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A kind of immunomagnetic beads and its preparation method and application

An immunomagnetic bead, cross-linking reaction technology, applied in instruments, measuring devices, scientific instruments, etc., can solve the problems of harsh reaction conditions, complex modification process, harsh operating conditions, etc., to improve stability and increase antibody loading. Effect

Inactive Publication Date: 2021-04-30
BEIJING UNIV OF AGRI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] The preparation method of traditional immunomagnetic beads is complicated and the operating conditions are harsh. It is necessary to modify special functional groups on the surface of paramagnetic substances in order to bind antibodies or antigens, and the modification process is complicated and the reaction conditions are harsh.

Method used

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  • A kind of immunomagnetic beads and its preparation method and application
  • A kind of immunomagnetic beads and its preparation method and application
  • A kind of immunomagnetic beads and its preparation method and application

Examples

Experimental program
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Effect test

Embodiment 1

[0049] an immunomagnetic bead

[0050] Step 1: Preparation of Fe 3 o4 nano magnetic core

[0051] Weigh 20.0g of ferrous sulfate heptahydrate and dissolve it in 140.0mL of deionized water, heat in a water bath to 90°C and react for 1h under the condition of mechanical stirring at a speed of 400r / min. At the same time, another 1.6 g of potassium nitrate and 11.2 g of potassium hydroxide were dissolved in 60.0 mL of deionized water, and the prepared solution was added dropwise (1 mL / min) into the ferrous ion solution with a constant flow pump. Continue to stir at 90°C for 2 hours after the addition to obtain a black turbid solution, which is washed several times with deionized water and dried. The black solid is ferric oxide, and its SEM image is as follows figure 1 and 2 , Fe 3 o 4 The particle size of the nano-magnetic nuclei is between 100 and 300nm, the spheres of the magnetic nuclei are unstable, and some of the magnetic nuclei are irregular. Due to the relatively act...

Embodiment 2

[0092] Its difference with embodiment 1 is that AMP&ZnCl 2 The reaction conditions for hydrogel-coated magnetic beads are different, as follows.

[0093] Weigh 0.3 g of adenosine monophosphate disodium salt and dissolve it in 20 mL of HEPES buffer (10 mM, pH 7.4), then add 200 mg of ferric oxide, and stir thoroughly. Add 150mL of paraffin wax mixture containing 6.5% Span 80 dropwise under stirring, form an emulsified system after stirring at 500r / min, continue stirring for 30min and heat to 40°C in a water bath, then add 3mL of glutaraldehyde solution (C 5 h 8 o 2 , volume fraction 25%), constant temperature cross-linking reaction 30min. Lower the temperature, add 2mL 1mg / mL ZEA monoclonal antibody dropwise (1mL / min) into the reaction system with a constant flow pump at room temperature, stir well, then add 10mL 50mM ZnCl dropwise (1mL / min) with a constant flow pump 2 Solution, cross-linking reaction at room temperature for 2h.

[0094] After the reaction, wash several ti...

Embodiment 3

[0097] Its difference with embodiment 1 is that AMP&ZnCl 2 The reaction conditions for hydrogel-coated magnetic beads are different, as follows.

[0098] Weigh 0.3g of adenosine monophosphate disodium salt and dissolve in 20mL of HEPES buffer (10mM, pH 7.4), then add 200mg of ferric oxide, and stir thoroughly. Add 150mL of paraffin wax mixture containing 6.5% Span 80 dropwise under stirring, form an emulsified system after stirring at 500r / min, continue stirring for 30min and heat to 45°C in a water bath, then add 3mL of glutaraldehyde solution (C 5 h 8 o 2 , volume fraction 25%), constant temperature cross-linking reaction 25min. Lower the temperature, add 2mL 1mg / mL ZEA monoclonal antibody dropwise (1mL / min) into the reaction system with a constant flow pump at room temperature, stir well, then add 10mL 50mM ZnCl dropwise (1mL / min) with a constant flow pump 2 Solution, cross-linking reaction at room temperature for 2h.

[0099] After the reaction, wash several times wit...

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Abstract

The invention provides an immunomagnetic bead and its preparation method and application. The preparation method of immunomagnetic beads: mix adenosine monophosphate or its metal salt, aqueous phase buffer, ferric oxide, paraffin and Span 80, and stir to form an emulsified system; the volume ratio of paraffin and Span 80 is 7: 93~100; the mass ratio of adenosine monophosphate or its metal salt to ferric oxide is 0.1~0.3:0.2; add glutaraldehyde to the emulsification system, and the first step of cross-linking reaction occurs at constant temperature; then add the antibody And zinc salts, a second step of cross-linking reaction occurs, and then the solids are separated with a magnet. This preparation method takes advantage of the properties of metal-organic gels to immobilize proteins, and immobilizes antibodies and ferric oxide in the metal-organic gel solid system at the same time. The reaction conditions used are mild, and no special operations or complicated pretreatment procedures are required. Therefore, , the preparation method expands favorable conditions for the wide application of immunomagnetic beads.

Description

technical field [0001] The invention relates to the technical field of biological detection, in particular to an immunomagnetic bead and its preparation method and application. Background technique [0002] Immunomagnetic beads are conjugates of superparamagnetic substances and antibodies, which can be used to bind corresponding antigens or antibodies. Because immunomagnetic beads can move directionally under the attraction of an external magnetic field, they can be used for the separation and detection of targets, and achieve the purpose of purifying genes, proteins, cells, microorganisms, etc. Due to the rapidity and specificity of the immune response, immunomagnetic beads have better sensitivity and specificity than conventional detection or separation methods. [0003] The preparation method of traditional immunomagnetic beads is complicated and the operating conditions are harsh. It is necessary to modify special functional groups on the surface of paramagnetic substan...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N33/531G01N33/543
CPCG01N33/531G01N33/54326
Inventor 丁轲孙毅蒙韩涛陈湘宁
Owner BEIJING UNIV OF AGRI
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