Preparation method of double-ratio biosensor based on amplification linear ranges of aptamers with different structures and application

A biosensor, linear range technology, applied in the field of biosensors, can solve problems such as limiting the linear range, and achieve the effects of high reliability, large amount of information, and good stability

Active Publication Date: 2021-01-29
JIANGSU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, such sensors still have a potentially significant limitation: single-point binding produces a fixed linear curve, limiting their useful linear range

Method used

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  • Preparation method of double-ratio biosensor based on amplification linear ranges of aptamers with different structures and application
  • Preparation method of double-ratio biosensor based on amplification linear ranges of aptamers with different structures and application
  • Preparation method of double-ratio biosensor based on amplification linear ranges of aptamers with different structures and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0039] The preparation method of the linear aptamer-hairpin aptamer double-ratio electrochemical aptasensor comprises the following steps:

[0040] (1) Preparation of label-free electrochemical probe AQ-rGO composite:

[0041] First, anthraquinone (AQ) and reduced graphene oxide (rGO) were dissolved in ultrapure water to obtain AQ dispersion and rGO dispersion, and the concentration of AQ dispersion was 0.5 mg·mL -1 , the concentration of rGO dispersion is 0.05 mg·mL -1 and use a cell pulverizer to process the rGO dispersion for 30 minutes to improve its dispersibility in water; then mix the AQ dispersion with the rGO dispersion at a volume ratio of 3:10, magnetically stir for 18 hours and wash with centrifugal water at a centrifugal speed of 8000rpm. After 15 minutes, the AQ-rGO composite material was obtained, dispersed in ultrapure water to obtain the AQ-rGO composite material dispersion, and stored at room temperature for later use;

[0042] (2) Take a glassy carbon elec...

Embodiment 2

[0048] Performance analysis of a linear aptamer-hairpin aptamer-based dual-ratio electrochemical aptasensor for detection of AFB1:

[0049] In the ratiometric electrochemical aptasensor based on hairpin aptamer ( figure 2 A), linear aptamer-hairpin aptamer dual-ratio electrochemical aptasensor ( figure 2 B) The surface is sequentially modified with different concentrations of AFB1, the incubation time at room temperature is 80min, and then the electrode is cleaned with Tris-HCl (pH=7.4) solution, and the performance curves of the two types of sensors are obtained respectively. The comparison shows that the introduction of linear aptamers The detection linear range of the sensor is significantly improved, such as image 3 As shown in Figures A and B; the sensor is the working electrode, the saturated Ag / AgCl electrode is the reference electrode, and the platinum wire electrode is the counter electrode. The electrochemical signal is recorded and detected by the electrochemica...

Embodiment 3

[0051] Detection of AFB1 in peanut by a linear aptamer-hairpin aptamer-based dual-ratio electrochemical aptasensor:

[0052] Peanuts are used as actual samples. Spray the AFB1 standard solution on the surface of peanut samples, shake vigorously for 60 minutes, extract 4g of peanuts with 10mL methanol aqueous solution (60%), then centrifuge at 8000rpm for 15 minutes; take the upper layer solution and wash it with 0.22μm Filter treatment to remove impurities and store the resulting actual sample solution for later use;

[0053] Actual sample solutions of different concentrations were modified on the surface of the aptamer sensor in Example 1, and the incubation time was 80 min at room temperature, and then the electrode was cleaned with Tris-HCl (pH=7.4) solution, the sensor was used as the working electrode, and the Ag / AgCl electrode was saturated As the reference electrode, the platinum wire electrode is the counter electrode, and the electrochemical signal is recorded and det...

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Abstract

The invention belongs to the technical field of biosensors, and particularly relates to a preparation method of a double-ratio electrochemical aptamer sensor based on amplification linear ranges of alinear aptamer and a hairpin aptamer. The method comprises the steps of forming an AQ-rGO composite material through the non-covalent action of AQ and rGO; and fixing the sulfydryl modified hairpin aptamer and linear aptamer on a sensing interface by using AuNPs; wherein one part of the hairpin aptamer comprises an aptamer sequence and carries an electrochemical molecule methylene blue, and the linear aptamer carries a ferrocene electrochemical signal molecule, so that a novel double-ratio electrochemical aptamer sensor is constructed; compared with the hairpin aptamer, the binding effect of the linear aptamer and a target AFB1 of the linear aptamer is easier to generate, so that the concentration of the target AFB1 causing the hairpin aptamer to carry MB signal molecules to change can beused as a concentration point for the qualitative analysis of the AFB1, the qualitative analysis of the AFB1 in food is realized, the detection sensitivity is high, and the stability is good.

Description

technical field [0001] The invention belongs to the technical field of biosensors, and in particular relates to a preparation method and application for amplifying the linear range of a double-ratio electrochemical aptamer sensor based on a linear aptamer-hairpin aptamer, which can be used for aflatoxin B1 ( Detection analysis of AFB1). Background technique [0002] Aflatoxin B1 (AFB1) is the most serious carcinogen, mutagen and teratogen, and has been widely found in many crops, such as peanut, corn and cotton. AFB1 is classified as category I by the International Agency for Research on Cancer. AFB1 is resistant to heat and chemical treatments and is ubiquitous in agricultural production and food processing. Many countries or regions have set the maximum allowable level of AFB1 in food and feed, usually 2ng mL -1 -20ng mL -1 . Therefore, it is urgent to develop a simple and sensitive strategy to detect AFB1 to ensure food safety. High-performance liquid chromatography...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N27/30G01N27/327G01N27/416
CPCG01N27/308G01N27/327G01N27/4163
Inventor 李玉叶由天艳刘东李文佳朱成喜
Owner JIANGSU UNIV
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