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Magnetic material for efficiently detecting circulating tumor cell and preparation method of magnetic material

A technology for tumor cells and magnetic materials, which is applied in the field of magnetic materials for efficient detection of circulating tumor cells and its preparation, can solve the problems of functionalization strategy limitations, expression differences, etc., and achieve narrow particle size distribution, high saturation magnetization, and magnetic properties. Good responsiveness and biocompatibility

Active Publication Date: 2018-03-30
SICHUAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, clinical studies have shown that the expression of EpCAM on the surface of different CTCs is significantly different, and even some tumor cells with invasive ability do not express it, which leads to obvious limitations of this type of functionalization strategy

Method used

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  • Magnetic material for efficiently detecting circulating tumor cell and preparation method of magnetic material
  • Magnetic material for efficiently detecting circulating tumor cell and preparation method of magnetic material
  • Magnetic material for efficiently detecting circulating tumor cell and preparation method of magnetic material

Examples

Experimental program
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Effect test

Embodiment 1

[0038] Example 1 Synthesis of rhodamine (RhB)-labeled HA-SH

[0039]Dissolve hyaluronic acid HA (Mw=100KD, 806mg, 2mmol) in 200mL PBS buffer (0.01mol / L, pH=7.4). After the HA is completely dissolved, add 1-(3-dimethylaminopropyl )-3-ethylcarbodiimide EDC (1.15g, 6mmol) and 1-hydroxybenzotriazole HOBt (0.81g, 6mmol), then placed on a magnetic stirrer for magnetic stirring for 2h, then weighed cystamine salt Acid acid (1.35 g, 6 mmol) was added to the reaction system, and the reaction was carried out overnight with magnetic stirring. The obtained reaction solution was dialyzed to remove unreacted reactants, and dialyzed against deionized water for 24 hours. Add dithiothreitol DTT (1.54 g, 10 mmol) into the dialyzed reaction solution to continue the reaction for 24 hours, then transfer the reaction solution to a dialysis bag, and dialyze with deionized water for 2 days to remove unreacted DTT to obtain HA -SH.

[0040] RhB-labeled HA-SH can be obtained by grafting RhB on HA-SH ...

Embodiment 2

[0042] Example 2 Preparation of Superparamagnetic Fluorescent HA-SH Capsules (magneticcapsules, MCs)

[0043] Take 10mg Fe 3 o 4 Nanoparticles (Fe 3 o 4 NPs) were dispersed in 1 mL of chloroform, 10 mg of RhB-HA-SH was dissolved in 25 mL of deionized water, and the two were mixed and placed in an ice bath. Use a cell disruptor to sonicate 3 times, each treatment for 5min (570W), add 1mL H for the third treatment 2 o 2 To promote the cross-linking of sulfhydryl groups, the resultant is subjected to magnetic separation (magnetic separation is to enrich the sample on a magnet), discard the supernatant after the magnetic separation is completed, and wash several times with PBS buffer solution, and finally adjust the concentration to 1 mg / mL to prepare superparamagnetic fluorescent HA-SH capsules.

[0044] Among them, Fe 3 o 4 Nanoparticles are prepared by pyrolysis method, the specific process is as follows: after removing water and oxygen from the system, iron acetylace...

Embodiment 3

[0050] Example 3 Preparation of targeted functional superparamagnetic fluorescent HA-SH capsules (TMCs)

[0051] The synthesis process of TMCs is as follows Figure 4 shown. Take about 5 mL of MCs suspension, add 2.5 mL of PBS solution containing 1MEDC and 1M HOBt, shake slowly at 4 °C for 4 h to activate MCs, then discard the supernatant by magnetic separation and redisperse the activated MCs in PBS. 10mg PEG-FA was added and reacted at room temperature for 4h, then transferred to 4°C, and about 50μg anti-EpCAM antibody was added to continue the reaction for 6h. The obtained product was collected by magnetic separation, washed several times with PBS buffer to remove unreacted reactants, and stored at 4°C for future use.

[0052] The above reaction products were verified, and the verification process was as follows: an appropriate amount of TMCs and an appropriate amount of Goat Anti-Rabbit IgG H&L (FITC) were incubated at 4°C for 30 min. After washing three times with PBS ...

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Abstract

The invention provides a magnetic material for efficiently detecting a circulating tumor cell and a preparation method of the magnetic material. The preparation method comprises the steps of: (1) allowing hyaluronic acid, 1-(3-dimethylamino propyl)-3-ethyl-carbodiimide hydrochloride, 1-hydroxybenzotriazole, cysteamine hydrochloride and dithiothreitol to react to prepare HA-SH, (2) grafting rhodamine onto HA-SH to prepare RhB-HA-SH, (3) allowing an Fe3O4 nanoparticle, RhB-HA-SH and H2O2 to react to prepare superparamagnetic fluorescent HA-SH, and (4) allowing superparamagnetic fluorescent HA-SH, 1-(3-dimethylamino propyl)-3-ethyl-carbodiimide hydrochloride, 1-hydroxybenzotriazole, PEG-FA and anti-EpCAM (epithelial cell adhesion molecule) to perform antibody reaction to prepare the magneticmaterial. The magnetic material is high in saturation magnetization, and good in magnetic response and biocompatibility, can achieve specific binding with the circulating tumor cell and has certain universality.

Description

technical field [0001] The invention belongs to the technical field of magnetic materials, and in particular relates to a magnetic material for efficiently detecting circulating tumor cells and a preparation method thereof. Background technique [0002] In recent years, malignant tumors have gradually become one of the major diseases that endanger human health. Its high morbidity and high fatality rate have attracted the attention of researchers in many fields, including doctors and scientific workers. Early detection of cancer-related cells / factors plays a vital role in the diagnosis, treatment and prognosis monitoring of cancer. Clinical practice has shown that malignant tumors in primary tissue generally do not directly lead to the death of patients, and tumor metastasis greatly reduces the survival rate of tumor patients. Therefore, early diagnosis and detection of tumor metastasis is of great importance to improve the survival rate of patients significance. [0003] ...

Claims

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Application Information

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IPC IPC(8): C09K11/06C09K11/02H01F1/00H01F1/42C12N5/09
Inventor 吴尧易强英马少华蓝芳周小熙
Owner SICHUAN UNIV
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