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An integrated particle-type immobilized enzyme biosensor and its preparation method and application

A biosensor and immobilized enzyme technology, applied in the direction of biological testing, material inspection products, etc., can solve the problems of rarely using MOF response characteristics, and achieve good reusability, simplified detection steps, and good selectivity.

Active Publication Date: 2021-07-27
SUN YAT SEN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

And the existing MOF-enzyme system only uses MOF as the protective layer and selective permeable layer of the enzyme, but rarely utilizes the designable response characteristics of MOF, such as fluorescence sensing, chemical catalysis, electrical conductivity, structural changes, etc.

Method used

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  • An integrated particle-type immobilized enzyme biosensor and its preparation method and application
  • An integrated particle-type immobilized enzyme biosensor and its preparation method and application
  • An integrated particle-type immobilized enzyme biosensor and its preparation method and application

Examples

Experimental program
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Effect test

Embodiment approach

[0039] Application of 3,5-diethyl-1,2,4-triazole copper (I), or Hdetz and cuprous ions in the preparation of immobilized enzyme biosensors.

[0040] Preferably, the immobilized enzyme biosensor has both enzyme catalytic performance and optical oxygen sensing performance.

[0041] Preferably, the enzyme is an enzyme that generates oxygen or consumes oxygen during the catalytic reaction.

[0042] More preferably, the enzyme is glucose oxidase, uric acid oxidase, D-amino acid oxidase, L-amino acid oxidase, L-α-hydroxy acid oxidase, peroxidase and the forms of these enzymes with fluorescent labels .

[0043] Preferably, the peroxidase includes catalase and horseradish peroxidase.

[0044] Preferably, the immobilized enzyme biosensor is of particle type.

[0045] An immobilized enzyme biosensor, which contains 3,5-diethyl-1,2,4-triazole copper (I) and enzyme, wherein the enzyme is immobilized on 3,5-diethyl-1,2 , within the framework of 4-triazole copper (I); or it is made of e...

Embodiment 1

[0061] Example 1 Preparation method of an integrated particle-type immobilized enzyme biosensor

[0062] containing 0.25mmol Cu(NO 3 ) 2 Add 1gNaOH to 20mL solution of 0.25mmol ascorbic acid, quickly generate Cu 2 O nanoparticles, washed after centrifugation. Dissolve the resulting nanoparticles in 20 mL of ethanol and add a drop of hydrazine hydrate or ascorbic acid. Under nitrogen protection, the mixture was added to the ) in 15 mL of aqueous solution. After stirring overnight (12 h) at 800 rpm at room temperature, the product was collected by centrifugation at 9000 rpm for 3 minutes, washed three times with deionized water, and then lyophilized into powder to obtain the immobilized enzyme biosensor GOx@MAF-2.

Embodiment 2

[0063] Example 2 A preparation method of an integrated particle-type immobilized enzyme biosensor

[0064] The method is the same as in Example 1, the only difference being that GOx in Example 1 is replaced by rhodamine B isothiocyanate (RhB)-labeled GOx (denoted as RhB-GOx). The immobilized enzyme biosensor finally prepared in Example 2 was designated as RhB-GOx@MAF-2.

[0065] The specific preparation method of the above-mentioned RhB-GOx is as follows: 9 mg rhodamine B isothiocyanate (RhB) was dissolved in 4 mL DMSO, and added dropwise to 2 mL GOx solution (10 mg mL -1 , 0.5M pH 9.5 carbonate buffer). The solution was stirred at room temperature at 300 rpm in the dark for 6 hours. Excess RhB was removed by acetone precipitation at -80°C for 1 hour and then centrifuged, and the enzyme labeled with fluorescent molecules was dried to obtain RhB-GOx.

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Abstract

The invention discloses an integrated particle-type immobilized enzyme biosensor and its preparation method and application. The invention utilizes Hdetz, cuprous ions and enzymes to co-react under mild conditions, and constructs an optical oxygen sensor with optical oxygen sensing performance and high efficiency. Enzyme@MAF‑2, an integrated immobilized enzyme biosensor for enzyme catalytic activity. The sensor has the advantages of high stability of immobilized enzyme, good selectivity and reusability. Moreover, its integrated sensing form can directly respond to the substrate without adding other auxiliary reagents, which greatly simplifies the detection steps and reduces the detection cost. It is the first case of using MOF with optical sensing properties as an immobilized enzyme carrier. All-in-one biosensor.

Description

technical field [0001] The invention relates to the field of metal organic framework (MOF) materials and the field of biosensors, in particular to a preparation method and application of an integrated MOF-immobilized enzyme biosensor. Background technique [0002] Particle-based biosensors immobilize enzymes in micro / nanomaterials and play an important role in many fields. Free enzymes have poor stability, are easily inactivated, and are difficult to separate from the bottom / product in a homogeneous reaction, resulting in difficult product recovery and continuous production, which are limited in practical applications. However, particle-based biosensors have better stability, recyclability, and catalytic activity than free enzymes. Because of their small particle size, it is beneficial to realize the injection-type living body detection method and avoid the immune effect of the human body. Although particle-based biosensors have many advantages, they still face serious cha...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N33/50G01N33/52
CPCG01N33/50G01N33/52
Inventor 戴宗许宇智柳思扬张杰鹏邹小勇
Owner SUN YAT SEN UNIV
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