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An electrochemical sensor based on a dynamic sandwich structure and its preparation and application

A sandwich and electrochemical technology, applied in the field of bioelectrochemistry, can solve the problems of insufficient sensitivity and accuracy, achieve the effect of improving detection sensitivity, achieving gain effect, and facilitating the analysis of precious samples

Active Publication Date: 2018-04-10
NANJING UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Despite these advantages, the use of electrochemical techniques to study protein interactions is still exploratory and often suffers from insufficient sensitivity and precision

Method used

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  • An electrochemical sensor based on a dynamic sandwich structure and its preparation and application
  • An electrochemical sensor based on a dynamic sandwich structure and its preparation and application
  • An electrochemical sensor based on a dynamic sandwich structure and its preparation and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0037] Example 1: Dynamic sandwich structure electrochemical sensor to detect goat anti-digoxigenin immunoglobulin reaction process

[0038] Step one, fix the gold electrode with a diameter of 3mm upside down, mix 98% concentrated sulfuric acid and hydrogen peroxide in a ratio of 3:1 to prepare a piranha solution, and add 10μL piranha solution to completely cover the gold surface for 10 minutes Then rinse with a large amount of ultrapure water; the gold electrode was gently polished with 1.0μm alumina powder on silk cloth and 0.3μm alumina powder on the suede for 5 minutes each to make the electrode surface smooth to the mirror surface; rinse and polish with a lot of ultrapure water The gold electrode surface is immersed in ethanol and ultrapure water successively, and ultrasonically cleaned for 5 minutes; the surface of the electrode is blown dry with nitrogen, and 10 μL piranha solution is added dropwise to the electrode surface for 10 minutes, and then rinsed with ultrapure wat...

Embodiment 2

[0041] Example 2: Dynamic sandwich structure electrochemical sensor detection antibody-antibody interaction reaction process

[0042] image 3 It is a schematic diagram of dynamic sandwich structure electrochemical sensor for antibody-antibody interaction analysis. On the basis of Example 1, if rabbit anti-goat immunoglobulin (Ab2) that can interact with goat anti-digoxigen immunoglobulin (Ab1) is further introduced into the detection system, the concentration is set at 0.01 nM, 0.5 nM , 1.5nM, 5nM, 8nM, 20nM, 200nM, 2000nM, due to the formation of "DNAzyme-goat antibody-rabbit anti-goat antibody" complex structure volume further increases, making DNAzyme cutting the signal chain more slowly ( Figure 4 ), within the same time of action, the electrochemical signal is further improved. By comparing the electrochemical signal of a single goat antibody, you can analyze the presence of rabbit anti-goat antibody in the solution ( Figure 5 ).

Embodiment 3

[0043] Example 3: Electrochemical sensor combining dynamic sandwich structure and gold nanoparticles sensitively detects goat anti-digoxigenin immunoglobulin reaction process

[0044] As the volume of the adsorbent on DNAzyme is larger, the ability to cut the signal chain on the gold electrode is lower. If DNAzyme and gold nanoparticles are connected to form a super-large "DNAzyme-target-gold nanoparticle" composite structure, it will Greatly improve the analysis capability of the sensor.

[0045] Step 1: Use 100mMK of gold nanoparticle solution (3nM) covered with 20nm citric acid 2 CO 3 Adjust the solution to pH 9.0; add 10 μL of 1 mg / mL rabbit anti-goat antibody to 990 μL of the above gold nanoparticle solution, and incubate at room temperature for 2 hours; centrifuge at 13000 rpm for 15 minutes, discard the supernatant, and resuspend in 990 μL PBS (pH 8.0) , Repeat once to remove the antibody that is not bound to the gold nanoparticles in the solution; add 10 μL of 10% BSA solut...

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Abstract

The invention belongs to the technical field of bioelectrochemistry, and concretely relates to a dynamic sandwich structure-based electrochemical sensor, and a making method and an application thereof. The electrochemical sensor is composed of an electrode, a signal chain and a reaction system, the electrode adopts a gold electrode, the signal chain is a segment of single-stranded DNA, the 5' end of the DNA chain is modified with a mercapto group, the 3' end of the DNA chain is modified with methylene blue, the mercapto group and the surface of the gold electrode are self-assembled to form a gold-mercapto bond standing on the surface of the gold electrode, the reaction system contains DNAzyme and a necessary reaction buffer system, the DNAzyme can be complemented and paired to the signal chain and cuts the signal chain into two segments of DNA short strands, and the DNAzyme is marked with a target protein identification element, and can be used for quantitatively detecting proteins and interaction among the proteins. The electrochemical sensor is simple to operate, can be operated by operators without special training, and is suitable for being used in remote regions and resource-poor regions.

Description

1. Technical Field [0001] The invention belongs to the technical field of bioelectrochemistry, and specifically relates to an electrochemical sensor based on a dynamic sandwich structure and its preparation and application. 2. Background technology [0002] There are about 650,000 protein interaction pairs in the human proteome. The interaction of these protein molecules with each other controls most of the activities of the cell. Deciphering the interaction network of these protein molecules can not only provide a deeper understanding of how these proteins are. Perform functions and ultimately affect cell activity, and help people design more specific drug molecules to treat specific diseases. Therefore, in the field of biological basic research, there is an inherent urgent requirement for the development of sensitive and operator-friendly analytical methods to detect the interaction events between these proteins. [0003] At present, a variety of methods for studying the interac...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/527
CPCG01N27/26G01N33/68
Inventor 李根喜李超马洁桦胡晓璐
Owner NANJING UNIV
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