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Magnetic nanoparticle immobilized with anthrax lethal factor on surface as well as preparation method and application of magnetic nanoparticle

A technology of anthrax lethal factor and magnetic nanoparticles, applied in the field of functionalized magnetic nanoparticles, can solve the problems that hinder the practical application of HNP1-3 biomarkers, achieve good clinical reference prospects, and simplify the effect of enrichment, separation and detection process

Inactive Publication Date: 2014-08-13
THE NAT CENT FOR NANOSCI & TECH NCNST OF CHINA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Nevertheless, existing tedious separation techniques such as HPLC or ELISA detection methods for all biomarkers that are expensive and not widely available have greatly hindered the practical application of HNP1-3 as biomarkers

Method used

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  • Magnetic nanoparticle immobilized with anthrax lethal factor on surface as well as preparation method and application of magnetic nanoparticle
  • Magnetic nanoparticle immobilized with anthrax lethal factor on surface as well as preparation method and application of magnetic nanoparticle
  • Magnetic nanoparticle immobilized with anthrax lethal factor on surface as well as preparation method and application of magnetic nanoparticle

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0064] Example 1 Preparation of Surface Immobilized Anthrax Lethal Factor Magnetic Nanoparticles

[0065] (1) Fe 3 o 4 Preparation of Superparamagnetic Nanoparticles

[0066] Weigh 54.0 mg FeCl 3 ·6H 2 O and 212.72 mg FeCl 2 4H 2 O mixed and dissolved in 52 ml of water, stirred at 700 rpm until completely dissolved, and 5 ml of NH 3 ·H 2 The mixed solution of O and 43 milliliters of water (completely added within 3S), reacted under nitrogen protection for 30 minutes and then stopped the reaction. After the product was magnetically sucked and washed with water to clarify, 100 milliliters of water was added therein for dispersion and set aside.

[0067] The resulting Fe 3 o 4 Transmission electron microscope images of magnetic nanoparticles such as figure 2 As shown in a, its size is around 10 nm.

[0068] (2) Silica coated Fe 3 o 4 Superparamagnetic nanoparticles (hereinafter referred to as Fe 3 o 4 SiO 2 )

[0069] Get 50 milliliters of step (1) gained Fe ...

Embodiment 2

[0087] Example 2 Reversible combination of AMNP and HNP1

[0088] Using MALDI-TOF to analyze the reversible combination of AMNP and HNP1, the results are as follows Figure 7 Shown:

[0089] After the AMNP (activated) obtained in Example 1 is combined with HNP1, the Zn in the surface anthrax lethal factor is removed through EDTA chelation 2+ causing its inactivation, the inactivated AMNP will release all HNP1( Figure 7 , channel 1), resulting in no detectable HNP1 on AMNP particles ( Figure 7 , channel 2). Addition of Zn to inactivated AMNP 2+ , the inactivated AMNP will be reactivated and can be used for the selective adsorption of HNP1 for the second time, and the bound HNP1 can be detected by MALDI-TOF analysis ( Figure 7 , channel 3); these newly bound HNP1 will again lose Zn due to AMNP 2+ While (EDTA chelation removes Zn in anthrax lethal factor 2+ ) was released, and the resulting inactivated AMNP had no HNP1 detectable by MALDI-TOF ( Figure 7 , channel 4)...

Embodiment 3A

[0090] Example 3 Enrichment of AMNP to HNP1

[0091] Figure 8 In order to use 1 microliter of the AMNP solution obtained in Example 1, enrich the MALDI-TOF spectrum of AMNP-HNP1 obtained from 650 microliters of human serum solution (the concentration of HNP1 is 0.8 micrograms / ml), Figure 8 Shown: AMNP can be enriched to HNP1 from solution.

[0092] Figure 9 For another example of AMNP enriching HNP1 from solution, Figure 9 showed that: AMNP can be enriched to HNP1 from solution to form AMNP-HNP1 complex, but Fe 3 o 4 COOH cannot enrich HNP1 in solution.

[0093] Figure 10 is the MALDI-TOF detection map of AMNP enrichment of reduced HNP1 from a solution of GSH (ie glutathione) and TCEP HCl (ie tris(2-carbonylethyl) phosphate hydrochloride). Among them, the first line above is the MALDI-TOF detection map of reduced HNP1 in TCEP HCl solution; the lower two lines show the results of AMNP enriching reduced HNP1 from GSH and TCEP HCl solutions respectively. The results...

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Abstract

The invention discloses a magnetic nanoparticle immobilized with an anthrax lethal factor on a surface as well as a preparation method of the magnetic nanoparticle and application of the magnetic nanoparticle in enrichment of human neutrophil peptides. The magnetic nanoparticle immobilized with the anthrax lethal factor on the surface is obtained by connecting the anthrax lethal factor to the surface of the superparamagnetic nanoparticle with amino and / or carboxyl on surface by virtue of amino-carboxyl coupling reaction; the magnetic nanoparticle can quickly and specially enrich the human neutrophil peptides for further detection, so that application of the human neutrophil peptides as tumor prognosis biomarkers is convenient, and therefore, clinical application prospect is very good.

Description

technical field [0001] The invention relates to the technical field of functionalized magnetic nanoparticles, in particular to a magnetic nanoparticle whose surface is immobilized with anthrax lethal factor and its preparation method and application. Background technique [0002] The occurrence and development of tumors are directly related to the tumor microenvironment, so the research on the tumor microenvironment has attracted widespread attention of scientists. The expression of proteins and polypeptides in the tumor microenvironment is usually tumor-associated, and many proteins and polypeptides secreted by tumor cells or normal cells have been studied as potential tumor microenvironment biomarkers. [0003] Among them, the biomarkers produced by tumor cells are usually related to the mutation during tumor progression, while the biomarkers derived from inflammatory cells in the tumor microenvironment are usually from the source that does not change with the development ...

Claims

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Application Information

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IPC IPC(8): C07K17/14H01F1/00G01N1/34
Inventor 梁兴杰赵元元邹国漳李志鹏陈飞张旭李盛亮王义峰王东亮姜永刚甘雅玲
Owner THE NAT CENT FOR NANOSCI & TECH NCNST OF CHINA
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