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Fluorescent quantum dot nanoprobe, preparation method thereof and application of fluorescent quantum dot nanoprobe in detection of circulating tumor cells

A technology of fluorescent quantum dots and tumor cells, applied in the field of biological detection, can solve the problem that heavy metal elements are not suitable for biological detection, etc., and achieve the effects of good identification ability and stability, strong anti-interference ability, high sensitivity and anti-interference ability

Pending Publication Date: 2022-01-11
FUJIAN INST OF RES ON THE STRUCTURE OF MATTER CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0005] However, the quantum dots in the second near-infrared region that are mainly studied at present are II-VI and IV-VI semiconductors, such as CdSe, CdTe and PbSe, etc. Although they have good optical properties, the heavy metal elements (Cd 2+ and Pb 2+ etc.) are not suitable for applications such as biological detection

Method used

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  • Fluorescent quantum dot nanoprobe, preparation method thereof and application of fluorescent quantum dot nanoprobe in detection of circulating tumor cells
  • Fluorescent quantum dot nanoprobe, preparation method thereof and application of fluorescent quantum dot nanoprobe in detection of circulating tumor cells
  • Fluorescent quantum dot nanoprobe, preparation method thereof and application of fluorescent quantum dot nanoprobe in detection of circulating tumor cells

Examples

Experimental program
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Effect test

preparation example 1

[0082] Based on CuInSe 2 The preparation method of the circulating tumor cell fluorescent detection probe of quantum dots is as follows:

[0083] 1) Weigh 10mg of oil-soluble CuInSe 2 Quantum dots, and dissolved in 10mL chloroform, then added 40mg of DSPE-PEG-COOH, stirred for 10min, the mixed liquid was evaporated to dryness with a rotary evaporator, then added 10mL ultrapure water, ultrasonically dissolved for 15min, to obtain water-soluble CuInSe 2 Quantum dots, stored at room temperature for later use.

[0084] 2) Weigh 20 mg of the water-soluble CuInSe prepared in step 1) 2 Quantum dots, 15mg NHS and 10mg EDC, added 5mL MES buffer (50mmol / L, pH 5.6), stirred at room temperature for 20min, centrifuged at 21000rpm for 10min, removed the supernatant, added 5mL PBS buffer (50mmol / L , pH=7.4), and then added 10 μL of anti-EpCAM antibody, placed on a rotary mixer, and reacted at 4° C. for 2 hours. Centrifuge at 21,000 rpm for 10 min, remove the upper layer, and disperse in ...

Embodiment 1

[0088] The feasibility of measuring the number of circulating tumor cells by the detection probe obtained in Preparation Example 1 is investigated, including the following steps:

[0089] 1) Coating: Dilute the capture probe anti-EpCAM antibody to 10 μg / mL with 0.05 mol / L carbonate buffer, add 100 μL of the dilution to each well of a 96-well plate, and incubate at 37°C for 1 hour , the liquid in the well was discarded, and then washed 3 times with PBS washing buffer.

[0090] 2) Blocking: add 300 μL of BSA solution (2% w / v) prepared with PBS buffer to each well, incubate at 37° C. for 1 hour, discard the liquid in the well, and wash 3 times with PBS washing buffer.

[0091] 3) Adding samples: culture breast cancer (MCF-7) cells with DMEM cell culture medium (containing 10wt% fetal bovine serum, 100 μg / mL penicillin and 100 μg / mL streptomycin), and after they grow to a certain number, use pancreatic Protease digestion was performed and the cells were harvested and made into a ...

Embodiment 2

[0096] Examination of selectivity of detection probes for circulating tumor cell types.

[0097] 1. the reagent used in this experiment and instrument are identical with embodiment 1, used CuInSe 2 The quantum dot circulating tumor cell fluorescence detection probe is consistent with that of Example 1.

[0098] 2. Examine CuInSe 2 The selectivity of quantum dot detection probes to circulating tumor cell types comprises the following steps:

[0099] 1) Coating: Dilute the capture probe anti-EpCAM antibody to 10 μg / mL with 0.05 mol / L carbonate buffer, add 100 μL of the dilution to each well of a 96-well plate, and incubate at 37°C for 1 hour , the liquid in the well was discarded, and then washed 3 times with PBS washing buffer.

[0100] 2) Blocking: add 300 μL of BSA solution (2% w / v) prepared with PBS buffer to each well, incubate at 37° C. for 1 hour, discard the liquid in the well, and wash 3 times with PBS washing buffer.

[0101] 3) Sample addition: Breast cancer (MCF-...

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Abstract

The invention belongs to the technical field of biological detection, and discloses a fluorescent quantum dot nanoprobe, a preparation method thereof and application of the fluorescent quantum dot nanoprobe in detection of circulating tumor cells. The detection probe comprises a water-soluble quantum dot modified by an antibody; the antibody can specifically bind to a circulating tumor cell surface marker, such as an anti-EpCAM antibody, an anti-EGFR antibody, an anti-cytokeratin CK-8, CK-18 and CK-19 antibody, an anti-tumor cell glycoprotein (TAG) antibody or an anti-lactoglobulin antibody; more preferably, an anti-EpCAM antibody; the water-soluble quantum dots are selected from near-infrared two-region fluorescent water-soluble non-toxic quantum dots. An antibody-modified water-soluble near-infrared two-region fluorescent non-toxic quantum dot material is used as a fluorescent probe, and the number of circulating tumor cells is detected by virtue of the change of the fluorescence intensity of the probe through an antigen-antibody specific recognition effect.

Description

technical field [0001] The invention belongs to the technical field of biological detection, and in particular relates to a fluorescent quantum dot nanoprobe, a preparation method thereof and an application for detecting circulating tumor cells. Background technique [0002] Circulating tumor cells refer to all kinds of tumor cells that enter the peripheral blood after the primary tumor or metastatic tumor breaks away from the base. Circulating tumor cells have been widely concerned by scientists since they were first discovered in the peripheral blood of cancer patients by Australian scientist Thomas Ashworth in 1869. More and more studies have shown that circulating tumor cells are closely related to cancer metastasis and can be used as markers of tumor metastasis and recurrence. Therefore, monitoring the number and change trend of circulating tumor cells is of great significance for monitoring the change trend of tumors, evaluating the effect of treatment, early diagnosi...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N21/64G01N33/569G01N33/574C09K11/02C09K11/88B82Y20/00B82Y30/00B82Y40/00
CPCG01N21/6428G01N21/6486G01N33/56966G01N33/57415G01N33/574C09K11/02C09K11/881B82Y20/00B82Y30/00B82Y40/00
Inventor 涂大涛廉纬陈学元周山勇刘䶮韩思远余少桦
Owner FUJIAN INST OF RES ON THE STRUCTURE OF MATTER CHINESE ACAD OF SCI
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