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Nano-grade fluorescent microspheres, preparation method thereof, and purpose thereof as markers used in immunity chromatography method

A nano-fluorescent microsphere and microsphere technology, which is applied in the preparation of microspheres, microcapsule preparations, fluorescence/phosphorescence, etc., can solve the problems of low detection sensitivity, inability to accurately quantify, and increase the cost of detection equipment in colloidal gold immunochromatography. , to achieve the effects of improving detection sensitivity, reducing detection and analysis costs, and good monodispersity

Inactive Publication Date: 2012-07-25
SUZHOU NANO HEALTH BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
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AI Technical Summary

Problems solved by technology

[0004]However, the colloidal gold immunochromatography technique has a low detection sensitivity, and in actual detection, only qualitative or semi-quantitative testing can be carried out, but cannot be accurately quantified
At present, relevant patents have reported immunochromatographic detection using fluorescent nanoparticles as markers. For example, a patent with publication number CN1645146A discloses an immunochromatographic method using fluorescent rare earth nanoparticles as markers and its detection test paper. The preparation of strips; the patent with publication number CN1866012A discloses a quantitative and rapid immunoassay method and its special device, which uses fluorescent substance chelate Eu 3+ , SrTi 3 + , Tb 3+ , Dy 3+ are combined with organic polymer nanoparticles to prepare fluorescent particles, which are quantified by fluorescence detection, but these nanoparticles The fluorescent microspheres are all excited under the ultraviolet light source, and the setting of the ultraviolet light source increases the cost of the detection instrument

Method used

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  • Nano-grade fluorescent microspheres, preparation method thereof, and purpose thereof as markers used in immunity chromatography method
  • Nano-grade fluorescent microspheres, preparation method thereof, and purpose thereof as markers used in immunity chromatography method

Examples

Experimental program
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Effect test

Embodiment 1

[0027] Example 1 Preparation of nano fluorescent microspheres whose surface functional groups are carboxyl groups

[0028] According to this embodiment, the preparation of nano fluorescent microspheres comprises the following steps:

[0029] (1) Preparation of polymethyl methacrylate microspheres with carboxyl groups on the surface

[0030] Take 5ml of polymethyl methacrylate particles (2.7%, 0.324μm) produced by Polysciences, wash and centrifuge, suspend in 10ml ultrapure water for ultrasonication, add 3mg of methacrylic acid monomer (used to form carboxyl groups on the surface of the microspheres) ) in 1ml water, mix for two minutes, add 0.5ml water containing 15mg polyvinyl alcohol, stir for 2min, then add 0.5ml water containing 8mg sodium lauryl sulfate, mix for 2min, then add 5ml ethanol, stir for 5min to get Polymethyl methacrylate microspheres.

[0031] (2) Preparation of fluorescent nanospheres

[0032] Slowly add 750 μl of dichloromethane containing 1 m...

Embodiment 2

[0033] Example 2 Characterization of nano fluorescent microspheres

[0034] Take 10 μl of fluorescent nanometer microspheres prepared in Example 1, suspend them in 750 μl of water for detection, collect the laser spectrum of the microspheres at 610 nm, and collect the emission spectrum at 580 nm, see figure 1 .

[0035] Scanning electron microscope test was carried out on the microspheres, see the spectrogram figure 2 , the particle size of the microspheres is between 370 and 390nm, and the particle size is uniform.

Embodiment 3

[0036] Example 3 Nano fluorescent microspheres are suitable for the preparation of quantitative immunochromatographic detection test strips

[0037] Can adopt the preparation method of conventional immunochromatographic detection test strip to prepare, specifically comprise the following steps:

[0038] (1) Preparation of fluorescent microsphere pads: Spray the fluorescent microspheres labeled with the monoclonal antibody to be tested on the glass fiber membrane (30×0.8 cm) at an amount of 4 μL / cm, and dry in vacuum at 25°C for 1~2h , put in a dry environment for later use.

[0039] (2) Preparation of nitrocellulose membrane (NC membrane): Use 0.01 M pH 7.4 PBS (phosphate buffer saline, which contains 5% sucrose and 0.05% Tween-20) to adjust the secondary antibody of the antibody to be tested (concentration: 0.4mg / mL), spray the resulting solution on the NC membrane to form a detection line; use 0.01M pH 7.2 PBS (phosphate buffer saline, which contains 3% sucrose and ...

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Abstract

The invention discloses nano-grade fluorescent microspheres, a preparation method thereof, and a purpose thereof. The nano-grade fluorescent microspheres have core-shell structures, and have particle sizes of 200-500nm. The cores are polymer microspheres containing fluorescence molecules, and the shells have one or more functional groups modified on the outer surfaces of the polymer microspheres. The polymer microspheres are polymethyl methacrylate microspheres, or microspheres formed by copolymers produced by copolymerization of methyl methacrylate and other monomers, or a mixture thereof. The weight of the methyl methacrylate units at least accounts for 50% of the total weight of the nano-grade fluorescent microspheres, and the weight of the fluorescence molecules accounts for 0.05-5% of the total weight of the nano-grade fluorescent microspheres. The nano-grade fluorescent microspheres provided by the invention are advantaged in stable particle sizes. The spectrum of the nano-grade fluorescent microspheres is suitable for visible emission lights. The immunity chromatography method employing the nano-grade fluorescent microspheres as markers is convenient, fast, and highly sensitive. With the method, quantitative detection can be carried out.

Description

technical field [0001] The invention relates to a nano fluorescent microsphere suitable for use as a marker of immunochromatography, a preparation method and application thereof. Background technique [0002] In biological and medical testing, immunoassays are often used, including radioimmunoassays, enzyme-linked immunoassays, and immunochromatography. Radioimmunoassay and enzyme-linked immunoassay require expensive equipment, professional operators and complicated operation steps, and it is difficult to obtain test results quickly. Immunochromatography is often used for rapid qualitative or semi-quantitative detection because of its simple operation, rapidity and low cost. [0003] Immunochromatography technology is a unique immunoassay method that appeared in the early 1980s. It usually uses strip-shaped fiber chromatography materials as the solid phase, and the sample solution swims on the chromatography strip through capillary action, and at the same time makes the sam...

Claims

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Application Information

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IPC IPC(8): C08L51/00C08K5/18C09K11/06C08F265/06C08F2/22B01J13/14G01N33/533G01N21/64
Inventor 李超张鹏夏志道
Owner SUZHOU NANO HEALTH BIOTECH
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