A dual-mode separation immunosensor based on enzyme-induced bioetching and its preparation method

An immune sensor and separate technology, applied in the field of immune sensors, can solve the problems of long operation time, inaccurate detection results, and high cost

Active Publication Date: 2021-09-21
SOUTH CHINA AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

These detection methods are often limited in practical applications due to problems such as inaccurate detection results, long operation time, low sensitivity, high cost, or the need for professional operators.

Method used

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  • A dual-mode separation immunosensor based on enzyme-induced bioetching and its preparation method
  • A dual-mode separation immunosensor based on enzyme-induced bioetching and its preparation method
  • A dual-mode separation immunosensor based on enzyme-induced bioetching and its preparation method

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Experimental program
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Effect test

Embodiment 1

[0035] A preparation method based on enzyme-induced bioetching dual-mode separation immunosensor, comprising the following steps:

[0036] S1. Preparation of CdS / ZnO NRs / r-GO: first synthesize graphene oxide hydrogel (GO), and prepare cadmium sulfide / zinc oxide nanorod array CdS / ZnO NRs on the reduced graphene oxide r-GO to obtain CdS / ZnO NRs / r-GO composites;

[0037] Preparation of HRP-liposome-Ab 2 Complex: preparation of liposome-encapsulated horseradish peroxidase HRP by thin-film dispersion method,

[0038] The HRP-liposome complex was obtained, and the HRP-liposome complex was combined with the second antibody Ab by glutaraldehyde cross-linking method 2 connect,

[0039] HRP-Liposome-Ab 2 Complex;

[0040] Preparation of gold nanobicones: Au NBPs prepared by seed growth method;

[0041] The specific operation for preparing CdS / ZnO NRs / r-GO is as follows: add 160mL concentrated sulfuric acid into a dry three-necked flask, slowly add 4g graphite powder and 14g potas...

Embodiment 2

[0050] Example 2 Characterization of CdS / ZnO NRs / r-GO composites

[0051] Synthesis of graphene oxide hydrogel (GO): Add 160mL concentrated sulfuric acid into a dry three-necked flask, slowly add 4g graphite powder and 14g potassium permanganate under stirring in an ice-water bath, and keep stirring the mixture at 35°C for 24h . After the reaction was completed, dilute with 400 mL of ice water, and then add hydrogen peroxide until the color of the mixture did not change and no bubbles were generated. Stirring was continued for 2 h to remove unreacted potassium permanganate, and then centrifuged at 5000 rpm for 3 minutes. Centrifuge and wash with 300mL HCl (1mol / L) three times and then wash with water until the supernatant is neutral. The precipitate after centrifugation was dialyzed for one week, and the product was divided into two equal parts, which were centrifuged and washed with water and ethanol respectively, and finally dispersed in water or ethanol and stored for late...

Embodiment 3

[0054] Example 3 HRP-liposome-Ab 2 Characterization of the complex

[0055] The liposome-encapsulated horseradish peroxidase (HRP) complex is prepared by a film dispersion method, and the process is briefly described as follows: Weigh dipalmitoylphosphatidylcholine (DPPC), cholesterol and dipalmitoylphosphatidylethanolamine (DPPE ) in a total of 30 mg (molar ratio 10:10:1), mixed and dissolved in 4 mL of chloroform / methanol mixture (volume ratio 6:1), and then transferred to a round-bottomed flask by rotary evaporation at 45 ° C to form a uniform layer Lipid film, then add 2 mL of phosphate buffer solution (PBS, 0.01 M, pH 7.4) containing 2.5 mg / mL HRP, and incubate at 35 °C for 2 h. The mixture was then sonicated for 5 min in an ice-water bath, repeating three cycles. Finally, the prepared HRP-liposome complex was centrifuged to remove unencapsulated HRP.

[0056] The complexes prepared above were combined with the second antibody (Ab) by glutaraldehyde cross-linking metho...

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Abstract

The invention discloses a dual-mode separation immunosensor based on enzyme-induced bioetching, covering / preparing cadmium sulfide / zinc oxide nanorod array CdS / ZnO NRs on the surface of three-dimensional reduced graphene oxide r-GO as a photoelectrode; using gold Nanobiconic Au NBPs as multicolor chromogenic substrates; horseradish peroxidase HRP was used to link photoelectrochemical immunoassay and colorimetric detection, in which Cd / ZnO NRs / r-GO was bioprinted through HRP-induced enzyme-catalyzed reaction Corrosion, thereby forming a photocurrent change, HRP catalyzed oxidation of hydrogen peroxide produced hydroxyl radicals for bio-etching Au NBPs to form gold nanoparticles of different sizes and shapes, thereby showing color changes and blue shift of LSPR peaks, the method of the present invention utilizes lipid Plastids encapsulate a large amount of HRP and load more Ab 2 To effectively amplify the response signal and further improve the accuracy of detection.

Description

technical field [0001] The invention belongs to the technical field of immunosensors, and more specifically relates to an enzyme-induced bioetching-based dual-mode separation immunosensor and a preparation method thereof. Background technique [0002] Photoelectrochemical (PEC) detection, as an emerging and rapidly developing detection technology, has attracted widespread attention. Compared with the traditional ELISA, PEC detection uses light as the excitation light source and photocurrent as the detection signal. Thanks to two different forms of energy conversion, the PEC immunoassay has a lower detection limit and higher sensitivity. In addition, relatively simple and low-cost devices are more conducive to the development of portable and miniaturized instruments. However, even though PEC detection has many of the advantages mentioned above, the traditional single-signal PEC immunoassay platform has been gradually difficult to meet the growing detection needs of people. ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N27/327G01N21/78G01N27/30G01N33/543
CPCG01N21/78G01N27/305G01N27/3278G01N33/5432
Inventor 刘英菊刘莹魏婕申浩然陈华明
Owner SOUTH CHINA AGRI UNIV
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