Electrochemical detection method for stem cells

An electrochemical and breast cancer stem cell technology, applied in the field of electrochemical sensing based on the directional recognition and detection of breast cancer stem cells based on polypeptide fibers, can solve the problems of low blood clearance efficiency, strong immunogenicity, poor tissue penetration, etc., and achieve high sensitivity High, specificity, broad application prospects

Active Publication Date: 2018-05-29
章毅 +8
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, antibodies have certain limitations in bioanalytical applications, such as: poor tissue penetration, strong immunogenicity, and low blood clearance efficiency

Method used

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  • Electrochemical detection method for stem cells
  • Electrochemical detection method for stem cells
  • Electrochemical detection method for stem cells

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0023] Example 1 Polypeptide chain P1 and its self-assembly

[0024] A polypeptide chain P1 modified with an azide group at the amino terminal was designed and synthesized, the sequence of which was: (NH2)Azide-KLVFFGGRLVSYNGIIFFLK(COOH). The sequence consists of three functional fragments, namely self-assembly functional fragment (KLVFF), spacer functional fragment (GG) and recognition functional fragment (RLVSYNGIIFFLK); the chemical synthesis of polypeptide chain P1 is completed by a professional peptide synthesis company.

[0025] Polypeptide chain P1 forms a polypeptide fiber by means of self-assembly functional fragments, and the azide group and recognition functional fragments are respectively located at both ends of the polypeptide fiber. The specific steps are as follows: take the stock solution of the polypeptide chain P1 in a microtube, dilute it with a certain solution, and react to make the P1 self-assemble to form a fibrous structure. The concentration of the po...

Embodiment 2

[0026] Example 2 Preparation of aptamer DNA functionalized electrode

[0027] In order to realize the capture of breast cancer stem cells, the nucleic acid aptamer DNA chain sequence should contain GGTGGTGGTGGTTGTGGTGGTGGTGG. Secondly, in order to realize the effective immobilization of the aptamer DNA chain on the electrode interface and the maintenance of capture activity, there should be multiple consecutive adenine bases at the 3' end of the sequence and a sulfhydryl group on the 6 carbon atoms (denoted as: C 6 -SH) modification. The chemical synthesis of nucleic acid aptamer DNA chain is completed by a professional DNA synthesis company.

[0028] Pretreatment of the gold electrode: First, the gold electrode is polished to a mirror surface with alumina powders with a particle size of 1 μm, 0.3 μm and 0.05 μm on the silk; then the gold electrode is placed in ethanol and ultrapure water for 3 minutes to remove residual Impurities; After rinsing with double distilled water,...

Embodiment 3

[0030] Example 3 Electrochemical sensing and detection of breast cancer stem cells

[0031] 90 μL of the sample solution containing the breast cancer stem cells to be tested interacted with the nucleic acid aptamer DNA functionalized electrode obtained in Example 2, and through the specific molecular recognition between the target cells and the nucleic acid aptamer DNA, the target breast cancer stem cells were placed on the electrode. Surface capture; the reaction time is 1 to 2 hours, and the reaction temperature is 30°C to 40°C.

[0032] The obtained electrodes capturing target breast cancer stem cells are reacted with 50 μL to 150 μL of polypeptide fibers at 30° C. to 40° C. for 1 hour to 2 hours, so as to realize the directional recognition of breast cancer stem cells by the polypeptide fibers.

[0033]Preparation of diphenylcyclooctyne (DBCO) functionalized silver nanoparticles: 100 mL of a mixed solution containing 0.4 mM silver nitrate and 0.4 mM trisodium citrate was p...

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Abstract

The invention discloses an electrochemical detection method for stem cells. The electrochemical detection method is characterized in that breast cancer stem cells are identified by polypeptide; the Nterminal of the polypeptide is modified with an azide group; an electrochemical signal is obtained by a copper-catalyzed azide-alkynyl Husigen cycloaddition reaction. According to the method providedby the invention, identification of the breast cancer stem cells is formed by designed and synthesized polypeptide and peptide fibers formed by self-assembly of the polypeptide, further silver nanoparticles with electrochemical response are introduced by means of click chemical reaction, and quantitative detection of target breast cancer stem cells is realized. The method has the advantages of high sensitivity and high specificity; no antibody needs to participate in, so the electrochemical detection method has a broad application prospect in the field of clinical diagnosis and the like.

Description

technical field [0001] The invention relates to an electrochemical sensing method for stem cells, in particular to an electrochemical sensing method for detecting breast cancer stem cells based on directional recognition of polypeptide fibers. Background technique [0002] As one of the most common malignant diseases in women, breast cancer has become a killer that threatens women's health. However, traditional clinical treatment methods cannot fundamentally eliminate the existence of cancer cells. A considerable part of cancer cells can escape the effects of chemotherapy and radiotherapy, resulting in disease recurrence and distant metastasis in patients. The theory of cancer stem cells (Cancer Stem Cells, CSCs) provides a new idea for the study of recurrence and metastasis of breast cancer. The theory points out that there is a small group of special cells in tumor tissue that are extremely rare but have the potential of self-renewal, proliferation and differentiation sim...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N27/327
CPCG01N27/26G01N27/30G01N27/3278
Inventor 章毅朱小立马骏伍婷王艺赵婧陈亮陆薇翟晓文张娟陈桂芳曹亚孙鹏刘立根盛慧明周渊峰
Owner 章毅
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