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Nano composite material, ESAT-6 electrochemical aptamer sensor and preparation and detection methods thereof

A nanocomposite material, ESAT-6 technology, applied in the field of electrochemical detection, can solve the problem of inability to effectively distinguish active tuberculosis patients from latent infection patients, low positive rate of sputum smear microscope examination, high bacterial content in sputum, etc. problem, achieve the effect of improving electron transfer ability, improving water solubility and stability, and high specificity

Active Publication Date: 2021-03-02
CHONGQING MEDICAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, MTB culture takes a long time; the positive rate of sputum smear microscopy is low and the requirements for bacterial content in sputum are high; the disadvantage of chest X-ray imaging is that it cannot provide etiological diagnosis and cannot detect potential infection; immunological methods: such as Tuberculin skin test, but it may have false positive results for people who have been vaccinated with BCG. Others, such as diagnostic kits, include Anda-TB ELISA diagnostic kits, TB-SA antibody test kits, etc., but the disadvantage is that currently Cannot effectively distinguish active TB patients from latently infected patients

Method used

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  • Nano composite material, ESAT-6 electrochemical aptamer sensor and preparation and detection methods thereof
  • Nano composite material, ESAT-6 electrochemical aptamer sensor and preparation and detection methods thereof
  • Nano composite material, ESAT-6 electrochemical aptamer sensor and preparation and detection methods thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0046] Preparation of PEI@NG@HP-Uio-66-NH 2 on Ce-MOF@Tb nanocomposite

[0047] Follow the steps below:

[0048] 1) Disperse 5mg of nitrogen-doped graphene (NG) in 10mL of absolute ethanol, ultrasonically disperse evenly, stir at room temperature, and then dropwise add 4mL of 1wt% polyethyleneimine (PEI) aqueous solution to the nitrogen-doped graphene dispersion solution, after stirring at room temperature for 1 h, the PEI@NG dispersion was obtained;

[0049] 2) 80mg ZrCl 4 and 2.4g of lauric acid were added to 20mL N, N-dimethylformamide (DMF) solution, ultrasonic at room temperature for 20min, and then 31mg of 2-aminoterephthalic acid (NH 2 -BDC), continue to ultrasonically disperse for 5min, transfer the above-mentioned solution to a reaction kettle for 12h at 120°C, cool, centrifuge, wash with DMF and ethanol and dry to obtain yellow HP-Uio-66-NH 2 Material;

[0050] 3) Take 40mg of HP-Uio-66-NH prepared in step 2) 2 powder, at the same time take 40mg of polyvinylpy...

Embodiment 2

[0055] Preparation of electrochemical DNA aptamer sensor for detection of ESAT-6

[0056] Follow the steps below: (The construction principle is as follows figure 1 shown)

[0057] 1) Treat the ESAT-6 binding aptamer (EBA) with 10 mM TES (pH=7.4) buffer solution at room temperature, and place it at 4°C for use;

[0058] 2) Wash the glassy carbon electrode with piranha washing solution (98% H 2 SO 4 / 30%H 2 o 2 =3:1, v / v) Rinse with ultrapure water after soaking for 30 minutes;

[0059] 3) Use the electrodes obtained in step 2) with 0.3 μm and 0.05 μm Al 2 o 3 The powder was polished to a mirror surface, and then the electrodes were ultrasonically treated in the order of ultrapure water, absolute ethanol, and ultrapure water for 5 minutes each, and dried at room temperature for later use.

[0060] 4) Place the electrode obtained in step 3) in 0.5M H 2 SO 4 Electrochemical activation was carried out in the medium, and the potential scan was -0.3~1.55V until a stable cycl...

Embodiment 3

[0065] Detection of ESAT-6 by Electrochemical DNA Aptamer Sensor

[0066] Using the electrochemical DNA aptamer sensor constructed in Example 2 to detect ESAT-6, follow the steps below:

[0067] 1. Draw the working curve

[0068] 1) Place the modified electrodes from Step 4) to Step 7) of Example 2 in a solution containing 0.1M Na 2 HPO 4 ,0.1M KH 2 PO 4 ,10mM KCl and 2mM MgCl 2 Characterized in 0.1M PBS (pH=7.0), and measured its electrochemical response signal, the results are as follows figure 2 Shown: (a) bare glassy carbon electrode; (b) dropwise PEI@NG@HP-Uio-66-NH 2 -on-Ce-MOF@Tb nanocomposite; (c) combined with EBA; (d) blocked by BSA.

[0069] 2) 10 μL of the target substance tuberculosis early secretion target antigen 6 of different concentrations was dropped onto the electrode of the aptamer sensor prepared in Example 2, and the current changes were measured respectively.

[0070] 3) According to the obtained current change value (such as image 3 Shown in...

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Abstract

The invention provides a nano composite material, which is a polyethyleneimine (PEI) functionalized nitrogen-doped graphene (NG) modified HP-Uio-66-NH2-on-Ce-MOF nano material, and an ESAT-6 electrochemical aptamer sensor prepared by using the nano composite material is used for quantitative detection of ESAT-6. The PEI@NG@HP-Uio-66-NH2-on-Ce-MOF @Tb nano composite material which is finally formedafter an electroactive substance toluidine blue (Tb) is loaded is used as a sensitive interface of the sensor; PEI@NG has good conductivity is used for signal amplification, and a double-MOF material, namely HP-Uio-66-NH2-on-Ce-MOF, has multiple porosities, a large specific surface area and excellent biological activity, and loads a large number of aptamers through hydrogen bonds, pi-pi accumulation and electrostatic interaction with the aptamers. Finally, Tb signal response changes are caused through specific binding of the aptamer and target objects with different concentrations, and quantitative detection of ESAT-6 is achieved. The electrochemical aptamer sensor prepared by the invention is successfully used for ultra-sensitive detection of ESAT-6.

Description

technical field [0001] The invention relates to the technical field of electrochemical detection, in particular to a nanocomposite material, an ESAT-6 electrochemical aptamer sensor, a preparation method and a detection method thereof. Background technique [0002] Tuberculosis (TB) is a fatal infectious disease caused by Mycobacterium tuberculosis (MTB). More than 10 million people become tuberculosis patients every year, which poses a serious challenge to international public health. MTB is the pathogen of tuberculosis, which mainly infects the lungs, but the infection can also affect other organs, such as bones, joints, kidneys and brains, posing a huge threat to the lives and health of the public. At present, asymptomatic infection and missed diagnosis of tuberculosis are the main causes of tuberculosis transmission and death. Therefore, it is necessary to develop a sensitive and rapid tuberculosis detection method to detect tuberculosis as early as possible, so as to gi...

Claims

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Application Information

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IPC IPC(8): G01N27/26G01N27/30G01N27/327
CPCG01N27/26G01N27/30G01N27/3278
Inventor 母昭德白丽娟谢娇娇
Owner CHONGQING MEDICAL UNIVERSITY
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