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Novel electrochemical biosensing method for detecting glucose oxidase and urease and application of novel electrochemical biosensing method

A glucose oxidase and biosensing technology is applied in the fields of functional biomaterials and biosensing, achieving good application prospects, realizing low-concentration glucose oxidase and urease activity analysis, and simple operation.

Active Publication Date: 2021-02-05
北京天宇恒泰科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, there is no relevant research on the monitoring of glucose oxidase and urease activity by combining the C-rich DNA long chain produced by TdT catalysis and the change law of impedance value induced by pH change at home and abroad.

Method used

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  • Novel electrochemical biosensing method for detecting glucose oxidase and urease and application of novel electrochemical biosensing method
  • Novel electrochemical biosensing method for detecting glucose oxidase and urease and application of novel electrochemical biosensing method
  • Novel electrochemical biosensing method for detecting glucose oxidase and urease and application of novel electrochemical biosensing method

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Experimental program
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Embodiment 1

[0038] Embodiment 1 Preparation of sensor

[0039] The technical solution adopted by the present invention to solve the above-mentioned technical problems is: a new electrochemical biosensing method for detecting glucose oxidase and urease and its application, the specific steps are as follows:

[0040] Preparation of Electrode 1:

[0041] The gold electrode is made of Al with a particle size of 0.05-1 μm 2 o 3 The powder was polished for 15 minutes, and then ultrasonically cleaned in ultrapure water for 3 times, 5 minutes each time. Then put the gold electrode in 0.2M sulfuric acid solution for cyclic voltammetry test, the scanning range is -0.3-1.6V, and the scanning speed is 0.01-0.1V / s, until a stable cyclic voltammetry curve is obtained, which is recorded as Electrode 1.

[0042] Preparation of Electrode 2:

[0043] 5 μL of thiol DNA solution with a concentration of 0.5 μM was dropped onto the surface of Electrode 1, and then placed in a refrigerator (4°C) overnight. ...

Embodiment 2

[0048] Example 2 Feasibility experiment of impedance switch

[0049] The invention builds an electrochemical AC impedance switch based on the reversible behavior of the i-motif structure under acidic and neutral conditions. see results figure 2 , when the glucose oxidase reaction solution is added, the impedance value will decrease, and after the urease reaction solution is added, the impedance value will increase. The stability of the sensor is further investigated through cycle tests. The results show that after 5 cycles of cycle tests, the sensor performance remains good.

Embodiment 3

[0050] Example 3 Detection of glucose oxidase

[0051] Prepare the sensor according to the above-mentioned Example 1, and control the final concentration of glucose oxidase to be: 0, 0.01, 0.02, 0.05, 0.08, 0.1, 0.2, 0.4, 0.8, 1, 2, 5, 8, 10, 20, 40, 50 U / L, and other steps are the same as the real-time example 1 above. Experimental results such as image 3 As shown, within a certain range, with the increase of glucose oxidase concentration, the electrochemical impedance value becomes smaller and smaller. The linear correlation equation of the electrochemical impedance of the sensor to the logarithmic value of the glucose oxidase concentration is Rct=-1424lgC GOx +6127,R 2 =0.9987, the linear range is 0.01-20U / L, and the detection limit is 0.0061U / L, indicating that the sensor can realize highly sensitive detection of glucose oxidase concentration.

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Abstract

The invention discloses a novel electrochemical biosensing method for detecting glucose oxidase and urease and application of the novel electrochemical biosensing method. The method comprises the following steps: firstly, modifying a 5'-sulfydryl DNA probe on a gold electrode, continuously catalyzing and polymerizing deoxycytidine triphosphate (dCTP) to a 3'-OH end of sulfydryl DNA through terminal deoxyribonucleic acid transferase (TdT) to form a longer C-rich DNA long chain, thus obtaining a larger electrochemical impedance value in 5.0mM [Fe(CN)6]3- / 4- through an electrochemical impedance method (EIS); dropwise adding glucose oxidase reaction liquid to the surface of the electrode, so that the C-rich DNA long-chain part is semi-protonated and forms an imotif structure, and the impedancevalue is reduced; dropwise adding urease reaction liquid to the surface of an electrode, wherein a C-rich DNA long chain is changed into a DNA single chain state from an imotif structure, so that theimpedance value is increased on the contrary, and glucose oxidase and urease sensitivity analysis is achieved based on the facts.

Description

technical field [0001] The invention relates to an electrochemical biosensor, in particular to a novel electrochemical impedance method and application for detecting glucose oxidase and urease, and belongs to the technical field of functional biological materials and biosensing. Background technique [0002] Diabetes is a public health problem worldwide. This metabolic disorder is caused by insulin deficiency and hyperglycemia. People with diabetes need to check blood sugar levels frequently to achieve better control of the disease. Glucose oxidase (GOx) is a flavoprotein containing flavin adenine dinucleotide FAD, which is composed of two subunits with the same amino acid sequence connected by disulfide bonds. This enzyme is capable of catalyzing the oxidation of β-D-glucose to δ-gluconolactone and oxygen peroxide in the presence of molecular oxygen, the former being hydrolyzed spontaneously to gluconic acid in water. Therefore, the development of a simple, label-free elec...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N27/327G01N27/02
CPCG01N27/3277G01N27/3271G01N27/02
Inventor 胡宇芳胡丹丹郭智勇王邃
Owner 北京天宇恒泰科技有限公司
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