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Preparation method and application of high-performance time resolved fluorescence microspheres

A time-resolved fluorescence, high-performance technology, applied in fluorescence/phosphorescence, analytical materials, material excitation analysis, etc., can solve the problems of large steric hindrance effect, insufficient fluorescence intensity and stability of time-resolved fluorescent microspheres, and achieve linearity Wide range, low steric hindrance effect, good stability

Inactive Publication Date: 2018-08-24
南京微测生物科技有限公司
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0007] The purpose of the present invention is to provide a preparation method of high-sensitivity time-resolved fluorescent microspheres that can improve the stability and fluorescence intensity of fluorescent microspheres and reduce steric hindrance effects, so as to solve the problem of fluorescence intensity of time-resolved fluorescent microspheres in the prior art. and insufficient stability, large steric hindrance effects, etc., to further improve the detection sensitivity and sample application range of time-resolved fluorescence immunochromatography

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  • Preparation method and application of high-performance time resolved fluorescence microspheres
  • Preparation method and application of high-performance time resolved fluorescence microspheres
  • Preparation method and application of high-performance time resolved fluorescence microspheres

Examples

Experimental program
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Embodiment 1

[0024] A method for preparing high-sensitivity time-resolved fluorescent microspheres capable of improving stability and fluorescence intensity and reducing steric hindrance effects, comprising the following steps:

[0025] (1) Preparation of highly stable polystyrene nanospheres

[0026] Dissolve 50mm of styrene monomer and 2.5mm of acrylic acid monomer in 50ml of deionized water containing 1.5mm of sodium dodecylsulfonate and 8mm of dextran (Mw: 25000), add it to a round bottom flask, and use a magnetic stirrer Stir evenly, then use high-purity nitrogen to remove the air in the round-bottomed flask, seal and heat to 75°C, add 3ml of 0.18mm potassium persulfate, seal the oxygen barrier and stir for 12 hours, then cool down to room temperature, and then use whatman Filter with 2v filter paper (pore size 8 μm), pass through a Sephadex G-200 molecular sieve column, collect the first eluted fraction and concentrate to a concentration of 100 mg / mL, add 0.05% sodium azide and store...

Embodiment 2

[0035] The influence of the dextran of embodiment 2 different concentrations and different molecular weights on microsphere stability

[0036] With reference to Example 1, other conditions and parameters were not adjusted, only the amount and molecular weight of dextran added in step (1) were changed, and then the final prepared fluorescent microspheres were diluted to 1 mg / ml with 30% methanol, Let stand overnight. Centrifuge at 20000RPM for 30min, take 100ul of the supernatant solution and add it to the microwell, and then use the AutoDELFIA-1235 automatic time-resolved fluorescence immunoassay analyzer produced by Perlin Elmer Life Sciences to test. If the fluorescence intensity in the supernatant is stronger, it means that The more serious the leakage of the fluorescent substance, the worse the stability of the microspheres, and vice versa, the stronger the stability of the fluorescent microspheres and the stronger the ability to resist the interference of the matrix.

[...

Embodiment 3

[0041] Embodiment 3 The influence of the doping of different ion species and its concentration on the fluorescence intensity

[0042] Referring to Example 1, other conditions and parameters were not adjusted, only the amount of ytterbium trichloride and lutetium trichloride added in step (2) was changed, and then the fluorescent microspheres finally prepared were added. Dilute to 1mg / ml with pure water, take 100ul and add it to a microwell plate, and then use the AutoDELFIA-1235 automatic time-resolved fluorescence immunoassay analyzer produced by Perlin Elmer Life Sciences to test, if the fluorescence intensity is stronger, it means chelation The better the fluorescence enhancement effect of the chelate, the higher the detection sensitivity will be; otherwise, the worse the fluorescence enhancement effect of the chelate, the lower the detection sensitivity will be.

[0043] Table 2 Effects of doping with different ion species and their concentrations on fluorescence intensity...

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Abstract

The invention relates to the technical field of detection and in particular relates to a preparation method and application of high-performance time resolved fluorescence microspheres. The preparationmethod comprises the following steps: (1) preparing high-stability polystyrene nano-microspheres; (2) preparing europium ion chelates doped with different ions; (3) preparing nano microspheres with high fluorescence intensity; (4) carrying out stereo-hindrance effect removing effect modification on the surfaces of the fluorescence microspheres. The time resolved fluorescence microspheres providedby the invention have the characteristics of high fluorescence intensity, low stereo-hindrance effect, good stability and the like; a time resolved fluorescence immunochromatography technology developed based on the microspheres has the advantages of high detection sensitivity, wide linear range, high precision, wide sample applicable range and the like, and hypersensitive, rapid and quantitativedetection and analysis of a substance to be detected can be realized.

Description

technical field [0001] The invention belongs to the technical field of biomedical detection, and in particular relates to a preparation method and application of high-performance time-resolved fluorescent microspheres. Background technique [0002] Time resolved fluorescent lateral flowimmunoassay (TRFLFI) is a rapid detection technology based on the combination of fluorescent nanosphere labeling technology and immunochromatographic analysis technology. Fluorescent nanospheres are used as tracer labels After a specific reaction occurs for proteins, polypeptides, hormones, antibodies, nucleic acid probes or biologically active cells, use a time-resolved fluorescence detector to measure the fluorescence intensity of the final conjugate, and calculate the reaction system based on the ratio of fluorescence intensity to relative fluorescence intensity. The concentration of the analyte is used to achieve the purpose of quantitative analysis. TRFLFI is gradually replacing enzyme-l...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/577G01N33/558G01N21/64
CPCG01N21/6408G01N21/6486G01N33/558G01N33/577
Inventor 肖理文徐秀赵皖曹建伟
Owner 南京微测生物科技有限公司
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