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On-step type label-free rapid fluorescence mercury ion detection method

A label-free, mercury ion technology, applied in the field of fluorescence detection, can solve the problems of limited wide application, large sample volume professional training, etc., and achieve the effect of sensitive method

Inactive Publication Date: 2017-09-01
GUANGZHOU MEDICAL UNIV +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, these traditional methods often require sophisticated instruments, complex sample pretreatment, large sample volume, and professional training for operations, which limit their wide application.

Method used

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  • On-step type label-free rapid fluorescence mercury ion detection method
  • On-step type label-free rapid fluorescence mercury ion detection method
  • On-step type label-free rapid fluorescence mercury ion detection method

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Experimental program
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Effect test

Embodiment 1

[0023] 1 Experimental method

[0024] 1.1 Reagents and materials ssDNA was designed by Mfold (http: / / unafold.rna.albany.edu / ?q=mfold / DNA-Folding-Form), synthesized by Sangon (Shanghai, China), and purified by high-performance liquid chromatography . The sequence of the mercury-specific ssDNA is: 5'-CTTCTTTCTTCCCCTTGTTTGTTG-3' (shown in SEQ ID NO.1). SG I (10,000×) was purchased from Invitrogen Biotechnology Co., Ltd (Shanghai, China) and stored at -20°C.

[0025] Hg 2+ , Pb 2+ , Ni 2+ , Zn 2+ , Ca 2+ , Cu 2+ , Fe 3+ , Cr 6+ and Cd 2+ The standard solutions were purchased from AccuStandard, Inc. (New Haven, USA), and the concentrations were 1mg mL -1 . The rest of the chemicals were analytically pure. The experimental water was ultrapure water. All experimental operations were performed at room temperature.

[0026] 1.2 Instrument fluorescence intensity (FIs) is measured by F-4500 fluorescence spectrophotometer (Hitachi, Japan), the measurement parameters are as ...

Embodiment 2

[0050] A one-step method for label-free fluorescence rapid detection of mercury ions, comprising the following steps:

[0051] (1) Add 200 μL of 100 nM mercury-specific ssDNA solution and 2 μL of 100× SG I fluorescent dye diluted with water to 6 centrifuge tubes, and then add 50 μL of 6 Hg selected from 5-1000 nM in a gradient. 2+ Aqueous solution, respectively 5, 20, 100, 400, 800 and 1000nM, mix well and incubate at room temperature for 0.5min;

[0052] (2) Using a fluorescence spectrophotometer, with 490nm as the excitation wavelength, the liquid obtained in step (1) is scanned at an emission wavelength of 535nm to obtain respective fluorescence intensity values, and prepare a fluorescence intensity response standard curve;

[0053] (3) Take 4 centrifuge tubes again, add 200 μL of 100 nM mercury-specific ssDNA solution and 2 μL of 100× SG I fluorescent dye diluted with water, then add 50 μL of the water sample to be tested filtered through a 0.45 μm microporous membrane, mi...

Embodiment 3

[0055] A one-step method for label-free fluorescence rapid detection of mercury ions, comprising the following steps:

[0056] (1) Add 200 μL of 100 nM Hg-specific ssDNA solution and 2 μL of 100× SGI fluorescent dye diluted with water to 8 centrifuge tubes, and then add 50 μL of 8 Hg selected from 5-1000 nM in a gradient. 2+ Aqueous solutions, respectively 5, 10, 20, 80, 160, 320, 640 and 1000nM, mix well and incubate at room temperature for 3min;

[0057] (2) Using a fluorescence spectrophotometer, with 490nm as the excitation wavelength, the liquid obtained in step (1) is scanned at an emission wavelength of 535nm to obtain respective fluorescence intensity values, and prepare a fluorescence intensity response standard curve;

[0058] (3) Take 5 centrifuge tubes again, add 200 μL of 100 nM mercury-specific ssDNA solution and 2 μL of 100× SG I fluorescent dye diluted with water, then add 50 μL of the water sample to be tested filtered through a 0.45 μm microporous membrane, m...

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Abstract

The invention discloses an on-step type label-free rapid fluorescence mercury ion detection method. On the basis that ssDNA (Single-Stranded Deoxyribonucleic Acid) can form a stable T-Hg<2+>-T compound with Hg<2+> and can be folded into a stable barrette structure double-chain DNA (dsDNA), a fluorescent dye SG I can be embedded into a small-groove structure area of a double-spiral structure of the dsDNA, relatively intense fluorescence can be stimulated, the fluorescence intensity is in positive correlation with concentration of Hg<2+>, and rapid detection on the Hg<2+> is achieved. The detection method disclosed by the invention has a lowest detection limit of 3nM which is less than the standard of the Environmental Protection Agency for the Hg<2+> in a water sample, is good in specificity, free of metal ion interference, good in repeatability, relatively low in detection cost, simple to operate and easy to popularize and apply to on-site Hg<2+> detection with a novel detection platform, can realize automation, and has great potential in emergency treatment on heavy metals, environment monitoring, risk evaluation and management and the like.

Description

technical field [0001] The invention relates to a one-step label-free fluorescence detection method for mercury ions, belonging to the technical field of fluorescence detection. Background technique [0002] Heavy metal pollution has become an important global problem. Human beings are seriously threatened by human health due to exposure to polluted natural water and drinking water, especially the highly toxic heavy metal mercury. Inorganic mercury, namely Hg and Hg 2+ Released into the environment by human or natural causes. Industrial sources of mercury include coal and gold mining, waste incineration, wood pulping, fossil fuel combustion, and chemical manufacturing, among others. Additional routes of human exposure include homes and workplaces, religious activities, and dental amalgam. In addition, Hg 2+ It is difficult to biodegrade and can accumulate in the body through biomagnification, which will cause different human diseases, such as renal failure, prenatal brai...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N21/64
CPCG01N21/6428
Inventor 刘楠李亚王玉张印红刘辉郝玉伟
Owner GUANGZHOU MEDICAL UNIV
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