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Staphylococcus aureus unmarked electrochemical aptamer sensor based on reduced graphene oxide-nanogold composite material

An aptamer sensor and Staphylococcus technology, applied in the field of electrochemistry, can solve the problems of few detection methods, achieve the effects of improving sensitivity, expanding detection signal, and improving stability

Inactive Publication Date: 2013-04-17
JIANGNAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

In recent years, the rapid detection of Staphylococcus aureus has made continuous progress. Antigen-antibody-based immunological methods such as enzyme-linked immunosorbent technique (ELISA), PCR, and loop-mediated isothermal amplification technology can all achieve rapid detection of Staphylococcus aureus. , but the target substances detected by these methods are mostly enterotoxin or a certain gene fragment secreted by Staphylococcus aureus, and there are relatively few detection methods directly targeting the whole bacteria

Method used

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  • Staphylococcus aureus unmarked electrochemical aptamer sensor based on reduced graphene oxide-nanogold composite material
  • Staphylococcus aureus unmarked electrochemical aptamer sensor based on reduced graphene oxide-nanogold composite material
  • Staphylococcus aureus unmarked electrochemical aptamer sensor based on reduced graphene oxide-nanogold composite material

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Experimental program
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Effect test

Embodiment 1

[0021] Dilute the Staphylococcus aureus solution into 8 concentration gradients sequentially with normal saline, numbered from 10 -1 ~10 -8 . At a constant temperature of 37°C, dilute the prepared modified capture probe with the working electrode to 10 -8 Concentration of bacterial solution incubation. After 30 min, the electrochemical impedance method was used for measurement in a three-electrode system. The working conditions are as follows: the electrolyte is 5mmol / LK containing 0.1mol / LKCl 3 [Fe(CN) 6 ] / K 4 [Fe(CN) 6 ](1:1), PH=7.4, the applied voltage is 0.24V, and the frequency range is 10 5 ~10 -1 HZ, the amplitude is 5mV. The same conditions, followed by 10 -7 ~10 -1 Impedance values ​​were measured after incubating at a constant temperature of 37°C for 30 min in the bacterial solution of the concentration. Gently rinse the electrode surface with ultrapure water before and after each measurement, and carefully blot dry with filter paper.

[0022] According...

Embodiment 2

[0024] Prepare the working electrode modified with Staphylococcus aureus aptamer according to the method described in the invention, and measure its resistance R 0 ;

[0025] Take 3 centrifuge tubes, add 900 μL of commercially available pure milk to each, and add the concentration of 10 2 cfu / mL, 10 3 cfu / mL, 10 4 Each 100 μL of cfu / mL Staphylococcus aureus standard substance was prepared with a concentration of 10 1 cfu / mL, 10 2 cfu / mL, 10 3 cfu / mL milk standard sample of Staphylococcus aureus;

[0026] According to the invention, under the constant temperature condition of 37°C, the working electrode is sequentially mixed at a concentration of 10 1 cfu / mL, 10 2 cfu / mL, 10 3cfu / mL Staphylococcus aureus milk samples were incubated for 30min, carefully rinsed and placed in an electrochemical workstation for measurement. The measurement conditions were as follows: the electrolyte was 5mM K containing 0.1mol / L KCl 3 [Fe(CN) 6 ] / K 4 [Fe(CN) 6 ](1:1), PH=7.4, the applie...

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Abstract

The invention discloses a method for detecting staphylococcus aureus based on reduced graphene oxide-nanogold composite material in combination with aptamer, belonging to the field of electrochemistry. According to the basic principle of the method, the reduced graphene oxide-nanogold composite material is used to modify a sulfhydrylation whole staphylococcus aureus capture probe by a layer by layer self-assembly method. When the probe is incubated in staphylococcus aureus liquid, effective binding sites of the probe can be bound with the staphylococcus aureus to wrap the staphylococcus aureus in a three-dimensional space structure, thus, the whole capture to target bacteria is realizes. The electron transmission on the surface of an electrode is blocked after the target bacteria is bound, so that a electrochemical resistance value increases, and quantitative determination on the staphylococcus aureus can be realized based on the change of the resistance value. By virtue of excellent electron transmission capability of graphene and large specific surface area of nanogold, detection signals can be amplified, and detection time can be shortened. The new method provided by the invention can be applied to whole bacteria detection on the staphylococcus aureus, can expand application of impedance-based electrochemical unmarked detection, and has the advantages of strong specificity, high sensitivity, cheapness, rapidness, and so on.

Description

technical field [0001] The invention mainly utilizes an aptamer sensor to electrochemically detect Staphylococcus aureus, and belongs to the field of electrochemistry. The feature is that the introduction of the reduced graphene oxide-nano gold composite material expands the detection signal and improves the detection sensitivity. At the same time, there is no need to introduce labels in the ultra-sensitive electrochemical impedance detection, so that the detection time is shorter and the cost is less. At the same time, it also provides a new method for the detection of whole bacteria of Staphylococcus aureus. Background technique [0002] Staphylococcus aureus is a Gram-positive bacterium belonging to the genus Staphylococcus and one of the most important pathogenic bacteria in the genus Staphylococcus. Staphylococcus aureus is a strong pathogen that can cause purulent infection after invading the human body, and can also cause acute enteritis by secreting enterotoxin, ma...

Claims

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Application Information

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IPC IPC(8): G01N27/30G01N27/327
Inventor 王周平贾飞段诺夏雨马小媛
Owner JIANGNAN UNIV
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