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Method for determining ATP content on basis of magnetic bead separation and DNA marker gold nanoparticle probe

A gold nanoparticle and DNA labeling technology, which is applied in the direction of measuring devices, material analysis and material analysis by electromagnetic means

Active Publication Date: 2016-04-20
凯惠睿智生物科技(上海)有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

In recent years, the research on new methods of biochemical analysis based on molecular aptamers has become one of the hotspots [ShukoorMI, AltmanMO, HanD, et al. Aptamer-nanoparticle assembly for logic-based detection [J]. ], however, the combination of nucleic acid aptamer technology and crystal violet (CV) aptamer electrochemical means to detect ATP has not been reported in the literature so far.

Method used

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  • Method for determining ATP content on basis of magnetic bead separation and DNA marker gold nanoparticle probe
  • Method for determining ATP content on basis of magnetic bead separation and DNA marker gold nanoparticle probe
  • Method for determining ATP content on basis of magnetic bead separation and DNA marker gold nanoparticle probe

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Embodiment 1

[0031] The impact of the amount of embodiment 1MB-DNA1 on the electrochemical signal

[0032] MB-DNA1 was used as a carrier for immobilizing probes. Firstly, the amount of probe solution was fixed, and the change of detection signal was investigated when the amount of MB-DNA1 solution was changed. The experimental results showed that the detection signal increased with the increase of the amount of MB-DNA1 solution; when the amount was greater than 75 μL, the signal change trend slowed down ( figure 2 (A)). Therefore, 75 μL was selected as the optimal dosage of MB-DNA1 solution in the experiment.

Embodiment 2

[0033] The impact of embodiment 2 probe dosage on electrochemical signal

[0034] When the amount of fixed MB-DNA1 solution was 75 μL, the effect of the amount of probe on the signal intensity was investigated. With the increase of the amount of probe, the detection signal gradually increased. When the amount of probe was 90 μL, the signal was the largest ( figure 2 (B)). Therefore, 90 μL was selected as the optimal amount of probe in the experiment.

Embodiment 3

[0035] Example 3 Effect of ATP and aptamer binding time on electrochemical signal

[0036] The experiment examined the effect of the binding time of ATP and aptamer on the signal intensity. The signal intensity increased rapidly with the increase of the binding time. The signal intensity reached the maximum at 40 minutes, and then stabilized. It can be seen that 40 minutes is enough for ATP and aptamer. body binding ( figure 2 (C)). Therefore, 40 min was selected as the optimal binding time in the experiment.

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Abstract

The invention belongs to the field of electrochemical sensors, and particularly relates to a method for determining the ATP content on the basis of magnetic bead separation and a DNA marker gold nanoparticle probe. The method comprises the steps that firstly, carboxylation magnetic beads (MB) are combined with amino-modified ATP aptamer DAN1 to generate an MB-DNA1 compound; CV aptamer DNA2 modified with SH and ATP aptamer complementary chain DNA3 are combined with nanogold, and CV is added to generate a probe CV / DNA2 / DNA3 / AuNPs; the MB-DNA1 compound is reacted with the probe, and magnetic beads modified with the probe are generated through the combination and complementation action of DNA1 and DNA3; a sample solution containing ATP is added into a magnetic bead solution modified with the probe, magnetic separation is conducted, and supernatant is taken; then the supernatant is dropwise added on an electrode modified with gold nanoparticles. The obtained electrode serves as a working electrode, and the working electrode, a reference electrode and an indicating electrode are inserted into an electrolyte solution to be subjected to electrochemical determination. Determination of the ATP content is achieved according to the intensity of an electrochemical signal.

Description

technical field [0001] The invention belongs to the field of electrochemical sensors, and in particular relates to a method for detecting ATP content based on magnetic bead separation and DNA-labeled gold nanoparticle probes. Background technique [0002] ATP (adenosine triphosphate) is an important substance for storing and supplying energy in human cells. ATP is an important energy source in the body, and is the structural basis and functional basis of organisms and life phenomena. It plays an irreplaceable role in material metabolism, body defense, blood coagulation, muscle contraction, cell information transmission, individual growth and development, and tissue repair. When the human body encounters strong stimuli, such as germ invasion, dying and other serious situations, ATP will be quickly converted into adenosine diphosphate, and at the same time, huge energy will be released, so that the various systems and organs of the body will quickly gain strong power. Theref...

Claims

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Application Information

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IPC IPC(8): G01N27/327G01N27/48
CPCG01N27/3277G01N27/3278G01N27/48
Inventor 混旭岳美娥宗迎夏
Owner 凯惠睿智生物科技(上海)有限公司
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