Method for capturing high-purity circulating tumor cells based on biological orthogonal chemical method

A tumor cell, bioorthogonal technology, applied in the field of biomedical materials, can solve the problems that antibodies or aptamers cannot achieve CTCs identification, cannot achieve CTCs capture and enrichment, storage and working conditions are harsh, and achieve good magnetic properties. Responsive behavior, protection from degradation, effect of good surface structural integrity

Active Publication Date: 2020-12-01
SICHUAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

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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  • Method for capturing high-purity circulating tumor cells based on biological orthogonal chemical method
  • Method for capturing high-purity circulating tumor cells based on biological orthogonal chemical method
  • Method for capturing high-purity circulating tumor cells based on biological orthogonal chemical method

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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, and the specific steps are as follows:

[0061] (1) Superparamagnetic Fe3O4 nanoparticles (Fe 3 o 4 NPs) preparation

[0062] FeCl 3 ·6H 2 O was dissolved in ethylene glycol, and sodium citrate (Na 3 CT), and finally ammonium acetate (NH 4 Ac), after magnetically 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, washed thoroughly with ethanol and deionized water four times, and finally magnetically separated to obtain Fe 3 o 4 Nanoparticles Superparamagnetic Fe3O4 nanoparticles. where 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 particle size analyzer, F...

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, and the specific steps are as follows:

[0094] (1) Superparamagnetic Fe3O4 nanoparticles (Fe 3 o 4 NPs) preparation

[0095] FeCl 3 ·6H 2 O was dissolved in ethylene glycol, and sodium citrate (Na 3 CT), and finally ammonium acetate (NH 4 Ac), after magnetically 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, washed thoroughly with ethanol and deionized water three times, and finally magnetically separated to obtain Fe 3 o 4 Nanoparticles Superparamagnetic Fe3O4 nanoparticles. where 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 DMEM complete medi...

experiment example

[0117] Example 1 (6) 3. The 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 combined with the tumor volume for mapping analysis.

[0118] Such as Figure 13 As shown, the CTCs in the blood of A549 tumor-bearing mice decreased with the increase of tumor volume, and the 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 biological orthogonal chemical method. The method comprises the following steps: s1, preparing superparamagnetic ferroferric oxide nanoparticles; s2, preparing a bionic magnetic vesicle; s3, performing functional modification on the bionic magnetic vesicles; s4, growing azide genes on the surfaces of thetumor cells; and s5, capturing the circulating tumor cells. After superparamagnetic ferroferric oxide nanoparticles and macrophages are incubated, bionic magnetic vesicles can be generated through mouse macrophages in a way similar to exosome release, and then functional modification is carried out; tumor cells generate functional groups on the surfaces of cell membranes in advance through intracellular metabolism of the tumor cells, the tumor cells are incubated with the functionalized magnetic vesicles, the two functional groups are combined through bioorthogonal chemistry, capture of circulating tumor cells is achieved, and the mode has the potential of being applied to 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 Applications(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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