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A method for detecting enterovirus cvb3

An enterovirus, purpose technology, applied in the field of detection of CVB3 based on in situ recombinase polymerase amplification photoelectrochemical biosensor

Active Publication Date: 2022-07-15
QINGDAO UNIV OF SCI & TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, it is challenging to develop rapid and sensitive molecular diagnostic techniques for CVB3

Method used

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  • A method for detecting enterovirus cvb3
  • A method for detecting enterovirus cvb3
  • A method for detecting enterovirus cvb3

Examples

Experimental program
Comparison scheme
Effect test

example 1

[0017] Example 1: Construction of PEC analysis detection method.

[0018] In the PEC sensor assay assay for CVB3, RPA was used to amplify the target. The PEC performance of the sensor construction process was investigated in 0.1 mM monosodium p-aminophenyl phosphate solution. The PEC signal of CPE is 35.3nA; the PEC signal of ZnSeNSs / AuNPs / BNNSs / CPE is 835.0nA; the PEC signal of Primer / ZnSeNSs / AuNPs / BNNSs / CPE (immobilized forward primer) is 713.1nA; Amplicon / Primer / The PEC signal of ZnSeNSs / AuNPs / BNNSs / CPE (amplified) is 682.0nA; SA-ALP / Amplicon / Primer / ZnSeNSs / AuNPs / BNNSs / CPE (connected to streptavidin-labeled alkaline phosphatase after amplification) The PEC signal is 3622.0 nA. in 5 mM [Fe(CN) containing 0.1 M KCl 6 ] 3- / 4- The impedance of the sensor assembly process was tested in solution. CPE showed smaller Ret values. When ZnSeNSs / AuNPs / BNNSs were modified onto CPE, the Ret value was increased compared with bare CPE. The Ret value increased after Primer was modif...

example 2

[0019] Example 2: Sample Detection

[0020] Monosodium p-aminophenyl phosphate (4-APP) was purchased from Shanghai Huicheng Biological Co., Ltd. (Shanghai, China). p-aminophenol, tris (Tris), mercaptoethylamine (MCH), tris(2-carboxyethyl)phosphine (TCEP), streptavidin-labeled alkaline phosphatase (SA-ALP) , N,N-dimethylformamide (DMF) was provided by Sangon Biotech Co., Ltd. (Shanghai, China). TwistAmp Basic RT-RPA kit (Twist-DX), recombinase polymerase buffer solution were purchased from Babraham (UK). Biotin-labeled reverse primer RP and thiolated forward primer FP were synthesized by Sangon Biotech Co., Ltd. (Shanghai, China). Boron nitride powder was purchased from Nanjing Pioneer Nano Co., Ltd.; zinc selenide was purchased from Maclean Biochemical Co., Ltd.

[0021] The sequences of the forward and reverse primers are as follows:

[0022] Forward primer FP: 5'-SH-AGTCCTCCGGCCCCTGAATGCGGCTAATCC-3'

[0023] Reverse primer RP: 5'-Biotin-GGATGGCCAATCCAATAGCTATATGGTAACAA-3'...

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Abstract

The invention belongs to the field of electrochemistry and analytical chemistry, and the purpose of the invention is to provide a method for detecting CVB3 based on in situ recombinase polymerase amplification photoelectrochemical biosensor. Using recombinase polymerase amplification, the amplified product is connected to the electrode in situ, the biotin on the amplified product interacts with the streptavidin-labeled alkaline phosphatase in the solution, and the alkaline phosphatase is regenerated. The substrate in the catalytic solution is hydrolyzed to monosodium p-aminophenyl phosphate to generate the free electron donor p-aminophenol. The p-aminophenol enhances the photoelectrochemical reaction of the modified electrode to generate an enhanced photoelectric signal to realize the detection of CVB3. This method It has high sensitivity and simple method.

Description

technical field [0001] The invention belongs to the field of analytical chemistry and photoelectrochemical sensors, in particular to a method for detecting CVB3 based on an in-situ recombinase polymerase amplification photoelectrochemical biosensor. Background technique [0002] Coxsackievirus B3 (CVB3) is a type of enterovirus with high morbidity and mortality. This small, single-stranded RNA virus causes a variety of symptoms. For example, hand, foot and mouth disease, severe acute flaccid paralysis, meningitis, and neonatal organ failure are common in children under five years of age. However, the development of rapid and sensitive molecular diagnostic techniques for CVB3 is challenging. The purpose of the present invention is to provide a method for detecting CVB3 based on in situ recombinase polymerase amplification photoelectrochemical biosensor. Using recombinase polymerase amplification (RPA), the amplified product is connected to the electrode in situ, and the bi...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/70C12Q1/6825C12Q1/6844
CPCC12Q1/701C12Q1/6825C12Q1/6844C12Q2521/507C12Q2522/101
Inventor 混旭王如昊林健健高杰邓陆平秦娜娜王明竞张慧
Owner QINGDAO UNIV OF SCI & TECH