Electrochemical sensor for detecting telomerase activity and method for manufacturing electrochemical sensor

An electrochemical and telomerase technology, applied in the field of electrochemical sensors and their preparation, can solve the problems of complex process, high telomerase concentration and low sensitivity, and achieve the effects of simple preparation method, mild reaction conditions and high sensitivity

Active Publication Date: 2015-09-09
QINGDAO UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, the most commonly used method for detecting telomerase activity is the TRAP method based on PCR amplification technology. The invention patent "a method for detecting telomerase activity" (patent No. 02116466.5) is based on PCR technology and reverse transcription of human telomerase The detection of a certain structural domain of the enzyme is used to determine the activity of telomerase, the detection signal is stable, and a large number of samples can be reproducibly operated at the same time, but the process is complicated and the sensitivity is low; the invention patent "a rapid detection of telomerase activity Kit and its application” (Patent No. 201210190716.3) us

Method used

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  • Electrochemical sensor for detecting telomerase activity and method for manufacturing electrochemical sensor
  • Electrochemical sensor for detecting telomerase activity and method for manufacturing electrochemical sensor

Examples

Experimental program
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Embodiment 1

[0043] (1) Take 20 mg of molybdenum disulfide powder and dissolve it in 1 mL of organic solvent N-methyl-2-pyrrolidone. After ultrasonic stripping for 2-3 hours, let the dispersion stand for 15-20 minutes. Peel off the complete large particles to obtain a single-layer or several-layer molybdenum disulfide nanosheet dispersion, take 1mL dispersion and 1ml, 1mmol·mL -1 The Thi solution was mixed and sonicated for 3 h at room temperature, and the supernatant was discarded to obtain MoS 2 - Thi nanocomposite, poured into 1 mL of water; mix 99 mL of deionized water with 1 mL of 1% (W / V) HAuCl 4 Put the solution into a three-necked flask, seal it, heat it under reflux under vigorous stirring, and boil it at about 105°C, then quickly add 2.5mL of 1% (W / V) trisodium citrate solution, and the color of the solution changes from light yellow to dark in about 1min. Red, continue heating under reflux, stop heating after 20min, stir and cool at room temperature (23-25°C) to obtain Au parti...

Embodiment 2

[0050] Embodiment 2: the mensuration of actual sample

[0051] (1) The preparation of the electrochemical sensor and the drawing of the working curve are the same as in Example 1.

[0052] (2) Take a certain amount of Hela cell (human cervical cancer cell) samples, centrifuge (10000rpm, 5min), wash twice with 5mL PBS solution, add 200μL cell lysate (10mmol·L -1 , Tris–HCl at pH=7.5, 1mmol·L -1 MgCl 2 Solution, 1mmol·L -1 EGTA, 0.1mmol·L -1 Benzamidine, 5mmol·L-1 β-mercaptoethanol, 0.5% CHAPS, 10% glycerol), ice bath for 1h, centrifuge at 4°C (20000rpm, 15min), take the supernatant, and store it at -80°C; take 4μL of the supernatant and 6μL of DNA chain extension material buffer Liquid (20mmol·L -1 , Tris-HCl at pH=8.3, 1.5mmol·L -1 MgCl 2 Solution, 1mmol·L -1 EGTA, 63mmol·L -1 KCl solution, 0.05% Tween 20, 0.2mmol L -1 dATP, 0.2mmol·L -1 dGTP, 0.2mmol L -1 dTTP) mixed, and sucked and dropped onto the surface of the gold electrode with a pipette gun, and reacted at ...

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Abstract

The invention discloses an electrochemical sensor for detecting telomerase activity and a method for manufacturing the electrochemical sensor. The method for manufacturing the electrochemical sensor mainly includes procedures of sequentially adsorbing DNA (deoxyribonucleic acid) strands S1 of thionine, Au@SiO<2> and G-rich structures in the surfaces of molybdenum disulfide nano-sheets and adding Hemin into the molybdenum disulfide nano-sheets to form MoS<2>-Thi-Au@Si<O>2/DNAzyme nano-composite materials; modifying TS (telomerase substrate) precursors on the surfaces of gold electrodes, extending the DNA strands by the aid of telomerase to form DAN strands S2, screwing the DNA strands S2 and the DNA strands S1 complementarily, and ultimately detecting the intensity of electrochemical signals of catalytic hydrogen peroxide oxidation ABTS to directly detect the telomerase activity. The electrochemical sensor and the method have the advantages that the electrochemical sensor manufactured by the aid of the method is simple in operation and high in sensitivity and stability and has an important significance on detecting cancerous cells.

Description

technical field [0001] The invention relates to an electrochemical sensor for detecting telomerase activity and a preparation method thereof, in particular to a G-quadruplex / DNAzyme-based electrochemical sensor for detecting telomerase activity and a preparation method thereof, belonging to The field of biological detection technology. Background technique [0002] At present, the incidence of cancer is increasing year by year due to various factors such as the aging of the population, the destruction of the ecological environment, and unhealthy lifestyles. However, the five-year survival rate of cancer patients in my country is only about 30%. The most fundamental reason is When the patient is diagnosed with cancer, he is already in the middle and late stage, thus delaying the best treatment time. Therefore, rapid and effective early and sensitive diagnosis is of great significance for timely detection of cell canceration. [0003] The role of telomerase in the process of ...

Claims

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Application Information

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IPC IPC(8): G01N27/30
Inventor 王宗花于建波毕赛杨敏张菲菲夏建飞桂日军夏延致
Owner QINGDAO UNIV
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