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Protein A coated magnetic nanocluster, preparation method and application thereof in capturing antibodies

A magnetic nanometer and nanocluster technology, which is applied in nanomagnetic materials and biological fields, can solve the problems of low magnetic responsiveness and difficulty in rapid adsorption, and achieve the effects of high biological activity, high saturation magnetization and small size

Active Publication Date: 2020-05-26
甘肃省科学院传感技术研究所
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the magnetic nanoparticles prepared by this method are only 5nm, the magnetic response is low, and it is difficult to achieve rapid adsorption under the control of an external magnetic field, which is limited to a certain extent.

Method used

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  • Protein A coated magnetic nanocluster, preparation method and application thereof in capturing antibodies
  • Protein A coated magnetic nanocluster, preparation method and application thereof in capturing antibodies
  • Protein A coated magnetic nanocluster, preparation method and application thereof in capturing antibodies

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0043] Example 1-Preparation of protein A-coated magnetic nanoclusters:

[0044] ① Weigh 0.5 mmol ferric chloride hydrate (FeCl 3 ·6H 2 O), 2.5 mmol ammonium acetate (NH 4 OAc) was dissolved in 5 mL of ethylene glycol, 1.39 mmol of polyacrylic acid (PAA) and 0.3 mmol of sodium citrate were added after ultrasonication for 30 min, and the ultrasonication was continued until the mixture turned bright red.

[0045] ② Pour the solution obtained in ① into a 10 mL microwave reaction tube, set the heating temperature to 240 °C, and the reaction time to 45 min for microwave reaction. After the reaction was complete, a black liquid was obtained. Alternate centrifugation washing with ethanol and deionized water three times, and low-pressure freeze-drying to obtain carboxylated ferric oxide nanoclusters (Fe 3 o 4 NCs), which were dispersed in water to form stock solution A (10mg / mL).

[0046] for the obtained Fe 3 o 4 The morphology, structure and functional group characterization...

Embodiment 2-5

[0055] Example 2-5, preparation of protein A-coated magnetic nanoclusters: the steps are the same as in Example 1, and the specific amounts of raw materials and reaction conditions are shown in Table 1.

[0056]

[0057] The average particle size of the clusters obtained under the conditions of Example 3 is about 80 nm, and the average particle size of the clusters obtained under the conditions of other embodiments is between 30-80 nm.

Embodiment 8

[0058] Example 8 - Detection of protein A-coated magnetic nanocluster capture antibody:

[0059] ① Mix two parts of 80μg fluorescein-labeled human immunoglobulin G (FAM-IgG) with 2mL PBS-dispersed magnetic nanoparticles (10mg / mL) and protein A-coated magnetic nanoparticles (10mg / mL). Incubate for 2 days, perform fluorescence test after magnetic separation;

[0060] ② The ability of protein A-coated magnetic nanoparticles to capture antibody human immunoglobulin G was detected by fluorescence spectroscopy, compared to the magnetic nanoparticles dispersed in 2mL PBS linked to fluorescein-labeled human immunoglobulin G (Fe 3 o 4 NCs-FAM-IgG) and protein A-coated magnetic nanoparticles dispersed in 2 mL PBS linked to fluorescein-labeled human immunoglobulin G (Fe 3 o 4 NCs-protein A-FAM-IgG) fluorescence intensity (fluorescence intensity comparison as shown in Figure 4 shown);

[0061] ③ The ability of protein A-coated magnetic nanoparticles to capture antibodies was analyze...

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Abstract

The invention discloses a protein A coated magnetic nanocluster, a preparation method and application thereof in capturing antibodies, which belong to the technical field of nano magnetic materials and biology. The preparation method comprises the following steps of: mixing ferric trichloride hydrate, ammonium acetate, ethylene glycol, polyacrylic acid and sodium citrate, carrying out ultrasonic treatment, preparing carboxyl-rich small-size ferroferric oxide nanoclusters by using a microwave-assisted polyol method, and then loading protein A on the surfaces of the magnetic clusters through activation to prepare the protein A coated magnetic nanoclusters. According to the method, a microwave-assisted heating method, a protein A directional connection antibody and fluorescence detection arefully combined, the high-performance protein A coated magnetic nanocluster is prepared, and a simple and effective method for detecting and capturing the antibody is designed. The protein A is used asan adapter protein to directionally connect an antibody and a magnetic nanocluster, and the specific binding technology has important value in the field of biomedicine.

Description

technical field [0001] The invention belongs to the field of nano-magnetic materials and biotechnology, and in particular relates to the preparation of protein A-coated magnetic nano-clusters and its application in capturing antibodies. Background technique [0002] Designing and constructing a successful targeted biohybrid magnetic nanoprobe is a major goal in the development of biomedicine, which lies at the interface between nanotechnology and molecular biology. The ultimate challenge is to develop reliable strategies for conjugating targeted biomolecules, especially monoclonal IgG, to magnetic nanoparticles. However, there is a common problem when binding, that is, the magnetic material will bind to the non-specific site of IgG, which will affect the targeting efficiency of the antibody. [0003] Protein A is a cell wall-associated protein on the surface of Staphylococcus aureus. It can reversibly bind to a variety of IgGs by binding to the Fc fragment of IgG (Fragment ...

Claims

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Application Information

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IPC IPC(8): B01J20/24B01J20/28B01J20/32G01N21/64
CPCB01J20/06B01J20/24B01J20/28009B01J20/3268G01N21/6428G01N21/6458G01N2021/6417G01N2021/6439
Inventor 刘一丹左显维韩根亮冯治棋何欣
Owner 甘肃省科学院传感技术研究所
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