Double enzyme sensor and its preparation and uses

A technology of enzyme sensor and phenol oxidase, which is applied in the field of double enzyme sensor and microbiological inspection of water body, can solve the problems of complex procedures, long detection cycle, complicated operation process, etc., and achieve the effect of short time, simple operation and high sensitivity

Inactive Publication Date: 2007-11-28
EAST CHINA NORMAL UNIVERSITY
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, these methods have the disadvantages of long detection period, complicated procedures, and various reagents required.
For example, according to the multi-tube fermentation method, Escherichia coli can ferment lactose to produce acid and CO 2 Coli water samples with different dilutions were cultured with lactose-containing medium, and the three inspection steps of initial fermentation test, plate separation and re-fermentation test were complicated. The operation process took more than 72 hours, and it was difficult to meet the requirements of rapid detection of E. coli. needs

Method used

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  • Double enzyme sensor and its preparation and uses
  • Double enzyme sensor and its preparation and uses
  • Double enzyme sensor and its preparation and uses

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0031] Example 1 Preparation of dual enzyme sensor

[0032] In the first step, the SnCl used 4 ·5H 2 O, indium chloride, and 37% hydrochloric acid are respectively 10.0g, 1.10g, 3.0mL, 6g methanol solution containing 25% tetrabutylammonium hydroxide; in the second step, in H 2 O 2 / NH 4 OH / H 2 The volume ratio of O is 1:1:5 (v / v) soaked for 0.5h, then soaked in toluene solution containing 0.5% 3-aminopropyltriethoxysilane for 10h, and then the electrode is soaked in 0.5 % Glutaraldehyde solution for 20 minutes, and finally immersed in citric acid-phosphate buffer solution containing 0.5% peroxidase for 10 minutes to prepare a dual-enzyme sensor.

Embodiment 2

[0033] Example 2 Rapid detection of Escherichia coli with the dual-enzyme sensor prepared in Example 1

[0034]In the first step, the nutrient broth concentration of salicylic acid and glucose is 4.0×10 -4 mol·L -1 , The incubation time is 2h; in the second step, the pH value of the citric acid-phosphate buffer solution is 5.0; in the third step, the response current value obtained from the actual sample corresponds to the E. coli concentration of 1.7×10 3 cfu / mL.

Embodiment 3

[0035] Example 3 Preparation of Dual Enzyme Sensor

[0036] In the first step, the SnCl used 4 ·5H 2 O, indium chloride, and 37% hydrochloric acid are respectively 10.2g, 1.33g, 3.5mL, 8g methanol solution containing 25% tetrabutylammonium hydroxide; in the second step, in H 2 O 2 / NH 4 OH / H 2 Soak in a solution with a volume ratio of 1:1:5 (v / v) for 1 hour, then soak in a toluene solution containing 1% 3-aminopropyltriethoxysilane for 12 hours, and then soak the electrode in 1% In glutaraldehyde solution for 30 minutes, and finally immersed in citric acid-phosphate buffer solution containing 1% peroxidase for 20 minutes, a dual-enzyme sensor was prepared.

[0037] Example 4 Rapid detection of Escherichia coli with the dual-enzyme sensor prepared in Example 3

[0038] In the first step, the nutrient broth concentration of salicylic acid and glucose is 5.0×10 -4 mol·L -1 , The incubation time is 2.5h; in the second step, the pH of the citric acid-phosphate buffer solution is 6.0; i...

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Abstract

The invention relates to a double-enzyme sensor and the equipment and the application, belonging to the technology field of double-enzyme sensor and microbiological test for water. The sensor comprises substrate of glass rod, nanometer indium tin oxide electrode, horseradish peroxidase and multiple phenoloxidase. The nanometer indium tin oxide electrode is attached on the surface of substrate of glass rod in film shape. The multiple phenoloxidase and horseradish peroxidase are attached on the surface of nanometer indium tin oxide electrode in turn respectively in film shape. On the substrate of glass rod attached by nanometer indium tin oxide electrode double-enzyme sensor is prepared as 3-animo propyl triethoxy silane is assembled by itself on the nanometer indium tin oxide electrode; Schiff base is formed by glutaraldehyde; and multiple phenoloxidase and horseradish peroxidase are bonded on the nanometer indium tin oxide electrode in chemical bond in turn. The double-enzyme sensor is used as working electrode and quick measurement for the concentration of Escherichia coli in water is realized by measuring the current through the working electrode in chronoamperometry.

Description

Technical field [0001] The invention relates to a dual-enzyme sensor and its preparation and application. The application refers to a method for using the sensor to quickly detect Escherichia coli. That is, the dual-enzyme sensor is used as a working electrode and the chronoamperometry is used to realize the rapid detection of the concentration of Escherichia coli in a water body. The method belongs to the technical field of dual-enzyme sensors and microbiological examination of water bodies. Background technique [0002] Escherichia coli is the most common flora that inhabits the intestines of humans and animals. However, when E. coli invades other organs or tissues outside the intestine, it can cause diseases such as urethritis, cystitis, cholecystitis, appendicitis, etc.; in infants, the elderly or patients with reduced immune function, E. coli infection can cause sepsis (up to 30% mortality). In addition, E. coli can also cause diarrhea in humans and cau...

Claims

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Application Information

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IPC IPC(8): G01N27/327C12Q1/10
Inventor 张文金利通耿萍唐辉张新爱朱伟王丹郑娇红安雅睿
Owner EAST CHINA NORMAL UNIVERSITY
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