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A method for capturing high-purity circulating tumor cells based on bioorthogonal chemistry

A tumor cell and bio-orthogonal technology, applied in the field of biomedical materials, can solve the problems that antibodies or aptamers cannot realize the recognition and labeling of CTCs, the capture and enrichment of CTCs, and the harsh storage and working conditions, etc., to achieve good biological Anti-fouling performance, good magnetic response behavior, and high recovery efficiency

Active Publication Date: 2022-03-01
SICHUAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

On the one hand, substances such as antibodies are expensive, fragile, and have harsh storage and working conditions; on the other hand, a single antibody or aptamer cannot recognize and mark all CTCs
For example, anti-epithelial cell adhesion molecule (anti-EpCAM) can recognize tumor cells with high expression of epithelial adhesion molecule, but for CTCs whose expression of EpCAM is down-regulated or derived from non-epithelial tumors, modifying anti-epithelial cell adhesion molecule (anti-EpCAM ) magnetic beads cannot effectively capture and enrich this type of CTCs

Method used

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  • A method for capturing high-purity circulating tumor cells based on bioorthogonal chemistry
  • A method for capturing high-purity circulating tumor cells based on bioorthogonal chemistry
  • A method for capturing high-purity circulating tumor cells based on bioorthogonal chemistry

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Embodiment 1

[0060] A preferred embodiment of the present invention provides a method for capturing high-purity circulating tumor cells based on a bioorthogonal chemical method. The specific steps are as follows:

[0061] (1) Superparamagnetic ferric oxide nanoparticles (Fe 3 O 4 NPs) preparation

[0062] FeCl 3 ·6H 2 O was dissolved in ethylene glycol, and sodium citrate (Na 3 CT), finally adding ammonium acetate (NH 4 Ac), after magnetic stirring for 1 h, the mixture was transferred to a stainless steel autoclave and kept at 200 °C for 16 h. After cooling, the sediment was collected by magnetic separation, and thoroughly washed with ethanol and deionized water four times, and finally magnetic separation was used to obtain Fe. 3 O 4 Nanoparticle superparamagnetic iron tetroxide nanoparticles. of which FeCl 3 ·6H 2 O, Na 3 CT, NH 4 The mass ratio of Ac is 2.89:1:8.26. Determination of particle size and surface potential of drug-loaded hybrid nanoparticles by Malvern laser par...

Embodiment 2

[0093] A preferred embodiment of the present invention provides a method for capturing high-purity circulating tumor cells based on a bioorthogonal chemical method. The specific steps are as follows:

[0094] (1) Superparamagnetic ferric oxide nanoparticles (Fe 3 O 4 NPs) preparation

[0095] FeCl 3 ·6H 2 O was dissolved in ethylene glycol, and sodium citrate (Na 3 CT), finally adding ammonium acetate (NH 4 Ac), after magnetic stirring for 1 h, the mixture was transferred to a stainless steel autoclave and kept at 220 °C for 18 h. After cooling, the sediment was collected by magnetic separation, and washed thoroughly with ethanol and deionized water three times. Finally, magnetic separation was used to obtain Fe. 3 O 4 Nanoparticle superparamagnetic iron tetroxide nanoparticles. of which FeCl 3 ·6H 2 O, Na 3 CT, NH 4 The mass ratio of Ac is 2.92:1:8.31.

[0096] (2) Preparation of biomimetic magnetic vesicles

[0097] Mouse macrophages (J774A.1) were cultured in...

experiment example

[0117] Example 1 (6) 3. Tumor-bearing mice were divided into two groups according to the tumor volume, and the number of CTCs corresponding to each mouse was counted, and the tumor volume was combined for graph analysis.

[0118] like Figure 13 As shown, CTCs in the blood of A549 tumor-bearing mice decreased with the increase of tumor volume, and CTCs in the blood of HepG2 tumor-bearing mice increased with the increase of tumor volume. Fe 3 O 4 MVs-DBCO efficiently captures CTCs through bioorthogonal chemistry, which is not affected by the heterogeneity of CTCs that may be brought about by tumor development.

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Abstract

The invention discloses a method for capturing high-purity circulating tumor cells based on a bio-orthogonal chemical method, comprising the following steps: S1. preparing superparamagnetic ferric iron tetroxide nanoparticles; S2. preparing bionic magnetic vesicles; S3. biomimetic magnetism Functional modification of vesicles; S4. Growth of azide gene on the surface of tumor cells; S5. Capture of circulating tumor cells. In the present invention, after incubating superparamagnetic iron ferric oxide nanoparticles with macrophages, the biomimetic magnetic vesicles can be produced by mouse macrophages through a pathway similar to exosome release, and then functionally modified; tumor cells are The functional groups are generated on the surface of the cell membrane through their own intracellular metabolism in advance, and incubated with the functionalized magnetic vesicles. The two functional groups are combined by bioorthogonal chemistry to capture circulating tumor cells. This method has the potential to be applied to Potential in liquid biopsy.

Description

technical field [0001] The invention belongs to the technical field of biomedical materials, and in particular relates to a method for capturing high-purity circulating tumor cells based on a bioorthogonal chemical method. Background technique [0002] Circulating Tumor Cells (CTCs) refer to all kinds of tumor cells existing in peripheral blood, and mainly refer to tumor cells that separate from the primary tumor or metastases of solid tumors and enter the peripheral blood circulation spontaneously or due to diagnosis and treatment. After escaping the immune killing of the body and surviving, CTCs can be transported to the distant tissues with blood dissemination and exudate. Under the action of various growth factors, they can adapt to the new microenvironment and form new metastases in other parts. Circulating tumor cells are very rare in peripheral blood, and there is a certain degree of heterogeneity in morphology and type. For example, tumor cells will undergo epithelia...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/09
CPCC12N5/0694C12N2509/00
Inventor 易强英康珂吴尧张宇佳朱南行李国浩
Owner SICHUAN UNIV
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