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Unmarked AuNPs-Thi electrochemical immunosensor for detecting transgenic protein and a preparation method thereof

An immune sensor and genetically modified technology, which is applied in the direction of material electrochemical variables, scientific instruments, instruments, etc., can solve the problems of poor parallelism of immune sensors, achieve the effects of shortening the test time, reducing the amount of antibodies, and increasing the amount of biological immobilization

Pending Publication Date: 2021-11-16
SHANGHAI ACAD OF AGRI SCI +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0011] Wang Z et al. (A novel oriented immunosensor based on AuNPs-thionine-CMWCNTs and staphylococcal protein A for interleukin-6 analysis complicated biological samples[J].Analytica Chimica Acta.2020,1140:145-152) developed an immunosensor based on AuNPs / Thi / The immunosensor of CMWCNTs (carboxylated multi-walled carbon nanotubes) is used for highly sensitive detection of interleukin-6 (IL-6). In this scheme, three kinds of materials are selected to form a composite material modified electrode, which can also achieve the goal of highly sensitive detection. However, multi-walled carbon nanotubes are prone to aggregation when modifying electrodes, which may lead to poor parallelism of the constructed immunosensor

Method used

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  • Unmarked AuNPs-Thi electrochemical immunosensor for detecting transgenic protein and a preparation method thereof
  • Unmarked AuNPs-Thi electrochemical immunosensor for detecting transgenic protein and a preparation method thereof
  • Unmarked AuNPs-Thi electrochemical immunosensor for detecting transgenic protein and a preparation method thereof

Examples

Experimental program
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Effect test

Embodiment 1

[0052] Example 1 Construction of a label-free AuNPs-Thi electrochemical immunosensor for detecting PAT protein

[0053] The working electrode of the label-free electrochemical immunosensor is a glassy carbon electrode, and gold nanoparticles AuNPs are electrodeposited on the surface of the glassy carbon electrode, and thionine Thi is stably adsorbed on the surface of the AuNPs through electrostatic adsorption to form an AuNPs-Thi film; the PAT-mAb antibody is Covalently bound to the AuNPs-Thi membrane, BSA as a blocking agent, see figure 1 , its construction process is as follows:

[0054] 1) The working electrode is polished and cleaned according to the steps, and the cleaned glassy carbon electrode is soaked in 1% HAuCl 4 In the solution, gold nanoparticles are electrodeposited on the surface of the electrode by CV method, the potential range is: 0V ~ 1.1V, and the scan rate is 50mV / s;

[0055] 2) Thi (5 μL) was modified on the surface of the glassy carbon electrode GCE by...

Embodiment 2

[0066] Detection performance analysis of the immunosensor constructed in Example 2

[0067] 1. Sample processing and determination method

[0068] The transgenic crop seed standards to be detected (see Table 2) were extracted with 0.01M PBS buffer (pH 7.4) at a mass-to-volume ratio of 1:3.

[0069] Table 2 List of GM crop seed powder standard products used

[0070]

[0071] After adding the seed powder into PBS, shake vigorously for 3-5 minutes, then centrifuge at 6000rpm for 5 minutes, collect the supernatant in layers, and dilute it into transgenic protein solutions of different concentrations for detection. The supernatant of corresponding non-transgenic crops is used as the experimental blank control. The actual seed samples were ground into powder by a food grinder or a mortar, added double distilled water at a ratio of 1:5, oscillated and mixed, centrifuged at 8000rpm for 5min, and the supernatant was taken for detection.

[0072] Differential pulse voltammetry (Di...

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Abstract

The invention discloses an unmarked AuNPs-Thi electrochemical immunosensor for detecting transgenic protein and a preparation method thereof. The preparation method comprises the following steps: electrodepositing gold nanoparticles AuNPs on the surface of a glassy carbon electrode, and stably adsorbing thionine Thi on the surface of the AuNPs to form an AuNPs-Thi film; and binding the transgenic protein-mAb antibody covalently to the AuNPs-Thi membrane, and then using BSA (bovine serum albumin) for sealing. According to the method, after AuNPs is electrodeposited on the surface of the electrode, a proper amount of Thi is adsorbed, an electrochemical signal is remarkably enhanced, an antibody of transgenic protein is connected to an AuNPs-Thi film through the amino cross-linking effect, the long-term stability and sensitivity of the sensor are facilitated, the method is simple and rapid, the amount of needed antibodies is small, the sensitivity is high, and the specificity and reproducibility are good; and the invention provides a novel method for electrochemical immunodetection of transgenic protein in transgenic crops.

Description

technical field [0001] The invention belongs to the technical field of food safety immune detection, and in particular relates to a label-free AuNPs-Thi electrochemical immunosensor for detecting transgenic proteins and a preparation method thereof. Background technique [0002] With the continuous development of genetic engineering technology, the planting area of ​​transgenic crops is expanding year by year, and its safety has become the focus of attention. Although my country has implemented mandatory labeling regulations on genetically modified crops and products, there are still many cases of illegal cultivation and sale of genetically modified crops. Therefore, the development of a simple, rapid and highly sensitive method for the identification of genetically modified crops is of great significance to the regulation of genetically modified crops and their products. [0003] In the detection of genetically modified crops, the detection based on the nucleic acid level ...

Claims

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Application Information

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IPC IPC(8): G01N27/30G01N27/327G01N27/48G01N33/68G01N33/577G01N33/553
CPCG01N27/308G01N27/3278G01N27/3277G01N27/48G01N33/68G01N33/577G01N33/54346G01N33/5438
Inventor 曾海娟王金斌杨倩雯
Owner SHANGHAI ACAD OF AGRI SCI
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