Composition for detecting breast cancer stem cells

A breast cancer stem cell and composition technology, which can be used in hybrid peptides, measuring devices, anti-receptor/cell surface antigen/cell surface determinant immunoglobulin, etc. The workload and difficulty of data analysis, the doping of wrong information, etc., achieve the effect of low technical level requirements, improving the accuracy of qualitative detection, and small amount of data analysis

Active Publication Date: 2021-06-04
章毅 +7
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At the same time, the amount of data generated by two-color flow cytometry analysis is large, and the probability of misinformation is correspondingly increased, which greatly increases the workload and difficulty of data analysis

Method used

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  • Composition for detecting breast cancer stem cells
  • Composition for detecting breast cancer stem cells
  • Composition for detecting breast cancer stem cells

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0057] Example 1 Electrochemical signal labeling of breast cancer stem cells based on CD44 antibody-nucleic acid conjugates

[0058] (a) Take 20 μL of DNA probes A and A at a concentration of 50 μM 1 Put it in a microtube, mix it evenly and put it under the condition of 25 ℃ for 2 hours to get AA 1 Double strand solution.

[0059] (b) Take 10 μL of the AA prepared in step (a) 1 The double-strand solution was placed in a microtube, and then 10 μL of 30 mM ascorbic acid solution, 10 μL of 2 mM copper sulfate solution, and 70 μL of 3-(N-morpholino)propanesulfonic acid (MOPS) buffer ( Containing 20mM MOPS, 300mM NaCl, pH 7.5), reacted at 25°C for 15 minutes, so that A 1 The polycytosine sequence at the 5' end of the chain (T 30 ) as a template to synthesize copper nanoparticles that can be used as electrochemical signal probes.

[0060] (c) Dissolve 5 μL of CD44 antibody with a concentration of 1 μM and 5 μL of Tetrazine-PEG5-NHS with a concentration of 5 μM in 100 μL of phos...

Embodiment 2

[0067] Electrochemical analysis of embodiment 2 breast cancer stem cells

[0068] (a) Take 20 μL of DNA probes A and A at a concentration of 50 μM 1 Put it in a microtube, mix it evenly and put it under the condition of 25 ℃ for 2 hours to get AA 1 Double-stranded solution; take 20 μL of DNA probes B and B at a concentration of 50 μM 1 Put it in a microtube, mix it evenly and put it at 25°C for 2 hours to get BB 1 Double-strand solution: Take 20 μL of DNA probes T and F with a concentration of 100 μM and place them in microtubes respectively to obtain T-strand and F-strand solutions.

[0069] (b) Take 10 μL of the AA prepared in step (a) 1 The double-strand solution was placed in a microtube, and then 10 μL of 30 mM ascorbic acid solution, 10 μL of 2 mM copper sulfate solution, and 70 μL of 3-(N-morpholino)propanesulfonic acid (MOPS) buffer ( Containing 20mM MOPS, 300mM NaCl, pH 7.5), reacted at 25°C for 15 minutes, so that A 1 The polycytosine sequence at the 5' end of t...

Embodiment 3

[0083] Example 3 Electrochemical Analysis of Breast Cancer Stem Cells Under the Coexisting Condition of Interfering Cells

[0084] In order to examine the specificity and anti-interference ability of this technology for the analysis of breast cancer stem cells, we divided 5×10 5 pcs, 5×10 4 and 5×10 3 breast cancer stem cells with 5×10 5 Three different molecular phenotypes (CD44 阳 CD24 阳 , CD44 阴 CD24 阳 , CD44 阴 CD24 阴 ) of interfering cells were mixed, and electrochemical analysis was carried out according to the steps in Example 2. The result is as Figure 5 As shown, the presence of three different molecular phenotypes of interfering cells will not cause changes in the peak current value of the electrochemical response of breast cancer stem cells, indicating that this technology uses antibody-nucleic acid conjugate-based selective electrochemical signal labeling and The background signal erasing strategy based on programmable DNA circuit can effectively eliminate...

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Abstract

The invention provides a composition for detecting breast cancer stem cells. The composition comprises a first antibody-nucleic acid conjugate specifically binding to cell surface CD44 protein, a second antibody-nucleic acid conjugate specifically binding to cell surface CD24 protein, a DNA single-chain T chain and a DNA single-chain F chain, wherein the T chain and a nucleic acid chain of the first antibody-nucleic acid conjugate are complementarily bound, the F chain and a nucleic acid chain of the second antibody-nucleic acid conjugate are complementarily bound, a programmed DNA loop is formed, signal markers on the surfaces of the breast cancer cells with CD44 positive and CD24 positive phenotypes are erased, and signal probes marked to the breast cancer stem cells are retained. The composition provided by the invention realizes identification and effective distinguishing of the breast cancer stem cells and the breast cancer cells, and has the characteristics of simple and convenient operation, accurate result, low technical level requirement, small data analysis amount and the like.

Description

technical field [0001] The invention relates to a composition for detecting cell typing, in particular to a composition constructed of several DNA sequences to realize the detection and separation of cells of different phenotypes. Background technique [0002] Breast cancer is one of the malignant diseases that seriously threaten women's health, and it is the second leading cause of female cancer death in the world after lung cancer. Breast cancer originates from the complex epithelial cells of the mammary gland, and cells from different sources are highly heterogeneous. In 2003, A 1 -Hajj et al first isolated CD44 from primary and metastatic breast cancer tissue according to the expression level of cell surface proteins 阳 CD24 阴 cell subgroups. This subpopulation differentiates slower than other breast cancer cells, but has stronger tumorigenic ability, and has similar self-renewal, proliferation and differentiation abilities as normal stem cells, so it is called breast...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K16/28G01N27/26C12Q1/02
CPCC07K16/2884C07K16/2896C07K19/00G01N33/5005G01N27/26Y02A50/30
Inventor 章毅伍婷赵婧陈亮曹亚
Owner 章毅
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