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Biosensor for detecting glutathione and fabrication method of biosensor

A biosensor, glutathione technology, applied in the field of biosensors, can solve the problems of complex procedures, difficult to generalize, time-consuming and poor sensitivity, and achieve the effects of low process cost, high sensitivity and easy operation

Active Publication Date: 2019-12-06
UNIV OF JINAN
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] There are many traditional GSH detection methods including spectrophotometry, high performance liquid chromatography, iodometric method, etc., however, they require complicated procedures, are time-consuming and have poor sensitivity, making it difficult to generalize
In order to overcome the above shortcomings, some GSH detection methods based on colorimetry and fluorescence have been developed, and these new technologies have brought great progress to the detection of GSH; however, further improvement is needed to achieve more sensitive and specific detection of GSH

Method used

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  • Biosensor for detecting glutathione and fabrication method of biosensor
  • Biosensor for detecting glutathione and fabrication method of biosensor
  • Biosensor for detecting glutathione and fabrication method of biosensor

Examples

Experimental program
Comparison scheme
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Embodiment 1

[0055] The preparation method of described chemiluminescence biosensor comprises the following steps:

[0056] The construction steps of composite probe P are as follows:

[0057]Add 18 μL of sterilized water, 3 μL of 10×PBS, 3 μL of 100 μM P0 probe, 3 μL of 100 μM P1 probe and 3 μL of 100 μM P2 probe into the pre-prepared sterile EP tube , shake for 30s, incubate at 95°C for 5min, slowly cool down to room temperature and hybridize as a probe, and store at -20°C for later use.

[0058] The operation steps of composite probe P modified to the surface of gold nanoparticles are as follows:

[0059] a. Take 1 mL nano-gold solution in a centrifuge tube, centrifuge for 15 min, and centrifuge the two tubes at the same time for later use. Centrifuge until the supernatant is colorless and transparent, remove the supernatant, and add 300 μL of sterile water to concentrate the nano-gold solution to 1 nM. Transfer to a 1 mL glass bottle and seal with tinfoil. After standing at room te...

Embodiment 2

[0069] The preparation method of described chemiluminescence biosensor comprises the following steps:

[0070] The construction steps of composite probe P are as follows:

[0071] Add 18 μL of sterilized water, 3 μL of 10×PBS, 3 μL of 100 μM P0 probe, 3 μL of 100 μM P1 probe and 3 μL of 100 μM P2 probe into the pre-prepared sterile EP tube , shake for 30s, incubate at 95°C for 5min, slowly cool down to room temperature and hybridize as a probe, and store at -20°C for later use.

[0072] The operation steps of composite probe P modified to the surface of gold nanoparticles are as follows:

[0073] a. Take 1 mL nano-gold solution in a centrifuge tube, centrifuge for 15 min, and centrifuge the two tubes at the same time for later use. Centrifuge until the supernatant is colorless and transparent, remove the supernatant, and add 300 μL of sterile water to concentrate the nano-gold solution to 1 nM. Transfer to a 1 mL glass bottle and seal with tinfoil. After standing at room t...

Embodiment 3

[0083] The preparation method of described chemiluminescence biosensor comprises the following steps:

[0084] The construction steps of composite probe P are as follows:

[0085] Add 18 μL of sterilized water, 3 μL of 10×PBS, 3 μL of 100 μM P0 probe, 3 μL of 100 μM P1 probe and 3 μL of 100 μM P2 probe into the pre-prepared sterile EP tube , shake for 30s, incubate at 95°C for 5min, slowly cool down to room temperature and hybridize as a probe, and store at -20°C for later use.

[0086] The operation steps of composite probe P modified to the surface of gold nanoparticles are as follows:

[0087] a. Take 1 mL nano-gold solution in a centrifuge tube, centrifuge for 15 min, and centrifuge the two tubes at the same time for later use. Centrifuge until the supernatant is colorless and transparent, remove the supernatant, and add 300 μL of sterile water to concentrate the nano-gold solution to 1 nM. Transfer to a 1 mL glass bottle and seal with tinfoil. After standing at room t...

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Abstract

The invention relates to the technical field of biosensors, in particular to a biosensor for detecting glutathione (GSH) by a gold nanoparticle-based DNA molecule machine. The biosensor comprises a hairpin probe HAP (stem-modified disulfide bond), a composite probe P, a P3 probe, heme, potassium ion, a target object GSH, nanometer gold and a buffer liquid, wherein the composite probe P is modifiedonto a surface of the nanometer gold by polyA modification. A hairpin structure is damaged by a cracking function of the target object GSH on the disulfide bond, a Walker nucleic acid chain is released, P2 can be replaced from the composite probe P through strand displacement reaction of a support point medium by the released Walker nucleic acid chain and the P3 probe, the P2 is a sequence rich with G-quadruplet, G-quadruplet / heme DNA enzyme is formed in the presence of the heme, so that an aptamer biosensor is built. The reaction of the sensor is finished only in one step, thus, the sensor has the advantages of rapid detection speed, low cost, low detection limit, high specificity and the like and is simple and convenient to operate.

Description

technical field [0001] The invention belongs to the technical field of biosensors, in particular to a biosensor for detecting glutathione by a DNA molecular machine based on gold nanoparticles, and also relates to a preparation method thereof. Background technique [0002] Glutathione is a tripeptide containing γ-amide bonds and sulfhydryl groups, composed of glutamic acid, cysteine ​​and glycine. Present in almost every cell of the body, widely distributed in eukaryotic and gram-negative bacteria, but almost absent in gram-positive bacteria such as lactic acid bacteria. Glutathione can help maintain normal immune system functions, and has anti-oxidation and integrated detoxification effects. The sulfhydryl group on cysteine ​​is its active group, which is easy to combine with certain drugs, toxins, etc., and has the ability to integrate Detoxification. As the most abundant non-protein molecule in cells, elevated levels of GSH are commonly seen in many types of human cance...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N21/64G01N21/76
CPCG01N21/6402G01N21/76
Inventor 王玉李莎莎刘素黄加栋张儒峰赵一菡瞿晓南张雪宋晓蕾王海旺王敬锋
Owner UNIV OF JINAN
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