5f, R1 ′ R2 ′ R3 ′ R4 ′ R5′ R6 ′ Each of the following is independently selected from hydrogen, halogen, straight-chain alkyl, branched-chain alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, cycloalkylene, heterocycloalkylene, cycloalkenyl, heterocycloalkenyl, alkoxy, alkylsilyl, aryl, alkyl-substituted aryl, alkoxy-substituted aryl, aryl-substituted aryl, fluoroalkyl-substituted aryl, fluoroaryl-substituted aryl, aralkyl, aryloxy, heteroaryl, alkyl-substituted heteroaryl, alkoxy-substituted heteroaryl, aryl-substituted heteroaryl, fluorine Alkyl-substituted heteroaryl, fluoroaryl-substituted heteroaryl, cyano, esters and their derivatives, alkyl ketones, alkyl esters, aryl esters, alkynyl, alkylamine, fluoroalkyl, fluoroaryl, polyalkylene, aniline-substituted fluorenyl, diphenylamino-substituted fluorenyl, triphenylamino-substituted fluorenyl, alkyl-substituted triphenylamino, alkyl-substituted diphenylamino, alkyl-substituted aniline, alkyl-substituted thiophenyl, borate ester, hydroxyl, amino, nitro and trimethylsilylphenoxy.

[0045] In one or more embodiments, in formulas III-5a, III-5b and III-5c, R1 ′ R2 ′ R3 ′ R4 ′ R5 ′ R6 ′ Each of the following substituents is independently selected from at least two substituents: the first substituent, the second substituent, the third substituent, and the fourth substituent; in formulas III-5d, III-5e, and III-5f, R1 ′ R2 ′ R3 ′ R4 ′ R5 ′ Each of the substituents is independently selected from at least two substituents selected from the first substituent, the second substituent, the third substituent, and the fourth substituent; The first substituent includes C6-C18 straight-chain alkyl, C6-C18 heteroalkyl, polyalkylene, C6-C18 esters and their derivatives, alkyl ketones, alkyl esters, alkylamines, borate esters, hydroxyl groups, amino groups, and nitro groups; the second substituent includes C4-C8 branched alkyl, aryl, alkyl-substituted aryl, aryl-substituted aryl, fluoroalkyl-substituted aryl, fluoroaryl-substituted aryl, aralkyl, aryloxy, heteroaryl, aniline-substituted fluorenyl, diphenylamino-substituted fluorenyl, triphenylamino-substituted fluorenyl, alkyl-substituted triphenylamino, alkyl-substituted diphenylamino, and alkyl-substituted aniline. The group includes C6-C18 cycloalkyl, heterocycloalkyl, cycloalkenyl, heterocycloalkenyl, and aryl esters; the third substituent includes hydrogen, halogen, C1-C5 straight-chain alkyl, C3 branched alkyl, C3-C5 cycloalkyl, C1-C6 alkoxy, cyano, and C2-C5 alkynyl; the fourth substituent includes fluoroalkyl, fluoroaryl, alkyl-substituted heteroaryl, alkoxy-substituted heteroaryl, aryl-substituted heteroaryl, fluoroalkyl-substituted heteroaryl, fluoroaryl-substituted heteroaryl, heterocyclic alkylene, cycloalkylene, alkyl-substituted thiophenyl, alkoxy-substituted aryl, alkylsilyl, and trimethylsilylphenoxy.

[0046] In one or more embodiments, the narrow-band organic light-emitting molecule is selected from one of the following structures: , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 ； Wherein, R is independently selected from hydrogen, halogen, straight-chain alkyl, branched-chain alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, cycloalkylene, heterocycloalkylene, cycloalkenyl, heterocycloalkenyl, alkoxy, alkylsilyl, aryl, alkyl-substituted aryl, alkoxy-substituted aryl, aryl-substituted aryl, fluoroalkyl-substituted aryl, fluoroaryl-substituted aryl, aralkyl, aryloxy, heteroaryl, alkyl-substituted heteroaryl, alkoxy-substituted heteroaryl, aryl-substituted heteroaryl. Fluoroalkyl-substituted heteroaryl, fluoroaryl-substituted heteroaryl, cyano, esters and their derivatives, alkyl ketones, alkyl esters, aryl esters, alkynyl, alkylamine, fluoroalkyl, fluoroaryl, polyalkylene, aniline-substituted fluorenyl, diphenylamino-substituted fluorenyl, triphenylamino-substituted fluorenyl, alkyl-substituted triphenylamino, alkyl-substituted diphenylamino, alkyl-substituted aniline, alkyl-substituted thiophenyl, borate ester, hydroxyl, amino, nitro and trimethylsilylphenoxy.

[0047] In one or more embodiments, R is independently selected from at least two substituents selected from a first substituent, a second substituent, a third substituent, and a fourth substituent; the first substituent includes C6-C18 straight-chain alkyl, C6-C18 heteroalkyl, polyalkylene, C6-C18 esters and their derivatives, alkyl ketones, alkyl esters, alkylamines, borate esters, hydroxyl groups, amino groups, and nitro groups; the second substituent includes C4-C8 branched alkyl, aryl, alkyl-substituted aryl, aryl-substituted aryl, fluoroalkyl-substituted aryl, fluoroaryl-substituted aryl, aralkyl, aryloxy, heteroaryl, aniline-substituted fluorenyl, diphenylamino-substituted fluorenyl, triphenylamino-substituted fluorenyl, alkyl The substituents include alkyl-substituted triphenylamino, alkyl-substituted diphenylamino, alkyl-substituted aniline, C6-C18 cycloalkyl, heterocycloalkyl, cycloalkenyl, heterocycloalkenyl, and aryl esters; the third substituent includes hydrogen, halogen, C1-C5 straight-chain alkyl, C3 branched alkyl, C3-C5 cycloalkyl, C1-C6 alkoxy, cyano, and C2-C5 alkynyl; the fourth substituent includes fluoroalkyl, fluoroaryl, alkyl-substituted heteroaryl, alkoxy-substituted heteroaryl, aryl-substituted heteroaryl, fluoroalkyl-substituted heteroaryl, fluoroaryl-substituted heteroaryl, heterocyclic alkylene, cycloalkylene, alkyl-substituted thiophenyl, alkoxy-substituted aryl, alkylsilyl, and trimethylsilylphenoxy.

[0048] In one or more embodiments, the narrow-band organic light-emitting molecule has a structure as shown in Formula III-5g or Formula III-5h: Formula III - 5g; Formula III-5h; In Formula III-5g and Formula III-5h, n1, n2, n3, n4, n5, n6, n7, n8, and n9 each independently represent zero or one or more groups; in Formula III-5g and Formula III-5h, R1, R2, R3, R4, R5, R6, R7, R8, and R9 each independently select from hydrogen, halogen, straight-chain alkyl, branched-chain alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, cycloalkylene, heterocycloalkylene, cycloalkenyl, heterocycloalkenyl, alkoxy, alkylsilyl, aryl, alkyl-substituted aryl, alkoxy-substituted aryl, aryl-substituted aryl, and fluoroalkyl-substituted aryl. , fluoroaryl-substituted aryl, aralkyl, aryloxy, heteroaryl, alkyl-substituted heteroaryl, alkoxy-substituted heteroaryl, aryl-substituted heteroaryl, fluoroalkyl-substituted heteroaryl, fluoroaryl-substituted heteroaryl, cyano, esters and their derivatives, alkyl ketones, alkyl esters, aryl esters, alkynyl, alkylamine, fluoroalkyl, fluoroaryl, polyalkylene, aniline-substituted fluorenyl, diphenylamino-substituted fluorenyl, triphenylamino-substituted fluorenyl, alkyl-substituted triphenylamino, alkyl-substituted diphenylamino, alkyl-substituted aniline, alkyl-substituted thiophenyl, borate ester, hydroxyl, amino, nitro and trimethylsilylphenoxy.

[0049] In one or more embodiments, in Formula III-5g, R1, R2, R3, R4, R5, and R6 are each independently selected from at least two substituents selected from the first substituent, the second substituent, the third substituent, and the fourth substituent; in Formula III-5h, R1, R2, R3, R4, R5, R6, R7, R8, and R9 are each independently selected from at least two substituents selected from the first substituent, the second substituent, the third substituent, and the fourth substituent. The first substituent includes C6-C18 straight-chain alkyl, C6-C18 heteroalkyl, polyalkylene, C6-C18 esters and their derivatives, alkyl ketones, alkyl esters, alkylamines, borate esters, hydroxyl groups, amino groups, and nitro groups; the second substituent includes C4-C8 branched alkyl, aryl, alkyl-substituted aryl, aryl-substituted aryl, fluoroalkyl-substituted aryl, fluoroaryl-substituted aryl, aralkyl, aryloxy, heteroaryl, aniline-substituted fluorenyl, diphenylamino-substituted fluorenyl, triphenylamino-substituted fluorenyl, alkyl-substituted triphenylamino, alkyl-substituted diphenylamino, and alkyl-substituted aniline. The group includes C6-C18 cycloalkyl, heterocycloalkyl, cycloalkenyl, heterocycloalkenyl, and aryl esters; the third substituent includes hydrogen, halogen, C1-C5 straight-chain alkyl, C3 branched alkyl, C3-C5 cycloalkyl, C1-C6 alkoxy, cyano, and C2-C5 alkynyl; the fourth substituent includes fluoroalkyl, fluoroaryl, alkyl-substituted heteroaryl, alkoxy-substituted heteroaryl, aryl-substituted heteroaryl, fluoroalkyl-substituted heteroaryl, fluoroaryl-substituted heteroaryl, heterocyclic alkylene, cycloalkylene, alkyl-substituted thiophenyl, alkoxy-substituted aryl, alkylsilyl, and trimethylsilylphenoxy.

[0050] In one or more embodiments, the narrow-band organic light-emitting molecule has the structure shown in Formula III-5i: Formula III-5i; In formula III-5i, R1, R2, R3, R4, and R5 are each independently selected from H and C1-C4 alkyl groups.

[0051] In one or more embodiments, the narrow-band organic light-emitting molecule is selected from one of the following structures: , , , , , , , , , , , , , , , , , .

[0052] In one or more embodiments, the narrow-band organic light-emitting molecule has the structure shown in Formula IV or Formula V: Formula IV, Formula V, In Formula IV, n1, n2, n3, and n4 each independently represent zero or one or more groups; in Formulas IV and V, R1, R2, R3, R4, and R are each independently selected from hydrogen, halogen, straight-chain alkyl, branched-chain alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, cycloalkylene, heterocycloalkylene, cycloalkenyl, heterocycloalkenyl, alkoxy, alkylsilyl, aryl, alkyl-substituted aryl, alkoxy-substituted aryl, aryl-substituted aryl, fluoroalkyl-substituted aryl, fluoroaryl-substituted aryl, aralkyl, aryloxy, heteroaryl, alkyl Alkyl-substituted heteroaryl, alkoxy-substituted heteroaryl, aryl-substituted heteroaryl, fluoroalkyl-substituted heteroaryl, fluoroaryl-substituted heteroaryl, cyano, esters and their derivatives, alkyl ketones, alkyl esters, aryl esters, alkynyl, alkylamine, fluoroalkyl, fluoroaryl, polyalkylene, aniline-substituted fluorenyl, diphenylamino-substituted fluorenyl, triphenylamino-substituted fluorenyl, alkyl-substituted triphenylamino, alkyl-substituted diphenylamino, alkyl-substituted aniline, alkyl-substituted thiophenyl, borate ester, hydroxyl, amino, nitro, and trimethylsilylphenoxy.

[0053] In one or more embodiments, in Formula IV, R1, R2, R3, and R4 are each independently selected from at least two substituents selected from the first substituent, the second substituent, the third substituent, and the fourth substituent; the first substituent includes C6-C18 straight-chain alkyl, C6-C18 heteroalkyl, polyalkylene, C6-C18 esters and their derivatives, alkyl ketones, alkyl esters, alkylamines, borate esters, hydroxyl groups, amino groups, and nitro groups; the second substituent includes C4-C8 branched alkyl, aryl, alkyl-substituted aryl, aryl-substituted aryl, fluoroalkyl-substituted aryl, fluoroaryl-substituted aryl, aralkyl, aryloxy, heteroaryl, aniline-substituted fluorenyl, diphenylamino-substituted fluorenyl, and triphenylamine. The substituents include fluorenyl, triphenylamino, diphenylamino, aniline, C6-C18 cycloalkyl, heterocycloalkyl, cycloalkenyl, heterocycloalkenyl, and aryl esters; the third substituent includes hydrogen, halogen, C1-C5 straight-chain alkyl, C3 branched alkyl, C3-C5 cycloalkyl, C1-C6 alkoxy, cyano, and C2-C5 alkynyl; the fourth substituent includes fluoroalkyl, fluoroaryl, alkyl-substituted heteroaryl, alkoxy-substituted heteroaryl, aryl-substituted heteroaryl, fluoroalkyl-substituted heteroaryl, fluoroaryl-substituted heteroaryl, heterocyclic alkylene, cycloalkylene, alkyl-substituted thiophenyl, alkoxy-substituted aryl, alkylsilyl, and trimethylsilylphenoxy.

[0054] In one or more embodiments, the narrow-band organic light-emitting molecule is selected from one of the following structures: , , , , .

[0055] In one or more embodiments, the narrow-band organic light-emitting molecule has the structure shown in Formula VI: Formula VI, In Formula VI, n is an integer of 0 ≤ n ≤ 10, R and R′ are each independently selected from hydrogen, alkyl, alkoxy, aryl, heteroaryl and halogen, Ar is aryl, and each X is independently selected from O, S, N and CR′′; when X is O or S, the Ar connected to X does not exist; R′′ is selected from hydrogen, alkyl, alkoxy, aryl, heteroaryl and halogen.

[0056] In one or more embodiments, the narrow-band organic light-emitting molecule has a structure as shown in Formula VI(a), Formula VI(b), or Formula VI(c): Formula VI(a), Formula VI(b) Formula VI(c), In formulas VI(a), VI(b) and VI(c), R and R′ are each independently selected from hydrogen, alkyl, alkoxy, aryl, heteroaryl and halogen, and Ar is aryl.

[0057] In one or more embodiments, the narrow-band organic light-emitting molecule has a structure as shown in Formula VII: Formula VII In Formula VII, n1, n2, n3, n4, n5, n6, n7, and n8 each independently represent zero or one or more functional groups; in Formula VII, each X is independently selected from O, NR9, S, and CR. 10 R 11 One of them, R1, R2, R3, R4, R5, R6, R7, R8, R9, R 10 R 11 Each is independently selected from hydrogen, C1-C6 alkyl, C1-C6 alkoxy, arylalkyl, heteroaryl, cyano, and halogen.

[0058] In one or more embodiments, the narrow-band organic light-emitting molecule is selected from one of the following structures: , , , , .

[0059] In one or more embodiments, the narrow-band organic light-emitting molecule has the structure shown in Formula VIII: Formula VIII; In Formula VIII, n1, n2, n3, and n4 each independently represent zero or one or more groups; in Formula VIII, R1, R2, R3, and R4 are each independently selected from hydrogen, alkyl, alkoxy, aryl, heteroaryl, halogen, cyano, and borate ester groups; X1 is selected from O, NR5, S, CR6R7; X2 is selected from N and CR8; and R5, R6, R7, and R8 are each independently selected from hydrogen, alkyl, alkoxy, aryl, heteroaryl, halogen, and cyano.

[0060] In one or more embodiments, the narrow-band organic light-emitting molecule has one of the following structures: , , .

[0061] In one or more embodiments, the narrow-band organic light-emitting molecule has a structure as shown in Formula VIII(a): VIII(a), In VIII(a), n1, n2, n3, and n4 each independently represent zero or one or more groups; in VIII(a), R1, R2, R3, and R4 are selected from hydrogen, alkyl, alkoxy, aryl, heteroaryl, halogen, and cyano.

[0062] In one or more embodiments, the narrow-band organic light-emitting molecule has one of the following structures: , , , , , , , , , .

[0063] In one or more embodiments, the narrow-band organic light-emitting molecule has a structure as shown in any one of Formulas IX-1 to IX-3: Equation IX-1; Equation IX-2; Equation IX-3; In formulas IX-1 to IX-3, n1, n2, n3, and n4 each independently represent zero or one or more groups; in formulas IX-1 to IX-3, R1, R2, R3, R4, R5, and R6 each independently select from hydrogen, halogen, straight-chain alkyl, branched-chain alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, cycloalkylene, heterocycloalkylene, cycloalkenyl, heterocycloalkenyl, alkoxy, alkylsilyl, aryl, alkyl-substituted aryl, alkoxy-substituted aryl, aryl-substituted aryl, fluoroalkyl-substituted aryl, fluoroaryl-substituted aryl, aralkyl, and aryloxy alkyl, heteroaryl, alkyl-substituted heteroaryl, alkoxy-substituted heteroaryl, aryl-substituted heteroaryl, fluoroalkyl-substituted heteroaryl, fluoroaryl-substituted heteroaryl, cyano, esters and their derivatives, alkyl ketones, alkyl esters, aryl esters, alkynyl, alkylamine, fluoroalkyl, fluoroaryl, polyalkylene, aniline-substituted fluorenyl, diphenylamino-substituted fluorenyl, triphenylamino-substituted fluorenyl, alkyl-substituted triphenylamino, alkyl-substituted diphenylamino, alkyl-substituted aniline, alkyl-substituted thiophenyl, borate ester, hydroxyl, amino, nitro and trimethylsilylphenoxy.

[0064] In one or more embodiments, in Formula IX-1, R1, R2, and R3 are each independently selected from at least two substituents selected from a first substituent, a second substituent, a third substituent, and a fourth substituent; in Formulas IX-2 and IX-3, R1, R2, R3, R4, R5, and R6 are each independently selected from at least two substituents selected from a first substituent, a second substituent, a third substituent, and a fourth substituent. The first substituent includes C6-C18 straight-chain alkyl, C6-C18 heteroalkyl, polyalkylene, C6-C18 esters and their derivatives, alkyl ketones, alkyl esters, alkylamines, borate esters, hydroxyl groups, amino groups, and nitro groups; the second substituent includes C4-C8 branched alkyl, aryl, alkyl-substituted aryl, aryl-substituted aryl, fluoroalkyl-substituted aryl, fluoroaryl-substituted aryl, aralkyl, aryloxy, heteroaryl, aniline-substituted fluorenyl, diphenylamino-substituted fluorenyl, triphenylamino-substituted fluorenyl, alkyl-substituted triphenylamino, alkyl-substituted diphenylamino, and alkyl-substituted aniline. The group includes C6-C18 cycloalkyl, heterocycloalkyl, cycloalkenyl, heterocycloalkenyl, and aryl esters; the third substituent includes hydrogen, halogen, C1-C5 straight-chain alkyl, C3 branched alkyl, C3-C5 cycloalkyl, C1-C6 alkoxy, cyano, and C2-C5 alkynyl; the fourth substituent includes fluoroalkyl, fluoroaryl, alkyl-substituted heteroaryl, alkoxy-substituted heteroaryl, aryl-substituted heteroaryl, fluoroalkyl-substituted heteroaryl, fluoroaryl-substituted heteroaryl, heterocyclic alkylene, cycloalkylene, alkyl-substituted thiophenyl, alkoxy-substituted aryl, alkylsilyl, and trimethylsilylphenoxy.

[0065] In one or more embodiments, the narrow-band organic light-emitting molecule is selected from one of the following structures: , , , , , , , , , , , , , , , , , , , , , , , , ; Wherein, R is independently selected from hydrogen, halogen, straight-chain alkyl, branched-chain alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, cycloalkylene, heterocycloalkylene, cycloalkenyl, heterocycloalkenyl, alkoxy, alkylsilyl, aryl, alkyl-substituted aryl, alkoxy-substituted aryl, aryl-substituted aryl, fluoroalkyl-substituted aryl, fluoroaryl-substituted aryl, aralkyl, aryloxy, heteroaryl, alkyl-substituted heteroaryl, alkoxy-substituted heteroaryl, aryl-substituted heteroaryl. Fluoroalkyl-substituted heteroaryl, fluoroaryl-substituted heteroaryl, cyano, esters and their derivatives, alkyl ketones, alkyl esters, aryl esters, alkynyl, alkylamine, fluoroalkyl, fluoroaryl, polyalkylene, aniline-substituted fluorenyl, diphenylamino-substituted fluorenyl, triphenylamino-substituted fluorenyl, alkyl-substituted triphenylamino, alkyl-substituted diphenylamino, alkyl-substituted aniline, alkyl-substituted thiophenyl, borate ester, hydroxyl, amino, nitro and trimethylsilylphenoxy.

[0066] In one or more embodiments, the narrow-band organic light-emitting molecule has a structure as shown in Formula X: Formula X, In formula X, n1, n2, and n3 each independently represent zero or one or more groups; in formula X, R1, R2, and R3 are selected from hydrogen, alkyl, alkoxy, aryl, heteroaryl, halogen, and cyano; X, X1, and X2 are selected from C, O, N, and S; and the chemical bonds between X and X1 and between X and X2 can be saturated or unsaturated.

[0067] In one or more embodiments, the narrow-band organic light-emitting molecule is selected from one of the structures shown below: , .

[0068] In one or more embodiments, the narrow-band organic light-emitting molecule has a structure as shown in Formula XI: Formula XI; In formula XI, n1, n2, n3, and n4 each independently represent zero or one or more groups; in formula XI, R1, R2, and R3 are each independently selected from hydrogen, alkyl, alkoxy, aryl, alkylaryl, heteroaryl, halogen, cyano, and diphenylamine.

[0069] In one or more embodiments, the narrow-band organic light-emitting molecule is selected from one of the following structures: , , ; Ar represents aryl.

[0070] In one or more embodiments, the narrow-band organic light-emitting molecule has a structure as shown in Formula XI: Formula XII; In equation XII, R1-R 10 Each is independently selected from H, C1-C5 alkyl groups, wherein the alkyl group is preferably a branched alkyl group, and more preferably a tert-butyl group.

[0071] In one or more embodiments, the structure of the narrow-band organic light-emitting molecule is as follows: .

[0072] In one or more embodiments, the narrow-band organic light-emitting molecule has a structure as shown in Formula XIII: Formula XIII; In one or more embodiments, in formula XIII, R1-R 12 Each is independently selected from H, C1-C5 alkyl groups, wherein the alkyl group is preferably a branched alkyl group, and more preferably a tert-butyl group.

[0073] In one or more embodiments, the structure of the narrow-band organic light-emitting molecule is as follows: .

[0074] In one or more embodiments, the narrow-band organic light-emitting molecule has a structure as shown in Formula XIV: Formula XIV; In formula XIV, R1-R4 are each independently selected from H, C1-C5 alkyl, aryl, and triphenylamino, wherein the alkyl is preferably branched alkyl, more preferably tert-butyl, and the aryl is preferably phenyl.

[0075] In one or more embodiments, the narrow-band organic light-emitting molecule is selected from one of the following structures: , .

[0076] In one or more embodiments, the narrow-band organic light-emitting molecule has a structure as shown in Formula XV: Formula XV; In formula XV, R1-R4 are each independently selected from aryl and fused-ring aryl groups, and R5-R8 are each independently selected from H and C1-C6 alkyl groups.

[0077] In one or more embodiments, the structure of the narrow-band organic light-emitting molecule is as follows: .

[0078] In one or more embodiments, the organic light-emitting molecule is selected from one of the following structures: , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , .

[0079] In one or more embodiments, the single-peak narrowband luminescent microsphere further includes a microsphere structure, wherein the organic luminescent molecules are located at one or more sites on the surface, interface, and interior of the microsphere structure.

[0080] In one or more embodiments, the microsphere structure comprises polymeric microspheres.

[0081] In one or more embodiments, the polymer microspheres are amphiphilic, and the surface of the polymer microspheres contains one or more groups selected from epoxy, carboxyl, amino, aldehyde, hydroxyl and amide groups.

[0082] In one or more embodiments, the polymer microspheres are styrene polymer microspheres, and the surface of the styrene polymer microspheres contains one or more groups selected from amino, amide, carboxyl and aldehyde groups.

[0083] In one or more embodiments, the styrene polymer comprises a homopolymer of styrene or a copolymer of styrene with a comonomer, said comonomer comprising olefins, alkynes, carboxylic acids containing carbon-carbon double bonds, acid anhydrides containing carbon-carbon double bonds, amides containing carbon-carbon double bonds, esters containing carbon-carbon double bonds, and derivatives thereof having one or more substituents selected from halogens, amino groups, amide groups, aldehyde groups, and carboxyl groups.

[0084] In one or more embodiments, the styrene polymer is a copolymer of styrene with one or more comonomers selected from aliphatic carboxylic acids containing carbon-carbon double bonds, aliphatic anhydrides containing carbon-carbon double bonds, aliphatic amides containing carbon-carbon double bonds, and aliphatic esters containing carbon-carbon double bonds.

[0085] In one or more embodiments, the styrene polymer is a copolymer of styrene with one or more comonomers selected from carboxylic acids of formula Y1-COOH and amides of formula Y2-CONH2, wherein Y1 and Y2 are each independently groups formed by replacing one or more carbon-carbon single bonds with carbon-carbon double bonds in a C2-C18 straight-chain, branched or cyclic aliphatic alkyl group.

[0086] In one or more embodiments, Y1 and Y2 are each independently groups formed by replacing one or more carbon-carbon single bonds of a C3-C8 straight-chain, branched, or cyclic aliphatic alkyl group with carbon-carbon double bonds.

[0087] In one or more embodiments, the styrene polymer is a copolymer of styrene with (meth)acrylic acid and / or (meth)acrylamide.

[0088] The present invention also provides a homogeneous detection reagent, which includes the single-peak narrow-band luminescent microspheres described in any embodiment herein.

[0089] In one or more embodiments, the single-peak narrow-band luminescent microspheres include a buffer, and the single-peak narrow-band luminescent microspheres are receptor microspheres of the homogeneous detection reagent.

[0090] In one or more embodiments, the homogeneous detection reagent further includes donor microspheres containing a photosensitizer.

[0091] In one or more embodiments, the homogeneous detection reagent comprises one or more single-peak narrowband luminescent microspheres, the one or more single-peak narrowband luminescent microspheres comprising a buffer, and the emission spectra of the multiple single-peak narrowband luminescent microspheres having zero overlap.

[0092] The present invention also provides a homogeneous detection kit, which includes the homogeneous detection reagent described in any embodiment herein.

[0093] The present invention also provides the use of the single-peak narrow-band luminescent microspheres described in any embodiment of the present invention in the preparation of optical probes, flow cytometry reagents, biomarkers, immunochromatographic reagents, immunochromatographic kits, or radiopharmaceuticals.

[0094] The present invention also provides optical probes, flow cytometry reagents, biomarkers, immunochromatographic assay reagents, immunochromatographic kits, or radiopharmaceuticals comprising single-peak narrow-band luminescent microspheres as described in any of the embodiments herein. Attached Figure Description

[0095] Figure 1 The emission spectrum of t3IDCz@PS microspheres in Example 1 is shown.

[0096] Figure 2 The emission spectrum of p3IDCz@PS microspheres in Example 2 is shown.

[0097] Figure 3 The emission spectra of tDIDCz and tDIDCz@PS microspheres in Example 13 are shown.

[0098] Figure 4 The morphology diagram of the tDIDCz@PS microspheres in Example 13 is shown.

[0099] Figure 5 The emission spectra of pICz and pICz@PS microspheres in Example 14 are shown.

[0100] Figure 6 The morphology diagram of pICz@PS microspheres in Example 14 is shown.

[0101] Figure 7 The emission spectra of ICz and ICz@PS microspheres in Example 22 are shown.

[0102] Figure 8 The morphology diagram of ICz@PS microspheres in Example 22 is shown.

[0103] Figure 9 The emission spectra of the organic light-emitting molecules and microspheres including the organic light-emitting molecules in Example 23 are shown.

[0104] Figure 10 The image shows the morphology of the microspheres containing organic light-emitting molecules in Example 23.

[0105] Figure 11 The emission spectra of the organic light-emitting molecules and microspheres including the organic light-emitting molecules in Example 24 are shown.

[0106] Figure 12 The image shows the morphology of the microspheres containing organic light-emitting molecules in Example 24.

[0107] Figure 13 The emission spectra of the organic light-emitting molecules and microspheres including the organic light-emitting molecules in Example 25 are shown.

[0108] Figure 14 The image shows the morphology of the microspheres containing organic light-emitting molecules in Example 25.

[0109] Figure 15 The emission spectra of the organic light-emitting molecules and microspheres including the organic light-emitting molecules in Example 26 are shown.

[0110] Figure 16 The image shows the morphology of the microspheres containing organic light-emitting molecules in Example 26.

[0111] Figure 17 The morphology of the t3IDCz@PS microspheres in Example 1 is shown.

[0112] Figure 18 The image shows the morphology of the p3IDCz@PS microspheres in Example 2.

[0113] Figure 19 The emission spectra of the microspheres in Examples 25, 26, and Comparative Example 1 are shown.

[0114] Figure 20 The emission spectrum of Eu(TTA)3-TPA-DPA-mCF3@PS microspheres in Comparative Example 6 is shown.

[0115] Figure 21 The image shows a transmission electron microscope (TEM) image of PS microspheres with a diameter of less than 100 nm from Embodiment 1 of the present invention.

[0116] Figure 22 Transmission electron microscopy (TEM) images of PS microspheres with diameters of 200-300 nm prepared in Example 4 (a) and Example 5 (b) of the present invention are shown.

[0117] Figure 23 The figures (a) prepared in Example 8 and (b) prepared in Example 10 of the present invention have a diameter of approximately 30 mm. Scanning electron microscope image of m-sized PS microspheres.

[0118] Figure 24 The image shows a scanning electron microscope (SEM) image of PS microspheres with a diameter of 50-60 μm prepared in Example 12 of the present invention. The scale bar is 300 μm.

[0119] Figure 25 The image shows a scanning electron microscope (SEM) image of PS microspheres with a diameter of 50-60 μm prepared in Example 15 of the present invention. The scale bar is 30 μm.

[0120] Figure 26The image shows a scanning electron microscope (SEM) image of PS microspheres with a diameter of 50-60 μm prepared in Example 17 of the present invention. The scale bar is 50 μm.

[0121] Figure 27 The image shows a scanning electron microscope (SEM) image of PS microspheres with a diameter of 50-60 μm prepared in Example 19 of the present invention. The scale bar is 30 μm.

[0122] Figure 28 The image shows a scanning electron microscope (SEM) image of PS microspheres with a diameter of approximately 300 μm prepared in Example 23 of the present invention. The scale bar is 80 μm.

[0123] Figure 29 The following diagram shows the immunochromatographic kit based on single-peak narrow-band luminescent microspheres prepared in Application Example 17 of the present invention: (a) front view and (b) internal structure diagram.

[0124] Figure 30 A schematic diagram of the multiple cell staining process in Application Example 18 of the present invention is shown.

[0125] Figure 31 This diagram illustrates the process of preparing a live biological tissue label in Application Example 19 of the present invention. Detailed Implementation

[0126] To enable those skilled in the art to understand the features and effects of the present invention, the terms and expressions used herein are explained and defined in general terms below. Unless otherwise specified, all technical and scientific terms used herein have the common meaning understood by those skilled in the art regarding the present invention, and in case of conflict, the definitions in this specification shall prevail.

[0127] The theories or mechanisms described and disclosed herein, whether right or wrong, should not in any way limit the scope of the invention, that is, the contents of the invention can be implemented without being limited by any particular theory or mechanism.

[0128] In this document, the terms “contains,” “includes,” “containing,” and similar terms encompass the meanings of “basically composed of” and “composed of.” For example, when this document discloses “A contains B and C,” “A is basically composed of B and C” and “A is composed of B and C” should be considered as having been disclosed in this document.

[0129] In this document, all features defined by numerical ranges or percentage ranges, such as numerical values, quantities, contents, and concentrations, are for the sake of brevity and convenience only. Accordingly, descriptions of numerical ranges or percentage ranges should be considered as covering and specifically disclosing all possible sub-ranges and individual numerical values ​​(including integers and fractions) within those ranges.

[0130] Unless otherwise specified, percentages refer to mass percentages and proportions refer to mass ratios in this article.

[0131] In this article, the sum of the percentages of all components in the composition is 100%.

[0132] In this document, when describing embodiments or examples, it should be understood that it is not intended to limit the invention to those embodiments or examples. Rather, all alternatives, modifications, and equivalents of the methods and materials described herein are covered within the scope of this invention.

[0133] For the sake of brevity, not all possible combinations of the technical features in each implementation scheme or embodiment are described herein. Therefore, as long as there is no contradiction in the combination of these technical features, the technical features in each implementation scheme or embodiment can be combined arbitrarily, and all possible combinations should be considered within the scope of this specification.

[0134] In this document, the various steps described in the method embodiments may be performed in sequence and / or in parallel. Furthermore, the method embodiments may include additional steps and / or omit the steps shown. The scope of the invention is not limited in this respect.

[0135] The term "comprising" and its variations as used herein are open-ended inclusion, meaning "including but not limited to". The term "based on" means "at least partially based on". The term "one embodiment" means "at least one embodiment". The term "another embodiment" means "at least one additional embodiment". The term "some embodiments" means "at least some embodiments". Definitions of other terms will be given in the description below.

[0136] As used herein, the term "single-peak narrow-band luminescent microsphere" refers to a microsphere incorporating a narrow-band emitter as the luminescent center. This can be achieved through covalent bonding, cross-linking, or physical encapsulation. The resulting microsphere has a size ranging from submicron to micrometer. The emitted microsphere exhibits a single peak with a full width at half maximum (FWHM) of no more than 35 nm. It should be noted that, compared to the narrow-band emission described in chromophores disclosed in the art, the single-peak narrow-band luminescent microsphere of this invention exhibits only narrow-band emission and lacks a broad spectrum. Furthermore, the single-peak narrow-band luminescent microsphere of this invention differs from existing narrow-band materials, which may include a main emission peak, secondary peaks, etc.

[0137] As used in this article, the term "microsphere" can refer to a structure that is roughly spherical, such as microspheres, ellipsoids, porous spheres, etc., with varying degrees of hollowness.

[0138] In this invention, the single-peak and narrow-band characteristics of the single-peak narrow-band luminescent microspheres enable them to exhibit better sensitivity in homogeneous detection.

[0139] The diameter of the single-peak narrowband luminescent microspheres of the present invention is in the nanometer to micrometer size range. In some embodiments, the diameter range of the single-peak narrowband luminescent microspheres is 50nm-500μm, for example 60nm-500μm, 80nm-500μm, 100nm-500μm, 120nm-500μm, 150nm-500μm, 200nm-500μm, 400nm-500μm, 600nm-500μm, 800nm-500μm, 1μm-500μm, 10μm- 500μm, 50μm-500μm, 100μm-500μm, 200μm-500μm, 400μm-500μm, 100nm-400μm, 250nm-350μm, 5 00nm-400μm, 800nm-200μm, 1μm-200μm, 10μm-200μm, 50μm-300μm, 100μm-300μm, 100μm-200μm.

[0140] Existing single-peak narrowband luminescent microspheres are generally quantum dot microspheres. Quantum dot microspheres suffer from the problem of excessively small size, typically less than 20 nm, or even less than 10 nm, limiting their widespread application in biological and detection fields. Furthermore, the preparation of single-peak narrowband quantum dot microspheres is challenging due to the poor solubility of quantum dots, making the coating process difficult and complex. In contrast, the single-peak narrowband luminescent microspheres of this invention incorporate organic light-emitting molecules, which are coated within the microspheres. The microspheres are in the nanometer to micrometer range in size, making them suitable for applications in the biological field, while also offering a simple preparation process.

[0141] In some embodiments, the photoluminescence lifetime of the single-peak narrowband luminescent microspheres is in nanoseconds. In this invention, a photoluminescence lifetime of nanoseconds means a photoluminescence lifetime of 1 ns to 999 ns.

[0142] In some embodiments, the half-width at half-maximum (WHM) of the emission spectrum of the single-peak narrow-band luminescent microsphere is no greater than 35 nm, for example, no greater than 34 nm, no greater than 33 nm, no greater than 32 nm, no greater than 31 nm, no greater than 30 nm, no greater than 29 nm, no greater than 28 nm, no greater than 26 nm, no greater than 24 nm, no greater than 22 nm, no greater than 20 nm, no greater than 18 nm, no greater than 16 nm, no greater than 14 nm, no greater than 12 nm, no greater than 10 nm, no greater than 8 nm, no greater than 6 nm, and no greater than 4 nm.

[0143] In some embodiments, the peak width at 1% intensity of the highest peak of the emission spectrum of the single-peak narrow-band luminescent microsphere is no greater than 150 nm, for example, no greater than 140 nm, no greater than 130 nm, no greater than 120 nm, no greater than 110 nm, no greater than 100 nm, no greater than 90 nm, no greater than 80 nm, no greater than 70 nm, no greater than 60 nm, no greater than 50 nm, no greater than 40 nm, no greater than 30 nm, no greater than 20 nm, no greater than 10 nm, and no greater than 5 nm.

[0144] Unless otherwise specified, the full width at half maximum (FWHM) in this article refers to the difference between the two wavelengths corresponding to half the peak intensity of the highest peak in the emission spectrum. In this article, if the number of emission peaks at an intensity greater than or equal to 5% of the highest peak value is one, the emission peak of the luminescent microsphere or luminescent molecule is considered a single peak.

[0145] In some embodiments, the emission spectrum of the single-peak narrow-band luminescent microspheres above 5% of the highest peak intensity does not exhibit double peaks and / or multiple peaks. Because baseline spikes, spikes (baseline rise), and irregular, sharp small-amplitude fluctuations within the normal range of baseline fluctuations can occur due to instrument errors or other errors, to obtain a highly reliable emission spectrum, the emission spectrum above 5% of the highest peak intensity should not exhibit double peaks and / or multiple peaks, thus matching the single-peak narrow-band requirement of this invention. In some embodiments, the organic luminescent molecules of this invention do not include europium (Eu)-containing luminescent molecules.

[0146] It should be noted that two scenarios exist in the actual emission spectra obtained. In one scenario, a secondary emission peak exists in the emission spectrum, independent of the main peak, with a peak value higher than the baseline of the region adjacent to the main peak (i.e., the lowest value connected to the main peak). The secondary emission peak has a complete peak shape and can be distinguished from the main peak, with a clear peak start point, peak value, and peak end point. In this invention, the emission spectrum forming the secondary emission peak is not defined as a single peak. However, in another scenario, a shoulder-shaped protrusion appears on the right side of the main peak. This protrusion is attached to the main peak, and its peak value is always lower than the baseline of the region adjacent to the main peak (the lowest value connected to the main peak). This spectral protrusion does not have an independent and complete peak shape; it only presents as a small arc protrusion extending from the downward trend of the main peak. It cannot be completely separated from the main peak and is integrated into the spectral shape of the downward trend of the main peak.

[0147] It is difficult to achieve single-peak narrowband luminescent microspheres with a full width at half maximum (FWHM) of no more than 35 nm, a single emission peak, and a particle size ranging from nanometers to micrometers in existing technologies. While reported narrowband polymer dots exhibit multi-peak emission, it is understandable that multi-peak applications place very high demands on material composition due to crosstalk between emission peaks, which significantly reduces the coded capacity of the material.

[0148] In some embodiments, the full width at half maximum (FWHM) of the emission spectrum of the narrow-band organic light-emitting molecules is no greater than 30 nm, for example, no greater than 29 nm, no greater than 28 nm, no greater than 26 nm, no greater than 24 nm, no greater than 22 nm, no greater than 20 nm, no greater than 18 nm, no greater than 16 nm, no greater than 14 nm, no greater than 12 nm, no greater than 10 nm, no greater than 8 nm, no greater than 6 nm, and no greater than 4 nm.

[0149] In some embodiments, the peak width at 1% intensity of the highest peak of the emission spectrum of the narrow-band organic light-emitting molecule is no greater than 140 nm, for example, no greater than 130 nm, no greater than 120 nm, no greater than 110 nm, no greater than 100 nm, no greater than 90 nm, no greater than 80 nm, no greater than 70 nm, no greater than 60 nm, no greater than 50 nm, no greater than 40 nm, no greater than 30 nm, no greater than 20 nm, and no greater than 10 nm.

[0150] It is understood that in this invention, the emission spectrum of the narrow-band organic light-emitting molecules themselves satisfies a full width at half maximum (FWHM) of no more than 30 nm, and a peak width at 1% intensity of the highest peak value of no more than 140 nm. The inventors discovered that after the narrow-band organic light-emitting molecules are coated in microspheres, their emission peak shape remains essentially unchanged, with only a very slight extension in a small portion of their FWHM, not exceeding 5 nm. Thus, after being coated in microspheres, the requirement that the FWHM of the microsphere's emission spectrum not exceed 35 nm is met.

[0151] It's important to note that the peak width at 1% of the intensity of the highest peak in the emission spectrum, also called the 1% peak width, is typically defined as the width of the emission spectrum on the wavelength (or frequency) axis when the spectral intensity drops to 1% of the highest peak intensity. Assuming the emission spectrum is plotted on the wavelength axis and the intensity axis on the intensity axis, first find the point with the highest intensity in the spectrum, i.e., the highest peak value. Then determine the intensity value corresponding to 1% of the highest peak intensity. Next, find the two wavelength positions corresponding to this 1% intensity value on the spectral curve (one to the left of the peak, and one to the right of the peak). The difference between these two wavelength positions is the peak width at 1% of the intensity of the highest peak in the emission spectrum. This parameter reflects the degree of spectral broadening in spectral analysis.

[0152] In this invention, the external stimulus includes at least one of light, chemical reaction, X-ray, ultrasound, radioactive irradiation, thermal stimulation, and mechanical stimulation. It can be a single external stimulus or a combination of any two, three, four, five, six, or seven external stimuli.

[0153] Understandably, although the emission peak shape and full width at half maximum (FWHM) remain unchanged after the organic light-emitting molecules are coated in microspheres, their brightness will decrease. This is understandable because the light emitted by the organic light-emitting molecules coated in microspheres needs to pass through the microspheres before being captured by the light-emitting detection device. This process involves refraction, reflection, scattering, and light absorption, resulting in a decrease in the number of emitted photons. Alternatively, when organic light-emitting molecules are coated at high concentrations in microspheres, the molecules are very close together, and an excited-state molecule may non-radiatively transfer energy to a neighboring ground-state molecule (even if the latter is not directly excited). If the acceptor molecule is non-radiatively relaxing (such as a quencher), or if the energy transfer between multiple molecules is ultimately captured by a non-radiative center, the overall luminescence efficiency will decrease.

[0154] In this invention, external stimuli can be applied to the single-peak narrow-band luminescent microspheres, including but not limited to light, chemical reaction, X-ray, ultrasound, radioactive irradiation, thermal stimulation, mechanical stimulation, etc., to increase the number of luminescent photons, thereby improving its luminescence brightness, so that the luminescence brightness is equal to or even higher than the luminescence intensity of the original organic luminescent molecules in the solution phase.

[0155] It should be noted that when the external stimulus is light alone, the single-peak narrow-band luminescent microspheres of the present invention can be used directly. When the external stimulus is a combination of light and other external stimuli mentioned in the context of the present invention, it is understood that additional photosensitizers and / or buffers are required.

[0156] For example, in some specific cases, X-rays can generate secondary electrons and holes through the photoelectric effect or Compton scattering. These charge carriers recombine to form a luminescent state, and energy is transferred resonantly to a donor microsphere coated with a photosensitizer. The photosensitizer is further excited to a triplet state, and energy is transferred to the ground-state oxygen (…) through energy transfer. 3 O2) is converted into singlet oxygen ( 1 O2). The inventors discovered that the optimized system showed an increase in luminescence intensity of nearly one order of magnitude, and time-resolved spectroscopy confirmed that its non-radiative energy transfer efficiency reached 100%, fully demonstrating the effectiveness of X-ray stimulation. 1 O2 yield has been greatly improved.

[0157] For example, in some specific cases, the external stimulus can be mechanical. Some organic light-emitting molecules contain anthracene endoperoxides, which can act as mechanoresponsive groups and react under mechanical force. The inventors discovered that mechanical stress can trigger a ring-reversal reaction of anthracene endoperoxides, generating a fluorescent anthracene nucleus while releasing singlet oxygen and increasing luminescence intensity. For example, in one possible example, anthracene endoperoxides and photosensitizers are coated together in microspheres as donor microspheres.

[0158] For example, in some specific cases, the external stimulus can be light. For instance, photosensitizers (porphyrins, BODIPY, etc.) are excited to a singlet state (S1) under specific wavelengths of light, and then transform into a triplet state (T1) through intersystem crossing. The triplet photosensitizer reacts with ground-state oxygen (such as triplet oxygen). 3 Energy transfer occurs in O2, making 3 O2 is converted into singlet oxygen ( 1 O2), the photosensitizer itself returns to the ground state.

[0159] For example, there could be reactive stimuli, such as chemical triggering (e.g., releasing singlet oxygen from aromatic peroxides), temperature stimulation (i.e., thermal stimulation), or radioactive irradiation, to stimulate molecules that meet the requirements, thereby promoting the production of singlet oxygen and thus increasing the overall luminescence intensity of organic light-emitting molecules in the microspheres.

[0160] Understandably, the combination of the above technical solutions can produce microspheres with single peaks, narrow bands, large size and high luminescence intensity. When such microspheres are applied, for example in homogeneous detection, the detection sensitivity is higher due to the brighter luminescence.

[0161] In some embodiments, the organic light-emitting molecule comprises a single light-emitting component of a conjugated π-electron system, and the organic light-emitting molecule has a planar rigid structure.

[0162] In some embodiments, the organic light-emitting molecule includes at least one substituent, the steric hindrance of which has a conformational free energy A of not less than 1.7 kcal / mol, for example not less than 4 kcal / mol.

[0163] Understandably, the conjugated π-electron system is the core determinant of the emission wavelength and efficiency of organic molecules. Long-range ordered π-conjugation can form discrete electronic energy levels, which is conducive to radiative transitions with concentrated energy distribution. A single component avoids the uneven energy transfer or peak overlap of different emission centers caused by multi-component systems (such as host-guest doping or multiple emitting groups), which would significantly broaden the spectrum. In multi-component systems (such as host-guest doping), energy transfer processes (such as Förster resonance energy transfer) may result in energy loss and spectral broadening due to fluctuations in intermolecular distance or differences in microenvironment. A single component eliminates such interference. During luminescence, molecular vibration or rotation dissipates energy through non-radiative pathways (vibrational relaxation), leading to spectral broadening and decreased quantum efficiency. Planar rigid structures (such as polycyclic aromatic hydrocarbons and rigid polycyclic molecules) can significantly restrict intramolecular motion, reduce the conversion of excited-state energy into thermal energy, thereby improving luminescence efficiency (reducing non-radiative decay) and sharpening emission peaks (weakening vibrational coupling and more concentrated energy level distribution). In addition, large steric hindrance is also beneficial for maintaining monomer luminescence and preventing concentration quenching and redshift broadening.

[0164] Therefore, through extensive experiments, the inventors discovered that in microspheres (such as polymer microspheres, silica microspheres, etc.), molecules are confined to a limited space, which makes them more prone to extremely high local concentrations and π-π stacking / quenching. Furthermore, if the microenvironment is inhomogeneous, it can lead to spectral shift, and light scattering effects within the microspheres can cause spectral broadening. To address these issues specifically, certain requirements must be met in the molecular structure design. In some embodiments of this invention, a single luminescent component is defined to avoid complex energy transfer. Simultaneously, the planar rigid structure reduces the impact of microenvironmental disturbances on energy levels, and the large steric hindrance groups can resist aggregation at high concentrations, thereby solving the technical challenges of encapsulating organic light-emitting molecules in microspheres.

[0165] In some preferred embodiments, the conjugated π-electron system contains at least two substituents selected from a first substituent, a second substituent, a third substituent, and a fourth substituent, wherein there is a mixed steric hindrance effect between the at least two substituents.

[0166] The luminescence performance of single-peak narrow-band luminescent microspheres affects their applications. Through extensive experiments, the inventors discovered that, while maintaining a single-peak narrow band, when there is a mixed steric hindrance effect among the substituents of the organic light-emitting molecules—that is, when the substituents of the organic light-emitting molecule's core are selected from at least two of the first, second, third, and fourth substituents—the configuration of the organic light-emitting molecule is optimized. This results in better solubility, easier entry into the microspheres, a greater number of organic light-emitting molecules entering the microspheres, brighter luminescence, and better overall luminescence performance, thus facilitating the application of the luminescent microspheres.

[0167] In this invention, the definitions of substituents are as follows: In the context of this invention, "hydrogen (H)" refers to a hydrogen atom at the junction, which usually indicates that the position has not been replaced by other groups.

[0168] In the context of this invention, "halogen" refers to elements in Group 17 (VIIA) of the periodic table, including fluorine (F), chlorine (Cl), bromine (Br), and iodine (I). The terms "halogen," "halogen," and "halogen group" are used interchangeably and refer to fluorine, chlorine, bromine, and iodine.

[0169] In the context of this invention, "straight-chain alkyl" and "branched-chain alkyl" generally refer to saturated chain groups consisting only of carbon (C) and hydrogen (H), with the general formula C1. n H 2n+1It can be a straight chain (such as n-butyl) or a branched chain (such as tert-butyl). Examples include methyl (-CH3), ethyl (-CH2CH3), n-propyl (-CH2CH2CH3, isopropyl (-CH(CH3)2), n-butyl (-CH2CH2CH2CH3), sec-butyl (-CH(CH3)CH2CH3), isobutyl (-CH2CH(CH3)2), tert-butyl (-C(CH3)3), pentyl, hexyl, heptyl, octyl, nonyl, decyl, and all their branched isomers. Preferred alkyl groups are those containing one to fifteen carbon atoms, including methyl, ethyl, propyl, 1-methylethyl, butyl, 1-methylpropyl, 2-methylpropyl, pentyl, 1-methylbutyl, 2-methylbutyl, 3-methylbutyl, 1,1-dimethylpropyl, 1,2-dimethylpropyl, 2,2-dimethylpropyl, etc. Alkyl groups may optionally be substituted. The alkyl group itself or combinations thereof may include straight-chain or branched alkyl groups, preferably C1-C6 alkyl groups, and more preferably C1-C4 alkyl groups.

[0170] In the context of this invention, "heteroalkyl" refers to a chain-like group similar to an alkyl group, but in which at least one carbon atom in the main chain is replaced by a heteroatom (such as O, N, S, Si, P, B, Se, etc.). These heteroatoms typically form stable bonds (such as ether-O-, amine-NR-, thioether-S-), for example, methoxyethyl (CH3OCH2CH2-), ethoxyethyl (CH3CH2OCH2CH2-), dimethylaminoethyl ((CH3)2NCH2CH2-), methylthiomethyl (CH3SCH2-), trimethylsilylethyl ((CH3)3SiCH2CH2-), etc. In some embodiments, the atom in the heteroalkyl group attached to the parent molecule is not a heteroatom. The heteroalkyl group may optionally be substituted.

[0171] In the context of this invention, "cycloalkyl" refers to a saturated cyclic group consisting only of carbon (C) and hydrogen (H). The ring can be monocyclic or polycyclic (fused ring, bridged ring, spirocyclic), for example, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, adamantyl, norbornyl, etc. Preferred cycloalkyl groups are those containing 3 to 12 cyclic carbon atoms, and include cyclopropyl, cyclopentyl, cyclohexyl, bicyclo[3.1.1]heptyl, spiro[4.5]decyl, spiro[5.5]undecyl, adamantyl, etc. Additionally, the cycloalkyl group may optionally be substituted.

[0172] In the context of this invention, "heterocyclic alkyl" generally refers to a cyclic group containing at least one heteroatom (such as O, N, S, Si, P, B, Se, etc.) and the ring is saturated (without double bonds or aromaticity). For example, heterocyclic alkyl groups can be tetrahydrofuranyl (e.g., 2-tetrahydrofuranyl), tetrahydropyranyl (e.g., 2-tetrahydropyranyl), piperidinyl (e.g., 1-piperidinyl, 2-piperidinyl, 3-piperidinyl, 4-piperidinyl), piperazineyl (e.g., 1-piperazineyl, 4-piperazineyl), morpholinyl (e.g., 4-morpholinyl), pyrrolidinyl (e.g., 1-pyrrolidinyl, 2-pyrrolidinyl), tetrahydrothiophenyl, octahydroquinolinyl, etc. Heterocyclic alkyl groups may optionally be substituted.

[0173] In the context of this invention, "cycloalkylene" refers to a cycloalkyl group formed by two adjacent carbon atoms on a ring forming a double bond with the parent molecule. Formally, it is equivalent to a divalent group formed by removing one H atom from each of two adjacent carbon atoms in a cycloalkane. For example, cycloalkylene can be cyclopropylene, cyclopentylene, cyclohexylene, etc.

[0174] In the context of this invention, "heterocyclic alkylene" refers to a structure in which two adjacent atoms on the ring (which can be carbon-carbon, carbon-heteroatom, or heteroatom-heteroatom) are connected to the parent molecule by a double bond. For example, heterocyclic alkylene can be tetrahydrofuranyl, piperidinyl, etc.

[0175] In the context of this invention, "cycloalkenyl" generally refers to a non-aromatic carbocyclic group containing at least one carbon-carbon double bond (C=C), with the bonding point typically located on a saturated carbon atom of the ring. Cycloalkenyl groups can be cyclohexenyl (e.g., 1-cyclohexenyl, 3-cyclohexenyl), cyclopentenyl, cyclooctadienyl, etc.

[0176] In the context of this invention, "heterocyclic alkenyl" generally refers to a non-aromatic heterocyclic group containing at least one double bond (which can be C=C, C=N, N=N, etc.). The bonding point is usually on a saturated or unsaturated carbon atom on the ring. For example, a heterocyclic alkenyl can be dihydrofuranyl, dihydropyranyl, 1,2,3,6-tetrahydropyridyl, 4,5-dihydrooxazolyl, etc.

[0177] In the context of this invention, "alkoxy" generally refers to an alkyl group linked to a parent molecule via an oxygen atom (-O-alkyl). For example, an alkoxy group can be methoxy (-OCH3), ethoxy (-OCH2CH3), n-propoxy (-OCH2CH2CH3), isopropoxy (-OCH(CH3)2), tert-butoxy (-OC(CH3)3), pentoxy, or hexoxy. "Alkoxy" includes straight-chain or branched alkoxy groups, preferably C1-C6 alkoxy groups, and more preferably C1-C3 alkoxy groups.

[0178] In the context of this invention, "alkylsilyl" refers to (alkyl) n-Si-, where n is 1, 2, or 3. The alkylsilyl group can be triisopropylsilyl (TIPS), trimethylsilyl ((CH3)3Si–, TMS), tert-butyldimethylsilyl ((CH3)2Si–(C(CH3)3), TBS), triethylsilyl ((CH2CH3)3Si–, TES), -CH3Si- (methylsilyl), -CH2CH3Si- (ethylsilyl), -(CH2CH3)2Si- (diethylsilyl). The alkylsilyl group can optionally be substituted, for example, (CH3)3C-Si–Ph2 (TBDPS), Ph–CH2–Si– (Bn-Si), CH3O–Ph–CH2–Si– (MTS).

[0179] In the context of this invention, "aryl" refers to an aromatic carbocyclic group. It is typically a benzene ring derivative, but also includes polycyclic aromatic hydrocarbons (naphthalene, anthracene, phenanthrene, etc.). For example, an aryl group can be phenyl (Ph, C6H5-), naphthyl (e.g., 1-naphthyl, 2-naphthyl), anthraceneyl (e.g., 9-anthrayl), phenanthrene, biphenyl (Ph-Ph-), terphenyl, etc. The term "aryl" includes monocyclic aromatic hydrocarbon groups and polycyclic aromatic ring systems. A polycyclic system can have two or more rings in which two carbons are shared by two adjacent rings (the rings are "fused"), wherein at least one of the rings is an aromatic hydrocarbon group, and the other rings can be cycloalkyl, cycloalkenyl, aryl, heterocyclic, and / or heteroaryl. Preferred aryl groups are those containing six to thirty carbon atoms, preferably six to twenty carbon atoms, more preferably six to twelve carbon atoms. Particularly preferred are aryl groups having six, ten, or twelve carbon atoms. Suitable aryl groups include phenyl, biphenyl, terphenyl, triphenylene, tetraphenylene, naphthalene, anthracene, phenanthrene, fluorene, pyrene, perylene, and azulene, with phenyl, biphenyl, terphenyl, triphenylene, fluorene, and naphthalene being preferred. Additionally, the aryl group may optionally be substituted.

[0180] In the context of this invention, "alkylaryl" generally refers to alkyl-substituted aryl groups (Aryl-Alkyl-), such as tolyl (CH3-C6H4-, ortho / meta / para), ethylphenyl (CH3CH2-C6H4-), xylyl ((CH3)2C6H3-), cumene ((CH3)2CH-C6H4-), 1-ethylnaphthyl, etc.

[0181] In the context of this invention, "arylalkyl" generally refers to aryl-substituted alkyl groups, such as benzyl (PhCH2-), phenethyl (PhCH2CH2-), 1-phenethyl (PhCH(CH3)-), and naphthylmethyl (Naph-CH2-).

[0182] In the context of this invention, "alkoxyaryl (alkoxy-substituted aryl)" generally refers to an aryl group that is substituted by one or more alkoxy groups, such as methoxyphenyl (Anisyl, CH3O-C6H4-), ethoxyphenyl, 2,4-dimethoxyphenyl, 3,4,5-trimethoxyphenyl, etc.

[0183] In the context of this invention, "heteroaryl" refers to an aromatic heterocyclic group. It contains at least one heteroatom (e.g., O, N, S, P, B) and conforms to Hückel's rule (typically having 4n+2 π electrons). For example, a heteroaryl can be pyridyl, furanyl, thiophene, etc., and their substituted derivatives. "Heteroaryl" includes monocyclic aromatic groups and polycyclic aromatic ring systems comprising at least one heteroatom. Heteratomes include, but are not limited to, O, S, N, P, and B. In many cases, O, S, or N are preferred heteroatoms. A monocyclic heteroaromatic system is preferably a monocyclic ring having 5 or 6 ring atoms, and the ring may have one to six heteroatoms. A heteropolycyclic system may have two or more rings in which two atoms are shared by two adjacent rings (the rings are "fused"), wherein at least one of the rings is a heteroaryl, and other rings may be cycloalkyl, cycloalkenyl, aryl, heterocyclic, and / or heteroaryl. A heteropolycyclic aromatic ring system may have one to six heteroatoms on each ring of the polycyclic aromatic ring system. Preferred heteroaryl groups are those containing three to thirty carbon atoms, preferably three to twenty carbon atoms, and more preferably three to twelve carbon atoms. Suitable heteroaryl groups include dibenzothiophene, dibenzofuran, dibenzoselenophene, furan, thiophene, benzofuran, benzothiophene, benzoselenophene, carbazole, indolocarbazole, pyridylindole, pyrrolodipyridine, pyrazole, imidazole, triazole, oxazole, thiazole, oxadiazole, oxtriazole, dioxazole, thiadiazole, pyridine, pyridazine, pyrimidine, pyrazine, triazine, oxazine, oxthiazine, oxadiazine, indole, benzimidazole, indazole, inoxazine, benzoxazole, benziisoxazole, benzothiazole, quinoline, isoquinoline, zoline, quinazole Phosphorus, quinoxaline, naphthidine, phthalazine, pteridine, xanthene, acridine, phenazine, phenothiazine, phenoxazine, benzofuran-pyridine, furan-dipyridine, benzothiophene-pyridine, thiophene-dipyridine, benzoselene-pyridine, and selenophene-dipyridine, preferably dibenzothiophene, dibenzofuran, dibenzoselene, carbazole, indolocarbazole, imidazole, pyridine, triazine, benzimidazole, 1,2-azaborane, 1,3-azaborane, 1,4-azaborane, borazine, and their aza analogs. Additionally, the heteroaryl group may optionally be substituted.

[0184] Among the aryl and heteroaryl groups listed above, triphenylene, naphthalene, anthracene, dibenzothiophene, dibenzofuran, dibenzoselenene, carbazole, indolocarbazole, imidazole, pyridine, pyrazine, pyrimidine, triazine, and benzimidazole, as well as their respective aza analogs, are of particular interest.

[0185] In the context of this invention, "aryloxy group" refers to an aryl group that is attached to the parent molecule via an oxygen atom (-O-aryl). For example, an aryloxy group can be phenoxy (-OPh), 1-naphthoxy, 4-biphenoxy, etc.

[0186] In the context of this invention, "cyano" refers to the -C≡N group.

[0187] In the context of this invention, "ethers and their derivatives" refers to groups containing a -COC- structure. In a broad sense, this includes alkoxy, aryloxy, and arylalkoxy groups (such as benzyloxy-OCH2Ph), for example, those contained in alkoxy, aryloxy, and heteroalkyl groups.

[0188] In the context of this invention, "esters and their derivatives" refers to groups containing the -C(=O)OR structure (carboxylic acid esters). Derivatives may refer to thioesters (-C(=O)SR), amides (-C(=O)NR2), etc.

[0189] In the context of this invention, "alkyl ketone" refers to a group in which a carbonyl (C=O) group is attached to one or two alkyl groups. That is, -C(=O)-alkyl or alkyl-C(=O)-, including acetyl (CH3C(=O)-), propionyl (CH3CH2C(=O)-), butyryl, isobutyryl, neopentyl ((CH3)3CC(=O)-), etc.

[0190] In the context of this invention, "alkyl ester" refers to a carboxylic acid ester group (-C(=O)OR), where R is an alkyl group. For example, an alkyl ester can be -C(=O)OCH3 (methyl ester), -C(=O)OCH2CH3 (ethyl ester), -C(=O)OCH(CH3)2 (isopropyl ester), -C(=O)OC(CH3)3 (tert-butyl ester), etc.

[0191] In the context of this invention, "aryl ester" refers to a carboxylic acid ester group (-C(=O)OR), where R is an aryl group, such as -C(=O)OPh (phenyl ester), -C(=O)O(4-NO2-Ph) (p-nitrophenyl ester), -C(=O)O(1-naphthyl), etc.

[0192] In the context of this invention, "alkynyl" refers to a chain group containing a carbon-carbon triple bond (C≡C), such as ethynyl (-C≡CH), propynyl (-CH2C≡CH, -C≡CCH3), butynyl (-CH2CH2C≡CH, -CH2C≡CCH3, -C≡CCH2CH3), etc.

[0193] In the context of this invention, "alkylamine" refers to a group formed by replacing one or two hydrogens on an amino group (-NH2) with an alkyl group. For example, alkylamines can be methylamino (CH3NH-), dimethylamino ((CH3)2N-), ethylamino (CH3CH2NH-), diethylamino ((CH3CH2)2N-), methylethylamino (CH3NHCH2CH3-), isopropylamino ((CH3)2CHNH-), etc., and also include -NH2 (primary amino group).

[0194] In the context of this invention, "fluoroalkyl" refers to an alkyl group in which one or more hydrogen atoms are replaced by fluorine (F) atoms. For example, fluoroalkyl groups can be trifluoromethyl (-CF3), difluoromethyl (-CHF2), fluoromethyl (-CH2F), pentafluoroethyl (-CF2CF3), 1,1,2,2-tetrafluoroethyl (-CHFCF3), 2,2,2-trifluoroethyl (-CH2CF3), perfluorobutyl (-CF2CF2CF2CF3), etc. In the context of this invention, "fluoroaryl" refers to an aryl group in which one or more hydrogen atoms are replaced by fluorine (F) atoms, such as fluorophenyl (F-C6H4-), difluorophenyl (F2C6H3-), trifluorophenyl (F3C6H2-), pentafluorophenyl (C6F5-), perfluoronaphthyl, etc.

[0195] In the context of this invention, "polyalkylene" refers to a compound composed of multiple alkylene units (-[CH2]). n -) Chains formed by linkages. This usually refers to oligomer segments with fewer repeating units. For example, polyalkylene groups can be polymethylene (-[CH2]) n -, n>1), polyoxyethylene (repeating unit is -CH2CH2O-), polyoxypropylene (repeating unit is -CH2CH(CH3)O-), etc., are often used as flexible spacer groups.

[0196] In the context of this invention, "alkyl (alkoxy, aryl, fluoroalkyl, fluoroaryl) substituted phenyl" refers to a phenyl ring in which one or more hydrogen atoms are substituted by an alkyl, alkoxy, aryl, fluoroalkyl, or fluoroaryl group, such as tolyl (methylphenyl), ethylphenyl, tert-butylphenyl, methoxyphenyl (anisyl), phenoxyphenyl, biphenyl (phenylphenyl), trifluoromethylphenyl, pentafluorophenyl, 3,5-di(trifluoromethyl)phenyl, 3,5-bis(tert-butyl)phenyl, 4-methoxy-3-methylphenyl, 2,4,6-triisopropylphenyl, etc.

[0197] In the context of this invention, "pyridyl" generally refers to a six-membered heteroaromatic ring containing one nitrogen atom and having 2-, 3-, and 4-linked isomers, such as 2-pyridyl, 3-pyridyl, and 4-pyridyl.

[0198] In the context of this invention, "alkyl (alkoxy, aryl, fluoroalkyl, fluoroaryl) substituted pyridine" generally refers to one or more hydrogen atoms on the pyridyl ring that are substituted by an alkyl, alkoxy, aryl, fluoroalkyl, or fluoroaryl group, such as 2-methylpyridinyl, 4-ethylpyridinyl, 2-methoxypyridinyl, 4-phenylpyridinyl, 5-trifluoromethylpyridinyl, 6-pentafluorophenylpyridinyl, 3,5-dimethylpyridinyl, and 4,4'-bipyridinyl.

[0199] In the context of this invention, "bipyridinyl" generally refers to two pyridine rings directly linked by a single bond. The most common isomers are 2,2'-bipyridine and 4,4'-bipyridine, such as 2,2'-bipyridin-5-yl and 4,4'-bipyridin-6-yl (referring to the connection point being on a specific carbon in the bipyridine molecule).

[0200] In the context of this invention, "alkyl (alkoxy, aryl, fluoroalkyl, fluoroaryl) substituted bipyridyl" generally refers to a bipyridyl molecule in which one or more hydrogen atoms are substituted by an alkyl, alkoxy, aryl, fluoroalkyl, or fluoroaryl group, such as 6,6'-dimethyl-2,2'-bipyridyl, 4,4'-di-tert-butyl-2,2'-bipyridyl, 4,4'-dimethoxy-2,2'-bipyridyl, 6-phenyl-2,2'-bipyridyl, 4,4'-bis(trifluoromethyl)-2,2'-bipyridyl, etc.

[0201] In the context of this invention, "tripyridyl" generally refers to three pyridine rings linked by a single bond in a linear configuration. The most common isomer is 2,2',6',2''-terpyridine (where the three pyridine rings are linked at positions 2 and 6). For example, it usually refers to the 4'- or 4-position of the 2,2',6',2''-terpyridyl group (meaning the connection point is on a specific carbon atom in the middle or terminal pyridine ring).

[0202] In the context of this invention, "alkyl (alkoxy, aryl, fluoroalkyl, fluoroaryl) substituted tripyridyl" generally refers to a tripyridyl molecule in which one or more hydrogen atoms are substituted by an alkyl, alkoxy, aryl, fluoroalkyl, or fluoroaryl group, such as 4'-methyl-2,2':6',2''-terpyridyl, 4,4'',4'''-tritert-butyl-2,2',6',2''-terpyridyl, 4'-(trifluoromethyl)-2,2':6',2''-terpyridyl, 4-phenyl-2,2',6',2''-terpyridyl, etc.

[0203] In the context of this invention, "furanyl" generally refers to a five-membered heteroaromatic ring containing one oxygen atom and having 2- and 3-linked isomers, such as 2-furanyl and 3-furanyl.

[0204] In the context of this invention, "alkyl (alkoxy, aryl, fluoroalkyl, fluoroaryl) substituted furanyl" generally refers to one or more hydrogen atoms on the furanyl ring being substituted by an alkyl, alkoxy, aryl, fluoroalkyl, or fluoroaryl group, such as 5-methylfuran-2-yl, 3-ethylfuran-2-yl, 5-methoxyfuran-2-yl, 5-phenylfuran-2-yl, 5-trifluoromethylfuran-2-yl, etc.

[0205] In the context of this invention, "thienyl" generally refers to a five-membered heteroaromatic ring containing a sulfur atom and having 2- and 3-linked isomers, such as 2-thienyl and 3-thienyl.

[0206] In the context of this invention, "alkyl (alkoxy, aryl, fluoroalkyl, fluoroaryl) substituted thiophene group" generally refers to one or more hydrogen atoms on the thiophene ring being substituted by an alkyl, alkoxy, aryl, fluoroalkyl, or fluoroaryl group, such as 3-methylthiophene-2-yl, 5-methylthiophene-2-yl, 5-ethylthiophene-2-yl, 5-methoxythiophene-2-yl, 5-phenylthiophene-2-yl, 5-(trifluoromethyl)thiophene-2-yl, and 3,4-diethylthiophene-2-yl.

[0207] In the context of this invention, "pyrrole" generally refers to a five-membered heteroaromatic ring containing one nitrogen atom and having 1-, 2-, and 3-linked isomers, such as 1-pyrrole (NH or N-substituted), 2-pyrrole, and 3-pyrrole.

[0208] In the context of this invention, "alkyl (alkoxy, aryl, fluoroalkyl, fluoroaryl) substituted pyrrole group" generally refers to one or more hydrogen atoms on the pyrrole ring that are substituted by an alkyl, alkoxy, aryl, fluoroalkyl, or fluoroaryl group, such as 1-methylpyrrole-2-yl, 1-phenylpyrrole-2-yl, 3-methylpyrrole-2-yl, 4-methylpyrrole-2-yl, 3,4-dimethylpyrrole-2-yl, 1-methyl-5-trifluoromethylpyrrole-2-yl, etc.

[0209] In the context of this invention, "pyrazolyl" generally refers to a five-membered heteroaromatic ring containing two adjacent nitrogen atoms and having 1-, 3-, 4-, and 5-linked isomers, such as 1-pyrazolyl (NH or N-substituted), 3-pyrazolyl, 4-pyrazolyl, and 5-pyrazolyl.

[0210] In the context of this invention, "alkyl (alkoxy, aryl, fluoroalkyl, fluoroaryl) substituted pyrazolyl" generally refers to one or more hydrogen atoms (including possible NH) on the pyrazolyl ring being substituted with alkyl, alkoxy, aryl, fluoroalkyl, or fluoroaryl, such as 1-methylpyrazol-3-yl, 1-phenylpyrazol-3-yl, 3,5-dimethylpyrazol-1-yl, 4-trifluoromethylpyrazol-1-yl, 3-methyl-5-phenylpyrazol-1-yl, etc.

[0211] In the context of this invention, "oxazolyl" generally refers to a five-membered heteroaromatic ring containing one oxygen atom and one nitrogen atom (adjacent), with 2-, 4-, and 5-linked isomers, such as 2-oxazolyl, 4-oxazolyl, and 5-oxazolyl.

[0212] In the context of this invention, "alkyl (alkoxy, aryl, fluoroalkyl, fluoroaryl) substituted oxazolyl" generally refers to one or more hydrogen atoms on the oxazolyl ring being substituted by an alkyl, alkoxy, aryl, fluoroalkyl, or fluoroaryl group, such as 2-methyl-4-oxazolyl, 4-methyl-5-oxazolyl, 2-phenyl-4-oxazolyl, 5-trifluoromethyl-2-oxazolyl, 4,5-diphenyl-2-oxazolyl, etc.

[0213] In the context of this invention, "thiazolyl" generally refers to a five-membered heteroaromatic ring containing one sulfur atom and one nitrogen atom (adjacent), with 2-, 4-, and 5-linked isomers, such as 2-thiazolyl, 4-thiazolyl, and 5-thiazolyl.

[0214] In the context of this invention, "alkyl (alkoxy, aryl, fluoroalkyl, fluoroaryl) substituted thiazolyl" generally refers to one or more hydrogen atoms on the thiazolyl ring being substituted by an alkyl, alkoxy, aryl, fluoroalkyl, or fluoroaryl group, such as 2-amino-4-thiazolyl, 4-methyl-5-thiazolyl, 2-phenyl-4-thiazolyl, 5-trifluoromethyl-2-thiazolyl, 4,5-dimethyl-2-thiazolyl, etc.

[0215] In the context of this invention, "imidazolyl" generally refers to a five-membered heteroaromatic ring containing two nitrogen atoms (separated by one carbon atom) and having isomers linked at the 1-, 2-, 4-, and 5- positions.

[0216] In the context of this invention, "alkyl (alkoxy, aryl, fluoroalkyl, fluoroaryl) substituted imidazolium" generally refers to one or more hydrogen atoms (including possible NH) on the imidazolium ring being substituted with alkyl, alkoxy, aryl, fluoroalkyl, or fluoroaryl, such as 1-methylimidazol-2-yl, 1-benzylimidazol-2-yl, 4-methylimidazol-2-yl, 2-phenylimidazol-4-yl, 4,5-diphenylimidazol-1-yl, 2-trifluoromethylimidazol-1-yl, etc.

[0217] In the context of this invention, "pyrazinyl" generally refers to a six-membered heteroaryl ring containing two nitrogen atoms (para position). It has 2-, 3-, 5-, and 6-linked isomers (2-, 6-position equivalent; 3-, 5-position equivalent), such as 2-pyrazinyl, 3-pyrazinyl (or 5-pyrazinyl, the same).

[0218] In the context of this invention, "alkyl (alkoxy, aryl, fluoroalkyl, fluoroaryl) substituted pyrazinyl" generally refers to one or more hydrogen atoms on the pyrazinyl ring being substituted by an alkyl, alkoxy, aryl, fluoroalkyl, or fluoroaryl group, such as 2-methylpyrazin-3-yl (or 2-methyl-5-pyrazinyl), 2,3-dimethylpyrazin-5-yl, 2-methoxypyrazin-3-yl, 2-phenylpyrazin-3-yl, 2-trifluoromethylpyrazin-3-yl, etc.

[0219] In the context of this invention, "benzoxazole" generally refers to a heteroaromatic ring formed by the fusion of a benzene ring and an oxazole ring. The linkage can be on either the benzene ring or the oxazole ring. Common examples include 2-benzoxazole (linked at the 2-position of the oxazole ring), such as 2-benzoxazole, 4-benzoxazole, 5-benzoxazole, 6-benzoxazole, and 7-benzoxazole.

[0220] In the context of this invention, "alkyl (alkoxy, aryl, fluoroalkyl, fluoroaryl) substituted benzoxazole group" generally refers to one or more hydrogen atoms on a benzoxazole group molecule that are substituted by an alkyl, alkoxy, aryl, fluoroalkyl, or fluoroaryl group, such as 5-methylbenzoxazole-2-yl, 6-methoxybenzoxazole-2-yl, 5-phenylbenzoxazole-2-yl, 5-trifluoromethylbenzoxazole-2-yl, etc.

[0221] In the context of this invention, "benzothiadiazole" generally refers to a heteroaromatic ring formed by the fusion of a benzene ring and a 1,2,5-thiadiazole ring. Common connection sites are at the 4- or 7-position (equivalent). For example, the benzothiadiazole group can be 2,1,3-benzothiadiazole-4-yl (or 7-yl) or 2,1,3-benzothiadiazole-5-yl (or 6-yl).

[0222] In the context of this invention, "benzothiadiazole group substituted with alkyl (alkoxy, aryl, fluoroalkyl, fluoroaryl)" generally refers to a benzothiadiazole group in which one or more hydrogen atoms are substituted with alkyl, alkoxy, aryl, fluoroalkyl, or fluoroaryl groups, such as 4-methyl-2,1,3-benzothiadiazole-5-yl, 5,6-dimethoxy-2,1,3-benzothiadiazole-4-yl, 4,7-diphenyl-2,1,3-benzothiadiazole-5-yl, 4-trifluoromethyl-2,1,3-benzothiadiazole-5-yl, etc.

[0223] In the context of this invention, "carbazole" generally refers to a heterocyclic system formed by the fusion of two benzene rings and one pyrrole ring, usually referring to an unsubstituted parent nucleus, such as 9H-carbazole.

[0224] In the context of this invention, "alkyl (alkoxy, aryl, fluoroalkyl, fluoroaryl) substituted carbazole" generally refers to one or more hydrogen atoms (including NH) on a carbazole molecule that are substituted with alkyl, alkoxy, aryl, fluoroalkyl, or fluoroaryl groups, such as 9-methyl-9H-carbazole-3-yl (N-methyl, attached at C3), 9-phenyl-9H-carbazole-3-yl, 9-ethyl-9H-carbazole-2-yl, 3,6-di-tert-butyl-9H-carbazole-9-yl (attached at N9), 3,6-dimethoxy-9H-carbazole-2-yl, 3-trifluoromethyl-9H-carbazole-9-yl, 9-(4-trifluoromethylphenyl)-9H-carbazole-3-yl, etc.

[0225] In the context of this invention, "carbazolium" generally refers to a carbazolium-derived group, with the linkage point on a nitrogen atom (N9) or a carbon atom (C1-C8), such as 9H-carbazol-9-yl (N-linked), 9H-carbazol-1-yl, 9H-carbazol-2-yl, 9H-carbazol-3-yl, 9H-carbazol-4-yl, etc. (C-linked).

[0226] In the context of this invention, "alkyl (alkoxy, aryl, fluoroalkyl, fluoroaryl) substituted carbazole group" generally refers to one or more hydrogen atoms (excluding the linkage point) on a carbazole group molecule that are substituted by an alkyl, alkoxy, aryl, fluoroalkyl, or fluoroaryl group. If the linkage point is N9, the H atom on N9 can also be substituted. Examples include 3,6-di-tert-butyl-9H-carbazole-9-yl (linked at N9), 9-methyl-9H-carbazole-3-yl (linked at C3), 9-ethyl-9H-carbazole-2-yl (linked at C2), and 1,8-dimethyl-9H-carbazole-9-yl (linked at N9), etc.

[0227] In the context of this invention, "fluorenyl" generally refers to a polycyclic aromatic hydrocarbon group formed by the fusion of two benzene rings through a five-membered ring (containing a methylene bridge). The bonding site can be at the 9-position (bridge carbon) or at the 1-, 2-, 3-, or 4-position on the benzene ring. Examples include 9H-fluoren-9-yl (bonded to the bridge carbon C9), 9H-fluoren-2-yl (bonded to the 2-position on the benzene ring), 9H-fluoren-3-yl, 9H-fluoren-4-yl, and 9H-fluoren-1-yl.

[0228] In the context of this invention, "alkyl (alkoxy, aryl, fluoroalkyl, fluoroaryl) substituted fluorenyl" generally refers to one or more hydrogen atoms (including H on the bridging carbon C9) of a fluorenyl molecule that are substituted by an alkyl, alkoxy, aryl, fluoroalkyl, or fluoroaryl group. Examples include 9-methyl-9H-fluoren-9-yl (C9 substituted by a methyl group), 9-phenyl-9H-fluoren-9-yl, 2,7-di-tert-butyl-9H-fluoren-9-yl, 9,9-dimethylfluoren-2-yl, 4-methoxyfluoren-9-yl, 2-trifluoromethylfluoren-9-yl, etc.

[0229] In the context of this invention, "triphenylamine-substituted fluorenyl" generally refers to a fluorenyl molecule (typically at the 2-, 3-, 4-, or 7-position on the benzene ring) being substituted with a triphenylamine group (-N(Ph)3). Sometimes it also refers to the 9-position being substituted with a triphenylamine group. Examples include 4-(diphenylamino)-9H-fluoren-9-yl (linkage point at C9), 9,9-dimethyl-2-(diphenylamino)-9H-fluoren-7-yl (linkage point at C7), etc. The structure of triphenylamine is... .

[0230] In the context of this invention, "diphenylamino-substituted fluorenyl" generally refers to a fluorenyl molecule (typically located on the benzene ring, such as at the 2-, 3-, 4-, or 7-position) where a diphenylamino group (-N(Ph)2) is substituted. Sometimes it also refers to a 9-position substitution with a diphenylamino group. Examples include 4-(diphenylamino)-9H-fluoren-9-yl and 9,9-dimethyl-2-(diphenylamino)-9H-fluoren-7-yl.

[0231] In the context of this invention, "aniline-substituted fluorenyl" generally refers to a fluorenyl molecule (typically at the 2-, 3-, 4-, 7- position on the benzene ring) being substituted with a diphenylamino group (-NHPh).

[0232] In the context of this invention, "alkyl-substituted triphenylamine" generally refers to a triphenylamine molecule in which one or more hydrogen atoms of the benzene ring are substituted with alkyl groups. For example, 4'-methyl-N-phenyl-N-[1,1'-biphenyl]-4-ylamino-4-yl (more commonly known as 4-methyldiphenylamino, but structurally referring to partial substitution with benzidine), and more commonly refers to -N(Ar)2 with an alkyl group on the benzene ring, such as -N(4-methylphenyl)2, -N(3,5-dimethylphenyl)2, -N(4-tert-butylphenyl)2, etc.

[0233] In the context of this invention, "alkyl-substituted diphenylamino" generally refers to a diphenylamino molecule in which one or more hydrogen atoms of the benzene ring are substituted with alkyl groups.

[0234] In the context of this invention, "alkyl-substituted thiophenyl" generally refers to a thiophenyl (-SPh) molecule in which one or more hydrogen atoms of the benzene ring are replaced by an alkyl group. Examples include 4-methylphenylthio (-S-C6H4-CH3(p)), 2,4,6-trimethylphenylthio (-S-Mes), 4-tert-butylphenylthio, etc.

[0235] In the context of this invention, "boronate group" generally refers to and the groups obtained by replacing hydrogen atoms with substituents (usually alkyl or alkylene groups) (e.g. ( ), which is connected to the parent molecule through boron atoms.

[0236] In this document, various groups (e.g., alkyl, cycloalkyl, heteroalkyl, heterocycloalkyl, alkenyl, cycloalkenyl, heteroalkenyl, ynyl, aralkyl, heterocycloyl, aryl and heteroaryl) are independently unsubstituted or independently substituted by one or more general substituents.

[0237] The terms "substituted" and "substituted" generally refer to a substituent other than hydrogen (H) bonded to the relevant position, such as carbon or nitrogen. For example, when R1 represents monosubstituted, one R1 is not hydrogen (i.e., substituted). Similarly, when R1 represents disubstituted, both R1s are not hydrogen. Likewise, when R1 represents unsubstituted, R1 can be, for example, hydrogen with available valences in the ring atom, such as the carbon atom in benzene and the nitrogen atom in pyrrole, or simply unsubstituted for ring atoms with fully saturated valences, such as the nitrogen atom in pyridine. The maximum possible number of substitutions in a ring structure will depend on the total number of available valences in the ring atoms.

[0238] In this invention, with The number n of substituents R on a ring structure represented in a similar form can be zero, one, or more, in which case R represents unsubstituted, monosubstituted, or polysubstituted. The maximum number of substitutions will depend on the total number of available valences in the ring atoms. When R represents polysubstituted, the substituents can be the same or different. For example, In this context, R can represent unsubstituted, monosubstituted, disubstituted, trisubstituted, tetrasubstituted, pentasubstituted, or hexasubstituted, and each R can be the same or different.

[0239] As used herein, “combination thereof” means that one or more members of the applicable list are combined to form a known or chemically stable arrangement that can be conceived by one of ordinary skill in the art from the applicable list. For example, halogens and alkyl groups can be combined to form haloalkyl substituents; and halogens, alkyl groups, and aryl groups can be combined to form haloaralkyl groups. In one instance, the term substitution includes a combination of two to four listed groups. In another instance, the term substitution includes a combination of two to three groups. In yet another instance, the term substitution includes a combination of two groups.

[0240] In some cases, a pair of adjacent substituents may optionally join or fuse into a ring. Preferred rings are five-, six-, or seven-membered carbon rings or heterocycles, including both cases where a portion of the ring formed by the pair of substituents is saturated and a portion of the ring formed by the pair of substituents is unsaturated. As used herein, “adjacent” means that the two substituents involved may be adjacent to each other on the same ring, or on two neighboring rings having two closest available substituted positions (such as the 2, 2' positions in biphenyl or the 1, 8 positions in naphthalene), provided that a stable fused ring system can be formed.

[0241] It should be noted that the substituents in the organic small molecule structures in this article follow the valence shell electron pair repulsion theory, hybrid orbital theory, and octet rule.

[0242] In some embodiments, the single-peak narrow-band luminescent microspheres further include a microsphere structure. In this invention, organic light-emitting molecules can be immobilized on the surface, and / or interface, and / or interior of the microspheres. Typically, organic light-emitting molecules are distributed inside the microspheres; however, in this invention, it is not excluded that organic light-emitting molecules are also distributed on the surface and / or interface of the microspheres.

[0243] In this invention, the microspheres can be polymer microspheres, inorganic microspheres, composite and hybrid microspheres, or natural derivative microspheres. In some specific examples, the microspheres can be polystyrene (PS) microspheres, polylactic acid-glycolic acid copolymer (PLGA) microspheres, polyethylene glycol (PEG) hydrogel microspheres, polymethyl methacrylate (PMMA) microspheres, polycaprolactone (PCL) microspheres, polyvinyl alcohol (PVA) microspheres, polyacrylamide (PAM) microspheres, poly(N-isopropylacrylamide) (PNIPAM) microspheres, polydopamine (PDA) microspheres, polyglycolic acid (PGA) microspheres, chitosan microspheres, sodium alginate microspheres, gelatin microspheres, cellulose microspheres, or polylysine (PLL) microspheres. In other specific examples, microspheres can be silica (SiO2) microspheres, magnetic silica microspheres, hydroxyapatite (HAP) microspheres, zinc oxide (ZnO) microspheres, titanium oxide (TiO2) microspheres, zirconium oxide (ZrO2) microspheres, gold nanoparticle (Au) composite microspheres, quantum dot (CdSe / ZnS, etc.) composite microspheres, polymer-silica hybrid microspheres (such as PS@SiO2), liposome microspheres, carbon dot composite microspheres, dendritic macromolecular microspheres, protein microspheres, starch microspheres, dextran microspheres, and hyaluronic acid microspheres.

[0244] In some embodiments, the microsphere structure comprises polymeric microspheres. In some embodiments, the polymeric microspheres are amphiphilic, and the surface of the polymeric microspheres may be modified with one or more groups selected from epoxy, carboxyl, amino, aldehyde, and hydroxyl groups.

[0245] In some embodiments, the polymer microspheres are styrene polymer microspheres, and the surface of the styrene polymer microspheres may contain one or more coupling groups selected from amino, amide, carboxyl and aldehyde groups.

[0246] In some embodiments, the styrene polymer comprises a copolymer of styrene with other copolymerizable monomers, including olefins, alkynes, olefinically unsaturated carboxylic acids or their anhydrides, amides or esters, and derivatives thereof having one or more substituents selected from halogens, amino groups, amide groups, aldehyde groups and / or carboxyl groups. "olefin" refers to a compound containing a carbon-carbon double bond.

[0247] In some embodiments, the styrene polymer is selected from copolymers of styrene with aliphatic olefinic unsaturated carboxylic acids or their anhydrides, amides or esters.

[0248] In some embodiments, the styrene polymer is a copolymer of styrene with an olefinically unsaturated carboxylic acid of the formula Y-COOH or an amide thereof, wherein Y is a group formed by replacing one or more carbon-carbon single bonds with carbon-carbon double bonds in a C2-C18 straight-chain, branched, or cyclic aliphatic alkyl group. In some embodiments, Y is a group formed by replacing one or more carbon-carbon single bonds with carbon-carbon double bonds in a C3-C8 straight-chain, branched, or cyclic aliphatic alkyl group. In some embodiments, the styrene polymer is a copolymer of styrene with (meth)acrylic acid and / or (meth)acrylamide.

[0249] This invention also provides applications for single-peak narrowband luminescent microspheres.

[0250] In some embodiments, single-peak narrow-band luminescent microspheres are used to prepare homogeneous detection reagents. It is understood that single-peak narrow-band luminescent microspheres can be used directly or indirectly to prepare homogeneous detection reagents; for example, other components can be added to single-peak narrow-band luminescent microspheres to prepare homogeneous detection reagents. In the context of this invention, the composition of the single-peak narrow-band luminescent microspheres is open-ended, rather than closed-ended, deterministic.

[0251] In some embodiments, the single-peak narrow-band luminescent microspheres used to prepare homogeneous detection reagents also include a buffer, wherein the buffer and organic luminescent molecules are co-encapsulated within the microspheres as acceptor microspheres for the homogeneous detection reagents.

[0252] In some embodiments, the homogeneous detection reagent also includes donor microspheres coated with a photosensitizer.

[0253] In some embodiments, the homogeneous detection reagent includes one or more single-peak narrow-band luminescent microspheres, which may be coated with a buffer, and the emission spectra of the multiple single-peak narrow-band luminescent microspheres coated with the buffer have zero overlap.

[0254] In some embodiments, single-peak narrow-band luminescent microspheres are used to prepare homogeneous detection kits. In some embodiments, the homogeneous detection kit includes the aforementioned homogeneous detection reagents.

[0255] Homogeneous detection methods, compared to other methods, do not require the separation of free / bound molecules, directly detecting the signal of the reaction system. In homogeneous systems, unbound microspheres and reaction buffers may generate non-specific signals. The single-peak characteristic ensures that the emission signal of the microspheres comes from only a single peak position, resulting in higher discrimination against background interference (such as the broad-spectrum autofluorescence of the buffer), reducing background noise, and improving the signal-to-noise ratio (S / N). For example, in homogeneous fluorescence resonance energy transfer (FRET) detection, the donor / acceptor signals of single-peak microspheres are free of extraneous peaks, avoiding errors in energy transfer efficiency calculation and improving quantitative accuracy. Even if the center wavelengths of broad-peak microspheres are different, the peak tails may overlap (e.g., a broad peak at 550 nm overlaps with a broad peak at 600 nm), while narrow-band microspheres (e.g., 550±15 nm, 600±15 nm) can be completely separated. This feature enables multiplex detection, such as simultaneous detection of inflammatory factors IL-6, IL-8, and TNF-α. Each factor corresponds to a narrow-band microsphere, and the instrument acquires signals individually through different wavelength channels, enabling the detection of multiple indicators in a single reaction and significantly improving detection efficiency (the throughput of the reagent kit is increased by 3-5 times).

[0256] It is understood that when the single-peak narrow-band microspheres mentioned in the context of this invention are used in homogeneous detection, additional photosensitizers and buffers are usually required.

[0257] The term "photosensitizer" refers to a substance that absorbs photons and transfers the absorbed light energy to reactants in various forms, without changing its own mass or chemical properties before and after the chemical reaction. Photosensitizers can transfer excited-state energy to reactants through multiple pathways, including free radical initiation, redox reactions, and energy transfer. In addition to fulfilling the functions of traditional photocatalysts and photoinitiators, photosensitizers have a wider absorption range, utilize a broader wavelength range of sunlight, and have higher absorption and conversion efficiency. Normal substrates rarely generate their triplet intermediates directly through photoexcitation. However, if a photosensitizer is added to the system, its ground-state molecules, after absorbing photons, will generate singlet excited-state molecules upon photoexcitation. These singlet excited-state molecules are more likely to generate triplet intermediates through intersystem crossing, and their long lifetimes are sufficient to induce other substrates to generate corresponding triplet intermediates. These advantages allow photochemical reactions involving photosensitizers to produce more reaction intermediates and facilitate effective reaction control. In some contexts, "photosensitizer" can also be understood as a light absorber, which can be a traditional photosensitizing reagent or other energy donor materials.

[0258] In this application, the single-peak narrowband light-emitting material without buffer mentioned in the context of this application is equivalent to an emitter. In some contexts, it can be understood as a "light-emitting agent" or "light-emitting unit". Generally, it refers to a substance that can ultimately emit energy in the form of light energy. Such substances have light-emitting groups and can produce fluorescence or phosphorescence.

[0259] The term "buffer" can be understood in some contexts as "photochemical storage agent" or "photoelectric energy storage unit." A buffer primarily reacts with singlet oxygen in a cycloaddition reaction to generate an unstable epoxy intermediate, which then forms an excited-state diketone structure, ultimately transferring energy to the emitter. However, in some scenarios, buffers and photochemical storage agents differ slightly. Photochemical storage agents can be understood as long-term storage of photochemical energy, while buffers can be understood as short-term storage of photochemical energy. Both buffers and photochemical storage agents can participate in photochemical reactions, but the photochemical reaction products of photochemical storage agents are more stable and can exist for a longer period at room temperature, thus allowing for long-term energy storage. Buffers, on the other hand, have unstable photochemical products that undergo bond breaking and recombination after a short period of storage, simultaneously realizing energy conversion and extraction / transfer processes, thus only buffering energy for a limited time.

[0260] In some embodiments, this invention additionally coats a buffer as a acceptor microsphere within a single-peak narrow-band luminescent microsphere, and coats a photosensitizer within the microsphere as a donor microsphere. Based on the distance effect of energy transfer between the donor and acceptor microspheres, the concentration information of the analyte is indicated by the intensity of the long-persistent luminescence signal. Simply put, the donor microsphere contains a photosensitizer, and the acceptor microsphere contains a buffer and a luminescent molecule. The acceptor microsphere approaches the donor microsphere through binding via an immune response or other means. A light source selectively excites the donor microsphere, then activates the acceptor microsphere through an energy transfer process, ultimately emitting a long-persistent luminescence signal, which is detected by an optical detector and used for result analysis. The energy transfer efficiency is negatively correlated with distance, and the effective range is within a distance scale of approximately 100 nm. Donor and acceptor microspheres that do not bind via immune responses emit weaker light because the distance exceeds this range. Functionally, the donor microsphere is a sensitizing sphere, and the acceptor microsphere is a luminescent sphere. The presence and concentration information of the target analyte in the sample are determined by detecting the intensity of the long-persistent luminescence signal. Homogeneous detection technology effectively avoids cumbersome steps such as elution and centrifugation. By combining with high-affinity antibodies, it can complete a series of steps from incubation to detection in a short time, achieving highly sensitive testing while reducing operating costs.

[0261] In some implementations, single-peak narrowband luminescent microspheres are used to prepare optical probes. In biological assays, it is often necessary to simultaneously identify multiple targets (e.g., simultaneous detection of tumor markers CA125, CEA, and AFP), requiring different optical probes for each target. If the probe signal is broadband (e.g., FWHM > 50 nm), the spectra of different probes will severely overlap, leading to signal crosstalk (e.g., the emission peak of probe A overlaps with the excitation peak of probe B, which may falsely trigger the signal of probe B). Narrowband-responsive microspheres, however, can achieve wavelength orthogonality, assigning each probe an independent narrowband wavelength channel (e.g., 488 nm, 532 nm, 640 nm), with no channel overlap, thereby increasing the number of targets that can be detected simultaneously by 3-5 times (traditional broadband probes are typically limited to 2-3 channels). Biological samples (e.g., cells, tissue fluid) commonly exhibit autofluorescence (e.g., collagen has a fluorescence range of 350-500 nm, broadband and with unstable intensity), making the signal of broadband probes easily submerged by background noise.

[0262] In some implementations, single-peak narrow-band luminescent microspheres are used in flow cytometry assays. The single-peak characteristic of these microspheres avoids the complex spectral structures of the main and secondary emission peaks. If the microsphere has secondary emission peaks (such as those generated by dye impurities or structural defects in some multi-peak microspheres), these secondary peaks may fall within the filter range of other channels (e.g., a secondary peak in a CD4-detecting microsphere may fall within the CD8 channel), directly leading to false positive signals. The single-peak characteristic ensures that all fluorescence energy of the microsphere is concentrated within a single wavelength range, without additional stray peaks interfering with other channels. This strictly limits the emission peak to the filter range of the target channel. For example, if the CD3 channel uses a 530±15nm filter (detection range 515-545nm), and the matched microsphere emission peak is set to 530nm with FWHM=30nm (covering 515-545nm), then its signal will be completely captured by this channel and will not spill over into the adjacent CD4 channel (e.g., 570±15nm, 555-585nm)—the emission ranges of the two channels are 10nm apart and do not overlap. If the emission range is also centered at 530nm, it will cover 505-555nm, overlapping with the 555-585nm range of the CD4 channel by 5nm, causing approximately 10% of the CD3 signal to be interfering with the CD4 channel. This forces the flow cytometer to correct this through a complex compensation algorithm, but overcompensation will introduce new errors (e.g., underestimating the true signal of CD4). The number of detection channels in a flow cytometer is limited by the total resources of the excitation wavelength and emission spectral range. For example, the effective emission range of a 488nm laser is usually 500-650nm. If a wide-peak microsphere (FWHM=50nm) is used, it can accommodate up to 3 channels (525nm, 575nm, 625nm, spaced 50nm apart to avoid overlap). However, a narrow-band microsphere (FWHM=30nm) can insert 5 channels in the same range (510nm, 550nm, 590nm, 630nm, spaced 40nm apart, since 30nm peak width + 40nm spacing = no overlap).

[0263] In some implementations, single-peak narrow-band luminescent microspheres are used to prepare biomarkers. Biomarkers can be used to specifically identify target molecules and output precise signals, especially in multi-target joint detection (such as tumor markers, pathogen antigens, etc.), where signal interference must be avoided to ensure qualitative / quantitative accuracy. Biomarkers are often made by labeling antibodies / nucleic acids with microspheres, and their presence is reflected by emission signals (such as fluorescence) after binding to the target molecule. The single-peak characteristic means that the emission signal of the microsphere comes from only a single peak, without extraneous peaks (such as luminescence from impurities remaining during synthesis, or non-specific luminescence caused by defects in the microsphere itself), thus eliminating "false positive signals." For example, if extraneous peaks exist in microspheres labeled with the N protein of the novel coronavirus, they may be misinterpreted as signals from binding to the S protein, while the single-peak characteristic ensures that the signal comes only from the specific binding of the N protein, improving the qualitative accuracy of the marker. In multi-target detection (such as simultaneously labeling tumor markers such as CEA, AFP, and CA125), different microspheres need to be distinguished by emission wavelength. The narrow-band characteristics result in an extremely narrow emission spectral range for each microsphere (e.g., 500±15nm, 600±15nm). Even with similar center wavelengths, the overlap is extremely low, significantly improving the resolution of multi-target signals. Compared to broad-peak microspheres (e.g., 50nm half-width), narrow-band microspheres can accommodate more targets within a limited wavelength range (e.g., 10 targets within 400-800nm), breaking through the throughput limitations of traditional single-target detection and enabling multi-target measurements within a single tube.

[0264] In some implementations, single-peak narrow-band luminescent microspheres are used to prepare immunochromatographic assay reagents. Immunochromatography often relies on optical signals (fluorescence, color development), while the test strip matrix (such as a nitrocellulose membrane) may exhibit autofluorescence (broad spectrum). Single-peak microspheres have a single signal peak, which can be filtered by setting specific wavelength channels in the instrument to filter matrix background (e.g., detecting only the 580nm peak, excluding the 400-500nm autofluorescence of the membrane), making the T-line / C-line signal more singular. For example, in whole blood testing, the absorption peak of hemoglobin may interfere with the broad-spectrum microsphere signal, while the signal peak of single-peak microspheres can avoid its absorption range, preventing false negatives caused by signal attenuation. Multi-indicator immunochromatography (such as simultaneous detection of three cardiac markers: troponin, myoglobin, and CK-MB) requires multiple T-lines on the same test strip, each T-line corresponding to a type of microsphere. The narrowband characteristic of microspheres results in significant differences in emission wavelengths across different T-lines (e.g., 520nm, 620nm, 720nm narrowband). The instrument can read the signals of each T-line separately through multiple channels, avoiding cross-interference between signals from adjacent bands. Traditional wide-peak microspheres may cause T-line 1 signals to be mistakenly counted as T-line 2 due to peak overlap, while narrowband microspheres can reduce the cross-interference rate from 20% to below 5%, improving the accuracy of multi-index joint detection.

[0265] In some implementations, single-peak narrowband luminescent microspheres are used in the field of radionuclide applications, such as in the preparation of radionuclide drugs and the detection of radionuclides. Single-peak microspheres can be coupled with radionuclides as "optical labels" (e.g., for tumor targeting: the radionuclide is used for treatment, and the microspheres are used for localization). The single-peak characteristic ensures that the optical signal (e.g., fluorescence) is free of clutter, avoiding interference from the optical signal with the detection of radionuclides (e.g., gamma-ray counting, PET counting). For example, in a radionuclide-optical dual-labeled probe, the single-peak fluorescence signal (e.g., 680 nm) will not be misidentified by the radionuclide gamma-ray counter, ensuring independent quantification of the two signals. The narrowband optical signal has an extremely narrow wavelength range, which can be completely separated from the detection window of the radionuclide (e.g., the gamma-ray energy range). For example, using... 18 When F (PET nuclide, energy 511 keV) is coupled with narrowband microspheres at 650±15 nm, the narrowband optical signal does not affect the energy resolution of PET, while the broad-peak microspheres may cause counting errors due to covering part of the gamma-ray energy range. This characteristic improves the compatibility of multimodal detection and makes the optical positioning and nuclide quantification results more consistent.

[0266] The present invention will be described below by way of specific embodiments. It should be understood that these embodiments are merely illustrative and are not intended to limit the scope of the invention. The methods, reagents, and materials used in the embodiments and comparative examples are conventional methods, reagents, and materials in the art, unless otherwise stated. The starting material compounds in the embodiments and comparative examples are all commercially available.

[0267] In the examples and comparative examples, the emission spectrum of the luminescent microspheres was tested as follows: the luminescent microspheres were diluted 10 times to obtain a luminescent microsphere stock solution. During the test, the luminescent microsphere stock solution was mixed evenly with distilled water and placed in a quartz cuvette. The emission spectrum of the luminescent microspheres was tested using an Edinburgh FS5 fluorescence spectrometer.

[0268] In the examples and comparative examples, the emission spectrum of the luminescent molecules was tested as follows: the luminescent molecules were prepared into a 1mM toluene solution (luminescent molecule stock solution). During the test, the luminescent molecule stock solution and distilled water were mixed evenly and placed in a quartz cuvette. The emission spectrum of each luminescent molecule was tested using an Edinburgh FS5 fluorescence spectrometer.

[0269] Test Example 1

[0270] Method for testing the diameter of luminescent microspheres.

[0271] Images of the luminescent microspheres were acquired using transmission electron microscopy (TEM) or scanning electron microscopy (SEM), and the dimensions of each microsphere were extracted from the images and statistically analyzed using ImageJ software.

[0272] Test Example 2

[0273] Method for testing the luminescence lifetime of luminescent microspheres.

[0274] The luminescence lifetime of the microspheres was measured using time-correlated single-photon counting (TCSPC) technology with the FLS1000. The specific process is as follows: During testing, the excitation and emission wavelengths of the sample were first determined through steady-state spectroscopy. Then, the microsphere powder was sandwiched between quartz plates or placed in a powder cell and fixed on a standard front surface sample holder. Subsequently, a pulsed picosecond laser diode or nanosecond flash lamp was selected as the light source according to the excitation wavelength. The emission monochromator was adjusted to the characteristic emission peak, and TCSPC mode was enabled. After starting the measurement in Fluoracle software, the system recorded the arrival time of a single emitted photon after each pulse excitation. The fluorescence decay curve was reconstructed through statistical analysis of a large number of events. Finally, the decay curve was deconvolved in the analysis software to eliminate the broadening caused by the instrument response function. Based on the multi-exponential decay characteristics of microspheres often exhibited due to defect states, a multi-exponential model was selected for fitting to obtain the luminescence lifetime of the microspheres.

[0275] When testing the luminescence lifetime of the luminescent microspheres, the wavelength of the excitation source had no effect on the emission spectrum or full width at half maximum (FWHM) of the microspheres. This is because, for conventional fluorescence and photoluminescence (Stokes fluorescence), the shape (FWHM) of the emission spectrum is independent of the excitation wavelength. Excited molecules rapidly return to the lowest energy level of the first excited singlet state through internal conversion and vibrational relaxation, and then emit light. This relaxation process "erases" the excess energy memory from the absorbed photons.

[0276] Example 1

[0277] Synthesis of t3IDCz@PS microspheres

[0278] The t3IDCz molecule was prepared according to the following synthetic route, with the following steps:

[0279] (1) Dissolve 4-chloro-3-fluoroaniline in N,N-dimethylformamide (DMF) and add it to a round-bottom flask. At room temperature, slowly add a solution of N-bromosuccinimide (NBS) dissolved in DMF using a dropping funnel. After 6 hours, pour excess sodium thiosulfate aqueous solution into the mixture to quench the remaining NBS. The crude product is extracted, concentrated, adsorbed onto silica gel, and purified by column chromatography with dichloromethane (MC):n-hexane (1:10) as the eluent to obtain a white powder, namely 2-bromo-4-chloro-5-fluoroaniline, with a yield of 94%.

[0280] (2) 2-Bromo-4-chloro-5-fluoroaniline, cuprous iodide (CuI), tert-butyl nitrite, and acetonitrile were added to a round-bottom flask, stirred, and heated to reflux. After 12 hours, the mixture was filtered through a diatomaceous earth / silica gel filter, and the product was adsorbed onto the silica gel. The adsorbed silica gel was then loaded onto a column and purified by column chromatography with pure hexane. After all steps, a white solid, namely 1-bromo-5-chloro-4-fluoro-2-iodobenzene, was obtained in a yield of 42%.

[0281] (3) The reaction was carried out using 1-bromo-5-chloro-4-fluoro-2-iodobenzene and 3,6-di-tert-butyl-1-(4,4,5,5-tetramethyl-1,3,2-dioxoboronyl-2-yl)-9H-carbazole as starting materials. After purification, a white powder, namely 1-(2-bromo-4-chloro-5-fluorophenyl)-3,6-di-tert-butyl-9H-carbazole, was obtained with a yield of 49%.

[0282] (4) Add 1-(2-bromo-4-chloro-5-fluorophenyl)-3,6-di-tert-butyl-9H-carbazole, CuI, 1,10-phenanthroline, K3PO4, and DMF to a round-bottom flask. Reflux the mixture and stir for 4 hours. After filtering with a diatomaceous earth / silica gel filter, adsorb the product onto silica gel and purify by column chromatography using a mixture of dichloromethane (MC) and n-hexane (1:10) as the eluent. A white powder product, 2,11-di-tert-butyl-6-chloro-5-fluoroindolo[3,2,1- jk Carbazole, with a yield of 90%.

[0283] (5) 2,11-di-tert-butyl-6-chloro-5-fluoroindole[3,2,1- jk Carbazole, 4,4,4',4',5,5,5',5'-octamethyl-2,2'-bi(1,3,2-dioxoborhexacyclopentane), potassium acetate (KOAc), tris(dibenzylacetone)palladium (Pd2(dba)3), X-Phos, and 1,4-dioxane were added to a round-bottom flask and mixed with a stir bar. The reaction was heated overnight, and after cooling, filtered through a diatomaceous earth / silica gel filter. The crude solution was thoroughly dried using a rotary evaporator and recrystallized from dichloromethane and methanol. The filtered product was dried and collected as a white powder, namely 2,11-di-tert-butyl-5-fluoro-6-(4,4,5,5-tetramethyl-1,3,2-dioxoborhexacyclopentane-2-yl)indolo[3,2,1- jk Carbazole, with a yield of 86%.

[0284] (6) 2,6-Dibromo-4-(tert-butyl)aniline, 2,11-di-tert-butyl-5-fluoro-6-(4,4,5,5-tetramethyl-1,3,2-dioxoboronyl-2-yl)indolo[3,2,1- jkCarbazole, Pd(PPh3)4, and 1,2-dimethoxyethane (DME) were added to a round-bottom flask. An aqueous solution of K3PO4 was poured into the mixture, and the mixture was heated to 60°C. After reacting for 2 hours, the mixture was extracted with dichloromethane and water. The extracted organic layer was concentrated and adsorbed onto silica gel. Column chromatography purification was performed using dichloromethane (MC):n-hexane (1:4) as the eluent. A white powdery purified product, 2-bromo-4-(tert-butyl)-6-(2,11-di-tert-butyl-5-fluoroindolo[3,2,1-] jk [Carbazole-6-yl)aniline, yield 49%.

[0285] (7) 2-bromo-4-(tert-butyl)-6-(2,11-di-tert-butyl-5-fluoroindole[3,2,1- jk Carbazole-6-yl)aniline, potassium tert-butoxide (t-BuOK), and dimethyl sulfoxide (DMSO) were mixed in a round-bottom flask and heated to 80°C. After 12 hours, the reaction was quenched with water for extraction and adsorption. The crude product after adsorption was purified by column chromatography using a mixed solvent of dichloromethane (MC):n-hexane (1:9). A yellow powder product, namely 13-bromo-2,5,11-tritert-butyl-14H-pyrrolo[3,2-b:4,5,1- j'k' Dicarbazole, with a yield of 80%.

[0286] (8) Except for the use of 13-bromo-2,5,11-tritert-butyl-14H-pyrrolo[3,2-b:4,5,1- in this step j'k' Except for using dicarbazole (2.25 mmol) as the starting material, the synthesis method was similar to that of 3,6-di-tert-butyl-1-(4,4,5,5-tetramethyl-1,3,2-dioxoborhecyclopentan-2-yl)-9H-carbazole. After all steps, a yellow powder, namely 2,5,11-tri-tert-butyl-13-(4,4,5,5-tetramethyl-1,3,2-dioxoborhecyclopentan-2-yl)-14H-pyrrolo[3,2-b:4,5,1-j'k']dicarbazole, was collected from the filter in 89% yield.

[0287] (9) Add the solution of 3,6-di-tert-butyl-1-(2,5-dibromo-4-iodophenyl)-9H-carbazole dissolved in tetrahydrofuran (THF), 2M K3PO4 aqueous solution, and Pd(PPh3)4 to a round-bottom flask, and start heating to about 100°C while stirring. Add the solution of 2,5,11-tri-tert-butyl-13-(4,4,5,5-tetramethyl-1,3,2-dioxoboronyl-2-yl)-14H-pyrrolo[3,2-b:4,5,1- j'k'The dicarbazole solution was slowly added to the mixture. After 12 hours, the crude product was extracted with dichloromethane and water, adsorbed onto silica gel, and purified by column chromatography using dichloromethane (MC):n-hexane (1:1) as the eluent. The purified product was collected as a thoroughly dried white product, namely 2,5,11-tri-tert-butyl-13-(2,5-dibromo-4-(3,6-di-tert-butyl-9H-carbazole-1-yl)phenyl)-14H-pyrrolo[3,2-b:4,5,1- j'k' Dicarbazole, with a yield of 45%.

[0288] (10) 2,5,11-tri-tert-butyl-13-(2,5-dibromo-4-(3,6-di-tert-butyl-9H-carbazole-1-yl)phenyl)-14H-pyrrolo[3,2-b:4,5,1- j'k' Dicarbazole, cuprous iodide (CuI), 1,10-phenanthroline, potassium phosphate (K3PO4), and N,N-dimethylformamide were added to a round-bottom flask. The reaction was carried out under stirring and reflux for 8 hours. After cooling to room temperature, the crude product was poured into chloroform and heated until completely dissolved. The diluted solution was filtered through diatomaceous earth / silica gel filter paper and then evaporated by rotary evaporator. The dried crude product was recrystallized from methanol. The above process was repeated three times. After drying in a vacuum oven, a yellow powder, t3IDCz, was obtained with a yield of 90%. NMR results were as follows: 1 HNMR (500MHz, THF-d8): δ 8.92 (s, 2H), 8.62 (s, 2H), 8.40 (s, 2H), 8.39 (d, J =0.9Hz, 2H), 8.29 (d, J =1.8Hz, 2H), 8.25 (d, J =0.9Hz, 2H), 8.11 (d, J =8.4Hz, 2H), 7.71(dd, J =8.5, 1.9Hz, 2H), 1.47 (s, 9H), 1.40 (s, 18H), 1.30 (s, 18H). 13 C NMR(126MHz, THF-d8): δ 147.0, 145.0, 144.8, 138.1, 135.8, 135.7, 130.8, 130.2,129.9, 124.5, 120.4, 119.1, 117.4, 117.4, 117.0, 112.2, 106.9, 106.8, 36.6,36.5, 35.4, 33.2, 33.1, 32.1, 30.4.

[0289] Preparation of styrene microspheres: Following the method in CN117603383A, styrene and glycidyl methacrylate were added to a three-necked flask, along with potassium persulfate, sodium chloride, and deionized water. The mixture was stirred, the cooling water was turned on, and an inert gas was introduced. The mixture was then heated at room temperature. After 8 hours of reaction, the reaction solution turned milky white. The emulsion and tetrahydrofuran were mixed at a volume ratio of 1:5 and swelled for 5 minutes. The mixture was then centrifuged and the supernatant was discarded. The microspheres were then washed once by alternating centrifugation with ethanol and water to obtain hollow polystyrene microspheres.

[0290] t3IDCz@PS microspheres were prepared by coating polystyrene microspheres with t3IDCz via a swelling method. The specific preparation process was as follows: Polystyrene microspheres were centrifuged to remove surfactants from the synthesis process. The solid polystyrene microspheres were then re-dissolved in 100 mL of ultrapure water and sonicated to form a dispersed phase. Next, 1 mL each of 2 wt% sodium dodecylbenzenesulfonate and 1 wt% ethylenediamine polyoxyethylene polyoxypropylene block polyether were added to the polystyrene microsphere aqueous solution and stirred. The luminescent molecules were dispersed in tetrahydrofuran solution to form a dispersed phase. After the solution preparation was complete, the styrene polymer phase was rapidly added to the aqueous phase, and the temperature was gradually increased to 50 °C, with continuous stirring for 10 h. The resulting luminescent microspheres were then centrifuged to remove excess luminescent molecules, washed twice with ultrapure water and ethanol, and stored in ultrapure water at room temperature in the dark for later use.

[0291] The full width at half maximum (FWHM) of the emission spectrum of the microspheres was measured to be 27 nm. See the microsphere emission spectrum for details. Figure 1 See the microsphere morphology diagram. Figure 17 The emission spectrum of the microspheres was tested. One emission peak was observed at 5% of the peak intensity, the peak width at 1% of the peak intensity was 119 nm, and the luminescence lifetime was 21 ns. The microsphere diameter was measured to be between 198 nm and 203 nm. The emission spectrum of the luminescent molecules was also tested, with a full width at half maximum (FWHM) of 21 nm and a peak width at 1% of the peak intensity of 107 nm.

[0292] Example 2

[0293] Synthesis of p3IDCz@PS microspheres

[0294] The p3IDCz molecule was prepared according to the following synthetic route, with the steps as follows:

[0295] The synthesis method of p3IDCz is similar to that of t3IDCz in Example 1, except that in step (9), 3,6-diphenyl-1-(2,5-dibromo-4-iodophenyl)-3,6-diphenyl-9H-carbazole is used instead of 3,6-di-tert-butyl-1-(2,5-dibromo-4-iodophenyl)-9H-carbazole. After the wet purification process, a green powder, namely p3IDCz, is obtained with a yield of 92%.

[0296] The preparation of p3IDCz@PS microspheres is described in Example 1. The full width at half maximum (FWHM) of the emission spectrum of the microspheres was measured to be 24 nm. The emission spectrum of the microspheres is shown in [reference needed]. Figure 2 See the microsphere morphology diagram. Figure 18 The emission spectrum of the microspheres was tested. One emission peak was observed at 5% intensity (or greater than or equal to the peak value). The peak width at 1% intensity was 117 nm, and the luminescence lifetime was 11.7 ns. The microsphere diameter was measured to be between 198 nm and 203 nm. The emission spectrum of the luminescent molecules was also tested. The full width at half maximum (FWHM) was 16 nm, and the peak width at 1% intensity was 105 nm.

[0297] Example 3

[0298] Synthesis of Na-sBN@PS microspheres

[0299] Na-sBN molecules were prepared according to the following synthetic route, with the steps as follows:

[0300] (1) 1-Bromo-2-chloro-3-fluorobenzene, 3,6-di-tert-butylcarbazole, Cs2CO3 and N,N-dimethylformamide were stirred at 155 °C for 12 hours. The white precipitate was collected by filtration, washed sequentially with water and methanol, and dried under vacuum to obtain white solid 1 with a yield of 98%.

[0301] (2) A mixture of 1,4-dibromonaphthalene, 4,4,5,5-tetramethyl-1,3,2-dioxaborane-2-dipinacol boronic acid ester, potassium acetate, [1,1'-bis(diphenylphosphine)ferrocene]palladium(II) dichloride, and N,N-dimethylformamide was stirred overnight at 90°C under an argon atmosphere and then cooled to room temperature. The mixture was extracted with dichloromethane / water and dried over sodium sulfate, with the solvent removed under vacuum. The residue was purified by column chromatography to give a white solid 2 in 84% yield.

[0302] (3) The solution containing white solid 1, white solid 2, tetrakis(triphenylphosphine)palladium and potassium carbonate was stirred at 100°C under an argon atmosphere for 12 hours. Then it was cooled to room temperature. The mixture was extracted with dichloromethane, dried over sodium sulfate, and the solvent was removed by vacuum rotary evaporation. The residue was washed with ethanol and dried to give crude product 3.

[0303] (4) Crude product 3 was dissolved in toluene, and a solution of tert-butyllithium in pentane was slowly added under argon atmosphere at -40°C. The mixture was then stirred at 60°C for 6 hours, followed by vacuum rotary evaporation to remove the pentane. Boron tribromide was then added at 0°C, and the mixture was stirred at room temperature for 0.5 hours. N,N-diisopropylethylamine was then added at 0°C, and the mixture was stirred at 160°C for 20 hours. The mixture was then cooled to room temperature, and the solvent was removed under vacuum. The residue was purified by column chromatography to give a yellow solid Na-sBN in 31% yield.

[0304] The NMR results are as follows: 1 H NMR (600 MHz, Chloroform-d): δ 9.11 (dd, J = 7.0, 1.3Hz, 1H), 8.93 (d, J = 1.8 Hz, 1H), 8.72 (d, J = 7.9 Hz, 1H), 8.55-8.44 (m,2H), 8.38 (dd, J = 8.3, 1.3 Hz, 1H), 8.35 (dd, J = 8.4, 4.2 Hz, 2H), 8.27 (d, J = 2.0 Hz, 1H), 8.16 (d, J = 1.8 Hz, 2H), 7.92 (t, J = 8.1 Hz, 1H), 7.88-7.79 (m, 2H), 7.79-7.68 (m, 3H), 7.68-7.62 (m, 2H), 7.57-7.47 (m, 4H), 1.64(s, 9H), 1.53 (s, 9H), 1.47 (s, 18H) ppm. 13C NMR (151 MHz, CDCl3): δ =145.54, 145.14, 143.17, 142.92, 142.67, 141.99, 141.88, 141.38, 139.26,138.89, 138.45, 138.30, 135.17, 133.29, 132.81, 131.65, 131.44, 130.34,129.84, 129.02, 128.76, 128.47, 127.33, 127.14, 126.48, 125.60, 124.81,124.56, 124.18, 123.85, 123.65, 121.93, 121.24, 117.60, 117.45, 116.45,114.26, 113.82, 109.45, 35.39, 34.96, 34.89, 32.40, 32.16, 31.98 ppm.

[0305] The Na-sBN product obtained in the last step of the above process was coated into polystyrene microspheres using a swelling method, following the steps in Example 1, to prepare Na-sBN@PS microspheres. The emission wavelength of the microspheres was 525 nm, and the full width at half maximum (FWHM) was 31 nm. The emission spectrum of the microspheres was tested, and the number of emission peaks at 5% or greater of the highest peak intensity was found to be one. The peak width at 1% of the highest peak intensity was 131 nm, and the luminescence lifetime was 11.9 ns. The diameter of the microspheres was measured to be between 198 nm and 203 nm. The emission spectrum of the luminescent molecules was tested, and the FWHM was found to be 29 nm, with a peak width at 1% of the highest peak intensity of 129 nm.

[0306] Example 4

[0307] Synthesis of AZA-BN@PS microspheres

[0308] The AZA-BN molecule was prepared according to the following synthetic route, with the steps as follows:

[0309] (1) Under an argon atmosphere, 3,6-di-tert-butylcarbazole was added to a two-necked round-bottom flask equipped with a reflux condenser. A DMF suspension of cesium carbonate was added to the reaction flask and stirred at room temperature for 30 minutes. Subsequently, 5-bromo-2-chloro-1,3-difluorobenzene was added to the reaction flask all at once. The mixture was stirred at 150°C for 12 hours. After the reaction was completed, the mixture was diluted with water and recrystallized from hot ethanol to give a white solid 1 (9,9'-(5-bromo-2-chloro-1,3-phenylene)bis(3,6-di-tert-butyl-9H-carbazole)) in 92% yield.

[0310] (2) Under an argon atmosphere, 9,9'-(5-bromo-2-chloro-1,3-phenylene)bis(3,6-di-tert-butyl-9H-carbazole), (2-cyanophenyl)boronic acid, tetra(triphenylphosphine)palladium, and potassium carbonate were added to a three-necked flask. A suitable amount of tetrahydrofuran and water were then added to the flask to dissolve the substances. The mixture was refluxed and heated for 8 hours. After the reaction was complete, the reaction mixture was diluted with water, and the crude product was recrystallized from hot ethanol to give a white solid 2, with a yield of 89%.

[0311] (3) Under a nitrogen atmosphere, a pentane solution of n-butyllithium was slowly added to a solution of tert-butylbenzene containing compound 2 at 0°C. The reaction mixture was stirred for 30 minutes, then heated to 60°C and stirred for 2 hours. Boron tribromide was added at -40°C, followed by stirring at room temperature for 0.5 hours. N,N-diisopropylethylamine was added at 0°C, and the reaction mixture was then heated to 120°C and stirred for 5 hours. After the reaction was completed, the reaction mixture was cooled to room temperature. The mixture was extracted with an aqueous sodium acetate solution and dichloromethane. The organic layers were combined, dried over magnesium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography and recrystallized from dichloromethane / diethyl ether. A yellow powder, AZA-BN, was obtained, with a yield of 20%. The NMR results were as follows: 1 H NMR (400 MHz, Chloroform-d): δ 9.22 (d, J = 16.9 Hz, 1H), 9.10 (d, J = 4.5 Hz, 2H), 8.86 – 8.67 (m, 1H),8.63-8.48 (m, 2H), 8.45 (s, 1H), 8.34 – 8.25 (m, 1H), 8.25-8.12 (m, 2H),7.87-7.64 (m, 3H), 7.59 (s, 1H), 7.35 (s, 1H), 3.18 (s, 2H), 1.80 (s, 2H),1.69 (d, J= 6.2 Hz, 20H), 1.57 (s, 9H), 1.51 (s, 9H), 0.90 (t, J = 7.3 Hz, 3H). 13 C NMR (101 MHz, Chloroform-d) δ 156.06, 145.10, 144.87, 144.40, 142.86,142.07, 141.32, 140.93, 140.61, 138.21, 132.55, 130.22, 129.84, 128.93,128.77, 128.27, 128.17, 127.19, 126.42, 126.34, 126.06, 125.64, 124.44,123.46, 123.30, 122.38, 121.06, 120.55, 118.61, 117.46, 115.64, 113.74, 100.95, 35.50, 35.35, 35.27, 34.94, 34.84, 32.34, 31.98, 30.90, 23.30, 14.18.

[0312] The product AZA-BN obtained in the last step above was coated into polystyrene microspheres using a swelling method as described in Example 1 to prepare AZA-BN@PS microspheres. The microspheres exhibited a single emission peak with a measured full width at half maximum (FWHM) of 30 nm. The emission spectrum of the microspheres was tested, revealing one emission peak at 5% intensity (greater than or equal to the peak value), a peak width of 134 nm at 1% intensity (the peak value at which the peak value was reached), and a luminescence lifetime of 13.4 ns. The microsphere diameter was measured to be between 285 nm and 300 nm. The emission spectrum of the luminescent molecules was also tested, showing a FWHM of 28 nm and a peak width of 118 nm at 1% intensity (the peak value at which the peak value was reached).

[0313] Example 5

[0314] Synthesis of Cz-DBCz@PS microspheres

[0315] The Cz-DBCz molecule was prepared according to the following synthetic route, with the steps as follows:

[0316] (1) At room temperature, 3,6-di-tert-butylcarbazole, 2-bromo-1,3-difluoro-5-iodobenzene and CsCO3 were dissolved in dry DMF. The reaction mixture was stirred at 150°C for 18 hours, followed by washing with water and filtration. The crude product was recrystallized to give a white solid, 9,9'-(2-bromo-5-iodo-1,3-phenylene)bis(3,6-di-tert-butyl-9H-carbazole), in 90% yield.

[0317] (2) Under a nitrogen atmosphere, 9,9'-(2-bromo-5-iodo-1,3-phenylene)bis(3,6-di-tert-butyl-9H-carbazole) and 1-(4,4,5,5-tetramethyl-1,3,2-dioxaborane-2-yl)-9H-carbazole were added to a 100 mL three-necked flask. Then, dimethoxyethane, an aqueous solution of K₂CO₃, and Pd(PPh₃)₄ were added to the flask. The mixture was heated to 80 °C and stirred. After stirring for 4 hours, the reactants were poured into brine and extracted three times with dichloromethane and water. The organic phase was dried over MgSO₄, and the solvent was evaporated under vacuum. The crude product was purified on silica gel with petroleum ether / dichloromethane (5:1, v / v) to give a white solid, namely 9,9'-(2-bromo-5-(9H-carbazole-1-yl)-1,3-phenylene)bis(3,6-di-tert-butyl-9H-carbazole), in 78% yield.

[0318] (3) A butyllithium-hexane solution was slowly added to a xylene solution of 9,9'-(2-bromo-5-(9H-carbazole-1-yl)-1,3-phenylene)bis(3,6-di-tert-butyl-9H-carbazole) at -30 °C. The mixture was maintained at -30 °C for 2 hours under a nitrogen atmosphere, followed by the addition of boron tribromide at -30 °C. The reaction mixture was then heated to room temperature and stirred for 1 hour. N,N-diisopropylethylamine was added at 0 °C, and the reaction mixture was stirred at 150 °C for 24 hours. After cooling to room temperature, the precipitate was separated by filtration and recrystallized in methanol / dichloromethane to give a pale yellow solid, Cz-DBCz, in 60% yield. The NMR results were as follows: 1 HNMR (400 MHz, Methylene Chloride-d2) δ(ppm): 8.84 (s, 1H), 8.59 (s, 2H), 8.35-8.25 (m, 3H), 8.18-8.06 (m, 3H), 7.96-7.92 (m, 2H), 7.76-7.72 (m, 2H),7.64-7.43 (m, 3H), 7.22 (s, 1H), 1.84 (s, 9H), 1.70 (s, 9H), 1.64 (s, 9H),1.54 (s, 9H). 13C NMR (151 MHz, Methylene Chloride-d2) δ (ppm): 145.62,144.42, 142.40, 129.31, 129.02, 128.93, 125.79, 121.86, 121.76, 121.39,121.03, 120.51, 120.32, 116.61, 114.30, 100.2, 35.31, 35.18, 35.09, 34.73,32.11, 32.00, 31.72.

[0319] The product Cz-DBCz obtained in the last step of the above process was coated into polystyrene microspheres using a swelling method as described in Example 1 to prepare Cz-DBCz@PS microspheres. The emission peak was a single peak with a measured full width at half maximum (FWHM) of 20 nm. The emission spectrum of the microspheres was tested, and the number of emission peaks at 5% or greater of the highest peak intensity was found to be one. The peak width at 1% of the highest peak intensity was 100 nm, and the luminescence lifetime was 5.8 ns. The microsphere diameter was measured to be between 285 nm and 300 nm. The emission spectrum of the luminescent molecules was also tested, and the measured FWHM was 17 nm, with a peak width at 1% of the highest peak intensity of 96 nm.

[0320] Example 6

[0321] Synthesis of Cz-DBTPA@PS microspheres

[0322] The Cz-DBTPA molecule was prepared according to the following synthetic route, with the steps as follows:

[0323] The synthesis method of Cz-DBTPA is similar to that of Cz-DBCz in Example 5, except that in step (1), 1,3-dibromo-2,5-dichlorobenzene is used instead of 2-bromo-1,3-difluoro-5-iodobenzene, and di-p-toluidine is used. Replace 3,6-di-tert-butylcarbazole; in step (2), replace 1-(4,4,5,5-tetramethyl-1,3,2-dioxoborane-2-yl)-9H-carbazole with 3,6-di-tert-butyl-1-(4,4,5,5-tetramethyl-1,3,2-dioxoborane-2-yl)-9H-carbazole. The precipitate was separated by filtration and recrystallized in methanol / dichloromethane to give a yellowish-brown solid, Cz-DBTPA, in 40% yield. NMR results are as follows: 1 H NMR (400 MHz, Methylene Chloride- d2) δ (ppm): 8.68 (s,1H), 8.53 (s, 1H), 8.48 (s, 1H), 8.40 (d, J = 8.6 Hz, 1H), 8.18-8.11 (m, 2H), 8.08 (d, J = 8.5 Hz, 1H), 8.02 (d, J = 8.6 Hz, 1H), 7.88 (s, 1H), 7.62-7.51(m, 3H), 7.41-7.17 (m, 6H), 6.95 (d, J = 8.0 Hz, 1H), 2.55-2.52 (m, 9H), 2.42(s, 3H), 1.44 (s, 9H), 1.38 (s, 9H). 13 C NMR (151 MHz, Methylene Chloride- d 2)δ (ppm): 148.72, 146.02, 145.29, 144.44, 140.30, 139.41, 139.13, 138.32,134.56, 134.48, 132.66, 132.43, 131.95, 131.01, 130.53, 129.96, 128.53,124.06, 124.01, 123.22, 118.10, 117.63, 117.53, 117.16, 115.54, 100.17,34.85, 33.50, 31.98, 31.69, 31.62, 29.75, 23.32, 22.74, 21.03, 20.80, 18.39, 13.92.

[0324] The synthesis of Cz-DBTPA@PS microspheres is described in Example 5. The emission peak is a single peak, and its full width at half maximum (FWHM) is measured to be 19 nm. The emission spectrum of the microspheres was tested, and the number of emission peaks at 5% or greater of the highest peak intensity was found to be one. The peak width at 1% of the highest peak intensity was 101 nm, and the luminescence lifetime was 5.1 ns. The diameter of the microspheres was measured to be between 398 nm and 412 nm. The emission spectrum of the luminescent molecules was tested, and the FWHM was measured to be 18 nm, with a peak width of 96 nm at 1% of the highest peak intensity.

[0325] Example 7

[0326] Synthesis of PhO-DBCz@PS microspheres

[0327] PhO-DBCz molecules were prepared according to the following synthetic route, with the steps as follows:

[0328] The synthesis method of PhO-DBCz is similar to that of Cz-DBCz in Example 5, except that in step (2), 1-(4,4,5,5-tetramethyl-1,3,2-dioxaborane-2-yl)-9H-carbazole is replaced with (2-hydroxy-4-tert-butylphenyl)boronic acid pinacol ester. The precipitate was separated by filtration and recrystallized in methanol / dichloromethane to obtain a light yellow solid, namely PhO-DBCz, with a yield of 52%. The NMR results are as follows: 1 H NMR (400 MHz, Methylene Chloride-d2): δ(ppm): 8.28 (s, 1H), 8.21-8.07 (m, 4H), 8.02 (s, 1H), 7.97-7.87 (m, 3H), 7.78(d, J = 7.0 Hz, 2H), 7.61 (s, 1H), 7.48 (d, J = 8.2 Hz, 1H), 1.80 (s, 9H), 1.71 (d, J = 9.6 Hz, 18H), 1.54 (d, J = 13.1 Hz, 18H). 13 C NMR (151 MHz,Methylene Chloride- d 2): δ (ppm): 153.57, 153.09, 145.28, 145.21, 144.46,143.90, 141.81, 141.37, 137.88, 128.57, 128.51, 127.25, 126.40, 124.39,124.30, 124.26, 123.54, 122.26, 121.60, 120.80, 120.22, 119.94, 117.77,117.24, 115.21, 97.56, 35.49, 35.18, 35.00, 34.80, 32.28, 31.95, 31.78, 31.74, 31.31.

[0329] The synthesis of PhO-DBCz@PS microspheres is described in Example 5. The emission peak is a single peak, with a measured full width at half maximum (FWHM) of 18 nm. The emission spectrum of the microspheres was tested, and the number of emission peaks at 5% or greater of the highest peak intensity was found to be one. The peak width at 1% of the highest peak intensity was 90 nm, and the luminescence lifetime was 5.1 ns. The microsphere diameter was measured to be between 398 nm and 412 nm. The emission spectrum of the luminescent molecules was also tested, with a measured FWHM of 18 nm and a peak width at 1% of the highest peak intensity of 84 nm.

[0330] Example 8

[0331] Synthesis of BN-DICz@PS microspheres

[0332] BN-DICz molecules were prepared according to the following synthetic route, with the steps as follows:

[0333] (1) Compound 1, 2-chloroaniline, Pd2(dba)3, dppf and sodium tert-butoxide were dissolved in toluene solution and stirred under nitrogen atmosphere and refluxed. After cooling to room temperature, the solvent was removed under reduced pressure. The residue was purified by silica gel column chromatography (using dichloromethane / petroleum ether (1:5, v / v) as eluent) to obtain target compound 2 as a pale yellow solid with a yield of 95.1%.

[0334] (2) Compound 2, potassium carbonate, palladium acetate, and tri-tert-butylphosphine tetrafluoroborate were dissolved in dimethylacetamide. The mixture was stirred and heated to 150°C for 6 hours under an argon atmosphere. After cooling to room temperature, the mixture was dispersed in 1 liter of water. The resulting precipitate was filtered and washed with running water. The residue was purified by silica gel column chromatography using dichloromethane / petroleum ether (1:4, v / v) as the eluent, finally yielding target compound 3 as a light green solid with a yield of 48.9%.

[0335] (3) Under a nitrogen atmosphere, compound 3, 9-(2-bromo-3-fluorophenyl)-3,6-di-tert-butyl-9H-carbazole, cesium carbonate, and 60 mL of DMF were added to a double-necked round-bottom flask equipped with a reflux condenser. After stirring and refluxing for 12 hours, the mixture was diluted with water, and the crude product was recrystallized from hot ethanol to give product 4 as a yellow powder, with a yield of 72%.

[0336] (4) A pentane solution of n-butyllithium was slowly added to a tert-butylbenzene solution containing compound 4 at 0°C under a nitrogen atmosphere. After stirring at 60°C for 24 hours, the pentane was removed by vacuum distillation. Subsequently, boron tribromide was added at -40°C, and the reaction mixture was stirred at room temperature for 0.5 hours. N,N-diisopropylethylamine was added at 0°C, and the reaction mixture was then heated to room temperature. After stirring at 120°C for 5 hours, the reaction mixture was cooled to room temperature. An aqueous solution of sodium acetate and dichloromethane were added to the reaction mixture. The aqueous layer was separated and extracted with dichloromethane. The organic layers were combined, dried with magnesium sulfate, filtered, and evaporated under reduced pressure. The crude product BN-DICz was purified by column chromatography (silica gel, petroleum ether / CH2Cl2 = 50:1) and recrystallized from CH2Cl2 and methanol to give a pale yellow powder, namely BN-DICz, in a yield of 22%. The NMR results were as follows: 1 H NMR (600MHz, Chloroform-d): δ 8.74 (d, J = 32.4 Hz, 1H), 8.56 (d, J = 17.1 Hz, 1H), 8.42 (d, J =38.2 Hz, 1H), 8.33-8.07 (m, 4H), 8.04-7.82 (m, 3H), 7.74 (d, J =38.7 Hz, 1H), 7.67- 5 7.33 (m, 8H), 7.12 (s, 1H), 1.46 (d, J = 29.9 Hz, 18H).

[0337] The synthesis of BN-DICz@PS microspheres is described in Example 1. The emission peak is a single peak, and its full width at half maximum (FWHM) is measured to be 20 nm. The emission spectrum of the microspheres was tested, and the number of emission peaks at 5% or greater of the highest peak intensity was found to be one. The peak width at 1% of the highest peak intensity was 120 nm, and the luminescence lifetime was 6.61 ns. The diameter of the microspheres was measured to be between 398 nm and 412 nm. The emission spectrum of the luminescent molecules was tested, and the FWHM was measured to be 18 nm, with a peak width at 1% of the highest peak intensity of 112 nm.

[0338] Example 9

[0339] This embodiment describes the synthesis of DBN-ICz@PS microspheres.

[0340] DBN-ICz molecules were prepared according to the following synthetic route, with the steps as follows:

[0341] The synthesis method of DBN-ICz is similar to that of BN-DICz in Example 8. Starting with compound 5, the synthesis was carried out according to steps (3) and (4) of Example 8. The crude product DBN-ICz was purified by column chromatography (silica gel, petroleum ether / CH2Cl2 = 50:1) and recrystallized from CH2Cl2 and methanol to obtain an orange powder, namely DBN-ICz, with a yield of 36%.

[0342] The synthesis of DBN-ICz@PS microspheres is described in Example 1. The emission peak is a single peak, and its full width at half maximum (FWHM) is measured to be 20 nm. The emission spectrum of the microspheres was tested, and the number of emission peaks at 5% or greater of the highest peak intensity was found to be one. The peak width at 1% of the highest peak intensity was 130 nm, and the luminescence lifetime was 5.6 ns. The diameter of the microspheres was measured to be between 285 nm and 300 nm. The emission spectrum of the luminescent molecules was tested, and the FWHM was measured to be 18 nm, with a peak width at 1% of the highest peak intensity of 110 nm.

[0343] Example 10

[0344] Synthesis of BF-PZ@PS microspheres

[0345] BF-PZ molecules were prepared according to the following synthetic route, with the steps as follows:

[0346] (1) p-tert-butylbenzaldehyde was mixed with freshly distilled pyrrole, degassed by purging with nitrogen, and then a catalytic amount of BF3·OEt2 was added. The mixture was stirred at room temperature. After the reaction was complete, dilute NaOH was added to terminate the reaction. The aqueous phase was extracted with dichloromethane, and the combined organic phases were dried over anhydrous Na2SO4. After filtration and evaporation, the crude product was purified by silica gel column chromatography to obtain a white powder, namely 2,2'-((4-(tert-butyl)phenyl)methylene)bis(1H-pyrrole), in a yield of 40%.

[0347] (2) 2,2'-((4-(tert-butyl)phenyl)methylene)bis(1H-pyrrole), 2,3-dichloro-5,6-dicyano-1,4-benzoquinone, and dry dichloromethane were added to a flask. The mixture was stirred under a nitrogen atmosphere at room temperature. Then, dry triethylamine and BF3·OEt2 were added sequentially. Stirring was continued. The aqueous phase was then extracted with dichloromethane, and the organic phase was dried over anhydrous sodium sulfate. After filtration and evaporation, the crude product was purified by silica gel column chromatography to give the BODIPY precursor as a red fluorescent solid with a yield of 40%.

[0348] (3) Under a nitrogen atmosphere, an oven-dried Schlenk tube was packed with 4-bromo-1H-pyrazole and dry THF. The solution was cooled to 0°C with stirring, and then 60% NaH dispersion was slowly added. After hydrogen release stopped, TrtCl was added in portions, and the mixture was stirred overnight at room temperature. After the reaction was complete, the mixture was quenched with water and extracted with DCM. The combined organic phases were dried with anhydrous Na2SO4, filtered, and the solvent was removed under reduced pressure. The crude product was recrystallized from hexane and DCM to give a white powdery product, namely 4-bromo-1-triphenylmethyl-1H-pyrazole, in 78% yield.

[0349] (4) Under nitrogen atmosphere, 4-bromo-1-triphenylmethyl-1H-pyrazole, p-tert-butylphenylboronic acid, tetrakis(triphenylphosphine)palladium, potassium carbonate, and 1,2-dimethoxyethane / water were added to a Schlenk tube. The reaction mixture was stirred at 85°C and concentrated on a rotary evaporator. The residue was dissolved in dichloromethane, washed with water and brine, dried with anhydrous sodium sulfate, filtered, and dried under reduced pressure. The crude product was recrystallized from hexane and dichloromethane to give a white powder, namely 4-(4-(tert-butyl)phenyl)-1-triphenylmethyl-1H-pyrazole, in 80.5% yield.

[0350] (5) Compound 4-(4-(tert-butylphenyl)-1-triphenylmethyl-1H-pyrazole was stirred in methanol / dichloromethane at 40°C with 1.25 M HCl. Toluene was then added, and the reaction mixture was concentrated. The crude product was recrystallized from hexane and dichloromethane to give a white powder, namely 4-(tert-butylphenyl)-1H-pyrazole, in 98.1% yield.

[0351] (6) Under a nitrogen atmosphere, BODIPY precursor, 4-(tert-butylphenyl)-1H-pyrazole, AgOAc, and dry dimethyl sulfoxide were added to a dry Schlenk tube equipped with a magnetic stir bar. The reaction mixture was stirred at 80°C for 12 hours. Subsequently, the solvent was removed under reduced pressure. The residue was dissolved in dichloromethane, filtered through a Celite pad, and washed with DCM. The filtrates were combined, concentrated, and purified by silica gel column chromatography to give a purple powder, namely BF-PZ, in 52% yield. NMR results were as follows: 1 HNMR (400 MHz, CDCl3): δ = 1.38 (s, 18H), 1.41 (s, 9H) 6.97 (d, J = 4.4 Hz, 2H), 7.05 (d, J = 4.4 Hz, 2H), 7.48 (d, J= 8.4 Hz, 4H), 7.52-7.56 (m, 4H), 7.61 (d, J = 8.0 Hz, 4H), 8.10 (s, 2H), 9.13 (s, 2H) ppm. 13 C NMR (100 MHz, CDCl3): δ = 31.4, 31.5, 34.8, 35.1, 113.0, 125.6, 125.9, 126.0, 126.1, 127.7,128.7, 130.6, 130.8, 131.1, 132.5, 141.4, 142.9, 150.4, 150.6, 154.0 ppm.

[0352] The synthesis of BF-PZ@PS microspheres is described in Example 1. The emission peak is a single peak, and its full width at half maximum (FWHM) is measured to be 26 nm. The emission spectrum of the microspheres was tested, and the number of emission peaks at 5% or greater of the highest peak intensity was found to be one. The peak width at 1% of the highest peak intensity was 135 nm, and the luminescence lifetime was 6.1 ns. The diameter of the microspheres was measured to be between 285 nm and 300 nm. The emission spectrum of the luminescent molecules was tested, and the FWHM was measured to be 25 nm, with a peak width at 1% of the highest peak intensity of 132 nm.

[0353] Example 11

[0354] Synthesis of MeBF-PZ@PS microspheres

[0355] MeBF-PZ molecules were prepared according to the following synthetic route, with the steps as follows:

[0356] The synthesis method of MeBF-PZ is similar to that of BF-PZ in Example 10, except that in step (1), 4-tert-butyl-2,6-dimethylbenzaldehyde is used instead of p-tert-butylbenzaldehyde. The crude product MeBF-PZ was purified by column chromatography to obtain a purple powder, i.e., MeBF-PZ, with a yield of 48.2%. The NMR results are as follows: 1 H NMR (400 MHz, CDCl3): δ = 1.37 (s,9H), 1.38 (s, 18H), 2.21 (s, 6H), 6.69 (d, J = 4.4 Hz, 2H), 6.99 (d, J = 4.4Hz, 2H), 7.15 (s, 2H), 7.48 (d, J = 8.4 Hz, 4H), 7.61 (d, J= 8.4 Hz, 4H), 8.09 (s, 2H), 9.15 (s, 2H) ppm. 13 C NMR (100 MHz, CDCl3): δ = 20.5, 31.49,31.52, 34.7, 34.8, 113.2, 124.7, 125.9, 126.0, 126.1, 128.1, 128.7, 129.5,129.7, 132.7, 136.7, 141.4, 142.4, 150.4, 150.6, 152.2 ppm.

[0357] The synthesis of MeBF-PZ@PS microspheres is described in Example 1. The emission peak is a single peak, and its full width at half maximum (FWHM) is measured to be 21 nm. The emission spectrum of the microspheres was tested, and the number of emission peaks at 5% or greater of the highest peak intensity was found to be one. The peak width at 1% of the highest peak intensity was 126 nm, and the luminescence lifetime was 6.5 ns. The diameter of the microspheres was measured to be between 285 nm and 300 nm. The emission spectrum of the luminescent molecules was also tested, and the FWHM was measured to be 21 nm, with a peak width of 125 nm at 1% of the highest peak intensity.

[0358] Example 12

[0359] Synthesis of MeBF-MePZ@PS microspheres

[0360] The MeBF-MePZ molecule was prepared according to the following synthetic route, with the steps as follows:

[0361] The synthesis method of MeBF-MePZ is similar to that of BF-PZ in Example 10, except that in step (1), 4-tert-butyl-2,6-dimethylbenzaldehyde is used instead of p-tert-butylbenzaldehyde; and in step (4), 2,6-dimethyl-4-tert-butylphenylboronic acid is used instead of p-tert-butylphenylboronic acid. The crude product MeBF-MePZ was purified by column chromatography to obtain a purple-red powder with a yield of 37.3%. The NMR results are as follows: 1 H NMR (400 MHz, CDCl3): δ = 1.36 (s, 18H), 1.37 (s, 9H), 2.17 (s, 12H), 2.20 (s, 6H), 6.69 (d, J = 3.6 Hz, 2H), 6.97 (d, J = 4.4 Hz,2H), 7.11 (s, 4H), 7.15 (s, 2H), 7.70 (s, 2H), 8.68 (s, 2H) ppm.13 C NMR (100MHz, CDCl3): δ = 20.4, 21.6, 31.49, 31.54, 34.5, 34.7, 113.1, 123.1, 124.6,124.7, 128.1, 128.4, 129.5, 129.8, 130.9, 132.6, 136.6, 137.3, 144.2, 150.5,150.7, 154.9 ppm.

[0362] The synthesis of MeBF-MePZ@PS microspheres is described in Example 1. The emission peak is a single peak, and its full width at half maximum (FWHM) is measured to be 22 nm. The emission spectrum of the microspheres was tested, and the number of emission peaks at 5% or greater of the highest peak intensity was found to be one. The peak width at 1% of the highest peak intensity was 126 nm, and the luminescence lifetime was 6.0 ns. The diameter of the microspheres was measured to be between 285 nm and 300 nm. The emission spectrum of the luminescent molecules was also tested, and the FWHM was measured to be 22 nm, with a peak width of 125 nm at 1% of the highest peak intensity.

[0363] Example 13

[0364] Synthesis of tDIDCz@PS microspheres

[0365] The tDIDCz molecule was prepared according to the following synthetic route, with the steps as follows:

[0366] (1) Sodium hydride was dissolved in DMF and added to a round-bottom flask. Then, 3,6-di-tert-butyl-9H-carbazole was dissolved in DMF and added to the round-bottom flask at room temperature. The mixture was stirred. Next, 1,5-dibromo-2,4-difluorobenzene was dissolved in DMF and slowly added to a nitrogen-filled flask. The solution was heated overnight at 60°C with stirring. After the reaction flask cooled to room temperature, deionized water was added to the reaction mixture to quench excess sodium hydride, and the mixture was allowed to stand. The crude product was obtained by filtration, and the filtrate was further purified by reprecipitation using a dichloromethane / methanol mixture. A white solid, 9,9'-(4,6-dibromo-1,3-phenyl)bis(3,6-di-tert-butyl-9H-carbazole), was finally obtained in 77.3% yield.

[0367] (2) Triphenylphosphine, 9,9'-(4,6-dibromo-1,3-phenyl)bis(3,6-di-tert-butyl-9H-carbazole), benzyltriethylammonium chloride, potassium carbonate, and palladium acetate were added to a round-bottom flask. DMAC was then added to the flask, and the mixture was stirred and heated to reflux. After cooling, deionized water was added to the reaction mixture, and the crude product was obtained by filtration. The crude product was washed with dichloromethane and methanol to obtain a pure white solid, tDIDCz, with a yield of 89.2%. The NMR results were as follows: 1 H NMR (500 MHz, CDCl3): δ8.77-8.77 (sd, J = 0.50 Hz, 1H), 8.18-8.17 (sd, J = 2.00 Hz, 2H), 8.16-8.16(sd, J = 1.00 Hz, 2H), 8.11-8.10 (sd, J = 1.00 Hz, 2H), 8.08-8.08 (sd, J =0.50 Hz, 1H), 7.91-7.89 (d, J = 8.50 Hz, 2H), 7.66-7.63 (dd, J 1 = 2.00 Hz, J 2 = 8.50 Hz, 2H) 1.61 (s, 18H), 1.53 (s, 18H). 13 C NMR (125 MHZ, CDCl3): δ146.9, 145.0, 143.8, 138.6, 137.2, 130.6, 125.3, 124.1, 119.9, 118.6, 118.1,117.4, 116.1, 115.9, 111.8, 96.1, 36.2, 35.2, 33.1, 32.2.

[0368] The synthesis of tDIDCz@PS microspheres is described in Example 1. The emission peak is a single peak, and the measured full width at half maximum (FWHM) is 21 nm. The emission spectrum is shown in [reference needed]. Figure 3 ,in Figure 3 (a) is the emission spectrum of the tDIDCz molecule. Figure 3 (b) is the emission spectrum of tDIDCz@PS microspheres. Figure 4This is a morphology image of the tDIDCz@PS microspheres. The emission spectrum of the microspheres was tested; the number of emission peaks at 5% or greater of the peak intensity was measured to be one, the peak width at 1% of the peak intensity was 123 nm, and the luminescence lifetime was 5.03 ns. The microsphere diameter was measured to be between 500 nm and 515 nm. The emission spectrum of the luminescent molecules was also tested; the full width at half maximum (FWHM) was measured to be 16 nm, and the peak width at 1% of the peak intensity was 105 nm.

[0369] Example 14

[0370] Synthesis of pICz@PS microspheres

[0371] pICz molecules were prepared according to the following synthetic route, with the steps as follows:

[0372] (1) Sodium hydride was dissolved in DMF and added to a round-bottom flask. Then, 3,6-di-tert-butyl-9H-carbazole was dissolved in DMF and added to the round-bottom flask at room temperature. The mixture was stirred. Next, 1,4-dibromo-2,5-difluorobenzene was dissolved in DMF and slowly added to a nitrogen-filled flask. The solution was heated overnight at 60°C with stirring. After the reaction flask cooled to room temperature, deionized water was added to the reaction mixture to quench excess sodium hydride, and the mixture was allowed to stand. The crude product was obtained by filtration, and the filtrate was further purified by reprecipitation using a dichloromethane / methanol mixture. The final product was a white solid, 9,9'-(2,5-dibromo-1,4-phenylene)bis(3,6-di-tert-butyl-9H-carbazole), in 77% yield.

[0373] (2) 9,9'-(2,5-dibromo-1,4-phenylene)bis(3,6-di-tert-butyl-9H-carbazole), triphenylphosphine, benzyltriethylammonium chloride, potassium carbonate, and palladium acetate were added to a round-bottom flask. DMAC was then added to the flask, and the mixture was stirred and heated under reflux for 6 hours. After cooling, deionized water was added to the reaction mixture, and the crude product was filtered and washed to obtain a pale yellow solid, pICz, with a yield of 74.3%.

[0374] The synthesis of pICz@PS microspheres is described in Example 1. The emission peak is a single peak, and its full width at half maximum (FWHM) is measured to be 20 nm. The emission spectrum is shown in [reference needed]. Figure 5 ,in Figure 5 (a) is the emission spectrum of the pICz molecule. Figure 5 (b) is the emission spectrum of pICz@PS microspheres. Figure 6This is a morphology image of pICz@PS microspheres. The emission spectrum of the microspheres was tested; the number of emission peaks at 5% or greater of the peak intensity was found to be one, the peak width at 1% of the peak intensity was 121 nm, and the luminescence lifetime was 5.51 ns. The microsphere diameter was measured to be between 500 nm and 515 nm. The emission spectrum of the luminescent molecules was also tested; the full width at half maximum (FWHM) was 13 nm, and the peak width at 1% of the peak intensity was 97 nm.

[0375] Example 15

[0376] Synthesis of pICz-TPA@PS microspheres

[0377] pICz-TPA molecules were prepared according to the following synthetic route, with the steps as follows:

[0378] For the synthesis of pICz, see Example 14.

[0379] Liquid bromine (Br2) was dissolved in chloroform (CHCl3) and then slowly added dropwise at 0°C to a suspension of o-nitrobenzoyl chloride containing pICz. The mixture was stirred at 0°C, then gradually heated to room temperature while continuing stirring. After the reaction was complete, a saturated sodium sulfite solution was added to the reaction mixture. The organic layer was separated and washed with excess water. The organic solvent was removed by distillation, and the product was recrystallized from chloroform / ethanol (CHCl3 / EtOH) to give the crude product, a yellow solid, pICz-Br, in 87.9% yield.

[0380] 4-(N,N-diphenylamino)phenylboronic acid, pICz-Br, tetrakis(triphenylphosphine)palladium, potassium carbonate, ethanol, and toluene were mixed in a round-bottom flask. The mixture was refluxed and heated under an argon atmosphere. The solvent was evaporated under reduced pressure, and the resulting residue was extracted with dichloromethane and washed with deionized water. The organic phase was collected, dried over sodium sulfate, and then evaporated to dryness. Purification yielded a yellow solid, pICz-TPA, in 51.7% yield. NMR results were as follows: 1 H NMR (600 MHz, Chloroform- d ): δ 8.11(s, 4H), 7.79 (d, J = 8.3 Hz, 4H), 7.55 (d, J = 8.3 Hz, 4H), 7.43-7.34 (m,20H), 7.12 (ddd, J = 8.4, 6.2, 2.2 Hz, 4H), 5.95 (d, J = 8.7 Hz, 2H), 1.51(s, 18H), 1.49 (s, 18H).13 C NMR (151 MHz, Chloroform- d ): δ148.52, 147.95,145.80, 144.46, 144.13, 137.96, 134.96, 133.07, 132.45, 129.99, 129.48,129.12, 125.36, 124.19, 123.40, 123.03, 122.06, 118.83, 118.20, 118.03,117.76, 115.94, 113.53, 35.74, 34.70, 32.74, 31.92.

[0381] The synthesis of pICz-TPA@PS microspheres is described in Example 1. The emission peak is a single peak, and its full width at half maximum (FWHM) is measured to be 21 nm. The emission spectrum of the microspheres was tested, and the number of emission peaks at 5% or greater of the highest peak intensity was found to be one. The peak width at 1% of the highest peak intensity was 121 nm, and the luminescence lifetime was 4.06 ns. The diameter of the microspheres was measured to be between 500 nm and 515 nm. The emission spectrum of the luminescent molecules was tested, and the FWHM was measured to be 14 nm, with a peak width at 1% of the highest peak intensity of 103 nm.

[0382] Example 16

[0383] Synthesis of BOBO-Z@PS microspheres

[0384] The BOBO-Z molecule was prepared according to the following synthetic route, with the steps as follows:

[0385] (1) 1,3-Dibromo-5-methoxybenzene, benzidine, Pd(OAc)2, P(t-Bu)3H·BF4, and t-BuONa were mixed in dry toluene and refluxed under nitrogen. After cooling to room temperature, the reaction mixture was added to water and then filtered. The product was extracted with toluene and dried over anhydrous Na2SO4. After filtration and solvent evaporation, the crude product was purified. After solvent evaporation, the resulting solid was recrystallized from CH2Cl2 / MeOH to give a white solid, namely compound 1, with a yield of 85%.

[0386] (2) Compound 1 was dissolved in dry CH2Cl2 and stirred. BBr3 was slowly added dropwise at 0°C under a nitrogen atmosphere. The mixture was stirred at room temperature. After cooling to 0°C, a large amount of methanol and water were added to the reaction mixture for quenching. The product was extracted with CH2Cl2 and dried with anhydrous Na2SO4. After filtration and evaporation, the crude product was purified to give a white solid, namely compound 2, with a yield of 55%.

[0387] (3) A mixture of compound 2, 1,5-dibromo-2,4-difluorobenzene, and potassium carbonate was stirred in triethylene glycol dimethyl ether under nitrogen protection at a temperature maintained at 180°C. After cooling to room temperature, the reaction mixture was added to water and then extracted with dichloromethane. The combined organic layers were washed with water and dried with anhydrous sodium sulfate. After filtration and evaporation, the crude product was purified by recrystallization from dichloromethane / methanol to give a white solid, namely compound 3, in a yield of 70%.

[0388] (4) Compound 3 was dissolved in tert-butylbenzene, and n-butyllithium was slowly added dropwise under low temperature and nitrogen atmosphere with stirring. After reacting at 60°C, boron tribromide was slowly added and stirring continued. Subsequently, 1,2,2,6,6-pentamethylpiperidine was added to the reaction mixture, and stirring was further carried out at 160°C. After cooling to room temperature, the reaction mixture was added to water and extracted with toluene. The combined organic layers were washed with water and dried with anhydrous sodium sulfate. After filtration and evaporation, the crude product was purified by silica gel column chromatography and then recrystallized from dichloromethane / methanol to give BOBO-Z as a light yellow solid with a yield of 20%. The NMR results were as follows: 1 HNMR (400 MHz, (CDCl2)2, δ ): 10.17 (s, 1H), 9.12 (dd, J = 8.0, 1.1 Hz, 2H),7.53-7.48 (m, 6H), 7.43 (t, J = 7.3 Hz, 2H), 7.38 (t, J = 6.9 Hz, 2H), 7.34(s, 1H), 7.29-7.24 (m, 12H), 7.16-7.14 (m, 8H), 7.11 (t, J = 7.3 Hz, 4H), 6.85 (d, J = 8.2 Hz, 2H), 6.72 (d, J = 1.8 Hz, 2H), 5.75 (d, J = 1.8 Hz, 2H). 13 C{1H} NMR (100 MHz, CDCl3, δ): 163.03, 159.15, 152.63, 148.12, 147.15, 146.71, 142.14, 141.66, 134.92, 131.35, 130.65, 130.03, 129.23, 128.35, 126.06, 124.16, 123.71, 120.16, 119.45, 117.42, 111.06, 104.69, 100.84, 98.70.

[0389] The synthesis of BOBO-Z@PS microspheres is described in Example 1. The emission peak is a single peak, and its full width at half maximum (FWHM) is measured to be 15 nm. The emission spectrum of the microspheres was tested, and the number of emission peaks at 5% or greater of the highest peak intensity was found to be one. The peak width at 1% of the highest peak intensity was 120 nm, and the luminescence lifetime was 3.2 ns. The diameter of the microspheres was measured to be between 500 nm and 515 nm. The emission spectrum of the luminescent molecules was also tested, and the FWHM was measured to be 15 nm, with a peak width at 1% of the highest peak intensity of 119 nm.

[0390] Example 17

[0391] Synthesis of BSBS-Z@PS microspheres

[0392] The BSBS-Z molecule was prepared according to the following synthetic route, with the following steps:

[0393] (1) A mixture of 1,3,5-tribromobenzene, diphenylamine, Pd2(dba)3, SPhos, and sodium tert-butoxide was refluxed in dry toluene under nitrogen protection for 12 hours. After cooling to room temperature, the reaction mixture was added to water and then filtered through a Celite filter. The product was extracted with dichloromethane and dried over anhydrous sodium sulfate. After filtration and evaporation, the crude product was purified to give a white solid, compound 4, in 31% yield.

[0394] (2) Compound 4 was dissolved in dry diethyl ether, and n-butyllithium was slowly added at -70°C under nitrogen atmosphere. After stirring at -70°C for 1 hour, sulfur powder was added, and the reaction mixture was stirred at room temperature for 12 hours. After adding water, the precipitate was filtered and washed with methanol to give a white solid, namely compound 5, in 82% yield.

[0395] (3) Compound 5, 1,5-dibromo-2,4-diiodobenzene, t-BuONa, Pd2(dba)3, and DPEPos were mixed in dry o-xylene. The mixture was refluxed under nitrogen for 48 hours. After cooling to room temperature, the reaction mixture was added to water and then filtered through a Celite filter. The product was extracted with dichloromethane and dried over anhydrous sodium sulfate. After filtration and evaporation, the crude product was purified. After evaporation of the solvent, the resulting product, compound 6, was recrystallized from dichloromethane / methanol and used directly in the next reaction without further purification.

[0396] (4) Compound 6 was dissolved in dry tert-butylbenzene, and n-butyllithium was slowly added dropwise under low temperature and nitrogen atmosphere with stirring. After reacting at 60°C for 2 hours, boron tribromide was slowly added, and stirring was continued at room temperature for another 2 hours. Subsequently, 1,2,2,6,6-pentamethylpiperidine was added to the reaction mixture, and stirring was further carried out at 160°C for 24 hours. After cooling to room temperature, the reaction mixture was added to water, extracted with toluene, washed, dried, filtered, and evaporated. The crude product was purified and then recrystallized from dichloromethane / methanol to give BSBS-Z as a light yellow solid with a yield of 14%. The NMR results were as follows: 1 H NMR (400 MHz, DMSO- d 6, δ ): 9.67 (s, 1H), 8.74 (dd, J = 7.8, 1.4 Hz, 2H), 7.87 (s, 1H), 7.57(t, J = 7.5 Hz, 4H), 7.51 (t, J = 7.8 Hz, 2H), 7.46 (t, J = 7.3 Hz, 2H),7.34-7.28 (m, 14H), 7.15 (t, J = 7.3 Hz, 4H), 7.09 (dd, J = 8.5, 1.1 Hz, 8H),6.72-6.70 (m, 4H), 5.93 (d, J = 1.8 Hz, 2H). 13 C NMR (100 MHz, CDCl3, δ):150.47, 148.33, 147.58, 146.74, 146.46, 145.95,142.31, 141.60, 135.74,131.26, 130.65, 130.06, 129.27, 128.33, 125.87, 124.18, 121.21, 120.18,116.88, 108.59, 104.40.

[0397] The synthesis of BSBS-Z@PS microspheres is described in Example 1, and their full width at half maximum (FWHM) was measured to be 22 nm. The emission spectrum of the microspheres was tested, and the number of emission peaks at 5% or greater of the highest peak intensity was found to be one. The peak width at 1% of the highest peak intensity was 127 nm, and the luminescence lifetime was 1.3 ns. The diameter of the microspheres was measured to be between 500 nm and 515 nm. The emission spectrum of the luminescent molecules was also tested, and the FWHM was measured to be 21 nm, with a peak width at 1% of the highest peak intensity of 127 nm.

[0398] Example 18

[0399] Synthesis of BOBS-Z@PS microspheres

[0400] The BOBS-Z molecule was prepared according to the following synthetic route, with the steps as follows:

[0401] (1) The synthesis steps of compound 5 in the synthetic route are as described in Example 17.

[0402] (2) A mixture of compound 5, 1,5-dibromo-2,4-difluorobenzene, and potassium carbonate was stirred in triethylene glycol dimethyl ether under a nitrogen atmosphere at a temperature maintained at 140°C. After cooling to room temperature, the reaction mixture was added to water and then extracted with dichloromethane. The combined organic layers were washed with water and dried with anhydrous sodium sulfate. After filtration and evaporation, the crude product was purified by silica gel column chromatography to obtain a white solid powder, namely compound 7, in a yield of 32%.

[0403] (3) A mixture of compound 7, compound 2 (the synthesis steps of compound 2 are described in Example 16) and potassium carbonate in triethylene glycol dimethyl ether was stirred at 140°C for 12 hours under a nitrogen atmosphere. After cooling to room temperature, the reaction mixture was added to water and then extracted with dichloromethane, washed and dried. After filtration and evaporation, the crude product was purified to give a white solid, namely compound 8, with a yield of 44%.

[0404] (4) Compound 8 was dissolved in dry tert-butylbenzene, and n-butyllithium was slowly added dropwise at 0°C under a nitrogen atmosphere with stirring. After reacting at 60°C for 2 hours, boron tribromide was slowly added, and stirring was continued at room temperature for another 2 hours. Subsequently, 1,2,2,6,6-pentamethylpiperidine was added to the reaction mixture, and stirring was further carried out at 160°C for 24 hours. After cooling to room temperature, the reaction mixture was added to water, extracted with toluene, washed, and dried. After filtration and evaporation, the crude product was purified and recrystallized to give BOBS-Z as a light yellow solid with a yield of 20%. The NMR results were as follows: 1 H NMR (400 MHz, DMSO-d6, δ ): 9.87(s, 1H), 8.87 (d, J = 6.9 Hz, 1H), 8.77 (d, J = 6.9 Hz, 1H), 7.62-7.46 (m,9H), 7.40-7.27 (m, 14H), 7.18–7.08 (m, 12H), 6.77-6.72 (m, 3H), 6.40 (d, J =1.8 Hz, 1H), 5.94 (d, J = 1.6 Hz, 1H), 5.75 (d, J = 1.4 Hz, 1H). 13 C{1H} NMR (100 MHz, CDCl3, δ ): 160.69, 158.94, 152.70, 150.43, 148.28, 148.03, 147.71, 147.17, 146.61, 146.47, 144.78, 142.49, 141.66, 141.59, 135.78, 134.94, 131.35, 131.31, 130.65, 130.09, 129.99, 129.27, 129.24, 128.34, 126.11, 125.89, 124.23, 124.17, 120.19, 120.16, 117.35, 117.00, 113.02, 108.62, 104.33, 100.62, 98.45.

[0405] The synthesis of BOBS-Z@PS microspheres is described in Example 1, and their full width at half maximum (FWHM) was measured to be 21 nm. The emission spectrum of the microspheres was tested, and the number of emission peaks at 5% or greater of the highest peak intensity was found to be one. The peak width at 1% of the highest peak intensity was 124 nm, and the luminescence lifetime was 1.0 ns. The diameter of the microspheres was measured to be between 500 nm and 515 nm. The emission spectrum of the luminescent molecules was also tested, and the FWHM was measured to be 20 nm, with a peak width at 1% of the highest peak intensity of 124 nm.

[0406] Example 19

[0407] Synthesis of BTPT@PS microspheres

[0408] BTPT molecules were prepared according to the following synthetic route, with the steps as follows:

[0409] Compound 1 was dissolved in acetic acid and hydrogen peroxide. After reacting at 65°C for 12 hours, the reaction mixture was extracted with dichloromethane, washed with brine, and further purified by column chromatography using dichloromethane / petroleum ether (4:1, v / v) as eluent to give a white solid, BTPT, in 92% yield. NMR results were as follows: 1 H NMR (400 MHz, CDCl3-d): δ8.36 (d, J = 7.7 Hz, 2H), 8.23-8.16 (m, 2H), 7.72-7.66 (m, 3H), 7.64 (ddd, J = 8.4, 7.1, 1.6 Hz, 2H), 7.54 (ddd, J = 8.2, 7.1, 1.3 Hz, 2H). 13 C NMR (100MHz, DMSO-d6): δ 139.1, 136.4, 135.1, 129.3, 128.9, 128.6, 127.6, 126.7,124.3, 123.0.

[0410] The synthesis of BTPT@PS microspheres is described in Example 1, and their full width at half maximum (FWHM) was measured to be 33 nm. The emission spectrum of the microspheres was tested, and the number of emission peaks at 5% or greater of the highest peak intensity was found to be one. The peak width at 1% of the highest peak intensity was 146 nm, and the luminescence lifetime was 13.7 ns. The diameter of the microspheres was measured to be 1 μm. The emission spectrum of the luminescent molecules was tested, and the FWHM was measured to be 25 nm, with a peak width at 1% of the highest peak intensity of 135 nm.

[0411] Example 20

[0412] Synthesis of Na-dBN@PS microspheres

[0413] Na-dBN molecules were prepared according to the following synthetic route, with the steps as follows:

[0414] The Na-dBN in this example was synthesized using the method described in Example 3, with a yield of 15%. The NMR results are as follows: 1 H NMR (500 MHz, Chloroform-d): δ = 9.19 (s, 2H), 9.01 (d, J = 1.8 Hz, 2H), 8.52 (d, J = 2.2 Hz, 4H), 8.30-8.25 (m, 4H), 8.18 (d, J = 8.7 Hz, 2H), 8.13 (d, J =7.8 Hz, 2H), 7.70 (t, J = 8.0 Hz, 2H), 7.60 (dd, J = 8.7, 2.1 Hz, 2H), 1.71(s, 18H), 1.56 (s, 18H) ppm. 13 C NMR (151 MHz, CDCl3): δ = 145.53, 145.07,142.60, 141.93, 141.87, 139.41, 138.29, 137.70, 134.72, 133.19, 132.84,130.24, 128.30, 127.19, 125.01, 124.50, 124.23, 122.17, 121.40, 117.96,117.42, 114.41, 113.89, 35.45, 34.98, 32.44, 32.02 ppm.

[0415] The method for coating PS microspheres with Na-dBN prepared in this embodiment is the same as in Example 1, resulting in Na-dBN@PS microspheres. The emission spectrum of these microspheres was measured to be single-peaked, with an emission wavelength of 629 nm and a half-maximum width (FWHM) of 34 nm. Testing the emission spectrum of the microspheres revealed one emission peak at 5% intensity (greater than or equal to the highest peak value), a peak width of 147 nm at 1% intensity (greater than the highest peak value), and a luminescence lifetime of 4.76 ns. The microsphere diameter was measured to be 1 μm. Testing the emission spectrum of the luminescent molecules revealed a FWHM of 30 nm and a peak width of 139 nm at 1% intensity (greater than the highest peak value).

[0416] Example 21

[0417] Synthesis of tPhBODIPY@PS-NH2 microspheres

[0418] tPhBODIPY was synthesized according to the published literature Adv. Optical Mater. 2020, 8, 2000483. and Chem. Commun. 2013, 49, 993. The synthetic route is as follows:

[0419] In the above route, 4,4''-di-tert-butyl-[1,1',3',1''-terphenyl]-5'-carboxaldehyde is condensed with 2,4-dimethylpyrrole, followed by oxidation with 2,3-dichloro-5,6-dicyano-1,4-benzoquinone (DDQ), and then triethylamine (Et3N) and boron trifluoride (BF3) are added to prepare the target molecule tPhBODIPY in the synthetic route.

[0420] The method for coating tPhBODIPY prepared in this example into PS-NH2 (aminated polystyrene microspheres) microspheres is the same as in Example 1, except that the PS microspheres were replaced with PS-NH2 microspheres to obtain tPhBODIPY@PS-NH2 microspheres. The emission spectrum of these microspheres was measured to be single-peaked, with an emission wavelength of 520 nm and a half-maximum width (FWHM) of 32 nm. Testing the emission spectrum of the microspheres revealed one emission peak at 5% intensity (greater than or equal to the highest peak value), a peak width of 137 nm at 1% intensity, and a luminescence lifetime of 7.56 ns. The microsphere diameter was measured to be between 285 nm and 295 nm. Testing the emission spectrum of the luminescent molecules revealed a FWHM of 28 nm and a peak width of 122 nm at 1% intensity.

[0421] Example 22

[0422] Synthesis of ICz@PS microspheres

[0423] The specific synthetic route for ICz molecules is as follows:

[0424] Carbazole, 1-bromo-2-fluorobenzene, cesium carbonate, and N,N-dimethylformamide were added sequentially to a flask. The mixture was stirred and heated to 130°C for 12 hours under a nitrogen atmosphere. After the reaction was complete, the system was cooled to room temperature, poured into water, filtered under reduced pressure, and the residue was washed with plenty of water. The residue was further slurried with methanol and filtered to obtain the intermediate in the synthetic route of this example. The intermediate synthesized in the previous step, benzyltriethylammonium chloride, potassium carbonate, triphenylphosphine, palladium acetate, and N,N-dimethylacetamide were then added sequentially to a flask. The mixture was stirred and heated to 150°C for 3 hours under nitrogen protection. After the reaction, the system was cooled to room temperature, poured into water, filtered under reduced pressure, and the residue was washed with plenty of water. The residue was separated and purified on a silica gel column using a dichloromethane:petroleum ether ratio of 1:20 (volume ratio) as eluent to obtain the target molecule ICz in 89.5% yield.

[0425] Following the method in Example 1, ICz was coated into polystyrene microspheres to obtain ICz@PS microspheres.

[0426] See emission spectrum test results Figure 7 ,in Figure 7 (a) is the emission spectrum of the organic light-emitting molecule ICz in toluene solution; Figure 7 (b) shows the emission spectrum of ICz@PS microspheres in toluene solution, with an emission peak at 375 nm, a full width at half maximum (FWHM) of 35 nm, and a luminescence quantum efficiency of 40%. Figure 8 The ICz@PS microspheres exhibit good uniformity, with a particle size range of 285 nm to 295 nm. The emission spectrum of the microspheres was tested, revealing one emission peak at 5% intensity (or greater than or equal to the peak value), a peak width of 110 nm at 1% intensity, and a luminescence lifetime of 12.5 ns. The emission spectrum of the luminescent molecules was also tested, showing a full width at half maximum (FWHM) of 21 nm and a peak width of 99 nm at 1% intensity.

[0427] Example 23

[0428] The synthetic route of the target molecule in this embodiment is as follows:

[0429] (1) Under an argon atmosphere, 1-bromo-2,6-difluorobenzene, 1-aminonaphthalene, potassium carbonate, and NMP were added to a flask, and the mixture was heated and stirred for 6 hours. After the reaction was cooled to room temperature, the solid was filtered out, washed with deionized water, and dried to obtain the product.

[0430] (2) Under an argon atmosphere, the product obtained in the first step, 3,6-di-tert-butylcarbazole, cesium carbonate, and anhydrous DMF were added to a flask, and the mixture was heated and stirred. After the reaction was cooled to room temperature, the solid was filtered out, washed with deionized water, and dried under reduced pressure to obtain the product.

[0431] (3) Under an argon atmosphere, the product obtained in step two and dried o-xylene were added to a flask. A solution of n-butyllithium was added dropwise at -30°C. After the addition was complete, the mixture was stirred at -30°C, then stirred again at room temperature. The temperature was then lowered to -30°C, and boron tribromide was added dropwise to the system. After the addition was complete, the mixture was stirred at room temperature. The temperature was lowered again to 0°C, and N,N-diisopropylethylamine was added dropwise to the reaction system. After the addition was complete, the temperature was raised to allow the reaction to proceed. After the reaction was cooled to room temperature, the solid precipitated in the system was filtered and washed with methanol. The crude product was separated by column chromatography to obtain the target molecule.

[0432] The target molecule was coated in polystyrene microspheres according to the method in Example 1 to obtain luminescent microspheres.

[0433] See emission spectrum test results Figure 9 ,in Figure 9 (a) is the emission spectrum of the luminescent molecule in toluene solution; Figure 9 (b) is the emission spectrum of the microsphere containing the luminescent molecules in toluene solution, showing that the emission peak is a single peak with a half-peak width of 30 nm. Figure 10 The microspheres containing the luminescent molecules exhibit good uniformity. The emission spectra of the microspheres were tested, revealing one emission peak at 5% or greater of the peak intensity, a peak width of 131 nm at 1% of the peak intensity, and a luminescence lifetime of 6.71 ns. The microsphere diameter was measured to be between 305 nm and 320 nm. The emission spectra of the luminescent molecules were also tested, showing a full width at half maximum (FWHM) of 28 nm and a peak width of 120 nm at 1% of the peak intensity.

[0434] Example 24

[0435] The synthetic route of the target molecule in this embodiment is as follows:

[0436] (1) Under argon atmosphere, 1,4-dibromo-2,6-difluorobenzene, 4-aminobromobenzene, potassium carbonate and NMP were added to a flask, and the mixture was heated and stirred. After the reaction was cooled to room temperature, the solid was filtered out, washed with deionized water, and dried under reduced pressure to obtain the product.

[0437] (2) Under argon atmosphere, the product obtained in step one, 3,6-di-tert-butylcarbazole, cesium carbonate and anhydrous DMF were added to a flask, and the mixture was heated and stirred. After the reaction was cooled to room temperature, the solid was filtered out, washed with deionized water, and dried under reduced pressure to obtain the product.

[0438] (3) Under argon atmosphere, the product obtained in step two and dried o-xylene were added to a flask. A solution of n-butyllithium was added dropwise at -30°C. After the addition was complete, the mixture was stirred at -30°C for 10 minutes, then brought to room temperature and stirred for two hours. The temperature was then lowered to -30°C, and boron tribromide was added dropwise to the system. After the addition was complete, the mixture was stirred at room temperature for 1 hour. The temperature was then lowered again to 0°C, and N,N-diisopropylethylamine was added dropwise to the reaction system. After the addition was complete, the temperature was raised to allow the reaction to proceed. After the reaction was cooled to room temperature, the solid precipitated in the system was filtered and washed with methanol. The crude product was separated by column chromatography to obtain the final product.

[0439] (4) Under argon atmosphere, add the product obtained in step 3 and dry tetrahydrofuran to a flask, and add n-butyllithium solution dropwise at -78°C. After stirring the reaction after the addition is complete, add the reactants dropwise. The reaction solution was brought back to room temperature and stirred. The reaction solution was diluted with ethyl acetate, washed three times with saturated sodium chloride aqueous solution, the organic phase was collected and dried over MgSO4, and the crude product was separated by column chromatography to obtain the final product.

[0440] The target molecule was coated in polystyrene microspheres according to the method in Example 1 to obtain luminescent microspheres.

[0441] See emission spectrum test results Figure 11 ,in Figure 11 (a) is the emission spectrum of the luminescent molecule in toluene solution; Figure 11 (b) is the emission spectrum of the microsphere containing the luminescent molecules in toluene solution, showing that the emission peak is a single peak with a half-peak width of 33 nm. Figure 12 The microspheres containing the luminescent molecules exhibit good uniformity. The emission spectra of the microspheres were tested, revealing one emission peak at 5% or greater of the peak intensity, a peak width of 122 nm at 1% of the peak intensity, and a luminescence lifetime of 7.81 ns. The microsphere diameter was measured to be between 285 nm and 305 nm. The emission spectra of the luminescent molecules were also tested, showing a full width at half maximum (FWHM) of 22 nm and a peak width of 112 nm at 1% of the peak intensity.

[0442] Example 25

[0443] The synthetic route of the target molecule in this embodiment is as follows:

[0444] An anhydrous N,N-dimethylformamide solution containing 3,6-di-tert-butyl-9H-carbazole was slowly added dropwise to an anhydrous N,N-dimethylformamide solution containing potassium tert-butoxide. After stirring at 25°C for 2 hours, an anhydrous N,N-dimethylformamide solution containing 2-bromo-1,3-difluorobenzene was added dropwise over 15 minutes. The solution was then heated to 158°C and stirred for 24 hours before being cooled in ice water. The solution was extracted three times consecutively with dichloromethane. The combined organic layers were dried over anhydrous sodium sulfate, filtered, and concentrated under vacuum. Further purification was achieved by column chromatography (eluent: petroleum ether) to obtain a white solid intermediate 1.

[0445] Under a nitrogen atmosphere and at -75°C, a hexane solution of n-butyllithium was slowly added to a toluene solution containing intermediate 1. The temperature was then slowly raised to 68°C, and the mixture was stirred for 3 hours. The hexane was then removed under vacuum. The temperature was then lowered to -48°C, boron tribromide was added, and the mixture was stirred at 25°C for 8.5 hours. N,N-diisopropylethylamine was then added at 8°C, and the solution was heated to 128°C and stirred for 12 hours. The solution was then cooled in ice water and extracted three times consecutively with dichloromethane. The combined organic layers were dried over anhydrous sodium sulfate, filtered, and concentrated under vacuum. Further purification was achieved by column chromatography (eluent: petroleum ether) to obtain the target molecule.

[0446] The target molecule was coated in polystyrene microspheres according to the method in Example 1 to obtain luminescent microspheres.

[0447] See emission spectrum test results Figure 13 ,in Figure 13 (a) is the emission spectrum of the organic light-emitting molecule in toluene solution in this embodiment; Figure 13 (b) is the emission spectrum of the single-peak narrow-band luminescent microsphere in toluene solution in this embodiment, and its emission peak is a single peak. Figure 14 The single-peak, narrow-band luminescent microspheres exhibit good uniformity. The emission spectrum of the microspheres was tested, showing a full width at half maximum (FWHM) of 33 nm. There was one emission peak at 5% of the maximum intensity, and the peak width at 1% of the maximum intensity was 121 nm. The luminescence lifetime was 8.5 ns. The microsphere diameter was measured to be between 285 nm and 305 nm. The emission spectrum of the luminescent molecules was also tested, showing a FWHM of 22 nm and a peak width at 1% of the maximum intensity of 112 nm.

[0448] Example 26

[0449] The synthetic route of the target molecule in this embodiment is as follows:

[0450] The synthesis of the target molecule in this embodiment is the same as in Example 25, except that 3,6-di-tert-butyl-9H-carbazole in the synthetic route is replaced with 3,6-diphenyl-9H-carbazole.

[0451] The target molecule was coated in polystyrene microspheres according to the method in Example 1 to obtain luminescent microspheres.

[0452] See emission spectrum test results Figure 15 ,in Figure 15 (a) is the emission spectrum of the organic light-emitting molecule in toluene solution in this embodiment. Figure 15 (b) is the emission spectrum of the single-peak narrow-band luminescent microsphere in toluene solution in this embodiment, and its emission peak is a single peak. Figure 16 The single-peak, narrow-band luminescent microspheres exhibit good uniformity. The emission spectrum of the microspheres was tested, showing a full width at half maximum (FWHM) of 34 nm. There was one emission peak at 5% intensity (≥5% of the peak value), and the peak width at 1% intensity of the peak value was 134 nm. The luminescence lifetime was 11.1 ns. The microsphere diameter was measured to be between 285 nm and 305 nm. The emission spectrum of the luminescent molecules was also tested, showing a FWHM of 21 nm and a peak width at 1% intensity of the peak value of 123 nm.

[0453] Example 27

[0454] This embodiment provides t Synthesis of Bu-TiTAT@PS microspheres.

[0455] t For the synthesis steps of Bu-TiTAT, please refer to Chemical Engineering Journal 466 (2023) 143423. t NMR results of Bu-TiTAT: 1 H NMR (400 MHz, CD2Cl2) δ (ppm): 8.77-8.68 (m,1H), 8.01 (dd, J = 71.9, 17.5 Hz, 4H), 7.56 (dt, J = 7.7, 3.8 Hz, 3H), 7.49(t, J = 7.4 Hz, 1H), 7.38 (dd, J = 8.4, 5.4 Hz, 3H), 7.26 (dt, J = 28.6, 9.7Hz, 5H), 5.55 (dd, J = 18.2, 8.1 Hz, 1H), 1.46 - 1.39 (m, 27H).

[0456] t The structure of Bu-TiTAT is as follows:

[0457] t The synthesis of Bu-TiTAT@PS microspheres is described in Example 1.

[0458] First, the above-prepared... t Emission spectroscopy of the Bu-TiTAT molecule was performed, revealing a double peak shape. The measured half-width at half-maximum (WHM) was 18 nm, and the peak width at 1% intensity was 125 nm. Further analysis of the prepared... t The emission spectrum of Bu-TiTAT@PS microspheres was tested. The half-width of the microspheres was 21 nm, the peak width at 1% intensity of the highest peak was 125 nm, the number of emission peaks at ≥5% intensity of the highest peak was 2, the diameter of the microspheres was 170 μm, and the luminescence lifetime of the microspheres was in the nanosecond range.

[0459] Example 28

[0460] This embodiment provides t Synthesis of Bu-TTAT@PS microspheres.

[0461] t The synthesis steps of Bu-TTAT can be found in Chemical Engineering Journal 466 (2023) 143423. NMR results: 1 H NMR (400 MHz, CD2Cl2) δ (ppm): 8.38 (d, J = 4.2 Hz, 3H), 8.10 -7.98 (m, 9H), 7.45 (d, J = 6.7 Hz, 3H), 1.55 (s, 27H), 1.46 (s, 27H).

[0462] t The structure of Bu-TTAT is as follows:

[0463] t The synthesis of Bu-TTAT@PS microspheres is described in Example 1.

[0464] First, the above-prepared... t Emission spectroscopy of the Bu-TTAT molecule was performed, revealing a shoulder-shaped peak with a measured half-width of 30 nm and a peak width of 98 nm at 1% intensity. Further analysis of the prepared... tThe emission spectrum of Bu-TTAT@PS microspheres was tested. The half-width of the microspheres was 35 nm, the peak width at 1% intensity of the highest peak was 104 nm, the number of emission peaks at ≥5% intensity of the highest peak was 1, the diameter of the microspheres was 130 μm, and the luminescence lifetime of the microspheres was in the nanosecond range.

[0465] Example 29

[0466] This embodiment provides the synthesis of ACDID-Ph@PS microspheres.

[0467] The synthesis procedure for ACDID-Ph can be found in Adv. Optical Mater. 2020, 8, 2000480. NMR results: 1 H NMR (500 MHz, CDCl3) δ: 8.39 (d, J = 8.1 Hz, 1H), 7.91 (d, J = 7.9 Hz, 1H), 7.82 (d, J = 8.1 Hz, 2H), 7.71 (t, J = 7.5 Hz, 2H), 7.61 – 7.37 (m, 7H), 7.19 (t, J = 7.7 Hz, 1H), 7.02 – 6.79 (m, 3H), 6.13 (s, 1H), 2.29 (s, 6H). 13 CNMR (126 MHz, CDCl3): δ 139.00 (s), 138.58 (s), 131.95 (s), 131.69 (s), 129.79 (d, J = 11.4 Hz), 128.36 (s), 127.63 (s), 126.70 (s), 126.23 (s), 123.02 (s), 121.51 (s), 121.22 (s), 119.77 (s), 116.87 (s), 114.38 (s),112.35 (s), 112.05 (s), 108.45 (s), 34.51 (d, J = 13.1 Hz).

[0468] The structure of ACDID-Ph is as follows:

[0469] The synthesis of ACDID-Ph@PS microspheres is described in Example 1. First, the emission spectrum of the prepared ACDID-Ph molecules was measured, revealing a half-width at half-maximum (WHM) of 30 nm and a peak width at 1% intensity of the highest peak of 103 nm. Next, the emission spectrum of the prepared ACDID-Ph@PS microspheres was measured, showing a WHM of 33 nm, a peak width at 1% intensity of the highest peak of 106 nm, and one emission peak at ≥5% intensity of the highest peak. The microsphere diameter was 120 μm, and the luminescence lifetime was in the nanosecond range.

[0470] Example 30

[0471] This embodiment provides the synthesis of ACDID-TPA@PS microspheres.

[0472] For the synthesis steps of ACDID-TPA, please refer to Adv. Optical Mater. 2020, 8, 2000480.

[0473] MRI results: 1 H NMR (500 MHz, CDCl3): δ 8.38 (d, J = 8.1 Hz, 1H), 7.92 (d, J = 7.3 Hz, 2H), 7.84 (d, J = 7.7 Hz, 1H), 7.55 (t, J = 7.6 Hz, 2H), 7.48 (t, J = 7.7 Hz, 2H), 7.44-7.31 (m, 9H), 7.31-7.18 (m, 8H), 7.11 (t, J = 7.3 Hz, 2H), 2.28 (s, 6H). 13 C NMR (126 MHz, CDCl3): δ 147.71 (d, J = 4.4 Hz), 139.07(s), 138.67 (s), 130.07-129.61 (m), 127.66 (s), 126.77 (s), 126.24 (s), 125.21 (d, J = 25.9 Hz), 125.10-124.94 (m), 123.80 (s), 123.00 (s), 121.57(s), 121.27 (s), 112.39 (s), 112.15 (s), 34.68 (s).

[0474] The structure of ACDID-TPA is as follows:

[0475] The synthesis of ACDID-TPA@PS microspheres is described in Example 1. First, the emission spectrum of the prepared ACDID-TPA molecules was measured, revealing a half-width at half-maximum (WHM) of 31 nm and a peak width at 1% intensity of the highest peak of 107 nm. Next, the emission spectrum of the prepared ACDID-Ph@PS microspheres was measured, showing a WHM of 35 nm, a peak width at 1% intensity of the highest peak of 108 nm, and one emission peak at ≥5% intensity of the highest peak. The microsphere diameter was 125 μm, and the luminescence lifetime was in the nanosecond range.

[0476] Example 31

[0477] This application provides p Synthesis of FLDID@PS microspheres.

[0478] p -For the synthesis steps of FLDID, please refer to Adv. Optical Mater. 2024, 12, 2300551. NMR results: 1 H NMR (500 MHz, CDCl3): δ 8.67 (s, 2H), 8.49 (s, 2H), 8.27 (d, J = 7.3 Hz, 2H), 8.15 (d, J = 7.3 Hz, 2H), 8.09 (s, 2H), 7.89 (d, J = 7.4 Hz, 2H), 7.69(t, J = 7.3 Hz, 2H), 7.54 (d, J = 7.3 Hz, 2H), 7.43 (t, J = 7.2 Hz, 2H), 7.36(t, J = 7.3 Hz, 2H), 1.75 (s, 12H).

[0479] p The structure of -FLDID is as follows:

[0480] p The synthesis of -FLDID@PS microspheres is described in Example 1. Firstly, the microspheres prepared above... pEmission spectroscopy of the -FLDID molecule was performed, revealing a half-width at half-maximum (WHM) of 24 nm and a peak width at 1% intensity of 87 nm. Further analysis of the prepared... p The emission spectrum of the FLDID@PS microspheres was tested. The half-width of the microspheres was 26 nm, the peak width at 1% intensity of the highest peak was 88 nm, the number of emission peaks at ≥5% intensity of the highest peak was 1, the diameter of the microspheres was 144 μm, and the luminescence lifetime of the microspheres was in the nanosecond range.

[0481] Example 32

[0482] This embodiment provides the synthesis of BN-ICz-2@PS microspheres.

[0483] The synthesis steps for BN-ICz-2 can be found in Angew. Chem. Int. Ed. 2022, 61, e202202380. NMR results: 1 H NMR (600 MHz, Methylene Chloride-d2): δ 8.57 (s, 1H), 8.45 (d, J =14.2 Hz, 2H), 8.34-8.22 (m, 4H), 8.18 (s, 3H), 8.13 (d, J = 7.5 Hz, 1H), 8.07(d, J = 8.1 Hz, 1H), 8.00 (s, 1H), 7.93 (d, J = 7.7 Hz, 1H), 7.90-7.78 (m,3H), 7.69-7.58 (m, 4H), 7.54 (d, J = 8.8 Hz, 2H), 7.46 (t, J = 7.7 Hz, 1H), 7.40 (t, J = 7.5 Hz, 1H), 7.30 (t, J = 7.2 Hz, 1H), 7.00 (t, J = 7.5 Hz, 1H), 1.53 (s, 9H), 1.45 (s, 18H), 1.31 (s, 9H).

[0484] The structure of BN-ICz-2 is as follows:

[0485] The synthesis of BN-ICz-2@PS microspheres is described in Example 1. First, the emission spectrum of the prepared BN-ICz-2 molecule was measured. The peak shape was a shoulder peak, the half-width at half-maximum (WHM) was 20 nm, and the peak width at 1% intensity of the highest peak was 105 nm. Next, the emission spectrum of the prepared BN-ICz-2@PS microspheres was measured. The microsphere's WHM was 23 nm, the peak width at 1% intensity of the highest peak was 103 nm, the number of emission peaks at ≥5% intensity of the highest peak was 1, the microsphere diameter was 230 μm, and the microsphere luminescence lifetime was in the nanosecond range.

[0486] Example 33

[0487] This embodiment provides the synthesis of IDCz-B-II@PS microspheres.

[0488] The synthesis procedure for IDCz-B-II can be found in Adv. Optical Mater. 2025, 13, 2500384. NMR results: 1 H NMR (400 MHz, (CDCl2)2): δ 10.02 (s, 2H), 8.62-8.58 (m, 4H), 8.52 (d, J = 8.2 Hz, 2H), 8.07 (t, J = 8.2 Hz, 1H), 7.82-7.78 (m, 10H), 7.61-7.54 (m,4H), 7.49 (t, J = 8.0 Hz, 2H), 7.44 (d, J = 7.8 Hz, 2H), 7.03 (t, J = 7.8 Hz, 2H), 6.32 (d, J = 8.2 Hz, 2H).

[0489] The structure of IDCz-B-II is as follows:

[0490] The synthesis of IDCz-B-II@PS microspheres is described in Example 1. First, the emission spectrum of the prepared IDCz-B-II molecule was measured. It showed a single peak shape, a half-width at half-maximum (WHM) of 16 nm, and a peak width at 1% intensity of the highest peak of 121 nm. Next, the emission spectrum of the prepared IDCz-B-II@PS microspheres was measured. The microspheres had a WHM of 19 nm, a peak width at 1% intensity of the highest peak of 124 nm, and one emission peak at ≥5% intensity of the highest peak. The microsphere diameter was 93 μm, and the luminescence lifetime was in the nanosecond range.

[0491] Example 34

[0492] This embodiment provides the synthesis of CzB4-oPh@PS microspheres.

[0493] The synthesis steps of CzB4-oPh can be found in J. Am. Chem. Soc. 2023, 145, 11504-11511. NMR results: 1 H NMR (500 MHz, CDCl3): δ 2.83 (s, 6H, b), 2.99 (s, 6H, s), 5.39 (s, 2H,k), 6.27 (s, 1H, p), 6.62-6.69 (6H, j, m, o), 6.88-6.92 (m, 8H, u, w), 7.04(d, J = 7.7 Hz, g), 7.26-7.33 (m, 8H, f, h, n), 7.36-7.44 (m, 4H, e, l), 7.47-7.53 (m, 8H, i, v), 8.20 (s, 2H, a), 8.24 (s, 2H, t), 8.80 (s, 2H, c), 9.10 (d, J = 7.5 Hz, 2H, d), 9.18 (s, 2H, r), 10.60 (s, 1H, q).

[0494] The structure of CzB4-oPh is as follows:

[0495] The synthesis of CzB4-oPh@PS microspheres is described in Example 1. First, the emission spectrum of the prepared CzB4-oPh molecule was measured. It was a single peak with a half-width at half-maximum (WHM) of 15 nm and a peak width at 1% intensity of the highest peak of 80 nm. Then, the emission spectrum of the prepared CzB4-oPh@PS microspheres was measured. The microspheres had a WHM of 18 nm, a peak width at 1% intensity of the highest peak of 85 nm, and one emission peak at ≥5% intensity of the highest peak. The microsphere diameter was 200 μm, and the luminescence lifetime was in the nanosecond range.

[0496] Example 35

[0497] This embodiment provides S Synthesis of -DOBNT@PS microspheres.

[0498] S The synthesis steps for -DOBNT can be found in Adv. Mater. 2022, 34, 2204253. NMR results: 1 H NMR (400 MHz, CDCl3): δ 8.93 (d, J = 6.9 Hz, 1H), 8.64 (s, 1H), 8.37-8.29 (m,2H), 8.19 (d, J = 7.1 Hz, 1H), 8.05 (d, J = 8.2 Hz, 1H), 7.64 (dt, J = 16.5,7.9 Hz, 2H), 7.58-7.52 (m, 1H), 7.41 (t, J = 7.4 Hz, 1H), 6.86 (d, J = 8.2Hz, 1H), 3.24-3.09 (m, 2H), 2.69-2.60 (m, 1H), 2.46 (dt, J = 17.4, 6.0 Hz,1H), 1.98-1.88 (m, 2H), 1.83-1.72 (m, 2H). 13C NMR (101 MHz, CDCl3): δ 159.16,154.98, 143.21, 142.78, 142.14, 139.66, 134.36, 133.57, 133.06, 131.96,126.89, 126.69, 125.19, 123.72, 123.62, 122.13, 121.90, 120.78, 114.32,109.82, 107.11, 30.07, 28.14, 23.35, 23.14.

[0499] S The structure of -DOBNT is as follows:

[0500] S The synthesis of -DOBNT @PS microspheres is described in Example 1. Firstly, the microspheres prepared above... S Emission spectroscopy of the -DOBNT molecule was performed, revealing a single peak with a half-maximum width (WHM) of 20 nm and a peak width of 125 nm at 1% intensity. Further analysis of the prepared... S The emission spectrum of the -DOBNT @PS microspheres was tested. The half-width of the microspheres was 23 nm, the peak width at 1% intensity of the highest peak was 128 nm, the number of emission peaks at ≥5% intensity of the highest peak was 1, the diameter of the microspheres was 230 μm, and the luminescence lifetime of the microspheres was in the nanosecond range.

[0501] Example 36

[0502] This embodiment provides V- Synthesis of DABNA@PS microspheres.

[0503] V- The synthesis procedure for DABNA can be found in Adv. Sci. 2023, 10, 2205070. NMR results: 1 H NMR (CDCl3, 400 MHz): δ 5.64-5.65 (s, 6H, d, o, p), 5.74 (s, 2H, k), 6.35 (t, J = 7.4 Hz, 2H, v), 6.48 (d, J= 8.8 Hz, 2H, t), 6.85-6.93 (m, 20H, a, c, l, n, u), 7.00-7.07 (m, 20H, b, g, j, m), 7.10-7.26 (m, 16H, e, f, h, i, q), 7.33 (t, J =7.6 Hz, 2H, s), 7.45 (t, J = 7.6 Hz, 4H, r), 9.00 (d, J = 6.8 Hz, 2H, w), 10.6 (s, 2H, x).

[0504] V- The structure of DABNA is as follows:

[0505] V- The synthesis of DABNA@PS microspheres is described in Example 1. Firstly, the microspheres prepared above... V- Emission spectroscopy of the DABNA molecule was performed, revealing a shoulder-shaped peak with a measured half-width at half-maximum (WHM) of 16 nm and a peak width at 1% intensity of 88 nm. Further analysis of the prepared... V- The emission spectrum of DABNA @PS microspheres was tested. The half-width of the microspheres was 18 nm, the peak width at 1% intensity of the highest peak was 91 nm, the number of emission peaks at ≥5% intensity of the highest peak was 1, the diameter of the microspheres was 350 μm, and the luminescence lifetime of the microspheres was in the nanosecond range.

[0506] Example 37

[0507] This embodiment provides V Synthesis of DABNA-F@PS microspheres.

[0508] V The synthesis steps for -DABNA-F can be found in Adv. Sci. 2023, 10, 2205070. NMR results: 1H NMR (CDCl3, 400 MHz): δ 5.71 (s, 2H, m or n), 5.75 (s, 2H, d or i), 5.79 (s, 2H, m or n), 5.85 (s, 2H, d or i), 6.36 (t, J = 7.4 Hz, 2H, t), 6.51 (d, J = 8.4 Hz, 2H,r), 6.76-6.80 (m, 8H, f, h), 6.88-6.91 (m, 8H, a, j, s), 6.96-7.17 (m, 28H,b, c, e, g, k, l), 7.24 (d, J = 9.2 Hz, 4H, o), 7.33 (d, J = 7.6 Hz, 2H, q), 7.45 (t, J = 7.6 Hz, 4H, p), 9.00 (d, J = 7.6 Hz, 2H, u), 10.6 (s, 2H, v).

[0509] V The structure of -DABNA-F is as follows:

[0510] V The synthesis of -DABNA-F @PS microspheres is described in Example 1. Firstly, the microspheres prepared above... V Emission spectroscopy of the -DABNA-F molecule was performed, revealing a single peak with a measured half-maximum width (WHM) of 17 nm and a peak width of 95 nm at 1% intensity. Further analysis of the prepared... V The emission spectrum of the DABNA-F @PS microspheres was tested. The half-width of the microspheres was 21 nm, the peak width at 1% intensity of the highest peak was 98 nm, the number of emission peaks at ≥5% intensity of the highest peak was 1, the diameter of the microspheres was 300 μm, and the luminescence lifetime of the microspheres was in the nanosecond range.

[0511] Comparative Example 1

[0512] The specific synthetic route for the target molecule in this comparative example is as follows:

[0513] The synthesis of the target molecule in this comparative example is described in Example 25, except that 3,6-di-tert-butyl-9H-carbazole in the synthetic route is replaced with carbazole.

[0514] Following the method described in Example 1, the target molecule was coated into polystyrene microspheres to obtain luminescent microspheres, and their full width at half maximum (FWHM) was measured. The results are shown in [reference needed]. Figure 19 . Figure 19 The emission spectrum of the luminescent microspheres in toluene solution in this comparative example is shown, with a single emission peak. The emission spectrum of the microspheres was tested, and the measured full width at half maximum (FWHM) was 48 nm. There was one emission peak at 5% intensity (≥5% of the peak value), and the peak width at 1% intensity of the peak value was 122 nm. The luminescence lifetime was 8.5 ns. The diameter of the microspheres was measured to be between 287 nm and 305 nm. The emission spectrum of the luminescent molecules was tested, and the measured FWHM was 33 nm, with a peak width at 1% intensity of the peak value of 114 nm.

[0515] pass Figure 19 Comparing the emission spectra of the microspheres in Examples 25, 26, and Comparative Example 1 in the same figure, it was found that due to the addition of sterically hindered groups (e.g., tert-butyl, phenyl), the full width at half maximum (FWHM) of Examples 25 and 26 was significantly reduced compared to Comparative Example 1. The relative reduction in FWHM was approximately 35% in one group and approximately 43% in the other. Overall, the reduction in FWHM was between 35% and 45%, which is quite significant. This indicates that adding sterically hindered groups can indeed narrow the FWHM of the single-peak narrow-band luminescent microspheres formed when organic light-emitting molecules are coated within them.

[0516] Comparative Example 2

[0517] Referring to the published literature Chem. Commun., 2013, 49, 993, the following synthetic route was used to synthesize borane-boron fluoride dipyrrole (borane-BODIPY) as shown in the formula:

[0518] The target compound was coated in PS spheres in the manner described in Example 1. It was found that it exhibited a double peak under 350 nm excitation, with the main emission peak located at 520 nm and the secondary emission peak located at 410 nm.

[0519] The emission peak shifts of the microspheres in different solvents were tested, including hexane, toluene, dichloromethane, chloroform, ethyl acetate, methanol, acetonitrile, and N,N-dimethylformamide. Emission spectra were measured. The results showed that the peak positions of the emission spectra exhibited a blue or red shift of approximately 10 nm, but the full width at half maximum (FWHM) remained unchanged. This indicates that the number of emission peaks for small organic molecules is primarily influenced by their structure. Although the target compound in this comparative example has a similar structure to the compound in the aforementioned BODIPY derivative examples, it was found to completely fail to meet the requirement of a narrow single-peak band.

[0520] The emission spectrum of the luminescent microspheres prepared in this comparative example was tested, and the full width at half maximum (FWHM) was measured to be 42 nm. There were two emission peaks at 5% intensity (≥5% of the peak value), and the peak width at 1% intensity (≥136 nm) of the peak value was observed. The luminescence lifetime was 7.6 ns. The microsphere diameter was measured to be between 285 nm and 295 nm. The emission spectrum of the luminescent molecules was also tested, and the FWHM was measured to be 38 nm, with a peak width at 1% intensity (≥125 nm).

[0521] Comparative Example 3

[0522] Referring to the published literature Chem. Commun., 2013, 49, 993, the following method was used to synthesize borane-boron fluoride dipyrrole (borane-BODIPY), and the synthetic steps are as follows:

[0523] The specific synthesis process of this comparative example can be found in Comparative Example 1. The obtained target compound was coated in PS spheres in the manner described in Example 1. It was found that it had two emission peaks at an excitation wavelength of 350 nm, with the main peak located near 520 nm and the secondary peak located near 410 nm. Compared with Comparative Example 2, the full width at half maximum (WHM) was slightly increased, indicating that the addition of alkyl substituents did not significantly affect its peak shape.

[0524] The emission spectrum of the luminescent microspheres prepared in this comparative example was tested, and the full width at half maximum (FWHM) was measured to be 43 nm. There were two emission peaks at 5% intensity (greater than or equal to the peak value), and the peak width at 1% intensity (greater than the peak value) was 138 nm. The luminescence lifetime was 7.8 ns. The microsphere diameter was measured to be between 284 nm and 296 nm. The emission spectrum of the luminescent molecules was also tested, and the FWHM was measured to be 37 nm, with a peak width at 1% intensity (greater than the peak value) of 123 nm.

[0525] Comparative Example 4

[0526] Preparation of AgInS2 / ZnS quantum dots

[0527] Refer to publicly available documents ACS Omega2024, 9, 39287-39295. Synthesis of AgInS2 / ZnS quantum dots: Under magnetic stirring, aqueous solutions of silver nitrate, mercaptoacetic acid, and ammonium hydroxide were added to water to form an aqueous solution. After adding 0.45 mL of ammonium hydroxide aqueous solution, the resulting turbid, pale yellow suspension became transparent. Further addition of 0.7 mL of indium chloride in nitric acid aqueous solution turned the solution colorless. Next, 1.0 M sodium sulfide (Na2S) aqueous solution was added while stirring. The pH of the mixture at this point was 8-9. The resulting solution was heated in a water bath at 90℃-95℃ for 30 minutes. Subsequently, the indium sulfide (AgInS2) core was coated with a zinc sulfide (ZnS) shell. Under vigorous stirring, aqueous mercaptoacetic acid was added to a zinc acetate (Zn(CH3COO)2, 1.0 M) in nitric acid (HNO3) aqueous solution. The mixture was then heated for an additional 30 minutes to allow the shell to grow. Finally, the synthesized quantum dots were collected, evaporated, and then isopropanol (in a ratio of 1:2) was added to precipitate them. The product was then collected by centrifugation.

[0528] The average size of the AgInS2 / ZnS quantum dots prepared by dynamic light scattering analysis was 7.8 nm ± 0.5 nm, and the scanning electron microscopy (SEM) results showed that the prepared quantum dots were spherical.

[0529] Comparative Example 5

[0530] The AgInS2 / ZnS quantum dots in Comparative Example 4 were coated into carboxylated polystyrene microspheres according to the method of patent application CN107699230A. The steps are as follows: The AgInS2 / ZnS quantum dots prepared in Comparative Example 4 were dissolved in toluene and washed multiple times, then centrifuged to precipitate the quantum dots, and finally dissolved in chloroform. The treated quantum dots were added to chloroform to obtain a chloroform solution of the quantum dots. Carboxylated polystyrene microspheres were dispersed in dichloromethane to obtain a microsphere dispersion. The microsphere dispersion was added to the chloroform solution of the quantum dots and dispersed evenly for later use, resulting in a quantum dot-microsphere blend. The quantum dot-microsphere blend was added to an aqueous solution of PVA and mixed with stirring. Finally, the prepared quantum dot microspheres were centrifuged, the supernatant was removed, and the mixture was washed with pure water.

[0531] Using the same method as in Comparative Example 4, the diameter of the quantum dot microspheres was determined to be 49 nm, and the fluorescence efficiency was 79%. The emission spectrum of the luminescent microspheres prepared in this comparative example was tested, and the full width at half maximum (FWHM) was measured to be 134 nm. There was one emission peak at 5% intensity (≥5% of the peak value), the peak width at 1% intensity of the peak value was 400 nm, and the luminescence lifetime was 300 ns. The emission spectrum of the AgInS2 / ZnS quantum dots was tested, and the FWHM was measured to be 100 nm, with a peak width at 1% intensity of the peak value of 300 nm.

[0532] Comparative Example 6

[0533] Synthesis of Eu(TTA)3-TPA-DPA-mCF3@PS microspheres

[0534] The synthetic route for TPA-DPA-mCF3 is as follows:

[0535] The synthesis of TPA-DPA-mCF3 was performed according to J. Mater. Chem. C, 2022, 10, 7201. The procedure was as follows: TPA-DPA-CHO was added to a glacial acetic acid solution of 1,10-phenanthroline-6,7-dione under stirring at room temperature. Ammonium acetate and mCF3-amine were then added to the reaction mixture. The resulting mixture was then stirred at 110°C for 12 hours. The reaction mixture was poured into a small amount of water, followed by the addition of ammonium hydroxide solution. The resulting solid was then filtered, dissolved in dichloromethane, dried over anhydrous sodium sulfate, and the solvent was evaporated to obtain the crude product. The obtained compound was purified by silica gel (100-200 mesh) column chromatography using chloroform containing 5% methanol as the eluent. After solvent evaporation, the residue was dissolved in a small amount of tetrahydrofuran (THF) solution, and excess hexane was added to form a pale yellow solid. After the solid settled, it was decanted, and this process was repeated three times to obtain the pale yellow solid compound. MRI results: 1 H-NMR Data (CDCl3, 400MHz): δ 9.24 (s, 1H), 9.18 (d, J = 7.7 Hz, 2H), 9.10 (s, 1H), 7.97 (d, J = 8.1 Hz, 1H), 7.81 (dd, J= 7.9, 4.5 Hz, 2H), 7.75 (d, J = 8.1 Hz, 2H), 7.58 (d, J = 8.6 Hz, 1H), 7.38 (m, J = 6.3 Hz, 3H), 7.28 (m, 6H), 7.13 (m, J = 7.7 Hz, 5H), 7.04 (m, J =15.7, 8.2 Hz, 6H), 6.89 (d, J = 8.8 Hz, 1H). 13C-NMR Data (CDCl3, 100MHz): δ148.80, 147.98, 147.70, 138.78, 132.36, 131.29, 130.11, 129.27, 127.59,127.08, 126.40, 126.23, 124.89, 124.25, 124.09, 123.74, 122.77, 122.77,122.46, 119.57, 77.35, 77.24, 77.04, 76.72, 67.10.

[0536] Eu(TTA)3(H2O)2 was dissolved in anhydrous THF and stirred continuously in a round-bottom flask until a clear solution was obtained. Then, a solution of the ligand (TPA-DPA-mCF3) dissolved in anhydrous THF was slowly added dropwise. The reaction mixture was stirred at 60°C for 6 hours under an inert atmosphere. The reaction progress was monitored by thin-layer chromatography (TLC). After the reaction was complete, the mixture was concentrated using a rotary evaporator, and the residue was dissolved with a minimum volume of THF. Excess n-hexane was slowly added along the wall of the round-bottom flask to this solution, causing a precipitate to form. The supernatant was carefully decanted, and the solid residue was collected and dried. The above precipitation purification steps were repeated several times to finally obtain a pure powder of the target complex Eu(TTA)3-TPA-DPA-mCF3.

[0537] Microspheres were prepared according to the method disclosed in patent application CN118480059A. A mixture of Eu(TTA)3-TPA-DPA-mCF3 dye, polystyrene microspheres and tetrahydrofuran was ultrasonically treated to obtain a clear solution. 5 mL of deionized water was rapidly injected into the mixture, and the mixture was heated to 50 °C and stirred for 2 h in a fume hood. Then, the mixture was concentrated by rotary evaporation and filtered through a membrane filter to prepare Eu(TTA)3-TPA-DPA-mCF3@PS microspheres.

[0538] The microspheres obtained above were excited at a wavelength of 330 nm. The results are shown in [reference needed]. Figure 20 The study found that the full width at half maximum (FWHM) of the Eu(TTA)3-TPA-DPA-mCF3@PS microspheres met the narrow band requirement, but they exhibited multiple emission peaks. The emission spectrum of the luminescent microspheres prepared in this comparative example was tested, revealing a FWHM of 20 nm. Multiple emission peaks were observed at 5% intensity (greater than or equal to the highest peak value), with a peak width of 180 nm at 1% intensity of the highest peak value. The luminescence lifetime was 0.65 ns. The microsphere diameter was measured to be between 580 nm and 700 nm. The emission spectrum of the luminescent molecules was also tested, revealing a FWHM of 16 nm and a peak width of 187 nm at 1% intensity of the highest peak value.

[0539] The luminescent microspheres prepared in Examples 25-28 and Comparative Example 6 were tested.

[0540] Detection method for standard samples: Combine 20 μL of the standard sample with reagent 1 (100 μL), reagent 2 (20 μL), and reagent 3 (20 μL), incubate at 37°C with shaking for 10 minutes, and then take the reading. The standard sample is commercially available IL-6 (concentration 0-5000.0 pg / mL), reagent 1 is donor microsphere solution, reagent 2 is acceptor microsphere solution, and reagent 3 is PBS buffer.

[0541] Preparation method of donor microspheres: 0.1 g of carboxylated polystyrene spheres with a particle size of 100 nm were dissolved in 100 mL of ultrapure water and ultrasonically dispersed. Then, 0.5–5 mL each of 2 wt% sodium dodecylbenzenesulfonate and 1 wt% ethylenediamine polyoxyethylene polyoxypropylene block polyether were added to the dispersion and stirred to form an aqueous solution. A photosensitizer was dispersed in 10 mL of tetrahydrofuran to form an organic phase, which was then rapidly added to the above aqueous solution. The temperature was gradually increased to 50 °C and stirred continuously for 10 h. The reacted carboxylated polystyrene spheres were centrifuged, washed twice with ultrapure water and ethanol, and stored in ultrapure water to prepare energy donor microspheres, which were then stored at room temperature in the dark for later use. The donor microspheres were centrifuged at high speed, washed three times with PBS buffer, and finally adjusted to a solid content of 0.1%. The microspheres were ultrasonically dispersed to ensure uniform dispersion. 500 μL of the above dispersion was added to 500 μL of MES and 10 mg of EDC, and the reaction was carried out at room temperature for 2 hours. After the reaction, the mixture was centrifuged and washed, and then reconstituted in 500 μL of PBS buffer. 0.01 mg of IL-6-Ab1 monoclonal antibody was added to each buffer, and the mixture was reacted at room temperature for 24 hours. After the reaction, the mixture was stirred at 16000 rpm, centrifuged and washed, and then reconstituted in 500 μL of PBS buffer. 10 mg of BSA was added, and the mixture was reacted at room temperature for 8 hours. After the reaction, the mixture was centrifuged and washed to obtain 1 mg of energy donor microspheres coated with IL-6-Ab1 antibody. These microspheres were reconstituted in 2.0 mL of PBS buffer to prepare donor fraction II-1 containing the photosensitizer, and stored at 4 °C for later use.

[0542] Preparation method of receptor microspheres: Referring to the preparation method of donor microspheres, the difference is that the luminescent molecules and photochemical buffers of this embodiment are used instead of the photosensitizer and dispersed in 10 mL of tetrahydrofuran. IL-6-Ab2 monoclonal antibody is conjugated to the energy receptor microspheres and reacted at room temperature for 8 hours. After the reaction, the mixture is stirred at 35,000 rpm, with other conditions remaining the same. The final receptor component II-2 is obtained.

[0543] Detection method for whole blood samples: Add 5 μL of whole blood sample to be tested, diluent (PBS buffer), the recipient microsphere solution and donor microsphere solution prepared above to the same reaction vessel in sequence, stir well to prepare an immunoreaction system, and incubate for 5 min; irradiate the reaction vessel with 680 nm excitation light, collect the luminescence signal value, and substitute the luminescence signal value into the corresponding standard curve to obtain the concentration of IL-6 in the sample to be tested.

[0544] The luminescence signal values ​​of the luminescent microspheres prepared in Examples 25-28 and Comparative Example 6 were tested, and the results are shown in Table 1 below.

[0545] Table 1: Luminescence signal values ​​of the luminescent microspheres in Examples 25-28 and Comparative Example 6

[0546] Note: The unit of the light emission signal value in Table 1 is "1".

[0547] As shown in Table 1 above, the detection limit of the prior art in Comparative Example 6 is 1.00 pg / mL, the detection limit of the prior art in Example 25 is 1.00 pg / mL, and the detection limit of the prior art in Examples 26 and 27 is 2.00 pg / mL. The addition of external stimuli, such as X-rays and light, increases the production of singlet oxygen, thus increasing the luminescence intensity and raising the detection limit to 0.50 pg / mL. This means that the detection limit of the single-peak narrow-band luminescent microspheres used in this example for homogeneous detection is increased by at least four times, resulting in a significant breakthrough in detection sensitivity. A detection limit of 0.5 pg / mL can capture ultra-low abundance biomarkers that are undetectable by traditional methods. For example, in the early stages of tumors, the concentration of microRNA (miRNA) in ctDNA or exosomes is often below 1 pg / mL. This breakthrough makes ultra-early screening for diseases such as lung cancer and colorectal cancer possible. For targeted therapies (such as anti-PD-1 antibody therapy), dynamic monitoring of blood drug concentrations and tumor markers (such as ctDNA mutation burden) requires a detection limit below 1 pg / mL. User technology can provide real-time feedback on treatment effectiveness, guiding adjustments to individualized medication regimens.

[0548] Comparative Example 7

[0549] The synthetic route of the target molecule in this comparative example is as follows:

[0550] The synthesis method of the target molecule in this comparative example is similar to that in Example 10, except that: in step (1), benzaldehyde is used instead of p-tert-butylbenzaldehyde; and in step (4), phenylboronic acid is used instead of p-tert-butylbenzaldehyde.

[0551] Following the method in Example 1, the target molecule was coated into polystyrene microspheres to obtain luminescent microspheres. The emission spectrum of the microspheres prepared in this comparative example was tested, and the full width at half maximum (FWHM) was 47 nm. There was one emission peak at 5% intensity (≥5% of the peak value), and the peak width at 1% intensity of the peak value was 130 nm. The luminescence lifetime was 6.5 ns. The microsphere diameter was measured to be between 280 nm and 305 nm. The emission spectrum of the luminescent molecule was also tested, and the FWHM was 45 nm, with a peak width at 1% intensity of the peak value of 125 nm.

[0552] Comparative Example 8

[0553] The synthetic route of the target molecule in this comparative example is as follows:

[0554] The synthesis method of the target molecule in this comparative example is similar to that in Example 1.

[0555] Following the method in Example 1, the target molecule was coated into polystyrene microspheres to obtain luminescent microspheres. The emission spectrum of the microspheres prepared in this comparative example was tested, and the full width at half maximum (FWHM) was 42 nm. There was one emission peak at 5% intensity (≥5% of the peak value), the peak width at 1% intensity of the peak value was 120 nm, and the luminescence lifetime was 13.0 ns. The microsphere diameter was measured to be between 195 nm and 206 nm. The emission spectrum of the luminescent molecule was also tested, and the FWHM was 39 nm, with a peak width at 1% intensity of the peak value of 108 nm.

[0556] Application Example 1

[0557] The pICz-TPA@PS microspheres prepared in Example 15 were processed and added to a buffer in the prior art as receptor microspheres.

[0558] The photosensitizer in the existing homogeneous detection system was coated in polystyrene microspheres as donor microspheres.

[0559] N-hydroxythiosuccinimide was added to both the donor and recipient microsphere systems and vortexed. Then, an equal volume of 1-ethyl-(3-dimethylaminopropyl)carbodiimide was added and shaken. After activation, the solution was centrifuged and the supernatant was discarded. 2-(N-morpholino)ethanesulfonic acid was added and the mixture was sonicated. Antibodies were added to the sonicated mixture for labeling. IL-6-Ab1 antibody was added to the donor microspheres and IL-6-Ab2 antibody was added to the recipient microspheres. The mixture was mixed in the dark, centrifuged, and the supernatant was discarded. Bovine serum albumin was added to the precipitate for blocking. The mixture was sonicated in the dark, centrifuged, and the supernatant was discarded. Borate buffer was added to the precipitate, and the mixture was sonicated and stored.

[0560] 2 μL of the serum sample to be tested and 98 μL of diluent were added sequentially to the same reaction vessel, stirred thoroughly to prepare an immunoassay system, and incubated for 5 min. The reaction vessel was then irradiated, the excitation light was turned off, and the light signal value was collected. The light signal value was then substituted into the light signal-analyte concentration standard curve to obtain the concentration value of the analyte.

[0561] Application Example 2

[0562] Referring to Application Example 1, the difference is that the pICz-TPA@PS microspheres are replaced with the Cz-DBCz@PS microspheres in Example 5 for preparing receptor microspheres.

[0563] Application Example 3

[0564] Referring to Application Example 1, the difference is that the pICz-TPA@PS microspheres are replaced with PhO-DBCz@PS microspheres from Example 7 for preparing receptor microspheres.

[0565] Application Example 4

[0566] Referring to Application Example 1, the difference is that X-rays are applied simultaneously as an external stimulus during excitation light irradiation.

[0567] Application Example 5

[0568] Referring to Application Example 1, the only difference is that the luminescent molecules in the receptor microspheres are replaced with the luminescent agent in the homogeneous detection system of the prior art, while everything else remains the same. The detection marker is IL-6. Its concentration is detected to obtain the test results.

[0569] Table 2: Test Results of Application Examples 1-5

[0570] As shown in Table 2 above, the detection limit (LOD) of Application Example 5 using the luminescent agent from the prior art is 3.00 pg / mL, the LOD of Application Example 1 is 1.00 pg / mL, and the LODs of Application Examples 2 and 3 are 1.50 pg / mL. In Application Example 4, where X-rays and light are combined as external stimuli, the increased singlet oxygen production leads to increased luminescence intensity, resulting in a LOD of 0.50 pg / mL. This means that the detection limit of the single-peak narrow-band luminescent microspheres of the present invention, when used in homogeneous detection, can be increased by 6 times, thus achieving a significant breakthrough in detection sensitivity. A detection limit of 0.5 pg / mL can capture ultra-low abundance biomarkers that are undetectable by conventional methods. For example, in the early stages of tumors, the concentration of microRNAs (miRNAs) in ctDNA or exosomes is often below 1 pg / mL. This breakthrough makes ultra-early screening for diseases such as lung cancer and colorectal cancer possible. For targeted therapies (such as anti-PD-1 antibody therapy), dynamic monitoring of blood drug concentrations and tumor markers (such as ctDNA mutation burden) requires a detection limit below 1 pg / mL. User technology can provide real-time feedback on treatment effectiveness, guiding adjustments to individualized medication regimens.

[0571] Application Example 6-15

[0572] The concentration of the biomarker was tested in accordance with Application Example 1, except that the microspheres in Table 3 were replaced with the Cz-DBCz@PS microspheres in Example 5 to prepare the receptor microspheres.

[0573] Table 3: Microspheres used in Application Example 6-15

[0574] The luminescence signal values ​​of the microspheres in Application Examples 6-15 were tested using the same methods as those used in Examples 25-28 and Comparative Example 6.

[0575] Table 4: Luminescence signal values ​​of microspheres in Application Example 6-15

[0576] Note: The unit of the luminous signal values ​​in Table 4 is "1".

[0577] As can be seen from Table 4, the experimental results of Application Examples 6-15 show that the single-peak narrow-band luminescent microspheres of the present invention have the advantage of high sensitivity in homogeneous detection.

[0578] Application Example 16: Multiplex Nanoflow Cytometry Detection Based on Single-Peak Narrow-Band Emitting Microspheres

[0579] This application example utilizes the single-peak narrow-band luminescent microspheres mentioned above in nanoflow cytometry to detect exosomal protein markers CD151, TTF-1, and SFTPB proteins.

[0580] Three types of single-peak narrow-band luminescent microspheres were selected. In this application example, the microspheres from Examples 2, 5, and 7 were combined. The emission half-width (FWHM) of these three examples was less than 20 nm. The luminescent microspheres from the above examples were antibody-conjugated, and then plasma exosomes were extracted for sample incubation and flow cytometry detection.

[0581] Specifically, in the microsphere antibody conjugation stage, three types of single-peak narrow-band luminescent microspheres were taken, washed twice with MES buffer at pH 6.0 and resuspended, then activated with EDC and NHS at room temperature in the dark. After centrifugation and discarding the supernatant, Anti-CD151 antibody (10418-1-AP), Anti-TTF-1 antibody (ab227652), and Anti-SFTPB antibody (13664-1-AP) were added respectively, and conjugated at room temperature in the dark for 2 hours. Subsequently, BSA solution with a final concentration of 1% was added for blocking for 30 min, washed three times with PBS filtered through a filter membrane, and resuspended in 1 mL of storage solution. The microspheres were then stored at 4°C in the dark for later use. Plasma exosome extraction was strictly performed according to the published method. Peripheral blood samples were collected from clinically diagnosed non-small cell lung cancer patients and healthy individuals undergoing physical examinations. Blood cells were removed by centrifugation at 4°C, and the supernatant was centrifuged again to remove cell debris and impurities. Plasma samples were then aliquoted and added to centrifuge tubes. After centrifugation at 4°C and discarding the supernatant, the precipitate was resuspended in PBS. The ultracentrifugation and washing steps were repeated once, and the exosomes were finally resuspended in PBS. During the sample incubation and detection phase, the prepared exosome samples were taken, and luminescent microsphere working solutions containing three conjugated antibodies were added. PBS was added to bring the total volume to 100 μL, and the samples were incubated overnight at 4°C in the dark. After incubation, the samples were diluted with PBS and transferred to flow cytometry tubes for detection. The nanoflow cytometry detection parameters were set to 488 nm excitation light power of 10 mW, corresponding to the activation of the three detection channels at 525 nm, 605 nm, and 685 nm. Each sample was sampled for 20 s, and a threshold was set using isotype control microspheres to distinguish between positive and background signals.

[0582] Application Comparative Example 1

[0583] Comparative Example 1 of this application uses a multicolor direct fluorescent labeling scheme, with FITC (fluorescein isothiocyanate) labeling Anti-CD151, PE (phycoerythrin) labeling Anti-TTF-1, and APC (allophycocyanin) labeling Anti-SFTPB, and exosomes are directly detected after immunofluorescence staining.

[0584] The FITC structure is as follows:

[0585] In this comparative application, the microsphere coupling step is not required. The remaining exosome extraction process is the same as in Application Example 16. During the fluorescence staining stage, 10 μL of exosome sample is taken, 4% BSA is added, and the sample is blocked at room temperature for 10 min. Then, the corresponding working concentrations of FITC-labeled Anti-CD151, PE-labeled Anti-TTF-1, and APC-labeled Anti-SFTPB antibodies are added. The sample is incubated overnight at 4°C in the dark. After washing and resuspending with PBS, the sample is then tested. The detection channels and instrument parameters are the same as in Application Example 16.

[0586] Application Comparative Example 2

[0587] Comparative Example 2 of this application uses ordinary broadband polystyrene fluorescent microspheres (all with a half-width of more than 85 nm), coupled with the same three antibodies as in Application Example 16, and the incubation and detection procedures are completely consistent with Application Example 16.

[0588] The microsphere coupling, exosome extraction, incubation and detection procedures in this comparative application are exactly the same as those in Application Example 16, except that ordinary broadband fluorescent microspheres are used instead.

[0589] , , .

[0590] The emission half-width of D-1, D-2, and D-3 is greater than 80 nm.

[0591] Three types of single-labeled microsphere samples were cross-detected in three detection channels, and the signal crosstalk rate was calculated. In Application Example 16, the crosstalk rates for microspheres B and C detected in the 525 nm channel were 2.1% and 1.8%, respectively; the crosstalk rates for microspheres A and C detected in the 605 nm channel were 1.5% and 2.4%, respectively; and the crosstalk rates for microspheres A and B detected in the 685 nm channel were both below 2.0%. The signals from the three types of microspheres showed virtually no mutual interference, perfectly matching the spectral characteristics of a single peak and narrow band, enabling the differentiation of the three indicators. In Comparative Application Example 1, the spectral overlap of organic fluorescent dyes led to significant crosstalk. The crosstalk rate for the FITC signal reached 12.7% in the 605 nm channel, and the crosstalk rate for the PE signal reached 8.3% in the 685 nm channel, easily causing false positives for low-abundance samples. The crosstalk of the broadband microspheres in Comparative Example 2 is similar to that in Comparative Example 1. The crosstalk rate of the 525 nm channel to the 605 nm microsphere is 9.6%, and the 685 nm channel has a spillover signal of the 605 nm microsphere with a crosstalk rate of 7.2%, which makes it impossible to achieve accurate quantification of the three indicators.

[0592] Plasma samples from 10 patients with non-small cell lung cancer and 10 healthy individuals were selected. The methods described in Application Example 16, Comparative Example 1, and Comparative Example 2 were used for combined detection of three indicators. Application Example 16 showed that the proportion of CD151, TTF-1, and SFTPB-positive exosomes in the lung cancer group was significantly lower than that in the healthy group. The combined AUC of the three indicators reached 0.932, higher than the 0.898 in Comparative Example 1, and the intra-group coefficients of variation were all less than 5%, indicating better repeatability. Comparative Example 1 showed an increased false positive rate in the healthy group due to fluorescence crosstalk, with a combined AUC of only 0.814, indicating significant bias in the detection results of low-abundance early-stage lung cancer samples. Comparative Example 2 showed high dispersion in the detection results, with decreased differentiation between the healthy and lung cancer groups; the combined AUC was 0.807. In the detection of 10 stage I non-small cell lung cancer samples, the positive detection rate of Case 16 reached 90%, while the detection rates of Comparative Case 1 and Comparative Case 2 were 60% and 55%, respectively, which fully demonstrates that the high sensitivity of single-peak narrow-band microspheres has higher accuracy and sensitivity in early lung cancer screening.

[0593] Application Example 17: Immunochromatography Based on Single-Peak Narrow-Band Luminescent Microspheres

[0594] Take 1 mg of microspheres from Examples 36, 34, 10, or 20, wash three times with 0.05 mol / L MES buffer (pH=6.0), resuspend, and add freshly prepared EDC and NHS solution. Activate by shaking at room temperature for 30 min. Centrifuge to remove the supernatant, resuspend the microspheres in 0.05 mol / L borate buffer (pH=7.4), and add the corresponding target detection antibody (SAA, PCT, CRP, IL-6). React by shaking at room temperature for 2 h. Block with 1% BSA solution for 1 h; centrifuge to remove unbound antibody and blocking agent, resuspend in PBS buffer containing 0.5% BSA, and store at 4°C protected from light.

[0595] Capture antibodies targeting four different targets were sprayed onto an NC membrane using a coating instrument, forming four detection lines: T1 (SAA), T2 (PCT), T3 (CRP), and T4 (IL-6). Goat anti-mouse IgG was sprayed as the control line (C line), with a line spacing of 3 mm. The membrane was then vacuum-dried overnight at 37°C. (See also...) Figure 29 Four types of immunoglobulins were mixed in equal proportions and diluted to the working concentration with a reconstitution buffer containing sucrose and Tween-20. The mixture was then evenly sprayed onto the conjugate pad and dried at 37°C for 2 hours. The sample pad, conjugate pad, NC membrane, and absorbent pad were then sequentially pasted onto a PVC base plate with an overlap width of 2 mm. The mixture was then cut into 4 mm wide test strips using a strip cutter, inserted into the cartridge, and stored in a dry environment.

[0596] The target standard was diluted with negative human serum to create a series of concentration gradients. 100 μL of each sample was added to the sample well of the test strip and allowed to react at room temperature for 15 min. The fluorescence intensity of the T and C lines in the four emission channels was acquired using a multi-channel fluorescence reader, and the T / C ratio was calculated. A standard curve was fitted with the target concentration on the x-axis and the corresponding T / C ratio on the y-axis, and the limit of detection (LOD) was calculated according to the 3σ principle. Simultaneously, positive samples containing only a single target were added, and the signal values ​​of the four channels were read. The signal proportion of non-target channels, i.e., the crosstalk rate, was calculated. The results are shown in Tables 5 and 6.

[0597] Application Comparative Example 3

[0598] Comparative Example 3 of this application uses four broad-peak structures as examples, among which MeODPA-DiKTa has a peak value of 655 nm and FWHM is 71 nm, with the following structures: ; The peak value of FL-BTD-IDB (fluorenylbenzothiadiazole) is 575 nm, and the FWHM is 76 nm. The structure is as follows: ; The FL-BTD-Oar has a peak wavelength of 528 nm (green light) and an FWHM of 73 nm. Its structure is as follows: ; Twisted carbazole-benzophenone (CzBP) (blue light), peak at 452 nm, FWHM=77 nm, structure as follows: ; The four broad-peak structures described above were coated in polystyrene microspheres according to Example 1, and then SAA, PCT, CRP, and IL-6 were detected according to Application Example 17. The results are shown in Tables 5 and 6.

[0599] Table 5

[0600] Table 6

[0601] Application Example 18: Multiplex Cell Staining Based on Single-Peak Narrow-Band Luminescent Microspheres

[0602] This application example uses human pancreatic ductal adenocarcinoma (PDAC) tissue microarrays (containing 60 cancerous tissues and corresponding adjacent normal tissues) and PD-L1 gradient-expressing cell slices as detection samples. Four tumor targets—PanCK, CD8, CD68, and PD-L1—were selected to verify the advantages of single-peak narrow-band luminescent microspheres in multiplex immunofluorescence staining. This application example uses four specific secondary antibodies labeled with single-peak narrow-band luminescent microspheres (Examples 36, 34, 10, and 20) as detection reagents. The emission peaks of the four microspheres correspond to 480 nm, 525 nm, 580 nm, and 650 nm, respectively, with a half-width at half-maximum (HWHM) of no more than 35 nm. This allows for non-overlapping spectral arrangement in the visible light region, and they are matched with four primary antibodies: mouse anti-PanCK, rabbit anti-CD8, goat anti-CD68, and guinea pig anti-PD-L1.

[0603] First, the paraffin tissue microarray underwent routine dewaxing and hydration. After microwave antigen retrieval using pH 9.0 EDTA buffer, it was incubated with 3% hydrogen peroxide solution at room temperature in the dark to block endogenous peroxidase. Then, non-specific binding sites were blocked with 5% BSA at room temperature. After blocking, a mixed working solution of four primary antibodies was added directly, and the microarray was incubated overnight at 4°C in a humidified chamber in the dark. The next day, after thorough washing with PBS, a mixed working solution of four narrow-band luminescent microsphere-labeled secondary antibodies was added, and the microarray was incubated at room temperature in the dark for 1 hour. After multiple washes with PBS, the cell nuclei were counterstained with DAPI. Finally, the microarray was mounted with anti-fluorescence quenching mounting medium and images were acquired. The entire detection process requires only one antigen retrieval and one round of antibody incubation to simultaneously label four target sites, with a total operation time of approximately 9 hours, which can be completed within a single working day.

[0604] Application Comparative Example 4

[0605] This application strictly followed the given multiplex immunofluorescence staining protocol, using poly-HRP secondary antibody in combination with four tyramine fluorescent substrates: FITC, Cy3, Cy5, and Alexa Fluor 700, to detect the same four targets as in Application Example 18. The experiment required four cycles of single-target labeling, each cycle sequentially performing antigen retrieval, endogenous peroxidase blockade, non-specific blocking, single-target primary antibody incubation, HRP secondary antibody incubation, tyramine fluorescent substrate reaction, and high-temperature antibody elution. After completing all four staining cycles, DAPI nuclear counterstaining and mounting were performed. Due to the broad emission spectrum and significant channel crosstalk of traditional organic fluorescent dyes, images were ultimately acquired using a multispectral imaging system, and crosstalk was corrected using a spectral splitting algorithm before quantitative analysis.

[0606] The results showed that in Application Example 18, the cross-color rate of the four detection channels was less than 1%, and the signals of each channel could be separated without spectral splitting. The false positive rate of CD8 and PD-L1 double-positive cell subpopulations was reduced by 83% compared with the control example. In Application Example 4, due to the limitation of the broad emission peak of the organic fluorescent dye, the cross-color rate of each channel was generally in the range of 4% to 8%. Even after correction by the multispectral unmixing algorithm, a certain quantitative bias was still introduced, and the error was more significant in the co-localization analysis of double-positive cells.

[0607] Application Example 19: Biological Tissue Live Tags Based on Single-Peak Narrow-Band Luminescent Microspheres

[0608] This application example demonstrates the use of the single-peak narrow-band luminescent microspheres mentioned above in the field of biological tissue labeling, using the single-peak narrow-band luminescent microspheres mentioned above as a labeling reagent for spray-on narrow-band luminescent microspheres.

[0609] OATP (organic anion transport peptide) targeting ligands were modified on the surface of single-peak narrow-band luminescent microspheres to prepare a stable aqueous microsphere suspension. Following the atomization spraying method reported in LifeMedicine, 2023, Volume 2, Issue 1, lnac059, the microspheres were directly applied to the tumor cut surface. Unbound microspheres could be quickly washed away with physiological saline, leaving only microspheres that specifically bind to tumor cells, and then narrow-band luminescence signals were collected.

[0610] In this case, the single-peak narrow-band luminescent microspheres in Example 5 targeted OATP to label tumor tissue, the single-peak narrow-band luminescent microspheres in Example 7 targeted CD31 to label blood vessels, and the single-peak narrow-band luminescent microspheres in Example 8 targeted LYVE-1 to label lymphatic vessels were mixed and sprayed onto the tissue section in one go. The signals were simultaneously acquired by a multi-channel imaging system to obtain the tumor boundary and vascular distribution information in one go.

[0611] Application Comparative Example 5

[0612] The first three types of broad-peak luminescent microspheres of Application Example 3 were labeled according to the process of Application Example 19.

[0613] The test results show that the method reported in LifeMedicine, 2023, Volume 2, Issue 1, lnac059, uses a single luminescent channel and can only identify the tumor region, failing to simultaneously label key structures in surrounding normal tissue. Furthermore, the broad-peak luminescent microspheres used in Comparative Example 5 exhibit severe overlap and cross-color interference, making it impossible to distinguish multiple targets. The tumor / normal tissue signal-to-noise ratio of the small molecule self-assembled nanoprobes reported in LifeMedicine, 2023, Volume 2, Issue 1, lnac059, is approximately 4, while the tumor / normal tissue signal-to-noise ratio of the sprayed probe using the single-peak narrow-band luminescent microspheres in Example 19 can be improved to over 10. Moreover, the single-peak narrow-band luminescent microspheres in Example 19 are multi-colored without cross-color interference and can simultaneously label three targets.

[0614] Specific embodiments of the present invention have been described above. It should be understood that the present invention is not limited to the specific embodiments described above, and those skilled in the art can make various modifications or variations within the scope of the claims, which do not affect the essence of the present invention. The above preferred features can be used in any combination without conflict.

[0615] Although the present invention has been illustrated and described with reference to certain preferred embodiments, those skilled in the art should understand that the above description is a further detailed explanation of the invention in conjunction with specific embodiments, and it should not be considered that the specific implementation of the invention is limited to these descriptions. Those skilled in the art can make various changes in form and detail, including some simple deductions or substitutions, without departing from the spirit and scope of the invention. The embodiments of the invention have been described in detail above with reference to the accompanying drawings; however, the invention is not limited to the above embodiments, and various changes can be made within the scope of knowledge possessed by those skilled in the art without departing from the spirit of the invention.

Claims

1. A single-peak narrow-band luminescent microsphere, characterized in that, The single-peak narrowband luminescent microspheres comprise organic luminescent molecules, the diameter of the single-peak narrowband luminescent microspheres is 50 nm to 500 μm, and the half-width at half-maximum (WHM) of the emission spectrum of the single-peak narrowband luminescent microspheres is no greater than 35 nm.

2. The single-peak narrow-band luminescent microsphere as described in claim 1, characterized in that, The single-peak narrow-band luminescent microspheres have one or more of the following characteristics: The peak width at 1% intensity of the highest peak of the emission spectrum of the single-peak narrow-band luminescent microsphere is no greater than 150 nm, preferably no greater than 130 nm. In the emission spectrum of the single-peak narrowband luminescent microsphere, the emission spectrum at an intensity greater than or equal to 5% of the highest peak value does not show double peaks or more peaks; The luminescence lifetime of the single-peak narrowband luminescent microsphere is 1 ns to 999 ns; The organic light-emitting molecule is a narrow-band organic light-emitting molecule, and the half-width at half maximum (WHM) of the emission spectrum of the narrow-band organic light-emitting molecule is no greater than 30 nm; preferably, the peak width at 1% intensity of the highest peak value of the emission spectrum of the narrow-band organic light-emitting molecule is no greater than 140 nm; preferably, under external stimulation, the half-width at half maximum (WHM) of the emission spectrum of the narrow-band organic light-emitting molecule is no greater than 30 nm, and the peak width at 1% intensity of the highest peak value is no greater than 140 nm. Under external stimulation, the half-width at half-maximum (WHM) of the emission spectrum of the single-peak narrowband luminescent microsphere is no greater than 35 nm, and the peak width at 1% intensity of the highest peak is no greater than 150 nm; preferably, the single-peak narrowband luminescent microsphere also includes a component that is responsive to the external stimulation.

3. The single-peak narrow-band luminescent microsphere as described in claim 2, characterized in that, The external stimulus includes at least one selected from light, chemical reaction, X-ray, ultrasound, radioactive irradiation, thermal stimulation and mechanical stimulation, and the single-peak narrow-band luminescent microsphere emits light under the external stimulus; Preferably, the external stimulus is light, and the single-peak narrow-band luminescent microspheres produce fluorescence under light. Preferably, the external stimulus is light, and the single-peak narrow-band luminescent microspheres produce long afterglow luminescence under light. Preferably, the external stimulus is a chemical reaction, and the single-peak narrow-band luminescent microspheres produce chemiluminescence under the chemical reaction.

4. The single-peak narrow-band luminescent microsphere as described in claim 1, characterized in that, The organic light-emitting molecule contains only a single light group, the light group contains a conjugated π-electron system, and the organic light-emitting molecule has a planar rigid structure; Preferably, at least one substituent is present on the conjugated π-electron system, and the conformational free energy A of the substituent is ≥1.7 kcal / mol, preferably ≥4 kcal / mol; Preferably, the conjugated π-electron system has at least one substituent, which is selected from halogens, straight-chain alkyl, branched alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, cycloalkylene, heterocycloalkylene, cycloalkenyl, heterocycloalkenyl, alkoxy, alkylsilyl, aryl, alkyl-substituted aryl, alkoxy-substituted aryl, aryl-substituted aryl, fluoroalkyl-substituted aryl, fluoroaryl-substituted aryl, aralkyl, aryloxy, heteroaryl, alkyl-substituted heteroaryl, alkoxy-substituted heteroaryl, aryl-substituted heteroaryl, fluoroalkyl-substituted heteroaryl, fluoroaryl-substituted heteroaryl, cyano, esters and their derivatives, alkyl ketones. Alkyl esters, aryl esters, alkynyl groups, alkylamines, fluoroalkyl groups, fluoroaryl groups, polyalkylene groups, aniline-substituted fluorenyl groups, diphenylamine-substituted fluorenyl groups, triphenylamine-substituted fluorenyl groups, alkyl-substituted triphenylamine groups, alkyl-substituted diphenylamine groups, alkyl-substituted aniline groups, alkyl-substituted thiophenyl groups, borate ester groups, hydroxyl groups, amino groups, nitro groups, and trimethylsilylphenoxy groups; wherein the aryl group includes phenyl and fluorenyl groups, and the heteroaryl group includes pyridyl, bipyridyl, tripyridyl, furanyl, thiophenyl, pyrroleyl, pyrazolyl, oxazolyl, thiazolyl, imidazoleyl, pyrazinyl, benzoxazolyl, benzothiadiazolyl, and carbazoleyl groups; Preferably, the conjugated π-electron system contains at least two substituents selected from the first, second, third, and fourth substituents, and the substituents exhibit mixed steric hindrance effects. The first substituent includes C6-C18 straight-chain alkyl, C6-C18 heteroalkyl, polyalkylene, C6-C18 esters and their derivatives, alkyl ketones, alkyl esters, alkylamines, borate esters, hydroxyl groups, amino groups, and nitro groups. The second substituent includes C4-C8 branched alkyl, aryl, alkyl-substituted aryl, aryl-substituted aryl, fluoroalkyl-substituted aryl, fluoroaryl-substituted aryl, aralkyl, aryloxy, heteroaryl, phenylamino-substituted fluorenyl, diphenylamino-substituted fluorenyl, triphenylamino-substituted fluorenyl, alkyl-substituted triphenylamino, alkyl... The substituents include alkyl-substituted diphenylamino, alkyl-substituted aniline, C6-C18 cycloalkyl, heterocycloalkyl, cycloalkenyl, heterocycloalkenyl, and aryl esters; the third substituent includes hydrogen, halogen, C1-C5 straight-chain alkyl, C3 branched alkyl, C3-C5 cycloalkyl, C1-C6 alkoxy, cyano, and C2-C5 alkynyl; the fourth substituent includes fluoroalkyl, fluoroaryl, alkyl-substituted heteroaryl, alkoxy-substituted heteroaryl, aryl-substituted heteroaryl, fluoroalkyl-substituted heteroaryl, fluoroaryl-substituted heteroaryl, heterocycloalkylene, cycloalkylene, alkyl-substituted thiophenyl, alkoxy-substituted aryl, alkylsilyl, and trimethylsilylphenoxy; wherein the C6-C18 cycloalkyl includes adamantyl, and the alkylsilyl includes triisopropylsilyl.

5. The single-peak narrow-band luminescent microsphere as described in claim 2, characterized in that, The narrow-band organic light-emitting molecule has one or more of the following features (A) to (N): (A) The narrow-band organic light-emitting molecule has the structure shown in Formula I or Formula II: Formula I, Formula II; In Formula I and Formula II, each X is independently selected from fluorine, oxygen, cyano and fluoroalkoxy groups; In formula II, and Each is an independent structural unit, existing or not existing; when When it exists, it may be optionally replaced by one or more R2s; when When present, it may be optionally replaced by one or more R2'; In Formula II, n1 and n2 each independently represent zero or one or more groups; In Formulas I and II, R1, R2, R2′, R3, R3′, R4, and R4′ are independently selected from hydrogen, halogen, straight-chain alkyl, branched alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, cycloalkylene, heterocycloalkylene, cycloalkenyl, heterocycloalkenyl, alkoxy, alkylsilyl, aryl, alkyl-substituted aryl, alkoxy-substituted aryl, aryl-substituted aryl, fluoroalkyl-substituted aryl, fluoroaryl-substituted aryl, aralkyl, aryloxy, heteroaryl, alkyl-substituted heteroaryl, alkoxy-substituted Heteroaryl, aryl-substituted heteroaryl, fluoroalkyl-substituted heteroaryl, fluoroaryl-substituted heteroaryl, cyano, esters and their derivatives, alkyl ketones, alkyl esters, aryl esters, alkynyl, alkylamine, fluoroalkyl, fluoroaryl, polyalkylene, aniline-substituted fluorenyl, diphenylamino-substituted fluorenyl, triphenylamino-substituted fluorenyl, alkyl-substituted triphenylamino, alkyl-substituted diphenylamino, alkyl-substituted aniline, alkyl-substituted thiophenyl, borate ester, hydroxyl, amino, nitro and trimethylsilylphenoxy; Preferably, in Formula I, R1, R2, R2′, R3, R3′, R4, and R4′ are each independently selected from at least two substituents selected from the first, second, third, and fourth substituents; in Formula II, R1, R2, R2′, R4, and R4′ are each independently selected from at least two substituents selected from the first, second, third, and fourth substituents; the first substituent includes C6-C18 straight-chain alkyl, C6-C18 heteroalkyl, polyalkylene, C6-C18 esters and their derivatives, alkyl ketones, alkyl esters, alkylamines, borate esters, hydroxyl groups, amino groups, and nitro groups; the second substituent includes C4-C8 branched alkyl, aryl, alkyl-substituted aryl, aryl-substituted aryl, fluoroalkyl-substituted aryl, fluoroaryl-substituted aryl, and aralkyl. The substituents include: aryloxy, heteroaryl, aniline-substituted fluorenyl, diphenylamino-substituted fluorenyl, triphenylamino-substituted fluorenyl, alkyl-substituted triphenylamino, alkyl-substituted diphenylamino, alkyl-substituted aniline, C6-C18 cycloalkyl, heterocycloalkyl, cycloalkenyl, heterocycloalkenyl, and aryl esters; the third substituent includes hydrogen, halogen, C1-C5 straight-chain alkyl, C3 branched alkyl, C3-C5 cycloalkyl, C1-C6 alkoxy, cyano, and C2-C5 alkynyl; the fourth substituent includes fluoroalkyl, fluoroaryl, alkyl-substituted heteroaryl, alkoxy-substituted heteroaryl, aryl-substituted heteroaryl, fluoroalkyl-substituted heteroaryl, fluoroaryl-substituted heteroaryl, heterocycloalkylene, cycloalkylene, alkyl-substituted thiophenyl, alkoxy-substituted aryl, alkylsilyl, and trimethylsilylphenoxy; Preferably, the narrow-band organic light-emitting molecule has the structure shown in any one of Formula I (a1) to Formula I (a6): Formula I (a1), Equation I (a2), Equation I (a3), Equation I (a4), Formula I (a5), Formula I (a6); In formulas I(a1) to I(a6), n1, n2, n3, and n4 each independently represent zero or one or more groups; in formulas I(a1) to I(a6), R1′, R2, R2′, R3, R3′, R4, R4′, and R′ a 、R′ b 、R′ c and R′ d Each is independently selected from hydrogen, halogen, straight-chain alkyl, branched alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, cycloalkylene, heterocycloalkylene, cycloalkenyl, heterocycloalkenyl, alkoxy, alkylsilyl, aryl, alkyl-substituted aryl, alkoxy-substituted aryl, aryl-substituted aryl, fluoroalkyl-substituted aryl, fluoroaryl-substituted aryl, aralkyl, aryloxy, heteroaryl, alkyl-substituted heteroaryl, alkoxy-substituted heteroaryl, aryl-substituted heteroaryl, fluoroalkyl Alkyl-substituted heteroaryl, fluoroaryl-substituted heteroaryl, cyano, esters and their derivatives, alkyl ketones, alkyl esters, aryl esters, alkynyl, alkylamine, fluoroalkyl, fluoroaryl, polyalkylene, aniline-substituted fluorenyl, diphenylamino-substituted fluorenyl, triphenylamino-substituted fluorenyl, alkyl-substituted triphenylamino, alkyl-substituted diphenylamino, alkyl-substituted aniline, alkyl-substituted thiophenyl, borate ester, hydroxyl, amino, nitro and trimethylsilylphenoxy; Preferably, in formula I (a1), R1′, R2, R2′, R3, R3′, R4, and R4′ are each independently selected from at least two substituents selected from the first substituent, the second substituent, the third substituent, and the fourth substituent; in formula I (a2), R2, R2′, R3, R3′, R4, R4′, and R′ are each selected from at least two substituents selected from the first substituent, the second substituent, the third substituent, and the fourth substituent. a 、R′ b Each of the following substituents is independently selected from at least two substituents: a first substituent, a second substituent, a third substituent, and a fourth substituent; in formula I (a3), R2, R2′, R3, R3′, R4, R4′, and R′ a 、R′ b 、R′ c Each of the following substituents is independently selected from at least two substituents: a first substituent, a second substituent, a third substituent, and a fourth substituent; in formula I (a4), R2, R2′, R3, R3′, R4, R4′, and R′ a 、R′ b 、R′ c and R′ d Each of the following substituents is independently selected from at least two substituents: a first substituent, a second substituent, a third substituent, and a fourth substituent; in formula I (a5), R2, R2′, R3, R3′, R4, R4′, and R′ a 、R′ b Each of the following substituents is independently selected from at least two substituents: a first substituent, a second substituent, a third substituent, and a fourth substituent; in formula I (a6), R2, R2′, R3, R3′, R4, R4′, and R′ a 、R′ b 、R′ c Each of the substituents is independently selected from at least two substituents selected from the first, second, third, and fourth substituents; the first substituent includes C6-C18 straight-chain alkyl, C6-C18 heteroalkyl, polyalkylene, C6-C18 esters and their derivatives, alkyl ketones, alkyl esters, alkylamines, borate esters, hydroxyl groups, amino groups, and nitro groups; the second substituent includes C4-C8 branched alkyl, aryl, alkyl-substituted aryl, aryl-substituted aryl, fluoroalkyl-substituted aryl, fluoroaryl-substituted aryl, aralkyl, aryloxy, heteroaryl, aniline-substituted fluorenyl, diphenylamino-substituted fluorenyl, triphenylamino-substituted fluorenyl, and alkyl-substituted triphenylamine. The substituents include alkyl-substituted diphenylamino, alkyl-substituted aniline, C6-C18 cycloalkyl, heterocycloalkyl, cycloalkenyl, heterocycloalkenyl, and aryl esters; the third substituent includes hydrogen, halogen, C1-C5 straight-chain alkyl, C3 branched alkyl, C3-C5 cycloalkyl, C1-C6 alkoxy, cyano, and C2-C5 alkynyl; the fourth substituent includes fluoroalkyl, fluoroaryl, alkyl-substituted heteroaryl, alkoxy-substituted heteroaryl, aryl-substituted heteroaryl, fluoroalkyl-substituted heteroaryl, fluoroaryl-substituted heteroaryl, heterocycloalkylene, cycloalkylene, alkyl-substituted thiophenyl, alkoxy-substituted aryl, alkylsilyl, and trimethylsilylphenoxy. Preferably, the narrow-band organic light-emitting molecule has formula I (a 1-1 The structure shown is as follows: Ⅰ(a 1-1 )； Formula I (a) 1-1 In formula I (a), n1, n2, n3, and n4 each independently represent zero, one, or more groups; 1-1 In this context, R1, R2, R3, R4, R5, R1′, R2′, R3′, R4′, and R5′ are each independently selected from hydrogen, halogen, straight-chain alkyl, branched-chain alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, cycloalkylene, heterocycloalkylene, cycloalkenyl, heterocycloalkenyl, alkoxy, alkylsilyl, aryl, alkyl-substituted aryl, alkoxy-substituted aryl, aryl-substituted aryl, fluoroalkyl-substituted aryl, fluoroaryl-substituted aryl, aralkyl, aryloxy, heteroaryl, alkyl-substituted heteroaryl, alkoxy Alkyl-substituted heteroaryl, aryl-substituted heteroaryl, fluoroalkyl-substituted heteroaryl, fluoroaryl-substituted heteroaryl, cyano, esters and their derivatives, alkyl ketones, alkyl esters, aryl esters, alkynyl, alkylamine, fluoroalkyl, fluoroaryl, polyalkylene, aniline-substituted fluorenyl, diphenylamino-substituted fluorenyl, triphenylamino-substituted fluorenyl, alkyl-substituted triphenylamino, alkyl-substituted diphenylamino, alkyl-substituted aniline, alkyl-substituted thiophenyl, borate ester, hydroxyl, amino, nitro and trimethylsilylphenoxy; Preferably, Formula I (a 1-1 In this composition, R1, R2, R3, R4, R5, R1′, R2′, R3′, R4′, and R5′ are each independently selected from at least two substituents selected from the first substituent, the second substituent, the third substituent, and the fourth substituent; the first substituent includes C6-C18 straight-chain alkyl, C6-C18 heteroalkyl, polyalkylene, C6-C18 esters and their derivatives, alkyl ketones, alkyl esters, alkylamines, borate esters, hydroxyl groups, amino groups, and nitro groups; the second substituent includes C4-C8 branched alkyl, aryl, alkyl-substituted aryl, aryl-substituted aryl, fluoroalkyl-substituted aryl, fluoroaryl-substituted aryl, aralkyl, aryloxy, heteroaryl, aniline-substituted fluorenyl, and diphenylamino-substituted fluorenyl. The substituents include: triphenylamino-substituted fluorenyl, alkyl-substituted triphenylamino, alkyl-substituted diphenylamino, alkyl-substituted aniline, C6-C18 cycloalkyl, heterocycloalkyl, cycloalkenyl, heterocycloalkenyl, and aryl esters; the third substituent includes hydrogen, halogen, C1-C5 straight-chain alkyl, C3 branched alkyl, C3-C5 cycloalkyl, C1-C6 alkoxy, cyano, and C2-C5 alkynyl; the fourth substituent includes fluoroalkyl, fluoroaryl, alkyl-substituted heteroaryl, alkoxy-substituted heteroaryl, aryl-substituted heteroaryl, fluoroalkyl-substituted heteroaryl, fluoroaryl-substituted heteroaryl, heterocycloalkylene, cycloalkylene, alkyl-substituted thiophenyl, alkoxy-substituted aryl, alkylsilyl, and trimethylsilylphenoxy. Preferably, the narrow-band organic light-emitting molecule has one of the following structures: 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 ； (B) The narrow-band organic light-emitting molecule has a structure as shown in any one of Formula III-3 to Formula III-5: Formula III-3; Formula III-4; Formula III-5; In formulas III-3, III-4, and III-5, n1, n2, n3, n4, and n5 each independently represent zero or one or more groups; in formulas III-3, III-4, and III-5, X1 and X2 are nitrogen atoms, and R1, R2, R3, R4, and R5 are independently selected from hydrogen, halogen, straight-chain alkyl, branched-chain alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, cycloalkylene, heterocycloalkylene, cycloalkenyl, heterocycloalkenyl, alkoxy, alkylsilyl, aryl, alkyl-substituted aryl, alkoxy-substituted aryl, aryl-substituted aryl, and fluoroalkyl-substituted aryl. , fluoroaryl-substituted aryl, aralkyl, aryloxy, heteroaryl, alkyl-substituted heteroaryl, alkoxy-substituted heteroaryl, aryl-substituted heteroaryl, fluoroalkyl-substituted heteroaryl, fluoroaryl-substituted heteroaryl, cyano, esters and their derivatives, alkyl ketones, alkyl esters, aryl esters, alkynyl, alkylamine, fluoroalkyl, fluoroaryl, polyalkylene, aniline-substituted fluorenyl, diphenylamino-substituted fluorenyl, triphenylamino-substituted fluorenyl, alkyl-substituted triphenylamino, alkyl-substituted diphenylamino, alkyl-substituted aniline, alkyl-substituted thiophenyl, borate ester, hydroxyl, amino, nitro and trimethylsilylphenoxy; In Formulas III-3, III-4, and III-5, R1 and R5 at the adjacent positions of the carbon atom bonded to the nitrogen atom can be linked to form a nitrogen heterocycle; R2 and R3 at the adjacent positions of the carbon atom bonded to the nitrogen atom can be linked to form a nitrogen heterocycle; R3 and R4 at the adjacent positions of the carbon atom bonded to the nitrogen atom can be linked to form a six-membered heterocycle, wherein the para position of the nitrogen atom in the six-membered heterocycle can be substituted with one of oxygen, nitrogen, hydrogen, carbon, boron, or sulfur, and the para position of the nitrogen atom in the six-membered heterocycle can be bonded to hydrogen, C1-C8 alkyl, C1-C8 alkoxy, aryl, aralkyl, or heteroaryl. Preferably, in formulas III-3, III-4, and III-5, R1, R2, R3, and R5 are each independently selected from hydrogen, C1-C8 alkyl, C1-C8 alkoxy, aryl, aralkyl, and heteroaryl, and R4 is each independently selected from hydrogen, C1-C8 alkyl, C1-C8 alkoxy, aryl, aralkyl, heteroaryl, borate ester, and halogen. Preferably, in Formulas III-3, III-4, and III-5, R1, R2, R3, R4, and R5 are each independently selected from at least two substituents selected from the first, second, third, and fourth substituents; the first substituent includes C6-C18 straight-chain alkyl, C6-C18 heteroalkyl, polyalkylene, C6-C18 esters and their derivatives, alkyl ketones, alkyl esters, alkylamines, borate esters, hydroxyl groups, amino groups, and nitro groups; the second substituent includes C4-C8 branched alkyl, aryl, alkyl-substituted aryl, aryl-substituted aryl, fluoroalkyl-substituted aryl, fluoroaryl-substituted aryl, aralkyl, aryloxy, heteroaryl, aniline-substituted fluorenyl, and diphenylamino-substituted... The substituents include fluorenyl, triphenylamino-substituted fluorenyl, alkyl-substituted triphenylamino, alkyl-substituted diphenylamino, alkyl-substituted aniline, C6-C18 cycloalkyl, heterocycloalkyl, cycloalkenyl, heterocycloalkenyl, and aryl esters; the third substituent includes hydrogen, halogen, C1-C5 straight-chain alkyl, C3 branched alkyl, C3-C5 cycloalkyl, C1-C6 alkoxy, cyano, and C2-C5 alkynyl; the fourth substituent includes fluoroalkyl, fluoroaryl, alkyl-substituted heteroaryl, alkoxy-substituted heteroaryl, aryl-substituted heteroaryl, fluoroalkyl-substituted heteroaryl, fluoroaryl-substituted heteroaryl, heterocycloalkylene, cycloalkylene, alkyl-substituted thiophenyl, alkoxy-substituted aryl, alkylsilyl, and trimethylsilylphenoxy. Preferably, the narrow-band organic light-emitting molecule has the structure shown in formula III-5a, III-5b, III-5c, III-5d, III-5e, or III-5f: Formula III-5a Formula III-5b Formula III-5c Formula III-5d Formula III-5e Formula III-5f; In formulas III-5a, III-5b, III-5c, III-5d, III-5e, and III-5f, n1, n2, n3, n4, and n5 each independently represent zero or one or more groups; in formulas III-5a, III-5b, III-5c, III-5d, III-5e, and III-5f, R1 ′ R2 ′ R3 ′ R4 ′ R5 ′ R6 ′ Each of the following is independently selected from hydrogen, halogen, straight-chain alkyl, branched-chain alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, cycloalkylene, heterocycloalkylene, cycloalkenyl, heterocycloalkenyl, alkoxy, alkylsilyl, aryl, alkyl-substituted aryl, alkoxy-substituted aryl, aryl-substituted aryl, fluoroalkyl-substituted aryl, fluoroaryl-substituted aryl, aralkyl, aryloxy, heteroaryl, alkyl-substituted heteroaryl, alkoxy-substituted heteroaryl, aryl-substituted heteroaryl, fluorine Alkyl-substituted heteroaryl, fluoroaryl-substituted heteroaryl, cyano, esters and their derivatives, alkyl ketones, alkyl esters, aryl esters, alkynyl, alkylamine, fluoroalkyl, fluoroaryl, polyalkylene, aniline-substituted fluorenyl, diphenylamino-substituted fluorenyl, triphenylamino-substituted fluorenyl, alkyl-substituted triphenylamino, alkyl-substituted diphenylamino, alkyl-substituted aniline, alkyl-substituted thiophenyl, borate ester, hydroxyl, amino, nitro and trimethylsilylphenoxy; Preferably, in formulas III-5a, III-5b, and III-5c, R1 ′ R2 ′ R3 ′ R4 ′ R5 ′ R6 ′ Each of the following substituents is independently selected from at least two substituents: the first substituent, the second substituent, the third substituent, and the fourth substituent; in formulas III-5d, III-5e, and III-5f, R1 ′ R2 ′ R3 ′ R4 ′ R5 ′ Each of the substituents is independently selected from at least two substituents selected from the first, second, third, and fourth substituents; the first substituent includes C6-C18 straight-chain alkyl, C6-C18 heteroalkyl, polyalkylene, C6-C18 esters and their derivatives, alkyl ketones, alkyl esters, alkylamines, borate esters, hydroxyl groups, amino groups, and nitro groups; the second substituent includes C4-C8 branched alkyl, aryl, alkyl-substituted aryl, aryl-substituted aryl, fluoroalkyl-substituted aryl, fluoroaryl-substituted aryl, aralkyl, aryloxy, heteroaryl, aniline-substituted fluorenyl, diphenylamino-substituted fluorenyl, triphenylamino-substituted fluorenyl, and alkyl-substituted triphenylene. The group includes amino, alkyl-substituted diphenylamino, alkyl-substituted aniline, C6-C18 cycloalkyl, heterocycloalkyl, cycloalkenyl, heterocycloalkenyl, and aryl esters; the third substituent includes hydrogen, halogen, C1-C5 straight-chain alkyl, C3 branched alkyl, C3-C5 cycloalkyl, C1-C6 alkoxy, cyano, and C2-C5 alkynyl; the fourth substituent includes fluoroalkyl, fluoroaryl, alkyl-substituted heteroaryl, alkoxy-substituted heteroaryl, aryl-substituted heteroaryl, fluoroalkyl-substituted heteroaryl, fluoroaryl-substituted heteroaryl, heterocycloalkylene, cycloalkylene, alkyl-substituted thiophenyl, alkoxy-substituted aryl, alkylsilyl, and trimethylsilylphenoxy. Preferably, the narrow-band organic light-emitting molecule is selected from one of the following structures: 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 ； Wherein, R is independently selected from hydrogen, halogen, straight-chain alkyl, branched-chain alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, cycloalkylene, heterocycloalkylene, cycloalkenyl, heterocycloalkenyl, alkoxy, alkylsilyl, aryl, alkyl-substituted aryl, alkoxy-substituted aryl, aryl-substituted aryl, fluoroalkyl-substituted aryl, fluoroaryl-substituted aryl, aralkyl, aryloxy, heteroaryl, alkyl-substituted heteroaryl, alkoxy-substituted heteroaryl, aryl-substituted heteroaryl. , fluoroalkyl-substituted heteroaryl, fluoroaryl-substituted heteroaryl, cyano, esters and their derivatives, alkyl ketones, alkyl esters, aryl esters, alkynyl, alkylamine, fluoroalkyl, fluoroaryl, polyalkylene, aniline-substituted fluorenyl, diphenylamino-substituted fluorenyl, triphenylamino-substituted fluorenyl, alkyl-substituted triphenylamino, alkyl-substituted diphenylamino, alkyl-substituted aniline, alkyl-substituted thiophenyl, borate ester, hydroxyl, amino, nitro and trimethylsilylphenoxy; Preferably, R is independently selected from at least two substituents selected from the first substituent, second substituent, third substituent, and fourth substituent; the first substituent includes C6-C18 straight-chain alkyl, C6-C18 heteroalkyl, polyalkylene, C6-C18 esters and their derivatives, alkyl ketones, alkyl esters, alkylamines, borate esters, hydroxyl groups, amino groups, and nitro groups; the second substituent includes C4-C8 branched alkyl, aryl, alkyl-substituted aryl, aryl-substituted aryl, fluoroalkyl-substituted aryl, fluoroaryl-substituted aryl, aralkyl, aryloxy, heteroaryl, aniline-substituted fluorenyl, diphenylamino-substituted fluorenyl, triphenylamino-substituted fluorenyl, alkyl-substituted The substituents include triphenylamino, alkyl-substituted diphenylamino, alkyl-substituted aniline, C6-C18 cycloalkyl, heterocycloalkyl, cycloalkenyl, heterocycloalkenyl, and aryl esters; the third substituent includes hydrogen, halogen, C1-C5 straight-chain alkyl, C3 branched alkyl, C3-C5 cycloalkyl, C1-C6 alkoxy, cyano, and C2-C5 alkynyl; the fourth substituent includes fluoroalkyl, fluoroaryl, alkyl-substituted heteroaryl, alkoxy-substituted heteroaryl, aryl-substituted heteroaryl, fluoroalkyl-substituted heteroaryl, fluoroaryl-substituted heteroaryl, heterocycloalkylene, cycloalkylene, alkyl-substituted thiophenyl, alkoxy-substituted aryl, alkylsilyl, and trimethylsilylphenoxy. Preferably, the narrow-band organic light-emitting molecule has a structure as shown in Formula III-5g or Formula III-5h: Formula III - 5g; Formula III-5h; In Formula III-5g and Formula III-5h, n1, n2, n3, n4, n5, n6, n7, n8, and n9 each independently represent zero or one or more groups; in Formula III-5g and Formula III-5h, R1, R2, R3, R4, R5, R6, R7, R8, and R9 each independently select from hydrogen, halogen, straight-chain alkyl, branched-chain alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, cycloalkylene, heterocycloalkylene, cycloalkenyl, heterocycloalkenyl, alkoxy, alkylsilyl, aryl, alkyl-substituted aryl, alkoxy-substituted aryl, aryl-substituted aryl, and fluoroalkyl-substituted aryl. , fluoroaryl-substituted aryl, aralkyl, aryloxy, heteroaryl, alkyl-substituted heteroaryl, alkoxy-substituted heteroaryl, aryl-substituted heteroaryl, fluoroalkyl-substituted heteroaryl, fluoroaryl-substituted heteroaryl, cyano, esters and their derivatives, alkyl ketones, alkyl esters, aryl esters, alkynyl, alkylamine, fluoroalkyl, fluoroaryl, polyalkylene, aniline-substituted fluorenyl, diphenylamino-substituted fluorenyl, triphenylamino-substituted fluorenyl, alkyl-substituted triphenylamino, alkyl-substituted diphenylamino, alkyl-substituted aniline, alkyl-substituted thiophenyl, borate ester, hydroxyl, amino, nitro and trimethylsilylphenoxy; Preferably, in Formula III-5g, R1, R2, R3, R4, R5, and R6 are each independently selected from at least two substituents selected from the first, second, third, and fourth substituents; in Formula III-5h, R1, R2, R3, R4, R5, R6, R7, R8, and R9 are each independently selected from at least two substituents selected from the first, second, third, and fourth substituents; the first substituent includes C6-C18 straight-chain alkyl, C6-C18 heteroalkyl, polyalkylene, C6-C18 esters and their derivatives, alkyl ketones, alkyl esters, alkylamines, borate esters, hydroxyl groups, amino groups, and nitro groups; the second substituent includes C4-C8 branched alkyl, aryl, alkyl-substituted aryl, aryl-substituted aryl, fluoroalkyl-substituted aryl, and fluoroaryl-substituted... The aryl, aralkyl, aryloxy, heteroaryl, aniline-substituted fluorenyl, diphenylamino-substituted fluorenyl, triphenylamino-substituted fluorenyl, alkyl-substituted triphenylamino, alkyl-substituted diphenylamino, alkyl-substituted aniline, C6-C18 cycloalkyl, heterocycloalkyl, cycloalkenyl, heterocycloalkenyl, and aryl esters; the third substituent includes hydrogen, halogen, C1-C5 straight-chain alkyl, C3 branched alkyl, C3-C5 cycloalkyl, C1-C6 alkoxy, cyano, and C2-C5 alkynyl; the fourth substituent includes fluoroalkyl, fluoroaryl, alkyl-substituted heteroaryl, alkoxy-substituted heteroaryl, aryl-substituted heteroaryl, fluoroalkyl-substituted heteroaryl, fluoroaryl-substituted heteroaryl, heterocycloalkylene, cycloalkylene, alkyl-substituted thiophenyl, alkoxy-substituted aryl, alkylsilyl, and trimethylsilylphenoxy; Preferably, the narrow-band organic light-emitting molecule has the structure shown in Formula III-5i: Formula III-5i; In formula III-5i, R1, R2, R3, R4, and R5 are each independently selected from H and C1-C4 alkyl groups; Preferably, the narrow-band organic light-emitting molecule is selected from one of the following structures: 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 ； (C) The narrow-band organic light-emitting molecule has the structure shown in Formula IV or Formula V: Formula IV, Formula V, In Formula IV, n1, n2, n3, and n4 each independently represent zero or one or more groups; In Formulas IV and V, R1, R2, R3, R4, and R are each independently selected from hydrogen, halogen, straight-chain alkyl, branched-chain alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, cycloalkylene, heterocycloalkylene, cycloalkenyl, heterocycloalkenyl, alkoxy, alkylsilyl, aryl, alkyl-substituted aryl, alkoxy-substituted aryl, aryl-substituted aryl, fluoroalkyl-substituted aryl, fluoroaryl-substituted aryl, aralkyl, aryloxy, heteroaryl, alkyl-substituted heteroaryl, alkoxy-substituted heteroaryl, Aryl-substituted heteroaryl, fluoroalkyl-substituted heteroaryl, fluoroaryl-substituted heteroaryl, cyano, esters and their derivatives, alkyl ketones, alkyl esters, aryl esters, alkynyl, alkylamine, fluoroalkyl, fluoroaryl, polyalkylene, aniline-substituted fluorenyl, diphenylamino-substituted fluorenyl, triphenylamino-substituted fluorenyl, alkyl-substituted triphenylamino, alkyl-substituted diphenylamino, alkyl-substituted aniline, alkyl-substituted thiophenyl, borate ester, hydroxyl, amino, nitro and trimethylsilylphenoxy; Preferably, in Formula IV, R1, R2, R3, and R4 are each independently selected from at least two substituents selected from the first substituent, second substituent, third substituent, and fourth substituent; the first substituent includes C6-C18 straight-chain alkyl, C6-C18 heteroalkyl, polyalkylene, C6-C18 esters and their derivatives, alkyl ketones, alkyl esters, alkylamines, borate esters, hydroxyl groups, amino groups, and nitro groups; the second substituent includes C4-C8 branched alkyl, aryl, alkyl-substituted aryl, aryl-substituted aryl, fluoroalkyl-substituted aryl, fluoroaryl-substituted aryl, aralkyl, aryloxy, heteroaryl, aniline-substituted fluorenyl, diphenylamino-substituted fluorenyl, triphenylamino-substituted fluorenyl... The substituents include fluorenyl, alkyl-substituted triphenylamino, alkyl-substituted diphenylamino, alkyl-substituted aniline, C6-C18 cycloalkyl, heterocycloalkyl, cycloalkenyl, heterocycloalkenyl, and aryl esters; the third substituent includes hydrogen, halogen, C1-C5 straight-chain alkyl, C3 branched alkyl, C3-C5 cycloalkyl, C1-C6 alkoxy, cyano, and C2-C5 alkynyl; the fourth substituent includes fluoroalkyl, fluoroaryl, alkyl-substituted heteroaryl, alkoxy-substituted heteroaryl, aryl-substituted heteroaryl, fluoroalkyl-substituted heteroaryl, fluoroaryl-substituted heteroaryl, heterocycloalkylene, cycloalkylene, alkyl-substituted thiophenyl, alkoxy-substituted aryl, alkylsilyl, and trimethylsilylphenoxy. Preferably, the narrow-band organic light-emitting molecule is selected from one of the following structures: 、 、 、 、 ； (D) The narrow-band organic light-emitting molecule has the structure shown in Formula VI: Formula VI, In Formula VI, n is an integer of 0 ≤ n ≤ 10, R and R′ are each independently selected from hydrogen, alkyl, alkoxy, aryl, heteroaryl and halogen, Ar is aryl, each X is independently selected from one of O, S, N and CR′′, when X is O or S, the Ar connected to X does not exist, and R′′ is selected from hydrogen, alkyl, alkoxy, aryl, heteroaryl and halogen; Preferably, the narrow-band organic light-emitting molecule has a structure as shown in Formula VI(a), Formula VI(b), or Formula VI(c): Formula VI(a), Formula VI(b) Formula VI(c), In formulas VI(a), VI(b) and VI(c), R and R′ are each independently selected from hydrogen, alkyl, alkoxy, aryl, heteroaryl and halogen, and Ar is aryl; (E) The narrow-band organic light-emitting molecule has a structure as shown in Formula VII: Formula VII In formula VII, n1, n2, n3, n4, n5, n6, n7, and n8 each independently represent zero or one or more groups; In Equation VII, each X is independently selected from O, NR9, S, and CR. 10 R 11 One of them, R1, R2, R3, R4, R5, R6, R7, R8, R9, R 10 R 11 Each is independently selected from hydrogen, C1-C6 alkyl, C1-C6 alkoxy, arylalkyl, heteroaryl, cyano, and halogen; Preferably, the narrow-band organic light-emitting molecule is selected from one of the following structures: 、 、 、 、 ； (F) The narrow-band organic light-emitting molecule has the structure shown in Formula VIII: Formula VIII; In formula VIII, n1, n2, n3, and n4 each independently represent zero or one or more groups; In Formula VIII, R1, R2, R3, and R4 are each independently selected from hydrogen, alkyl, alkoxy, aryl, heteroaryl, halogen, cyano, and borate ester group; X1 is selected from one of O, NR5, S, and CR6R7; X2 is selected from one of N and CR8; and R5, R6, R7, and R8 are each independently selected from hydrogen, alkyl, alkoxy, aryl, heteroaryl, halogen, and cyano. Preferably, the narrow-band organic light-emitting molecule has one of the following structures: ， ， ； Preferably, the narrow-band organic light-emitting molecule has the structure shown in Formula VIII(a): Ⅷ(a)， In VIII(a), n1, n2, n3, and n4 each independently represent zero or one or more groups; in VIII(a), R1, R2, R3, and R4 are selected from hydrogen, alkyl, alkoxy, aryl, heteroaryl, halogen, and cyano. Preferably, the narrow-band organic light-emitting molecule has one of the following structures: 、 、 、 、 、 、 、 、 、 ； (G) The narrow-band organic light-emitting molecule has a structure as shown in any one of Formulas IX-1 to IX-3: Equation IX-1; Equation IX-2; Equation IX-3; In formulas IX-1 to IX-3, n1, n2, n3, and n4 each independently represent zero or one or more groups; in formulas IX-1 to IX-3, R1, R2, R3, R4, R5, and R6 each independently select from hydrogen, halogen, straight-chain alkyl, branched-chain alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, cycloalkylene, heterocycloalkylene, cycloalkenyl, heterocycloalkenyl, alkoxy, alkylsilyl, aryl, alkyl-substituted aryl, alkoxy-substituted aryl, aryl-substituted aryl, fluoroalkyl-substituted aryl, fluoroaryl-substituted aryl, aralkyl, and aryloxy alkyl, heteroaryl, alkyl-substituted heteroaryl, alkoxy-substituted heteroaryl, aryl-substituted heteroaryl, fluoroalkyl-substituted heteroaryl, fluoroaryl-substituted heteroaryl, cyano, esters and their derivatives, alkyl ketones, alkyl esters, aryl esters, alkynyl, alkylamine, fluoroalkyl, fluoroaryl, polyalkylene, aniline-substituted fluorenyl, diphenylamino-substituted fluorenyl, triphenylamino-substituted fluorenyl, alkyl-substituted triphenylamino, alkyl-substituted diphenylamino, alkyl-substituted aniline, alkyl-substituted thiophenyl, borate ester, hydroxyl, amino, nitro and trimethylsilylphenoxy; Preferably, in Formula IX-1, R1, R2, and R3 are each independently selected from at least two substituents selected from the first, second, third, and fourth substituents; in Formulas IX-2 and IX-3, R1, R2, R3, R4, R5, and R6 are each independently selected from at least two substituents selected from the first, second, third, and fourth substituents; the first substituent includes C6-C18 straight-chain alkyl, C6-C18 heteroalkyl, polyalkylene, C6-C18 esters and their derivatives, alkyl ketones, alkyl esters, alkylamines, borate esters, hydroxyl groups, amino groups, and nitro groups; the second substituent includes C4-C8 branched alkyl, aryl, alkyl-substituted aryl, aryl-substituted aryl, fluoroalkyl-substituted aryl, fluoroaryl-substituted aryl, aralkyl, and arylalkyl groups. The substituents include oxy-, heteroaryl, aniline-substituted fluorenyl, diphenylamine-substituted fluorenyl, triphenylamine-substituted fluorenyl, alkyl-substituted triphenylamine, alkyl-substituted diphenylamine, alkyl-substituted aniline, C6-C18 cycloalkyl, heterocycloalkyl, cycloalkenyl, heterocycloalkenyl, and aryl esters; the third substituent includes hydrogen, halogen, C1-C5 straight-chain alkyl, C3 branched alkyl, C3-C5 cycloalkyl, C1-C6 alkoxy, cyano, and C2-C5 alkynyl; the fourth substituent includes fluoroalkyl, fluoroaryl, alkyl-substituted heteroaryl, alkoxy-substituted heteroaryl, aryl-substituted heteroaryl, fluoroalkyl-substituted heteroaryl, fluoroaryl-substituted heteroaryl, heterocycloalkylene, cycloalkylene, alkyl-substituted thiophenyl, alkoxy-substituted aryl, alkylsilyl, and trimethylsilylphenoxy. Preferably, the narrow-band organic light-emitting molecule is selected from one of the following structures: 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 ； Wherein, R is independently selected from hydrogen, halogen, straight-chain alkyl, branched-chain alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, cycloalkylene, heterocycloalkylene, cycloalkenyl, heterocycloalkenyl, alkoxy, alkylsilyl, aryl, alkyl-substituted aryl, alkoxy-substituted aryl, aryl-substituted aryl, fluoroalkyl-substituted aryl, fluoroaryl-substituted aryl, aralkyl, aryloxy, heteroaryl, alkyl-substituted heteroaryl, alkoxy-substituted heteroaryl, aryl-substituted heteroaryl. , fluoroalkyl-substituted heteroaryl, fluoroaryl-substituted heteroaryl, cyano, esters and their derivatives, alkyl ketones, alkyl esters, aryl esters, alkynyl, alkylamine, fluoroalkyl, fluoroaryl, polyalkylene, aniline-substituted fluorenyl, diphenylamino-substituted fluorenyl, triphenylamino-substituted fluorenyl, alkyl-substituted triphenylamino, alkyl-substituted diphenylamino, alkyl-substituted aniline, alkyl-substituted thiophenyl, borate ester, hydroxyl, amino, nitro and trimethylsilylphenoxy; (H) The narrow-band organic light-emitting molecule has the structure shown in Formula X: Formula X, In formula X, n1, n2, and n3 each independently represent zero, one, or more groups; In formula X, R1, R2, and R3 are selected from hydrogen, alkyl, alkoxy, aryl, heteroaryl, halogen, and cyano, and X, X1, and X2 are selected from C, O, N, and S. The chemical bonds between X and X1 and between X and X2 can be saturated or unsaturated. Preferably, the narrow-band organic light-emitting molecule is selected from one of the structures shown below: 、 ； (I) The narrow-band organic light-emitting molecule has the structure shown in Formula XI: Formula XI; In formula XI, n1, n2, n3, and n4 each independently represent zero or one or more groups; In formula XI, R1, R2, and R3 are each independently selected from hydrogen, alkyl, alkoxy, aryl, alkylaryl, heteroaryl, halogen, cyano, and diphenylamino. Preferably, the narrow-band organic light-emitting molecule is selected from one of the following structures: 、 、 ； Where Ar is aryl; (J) The narrow-band organic light-emitting molecule has the structure shown in Formula XII: Equation XII; In equation XII, R1-R 10 Selected from H, C1-C5 alkyl groups, preferably branched alkyl groups, and more preferably tert-butyl groups; (K) The narrow-band organic light-emitting molecule has the structure shown in Formula XIII: Formula XIII; In equation XIII, R1-R 12 Selected from H, C1-C5 alkyl groups, preferably branched alkyl groups, and more preferably tert-butyl groups; (L) The narrow-band organic light-emitting molecule has a structure as shown in Formula XIV: Formula XIV; In formula XIV, R1-R4 are selected from H, C1-C5 alkyl, aryl, and triphenylamine, preferably branched alkyl, aromatic, and triphenylamine, and more preferably tert-butyl, phenyl, and triphenylamine; (M) The narrow-band organic light-emitting molecule has a structure as shown in Formula XV: Formula XV; In formula XV, R1-R4 are selected from aromatic hydrocarbons and polycyclic aromatic hydrocarbons, and R5-R8 are selected from H and C1-C6 alkyl groups; (N) The narrow-band organic light-emitting molecule is selected from one of the following structures: 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 。 6. The single-peak narrow-band luminescent microsphere as described in claim 1, characterized in that, The single-peak narrowband luminescent microsphere further includes a microsphere structure, wherein the organic luminescent molecules are located at one or more positions on the surface, interface, and interior of the microsphere structure; Preferably, the microsphere structure comprises polymeric microspheres; preferably, the polymeric microspheres are amphiphilic, and the surface of the polymeric microspheres contains one or more groups selected from epoxy, carboxyl, amino, aldehyde, hydroxyl, and amide groups; preferably, the polymeric microspheres are styrene polymer microspheres, and the surface of the styrene polymer microspheres contains one or more groups selected from amino, amide, carboxyl, and aldehyde groups; preferably, the styrene polymer comprises a copolymer formed by styrene and a comonomer, the comonomer comprising olefins, alkynes, carboxylic acids containing carbon-carbon double bonds, acid anhydrides containing carbon-carbon double bonds, amides containing carbon-carbon double bonds, esters containing carbon-carbon double bonds, and derivatives thereof having one or more substituents selected from halogens, amino, amide, aldehyde, and carboxyl groups; preferably, the styrene polymer is styrene and a copolymer formed from styrene and a copolymer of ... The styrene polymer is a copolymer of one or more comonomers selected from carboxylic acids containing carbon-carbon double bonds, aliphatic acid anhydrides containing carbon-carbon double bonds, aliphatic amides containing carbon-carbon double bonds, and aliphatic esters containing carbon-carbon double bonds; preferably, the styrene polymer is a copolymer of styrene with one or more comonomers selected from carboxylic acids represented by formula Y1-COOH and amides represented by formula Y2-CONH2, wherein Y1 and Y2 are each independently C2-C18 straight-chain, branched, or cyclic aliphatic alkyl groups in which one or more carbon-carbon single bonds are replaced by carbon-carbon double bonds; preferably, Y1 and Y2 are each independently C3-C8 straight-chain, branched, or cyclic aliphatic alkyl groups in which one or more carbon-carbon single bonds are replaced by carbon-carbon double bonds; preferably, the styrene polymer is a copolymer of styrene with (meth)acrylic acid and / or (meth)acrylamide.

7. A homogeneous detection reagent, characterized in that, The homogeneous detection reagent comprises any one of the single-peak narrow-band luminescent microspheres according to any one of claims 1-6.

8. The homogeneous detection reagent as described in claim 7, characterized in that, The homogeneous detection reagent has one or more of the following characteristics: The single-peak narrowband luminescent microspheres include a buffer, and the single-peak narrowband luminescent microspheres are receptor microspheres of the homogeneous detection reagent; The homogeneous detection reagent also includes donor microspheres containing photosensitizers; The homogeneous detection reagent includes one or more single-peak narrowband luminescent microspheres, the single-peak narrowband luminescent microspheres include a buffer, and the emission spectra of the single-peak narrowband luminescent microspheres have zero overlap.

9. A homogeneous detection kit, characterized in that, The homogeneous detection kit includes the homogeneous detection kit as described in claim 7 or 8.

10. Use of the single-peak narrow-band luminescent microspheres according to any one of claims 1-6 in the preparation of optical probes, flow cytometry reagents, biomarkers, immunochromatographic reagents, immunochromatographic kits, or radiopharmaceuticals.

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