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Method for quantitative determination of associated antigen by means of immunomagnetic beads and application thereof

A quantitative detection and immune magnetic bead technology, which is applied in the direction of measuring devices, instruments, scientific instruments, etc., can solve the problems of low sensitivity and inability to realize quantitative detection, and achieve high sensitivity, high signal value, and accurate detection results

Inactive Publication Date: 2016-12-14
北京乐普诊断科技股份有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] Commonly used detection methods for magnetic beads include immunoaggregation detection, optical detection, etc. These methods have disadvantages such as inability to achieve quantitative detection, low sensitivity, and the need for large-scale specialized detection equipment.

Method used

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  • Method for quantitative determination of associated antigen by means of immunomagnetic beads and application thereof
  • Method for quantitative determination of associated antigen by means of immunomagnetic beads and application thereof
  • Method for quantitative determination of associated antigen by means of immunomagnetic beads and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0048] In this embodiment, the use of immunomagnetic beads to detect D-Dimer dimer specifically includes the following steps:

[0049] (1) Choose carboxylated microspheres with a particle size of 3μm, balance the beads with 0.01M, pH 6 PBS buffer; add EDC / NHS (0.01M, pH 6 PBS buffer configuration) to make it in the system The concentration in the medium is 1mg / mL, the activation is 60min, the 0.01M, pH 6PBS buffer under the multifunctional magnetic separation rack is washed 2-3 times;

[0050] (2) Add D-Dimer coated antibody to the activated magnetic bead buffer (0.05M, pH7 MES buffer) system to make the concentration in the system 0.5mg / mL, and couple it for 4h at room temperature. Wash 2-3 times with 0.05M pH 6 MES buffer under the multifunctional magnetic separation rack; add 1% blocking solution BSA, and seal at room temperature for 40 minutes, and use 0.01M, pH 76PBS buffer under the multifunctional magnetic separation rack Wash and resuspend to obtain magnetic beads coupled ...

Embodiment 2

[0057] In this embodiment, using immunomagnetic beads to detect D-Dimer dimer antigen specifically includes the following steps:

[0058] (1) Choose carboxylated magnetic beads with a particle size of 1μm, balance the beads with 0.01M, pH 7 PBS buffer for 2-3 times; add EDC / NHS (0.01M, pH 7 PBS buffer configuration), Make the concentration in the system 0.5mg / mL, activate for 40min, wash 2-3 times with 0.01M, pH 6PBS buffer under the multifunctional magnetic separation rack to obtain activated magnetic beads;

[0059] (2) Add D-Dimer coating antibody to the activated magnetic bead buffer (0.05M, pH7 MES buffer) system to make the concentration in the system 1mg / mL, and couple for 2h at room temperature. Wash 2-3 times with 0.05M, pH6 MES buffer under the functional magnetic separation rack; add 1% blocking solution BSA, seal at room temperature for 60 minutes, and wash under the multifunctional magnetic separation rack with 0.01M, pH 7PBS buffer , Resuspend to obtain magnetic bead...

Embodiment 3

[0066] In this embodiment, using immunomagnetic beads to detect related antigens specifically includes the following steps:

[0067] (1) Choose carboxylated magnetic beads with a particle size of 200nm, balance the beads with 0.02M, pH 6 PBS buffer for 2-3 times; add EDC / NHS (0.02M, pH 6 PBS buffer configuration), Make the concentration in the system 0.1mg / mL, activate for 30 minutes, wash 2-3 times with 0.01M, pH 6PBS buffer under the multifunctional magnetic separation rack to obtain activated magnetic beads;

[0068] (2) Add D-Dimer coating antibody to the activated magnetic bead buffer (0.05M, pH6 MES buffer) system to make the concentration in the system 0.1mg / mL, and couple at room temperature for 4h, Wash 2-3 times with 0.05M pH 6 MES buffer under the multifunctional magnetic separation rack; add 1% blocking solution BSA, and block at room temperature for 40 minutes, and use 0.01M, pH 6PBS buffer under the multifunctional magnetic separation rack Wash and resuspend to obtai...

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Abstract

The invention provides a method for quantitative determination of an associated antigen by means of immunomagnetic beads. The method comprises the steps of coating an antibody with the beads in a coupling mode to obtain a bead solution of the coupled and coated antibody, mixing the bead solution of the coupled and coated antibody with a sample containing the associated antigen so that the antigen can be combined with the antibody coupled to the beads through specific immunity reaction, then adding an antibody solution marked with fluorescent microspheres so that specific immunity reaction can be conducted on the antibody marked with the fluorescent microspheres and the antigen on the beads, and detecting a fluorescence signal to obtain the content of the antigen. The detection method has the advantages that sensitivity is high, specificity is high, the problem that a certain kind of antibody is not coated firmly in certain occasions can be solved, quick detection of antigens can be achieved, stability is high, and detection results are accurate.

Description

Technical field [0001] The invention belongs to the field of biological detection, and relates to a method for quantitatively detecting related antigens by immunomagnetic beads and its application. Background technique [0002] Magnetic beads are a kind of high magnetic and stable magnetic material, with the advantages of small particle size, high uniformity, good suspension stability, super paramagnetism and so on. Its structure includes three parts: the core part is a magnetic material, and the most widely used iron and its oxides (Fe, Fe 3 O 4 , Fe 2 O 3 ); The core is wrapped with a layer of polymer solid particles (such as polyvinyl chloride, polystyrene, polyethyleneimine) as a carrier; the outermost layer is a functional base layer, such as hydroxyl (-COOH), amino (-NH) 2 ), aldehyde group (-CHO), carboxyl group (-COOH), etc., due to the different physical properties of the carrier microspheres, it can covalently bind different immunoligands, such as enzymes, cells, antibod...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/533
CPCG01N33/533
Inventor 张单单胡飞熊晶邱笑违余占江
Owner 北京乐普诊断科技股份有限公司
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