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Fluorescent biosensor for detecting kanamycin and preparation method and application thereof

A biosensor, kanamycin technology, applied in the field of biosensors, can solve the problems of cumbersome sample pretreatment or concentration process, cumbersome operation, difficult to popularize, etc., and achieve the effects of fast detection speed, good repeatability and low detection limit

Active Publication Date: 2019-03-29
UNIV OF JINAN
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Benefits of technology

This patented technology allows for quicker and more precise identification of drugs like catheterazole (CA) through their chemical structure called an azabicycloalkane ring system that binds with DNA molecules containing certain amino acids. These structures have been identified previously but they cannot be detected accurately because these techniques require expensive equipment or complicated procedures. By combining this technique with other sensors such as quantum dots, magnetic particles, atomic force microscopy, lasers, light scattering, Raman spectroscopies, and optically detectable tags, it becomes possible to create highly selective assays capable of quickly identifying drug candidates without complex operations.

Problems solved by technology

This patents describes an assays called High Performance Liquidchromatographic(HPLC), CapillarElectric Phase Time Resolved Fluid Specimensitive Microscope (CEPH). These techniques involve analyzing small amounts of drug solution containing different types of compounds like nitrates or quaternaries by measuring changes caused when certain chemical reactions occur between these substances. By comparing measurements from multiple samples they provide important clues about how much each type affects biological activity within cells.

Method used

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  • Fluorescent biosensor for detecting kanamycin and preparation method and application thereof
  • Fluorescent biosensor for detecting kanamycin and preparation method and application thereof
  • Fluorescent biosensor for detecting kanamycin and preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0060] A method for preparing a fluorescent biosensor for detecting kanamycin, comprising the following steps:

[0061] (1) Preparation of gold nanoparticles;

[0062] (2) Preparation of modified gold nanoparticles solution: modify Aptamer, Walker and Track on the surface of gold nanoparticles;

[0063] (3) Hybrid chain reaction: Mix the modified gold nanometer solution with the homogeneous reaction solution;

[0064] (4) Fluorescence detection.

[0065] In the step (1), the preferred preparation steps of gold nanoparticles are as follows:

[0066] (1) Install the required instruments and add 200mL ultrapure water into the three-necked flask (be careful not to let dust fall into the three-necked flask).

[0067] (2) Take 500uL (0.04g / mL) HAuCl4 in a single-package centrifuge tube, use a pipette gun to take 500uL and 200mL ultrapure water, stir and heat at a stirring speed of about 450 rpm, until boiled.

[0068] (3) Under the condition of stirring, take 3mL of 1% trisodium...

Embodiment 2

[0084] A method for preparing a fluorescent biosensor for detecting kanamycin, comprising the following steps:

[0085] (1) Preparation of gold nanoparticles;

[0086] (2) Preparation of modified gold nanoparticles solution: modify Aptamer, Walker and Track on the surface of gold nanoparticles;

[0087] (3) Hybrid chain reaction: Mix the modified gold nanometer solution with the homogeneous reaction solution;

[0088] (4) Fluorescence detection.

[0089] Steps (1), (2), and (4) are the same as in Example 1.

[0090] The main steps of the reaction process of step (3) are as follows: sterilized water, target substance (4 μL), modified nano-gold solution (6 μL), HAP1 (2 μL) (final concentrations were 0.1 nM, 0.2 nM, 0.3nM, 0.4nM, 0.5nM, 0.6nM), 10× buffer (buffer) (4 μL), restriction endonuclease IV (3 μL), signal probe (6 μL), added to In pre-prepared sterile EP tubes. Shake for 30 s and incubate for 2 h in a 37°C incubator.

[0091] The excitation wavelength of the fluore...

Embodiment 3

[0094] A method for preparing a fluorescent biosensor for detecting kanamycin, comprising the following steps:

[0095] (1) Preparation of gold nanoparticles;

[0096] (2) Preparation of modified gold nanoparticles solution: modify Aptamer, Walker and Track on the surface of gold nanoparticles;

[0097] (3) Hybrid chain reaction: Mix the modified gold nanometer solution with the homogeneous reaction solution;

[0098] (4) Fluorescence detection.

[0099] Steps (1), (2), and (4) are the same as in Example 1.

[0100] The main steps of the reaction process in step (3) are as follows: sterilized water, target substance (4 μL), modified nano-gold solution (6 μL), HAP1 (2 μL), HAP2 (2 μL), 10× buffer (3 μL), restriction endonuclease IV (3 μL), signal probe (6 μL) (final concentrations were 0.2 μM, 0.4 μM, 0.6 μM, 0.8 μM, 1.0 μM, 1.2μM, 1.4μM), added to the pre-prepared sterilized EP tube. Shake for 30 s and incubate for 2 h in a 37°C incubator.

[0101] The excitation wavelen...

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Abstract

The invention relates to the technical field of biosensor, in particular to a fluorescence biosensor based on amplification of a hybridization chain reaction. The problems that the specificity and sensitivity of the method for detecting the kanamycin is low, and the cost is high in the above prior art are solved. According to the fluorescence biosensor based on the amplification of the hybridization chain reaction, the cyclic amplification effect is achieved through the hybridization chain reaction between the endonuclease IV and the chain. The mixture is reacted homogeneously by fluorescenceresonance energy transfer of the fluorophores and the quenching group. The preparation method comprises the following steps of: preparing gold nanoparticles; modifying Walker and Track to the surfaceof gold nanoparticles; mixing the labeled gold nano solution with the homogeneous reaction solution; performing hybridization chain reaction, and detecting the fluorescence; using the specific recognition of the nucleic acid Aptamer, using the nucleic acid Aptamer for high specific detection of the target kanamycin; and using the amplification of the hybridization chain reaction to achieve signalamplification.

Description

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Claims

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Application Information

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Owner UNIV OF JINAN
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