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Immunomagnetic bead and preparation method thereof

A technology of immunomagnetic beads and magnetic microspheres, applied in biochemical equipment and methods, microsphere preparation, cell dissociation methods, etc., can solve the problems of inactivation of active substances such as proteins, protein cross-linking, and coating of unfavorable biological ligands and other problems, to achieve the effect of simple process steps, simple raw materials and low cost

Inactive Publication Date: 2017-02-01
SHANGHAI MAJORBIO BIO PHARM TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, this method also needs to add a strong reducing agent (such as sodium borohydride or sodium cyanoborohydride) during the reductive amination reaction, which is likely to cause inactivation of active substances such as proteins, and glutaraldehyde will cause magnetic microspheres and biological complexes. The cross-linking of the base will also lead to the internal cross-linking of the protein, which is not conducive to the coating of bioligands

Method used

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  • Immunomagnetic bead and preparation method thereof
  • Immunomagnetic bead and preparation method thereof
  • Immunomagnetic bead and preparation method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0031] (1) Preparation of magnetic nanoclusters: Preparation of magnetic nanoclusters Fe by co-precipitation method 3 o 4 1;

[0032] (2) Preparation of amino-modified magnetic microspheres: add 10 mg magnetic nanocluster Fe 3 o 4 1 solution, add 125 mg ammonia water, 30 mg tetraethyl orthosilicate and 30 mg (3-aminopropyl) triethoxysilane, react for 1 day, carry out magnetic separation, and wash each twice with ethanol and water alternately, and use 0.1 Amino-modified magnetic microspheres 1 with a concentration of 5 mmol / L were obtained after dispersion in a phosphate buffer solution with a pH of 7 to 8;

[0033] (3) Preparation of hydrazine-modified bioligands: Add 5 μL of SANH DMF solution (concentration: 5 mmol / L) to 100 μL fluorescently labeled CD45 antibody solution (concentration: 10 μmol / L), react at room temperature for 2 h , purified by an ultrafiltration column to obtain hydrazine-modified bioligand 1;

[0034] (4) Preparation of aldehyde-modified magnetic mic...

Embodiment 2

[0039] (1) Preparation of magnetic nanoclusters: Preparation of magnetic nanoclusters Fe by hydrothermal method 3 o 4 2;

[0040] (2) Preparation of amino-modified magnetic microspheres: add 10 mg magnetic nanocluster Fe 3 o 4 2, add 250 mg of ammonia water, 20 mg of tetraethyl orthosilicate and 10 mg of (3-aminopropyl) triethoxysilane, react for 2 days, carry out magnetic separation, and alternately wash twice with ethanol and water, and wash with 0.1 Amino-modified magnetic microspheres 2 with a concentration of 5 mmol / L were obtained after dispersion in a phosphate buffer solution with a pH of 7-8;

[0041] (3) Preparation of hydrazine-modified bioligands: Add 5 μL of SANH DMF solution (concentration: 2 mmol / L) into 100 μL BSA solution (concentration: 10 μmol / L), react at room temperature for 3 h, and filter with ultrafiltration column Purification to obtain hydrazine-modified bioligand 2;

[0042] (4) Preparation of aldehyde-modified magnetic microspheres: Add 5 μL o...

Embodiment 3

[0046] (1) Preparation of magnetic nanoclusters: Preparation of magnetic nanoclusters Fe by solvothermal method 3 o 4 3;

[0047] (2) Preparation of amino-modified magnetic microspheres: add 10 mg magnetic nanocluster Fe 3 o 4 Add 125 mg of ammonia water, 40 mg of tetraethyl orthosilicate and 5 mg of (3-aminopropyl) triethoxysilane to the solution of 3, conduct magnetic separation after 3 days of reaction, and alternately wash twice with ethanol and water, and wash with 0.1 Amino-modified magnetic microspheres 3 with a concentration of 5 mmol / L were obtained after dispersion in a phosphate buffer solution with a pH of 7 to 8;

[0048] (3) Preparation of hydrazine-modified magnetic microspheres: Add 2 μL of amino-modified magnetic microspheres 1 to 50 μL of SANH’s DMF solution (concentration: 2 mmol / L), react at room temperature for 24 h, and perform magnetic separation and purification. The magnetic microsphere 3 modified by the hydrazine group was obtained;

[0049] (4)...

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Abstract

The invention provides an immunomagnetic bead and a preparation method thereof. The immunomagnetic bead comprises a magnetic microsphere portion and a biological ligand portion. The structure formula of the immunomagnetic bead is A-NH-N=CH-B, wherein A and B refer to a magnetic microsphere and a biological ligand respectively, or A and B refer to a biological ligand and a magnetic microsphere respectively. The immunomagnetic bead has stable character and high magnetism, the magnetic microsphere and the biological ligand are coupled chemically, reaction has high specificity, internal crosslinking is avoided, and coupling efficiency is high; the preparation method is simple, reaction conditions are mild, and the biological ligand can keep biological activity; raw materials are easy to get, cost is low, process steps are simple, and high practicability is achieved.

Description

technical field [0001] The invention relates to the field of preparation of immunomagnetic beads, in particular to an immunomagnetic bead and a preparation method thereof. Background technique [0002] Immonumagnetic beads (IMB for short) are composed of magnetic microspheres and biological ligands. The combination of magnetic microspheres and biological ligands generally has physical adsorption and chemical coupling methods. Due to the short storage time of the prepared immunomagnetic beads, the easy fall off of the surface active substances of the magnetic microspheres, the poor stability and activity of the protein, the application of the physical adsorption method is limited. Chemical coupling is to use the active groups on the surface of the magnetic microspheres to covalently couple with the active groups of the bioligands, and coat the bioligands on the surface of the magnetic microspheres. The chemical coupling method has the advantages of large binding capacity, a...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K17/14C07K17/08C12N5/09B01J13/02H01F1/11
CPCC07K17/14B01J13/02C07K17/08C12N5/0693C12N2509/00H01F1/112
Inventor 蔡红东刘关陈昌岳张祥林
Owner SHANGHAI MAJORBIO BIO PHARM TECH