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Kit for detecting aflatoxin B1 and method for detecting aflatoxin B1

An aflatoxin and kit technology, applied in biochemical equipment and methods, measuring devices, fluorescence/phosphorescence, etc., can solve the problems of cumbersome, high instrument cost, enzymatic denaturation and degradation, etc.

Active Publication Date: 2021-05-18
QINGDAO AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Traditional ELISA requires time-consuming cultivation and specific elution conditions, and the enzymes involved in the experiment are prone to denaturation and degradation problems
In addition, the instrumental analysis method involves complex instruments, high cost, and requires cumbersome sample pretreatment, which is time-consuming and difficult to use on a large scale and meet the requirements of on-site testing.

Method used

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  • Kit for detecting aflatoxin B1 and method for detecting aflatoxin B1
  • Kit for detecting aflatoxin B1 and method for detecting aflatoxin B1
  • Kit for detecting aflatoxin B1 and method for detecting aflatoxin B1

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Embodiment 1

[0042] Detection principle of the present invention is:

[0043] S1, S2, S3, and S4 form DNA tetrahedral nanostructures (DTNs) through base complementary self-assembly, and there is a single-stranded extended sequence at the four vertices of the DTNs structure; aflatoxin B 1 The WA double-stranded structure composed of the nucleic acid aptamer A and its partially complementary nucleic acid sequence W; the single-stranded sequence of S1 and the 5' end of the W chain are both modified by sulfhydryl groups; The double-strand modification forms a DNA walker structure on the surface of gold nanoparticles (AuNPs).

[0044] When aflatoxin B 1 In the presence, aflatoxin B of the DNA walker structure 1 aptamer A and aflatoxin B 1 combined with the AuNPs of the DNA walker structure; the W chain is in a single-stranded state, and starts to walk on the surface of the AuNPs with base complementarity, and the E1 on the W chain binds to the complementary sequence (E2) on the S1 to form a ...

Embodiment 2

[0060] Use the kit of embodiment 1 to aflatoxin B 1 Method for detection:

[0061] (1) Filter the sample solution to be tested with a 0.22 μm filter membrane and dilute it 10 times; take 10 μL of the sample solution to be tested and add it to the PBS solution with DNAwalker structure; mix and incubate at 37°C for 0.5 h; then add the nicking endonuclease Nt.BsmAI , then mixed and reacted at 37°C for 0.5h, and then heated at 65°C for 20min to inactivate the enzyme.

[0062] (2) NaCl solution is added to the solution after the reaction in step (1), and the colloidal gold is precipitated in the salt particles to remove AuNPs; DTNs are contained in the supernatant.

[0063] (3) Add 5 μL of 10 μM H1 and 5 μL of 10 μM H2 to the supernatant obtained in step (2); mix and react at 37° C. for 0.5 h.

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Abstract

The invention discloses a kit for detecting aflatoxin B1 and a method for detecting aflatoxin B1, and belongs to the technical field of harmful substance detection. The kit for detecting the aflatoxin B1 comprises a DNA walker structure, a nicking incision enzyme, a hairpin structure H1 and a hairpin structure H2, according to the kit disclosed by the invention, a signal amplification mode of a hyperbranched fluorescent nano tree structure is constructed based on a DNA walker structure, so that a detection signal of aflatoxin B1 is enhanced, and the reaction speed is increased; and the high-sensitivity rapid detection of the aflatoxin B1 is realized.

Description

technical field [0001] The invention belongs to the technical field of harmful substance detection, in particular to a method for detecting aflatoxin B 1 kit and detection of aflatoxin B 1 Methods. Background technique [0002] Aflatoxins are secondary metabolites produced during the growth of Aspergillus flavus and Aspergillus parasiticus. They are highly toxic substances and are widely distributed in moldy grains and products, especially peanuts, peanut oil and their products. Of the more than 20 identified aflatoxins, aflatoxin B 1 (AFB 1 ) are considered to be the most toxic. In view of the serious impact of this mycotoxin on human health, governments and relevant organizations of various countries have formulated strict regulations and testing methods to limit the level of aflatoxin in food. [0003] The most widely used mycotoxin analysis techniques include high-performance liquid chromatography (HPLC), gas chromatography / mass spectrometry (GC / MS), and probe-based...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N21/64
CPCG01N21/6428G01N21/6486G01N2021/6432C12N15/115C12N2310/16C12N2310/113C12N2310/3517C12N2310/3519C12N2320/10C12Q1/682C12Q1/6823C12Q2525/205Y02A50/30C12Q2525/131C12Q2525/301C12Q2563/107C12Q2563/137C12Q2563/155C12Q2565/133C12Q1/6876
Inventor 杨庆利王琦吴薇赵方圆侯秀丹赵海燕朱英莲李兆杰
Owner QINGDAO AGRI UNIV
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