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Bacterial magnetic particle (BMP)-agglutinin complex and preparation method and application thereof

A composite and nano-magnetic technology, applied in the field of separation and purification, can solve the problems of poor particle size uniformity of nanoparticles, complicated preparation of epoxidized magnetic nanoparticles, etc., achieve good uniformity and dispersion, shorten preparation time, and improve efficiency. Effect

Inactive Publication Date: 2015-05-20
JIANGNAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the preparation of artificially synthesized epoxidized magnetic nanoparticles is cumbersome and the particle size uniformity of nanoparticles is poor.

Method used

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  • Bacterial magnetic particle (BMP)-agglutinin complex and preparation method and application thereof
  • Bacterial magnetic particle (BMP)-agglutinin complex and preparation method and application thereof
  • Bacterial magnetic particle (BMP)-agglutinin complex and preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0042] Example 1 Preparation of BMPs-conA complex

[0043] BMPs surface modification: Weigh 1 mg BMPs and heat in 100 μL 1% SDS solution at 100°C for 5 minutes to remove BMPs surface proteins; collect BMPs on a magnetic stand and wash 3 times with 100 mM phosphate (PB, pH=7.5) buffer; Afterwards, BMPs were resuspended in 20 mg / mL PE solution, ultrasonicated at 100 W for 10 min; after incubation in the dark for 2 h, the BMPs were washed three times as above. That is, the BMPs-PE complex is obtained.

[0044] BMPs-BS 3 Preparation of the complex: draw 900 μL of 4mM PB buffer to suspend 1 mg of the complex obtained in the previous step, and disperse it with 80W ultrasonic; weigh BS 3 0.572mg, fully dissolved in 100μL distilled water, then added to BMPs suspension, 37°C, 200rpm, reacted for 0.5h; the BMPs complex after the reaction of the magnetic frame collector, removed the supernatant, suspended in 100mM PB buffer, dispersed by 80w ultrasonic wave, and then used Collect by m...

Embodiment 2

[0050] Example 2 Preparation of BMPs-wheat germ agglutinin (WGA) complex

[0051] Prepare BMPs-wheat germ agglutinin (WGA) complexes as follows:

[0052] (1) BMPs surface modification: Weigh 1mg of BMPs, add 50uL of 2% CTAB solution, heat treatment at 60°C for 20min to remove the protein on the surface of BMPs, then centrifuge to collect BMPs, wash and resuspend in 1mg / mL Phosphatidylserine PS solution, 80W ultrasound for 1h, incubate in the dark for 24h, and wash 3 times with 40mM phosphate (PB, pH=7) buffer solution to obtain BMPs-PS complex;

[0053] (2) Preparation of BMPs-glutaraldehyde complex: Take 1 mg of BMPs-PS complex, suspend it with 100 uL of 10 mM buffer solution, disperse it by ultrasonic, then add glutaraldehyde solution containing 0.2 mg glutaraldehyde, and in 4 Shake at -40°C for 10-60 minutes, collect, and obtain the BMPs-glutaraldehyde complex;

[0054] (3) Preparation of BMPs-WGA complex: prepare 0.1mg / ml wheat germ agglutinin solution, add 1mg BMPs-glut...

Embodiment 3

[0055] Example 3 BMPs-lentil agglutinin (LCA)

[0056] Prepare BMPs-LCA complexes as follows:

[0057] BMPs surface modification: Weigh 1mg of BMPs, add 100mL of 0.5% CHAPS solution, shake for 2 hours to remove the protein on the surface of BMPs, then collect BMPs by centrifugation, wash and resuspend in 100mg / mL PE solution, 100W ultrasonic 30min, incubate in the dark, and wash 3 times with 120mM phosphate (PB, pH=9) buffer solution to obtain the BMPs-PE complex;

[0058] Preparation of BMPs-dimethyladipimide complex: Take 1mg of BMPs-PE complex, suspend it with 10mL of 6mM pH7 PB buffer, disperse it by ultrasonic, then add 10mg of dimethyladipimide The dimethyl adipimide solution of the amine was shaken and reacted at 4-40°C, and collected by centrifugation to obtain the BMPs-dimethyl adipimide complex;

[0059] Preparation of BMPs-LCA complex: Prepare 10 mg / ml lentil agglutinin solution, add 1 mg BMPs-dimethyladipimide complex to 10 mL wheat germ adipimide solution, suspe...

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Abstract

The invention discloses a bacterial magnetic particle (BMP)-agglutinin complex and a preparation method and application thereof, belonging to the technical field of separation and purification. The preparation method comprises the following steps: performing surface modification and transformation on BMPs, and adding a cross-linking agent and agglutinin for coupling, thereby obtaining the BMP-agglutinin complex. The surfaces of the BMPs are modified, the influence of self-proteins of the BMPs on subsequent application is eliminated, more primary amino is provided for agglutinin coupling due to the addition of PE, the coupling efficiency is improved, and the dispersity of the BMPs is improved; the cross-linking agent (BS3) serves as a connecting arm, so that covalent coupling is realized between the BMPs and the agglutinin, and an agglutinin recognition sugar chain area is not damaged; and moreover, the concentration of a buffer solution is regulated, the preparation time is shortened, and the coupling efficiency is improved. The complex disclosed by the invention is used for enriching and separating specific glycoproteins, and compared with traditional affinity chromatography, the method disclosed by the invention has the advantages of fewer steps, short time and the like.

Description

technical field [0001] The invention relates to a bacterial nano magnetosome-lectin and a preparation method and application thereof, belonging to the technical field of separation and purification. Background technique [0002] Sugars can be covalently combined with proteins or lipids to form carbohydrate complexes such as glycoproteins, proteoglycans, and glycolipids. They are important information molecules in the human body and play a role in specific recognition and mediation in many life and disease processes. effect. A small change in its structure or quantity may cause a change in its function, leading to the occurrence of various diseases, which has aroused people's great enthusiasm for glycomics research. With the advancement of science and technology and the development of biology, more and more technologies have been applied to the research of glycomics, such as lectin affinity chromatography technology, frontier affinity chromatography technology, high performa...

Claims

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Application Information

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IPC IPC(8): B01J20/281B01J20/30B01D15/38C07K14/47C07K14/77C07K1/22
CPCB01D15/3804B01J20/281B01J2220/52B01J2220/80C07K1/22C07K14/77
Inventor 关锋李想杨刚龙郭佳刘昌梅
Owner JIANGNAN UNIV
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