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DNA enzyme, application of DNA enzyme, copper ion detection test paper based on DNA enzyme and preparation method of copper ion detection test paper

A DNA enzyme and detection test paper technology, which is applied in the field of copper ion detection, can solve the problems of reducing the activity of reducing agents, affecting the stability and accuracy of detection results, etc., and achieves the effects of low detection limit, large infiltration range and improved detection efficiency.

Active Publication Date: 2021-06-29
NINGBO UNIV
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  • Description
  • Claims
  • Application Information

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Problems solved by technology

At present, reducing agents are used to reduce Cu 2+ reduced to Cu + , but the activity of the reducing agent (usually sodium ascorbate) will also decrease with time, which will affect the stability and accuracy of the test results to a certain extent

Method used

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  • DNA enzyme, application of DNA enzyme, copper ion detection test paper based on DNA enzyme and preparation method of copper ion detection test paper
  • DNA enzyme, application of DNA enzyme, copper ion detection test paper based on DNA enzyme and preparation method of copper ion detection test paper
  • DNA enzyme, application of DNA enzyme, copper ion detection test paper based on DNA enzyme and preparation method of copper ion detection test paper

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preparation example Construction

[0029] The present invention also provides a preparation method of divalent copper ion detection test paper, comprising the steps of:

[0030] (1) Soak and modify the hydrophobic region of the glass fiber filter paper in a mixed solution of methyltrichlorosilane-toluene and hydrochloric acid;

[0031] (2) Infiltrate the hydrophilic region of the glass fiber filter paper with a dye solution comprising 3-butyn-1-ol, 3-azido-7-hydroxycoumarin and DNase to obtain a copper ion detection test paper.

[0032] When using ordinary fiber test paper for detection, there will be a large-scale infiltration phenomenon after dropping the dye solution, resulting in uneven color distribution, so that the color development is not obvious, which reduces the detection sensitivity and detection efficiency.

[0033] In order to solve this problem, the present invention uses glass fiber filter paper as detection test paper, and described glass fiber filter paper is divided into hydrophobic zone and ...

Embodiment 1

[0061] This embodiment provides a copper ion detection test paper, the preparation method of the copper ion detection test paper is as follows:

[0062] (1) Use a puncher to punch a circular hole with a diameter of 0.7 cm on the glass fiber filter paper, use the hole as a hydrophilic area, and use the rest of the glass fiber filter as a hydrophobic area. Soak the hydrophobic area in a mixed solution of 20mM methyltrichlorosilane-toluene and 37% hydrochloric acid (the volume ratio when the two are mixed is 1:3) for 15min, take it out and dry it at 37°C for 20min to obtain a superhydrophobic The hydrophobic area of ​​the glass fiber filter paper; the DNA single strand shown in SEQ ID NO: 1 is denatured in R buffer, boiled in a thermal cycler for 5 minutes, and cooled to room temperature (22°C) within 10 minutes for refolding , to obtain the DNA enzyme of the present invention. Store the obtained DNase at 4°C for later use; in this example, the R buffer refers to Li-HEPES buffer...

Embodiment 2

[0065] This embodiment provides a method for detecting copper ions using the copper ion detection test paper prepared in Example 1.

[0066] (1) Preparation of copper ion standard solution: Prepare copper sulfate solution with water to form copper sulfate standard solutions with concentrations of 0, 10 μM, 20 μM, 30 μM, 40 μM, 50 μM, 60 μM, 70 μM, 80 μM, 90 μM, and 100 μM.

[0067] (2) Put the copper ion detection test paper into the copper ion standard solution of the above concentration, and react at 37°C for 30 minutes; take out the copper ion detection test paper, put it in an oven and dry it at 55°C for 10 minutes, and then put it in a UV light box During irradiation, draw the curve graph of the concentration and fluorescence intensity of the copper ion standard solution, and compare it with the control test paper without adding DNase, such as figure 1 shown, from figure 1 It can be seen that with the increase of the copper ion concentration, the fluorescence intensity o...

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Abstract

The invention belongs to the technical field of bivalent copper ion detection, and provides a DNA enzyme, application thereof, copper ion detection test paper based on the DNA enzyme and a preparation method of the copper ion detection test paper. The invention provides the preparation method of copper ion test paper. The preparation method comprises the following steps: (1) soaking and modifying a hydrophobic region of glass fiber filter paper in a mixed solution of methyl trichlorosilane-toluene and hydrochloric acid; and (2) infiltrating the hydrophilic region of the glass fiber filter paper with the dye liquor to obtain the copper ion detection test paper. The DNA enzyme provided by the invention has high catalytic click reaction efficiency on the copper ions, has a better signal amplification effect, and can be used for detecting the copper ions in water or food; the copper ion detection test paper provided by the invention has the advantages of high sensitivity, low detection line and high specificity, the detection limit is as low as 1.31 [mu]M, and the detection error is less than 5%.

Description

technical field [0001] The invention relates to the technical field of copper ion detection, in particular to a DNA enzyme and its application, a DNA enzyme-based copper ion detection test paper and a preparation method thereof. Background technique [0002] Copper is a trace element needed to maintain life activities and growth and development in almost all life forms. Unusual levels of free Cu 2+ It may act as a catalyst for the generation of oxygen and protein free radicals, leading to serious neurological diseases such as Alzheimer's disease, Parkinson's disease and Wilson's disease. Therefore, it is necessary to develop a sensitive, rapid and suitable for on-site detection of Cu 2+ detection methods to achieve practical emergency detection. Inhibit Cu from source 2+ The infringement on the human body is of great significance. For example, to detect Cu in food 2+ The content of Cu in food can help people to judge the safety factor of food. 2+ Screening can greatly...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N21/78G01N21/64C12N9/22C12Q1/34
CPCG01N21/78G01N21/643C12N9/22C12Q1/34G01N2021/775G01N2021/7759G01N2333/922
Inventor 干宁吴佳怡李天华曹玉廷吴大珍毕文超
Owner NINGBO UNIV
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