Aptamer probe for detecting two kinds of tumor makers and electrochemical biosensor and preparation method and application thereof

A technology of tumor markers and biosensors, applied in the direction of material electrochemical variables, scientific instruments, instruments, etc., can solve problems affecting the analytical performance of sensors

Active Publication Date: 2018-09-11
GUANGZHOU YUEWANG AGRI CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Whether the electrochemical aptamer sensor based on this principle is a signal-on type or a signal-off type, there is always a certain background current problem, which will affect the analytical performance of the sensor to a certain extent.

Method used

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  • Aptamer probe for detecting two kinds of tumor makers and electrochemical biosensor and preparation method and application thereof
  • Aptamer probe for detecting two kinds of tumor makers and electrochemical biosensor and preparation method and application thereof
  • Aptamer probe for detecting two kinds of tumor makers and electrochemical biosensor and preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0055] Example 1 Optimization of assembly time of the aptamer probe of the present invention

[0056] In this embodiment, the method for optimizing the assembly time of the aptamer probe includes the following steps:

[0057] (1) Polish the gold electrode on the suede in 0.3 μm and 0.05 μm alumina slurry until it becomes a mirror surface, rinse with distilled water, and then ultrasonicate in secondary water, ethanol, and secondary water for 5 minutes, and then Soak the electrode in Piranha solution for 15min, rinse with secondary water and place the gold electrode in 0.1M H 2 SO 4 The cyclic voltammetry scan was performed between -0.3V and +1.5V, and then the cleaned gold electrode was rinsed with secondary water.

[0058] (2) 20 μL of 3.8 μM aptamer probe was dropped onto the surface of the pretreated gold electrode, and self-assembled at room temperature for different times (10 min, 20 min, 40 min, 80 min, 120 min, 160 min, overnight).

[0059] (3) After the gold electrod...

Embodiment 2

[0062] Embodiment 2 Hpa II endonuclease digestion time optimization

[0063] In the present embodiment, the Hpa II endonuclease digestion time optimization method comprises the following steps:

[0064] (1) Polish the gold electrode on the suede in 0.3 μm and 0.05 μm alumina slurry until it becomes a mirror surface, rinse with distilled water, and then ultrasonicate in secondary water, ethanol, and secondary water for 5 minutes, and then Soak the electrode in Piranha solution for 15min, rinse with secondary water and place the gold electrode in 0.1M H 2 SO 4 The cyclic voltammetry scan was performed between -0.3V and +1.5V, and then the cleaned gold electrode was rinsed with secondary water.

[0065] (2) 20 μL of 3.8 μM aptamer probe was dropped onto the surface of the pretreated gold electrode, and self-assembled at room temperature for 2 h.

[0066] (3) After the gold electrode was washed with secondary water, 20 μL of 1 mM mercaptohexanol was dropped onto the surface of ...

Embodiment 3

[0070] Embodiment 3Hpa II endonuclease concentration optimization

[0071] In the present embodiment, the Hpa II endonuclease concentration optimization method comprises the following steps:

[0072] (1) Polish the gold electrode on the suede in 0.3 μm and 0.05 μm alumina slurry until it becomes a mirror surface, rinse with distilled water, and then ultrasonicate in secondary water, ethanol, and secondary water for 5 minutes, and then Soak the gold electrode in Piranha solution for 15min, rinse with secondary water and put the gold electrode in 0.1M H 2 SO 4 The cyclic voltammetry scan was performed between -0.3V and +1.5V, and then the cleaned gold electrode was rinsed with secondary water.

[0073] (2) 20 μL of 3.8 μM aptamer probe was dropped onto the surface of the pretreated gold electrode, and self-assembled at room temperature for 2 h.

[0074] (3) After the gold electrode was washed with secondary water, 20 μL of 1 mM mercaptohexanol was dropped onto the surface of ...

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Abstract

The invention discloses an aptamer probe for detecting two kinds of tumor makers and an electrochemical biosensor and a preparation method and application thereof. The aptamer probe comprises an aptamer sequence, the aptamer sequence and the tumor marker I are subjected to specific recognition and closely bound, the 5' end and the 3' end of the aptamer probe are subjected to complementary hybridization to make the aptamer probe form a hairpin structure, and the stem portion of the hairpin structure comprises a recognition sequence of the tumor maker II and a recognition site of restriction enzyme. The electrochemical biosensor is prepared by fixing the aptamer probe on the surface of a working electrode. By means of the aptamer probe for detecting the two kinds of tumor makers and the electrochemical biosensor and the preparation method and application thereof, parallel analysis of multiple kinds of tumor marker molecules of thrombin and M.Sss I DNA methylation transferase can be achieved; the biosensor is high in detection sensibility, the lower detection limit for the thrombin is 0.1 ng / mL, and the lower detection limit for the M.Sss I DNA methylation transferase is 0.04 U / mL.

Description

technical field [0001] The invention relates to the field of analytical chemistry, in particular to an aptamer probe for detecting two tumor markers, an electrochemical biosensor and its preparation method and application, which can be used for the analysis and detection of major disease markers such as tumors. Background technique [0002] Clinically, rapid and sensitive screening of malignant tumors often uses tumor markers as detection indicators. Tumor markers are a class of substances produced by tumor tissues and cells that are related to tumor formation and occurrence, mainly including tumor antigens, hormones, enzymes and their isozymes. For example, thrombin is a serine proteolytic enzyme formed from the thrombin precursor. It can catalyze fibrinogen into fibrin, promote blood coagulation and regulate coagulation. Prognosis and other aspects are of great significance. In addition, DNA methyltransferase can catalyze DNA methylation, which plays an important role in...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N27/327G01N27/48
CPCG01N27/3277G01N27/48
Inventor 张松柏李玉红成诗琦杨基峰胡霞沈广宇卢基林
Owner GUANGZHOU YUEWANG AGRI CO LTD
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