Nanometer fiber film sensor with function of rapid bacterium detection and preparation method thereof

A nanofiber membrane and nanofiber technology, applied in the direction of material excitation analysis, fluorescence/phosphorescence, etc., can solve the loss of biological luciferase for one-time use, increased use cost, weak interaction between luciferase and carrier, etc. problems, to achieve the effect of preventing large losses, saving costs, and simple design ideas

Active Publication Date: 2014-06-25
WUHAN TEXTILE UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, there is a lack of luciferase immobilization on an appropriate carrier for the bacterial sensor prepared by the bioluminescent method, or the interaction between the luciferase and the carrier is weak, which will easily cause the loss of the bioluciferase and one-time use, and the cost of use will increase.

Method used

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preparation example Construction

[0032] The preparation method comprises the following steps;

[0033] Preparation of a biosensor for rapid detection of bacteria:

[0034] The preparation of A polyolefin copolymer nanofibrous film, described preparation adopts the preparation method that Chinese patent publication number is CN 101553607A, is about to mix PVA-co-PE, cellulose acetate butyrate (CAB) according to mass ratio as 1: 4 Even and dry. Melting, blending and extruding through a twin-screw extruder, after collection, CAB is extracted by refluxing acetone at 60° C. for 72 hours to obtain nanofibers with a fiber diameter of ≤500 nm.

[0035] The nanofibers of PVA-co-PE extracted with acetone are dispersed in a certain solvent with a mass fraction of 1% to 5% to prepare a nanofiber suspension. Then, using high-pressure airflow forming technology, the nanofiber suspension is formed on various fiber fabrics.

[0036] Preparation of B active PVA-co-PE nanofibrous membrane

[0037] Place the PVA-co-PE nanof...

Embodiment 1

[0056]According to the above method, the prepared PVA-co-PE nanofiber membrane is placed in a container, mixed with a 2mol / L sodium hydroxide solution until completely soaked, and then the container is placed in a constant temperature water bath shaker at a constant temperature of 15°C , the time is 30 minutes, take it out, wash it, and dry it. Then mix the NaOH-treated nanofibrous membrane with a 1,4-dioxane solution with a mass fraction of 5% cyanuric chloride in a container until completely soaked, and place it in a constant temperature water bath shaker at a constant temperature of 15°C. , the time is 60 minutes, take out, wash and dry, and prepare a nanofiber membrane with active triazine on the surface.

[0057] Mix the above-mentioned active nanofiber membrane with 0.1g / L luciferase, 1mmol / L dithiothreitol, 1mmol / L EDTA, 1mmol / LTris-HCl buffer solution and stir evenly. The immersion temperature is 0°C. The immersion time was 12 hours, and then washed with 1 mmol / L tris...

Embodiment 2

[0059] According to the above method, the prepared PVA-co-PE nanofiber membrane is placed in a container, mixed with a concentration of 3mol / L sodium hydroxide solution until completely soaked, and then the container is placed in a constant temperature water bath shaker at a constant temperature of 25°C , the time is 20 minutes, take it out to clean and dry. Then mix the NaOH-treated nanofibrous membrane with a 1,4-dioxane solution with a mass fraction of 10% cyanuric chloride in a container until completely soaked, and place it in a constant temperature water bath shaker at a constant temperature of 20°C. , the time is 30 minutes, take out, wash and dry, and prepare a nanofiber membrane with active triazine on the surface.

[0060] Mix the above active nanofiber membrane with 1g / L luciferase, 3mmol / L dithiothreitol, 3mmol / L EDTA, 50mmol / L Tris-HCl buffer solution and stir evenly, the immersion temperature is 2°C, The immersion time was 18 hours, and then washed with 50 mmol / ...

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Abstract

The invention discloses a polyolefin nanometer fiber film sensor for rapid bacterium detection and a preparation method thereof. According to the invention, polyolefin copolymer is adopted as a substrate material, and the chemically-modified polyolefin nanometer fiber film biosensor is prepared by surface chemical modification, and solid phase synthetic process of bioactive molecules. The nanometer fiber film biosensor prepared by the method of the invention overcomes, with respect to performance, the disadvantages of time consumption, manpower consumption, poor accuracy for bacterium culture and bacterial colony counting in traditional bacterium detection, realizes rapid detection of ATP released by bacteria by using the luminescence principle between triphosadenine, luciferase, and fluorescein, and realizes rapid response to biological harmful substances. With the solid phase synthesis, bioactive enzymes are well fixed on the surface of the nanometer fiber film, which realizes repeated and circular detection of bacteria. The nanometer fiber film sensor is widely applicable to fields of biotechnology, medical health, food and medicine detection, water treatment, and the like.

Description

technical field [0001] The invention relates to a nanofiber membrane sensor with the function of rapidly detecting bacteria and a preparation method thereof. Background technique [0002] Biohazardous germs have always been a major hazard to human health, both in developing and developed countries. The problem of biological harmful germs is a widespread concern in more and more countries around the world. Therefore, research in this area is also in the ascendant, and the research on biosensors that can quickly detect bacteria has become a hot spot therein. [0003] At present, the methods for rapid detection of bacteria can be divided into bioelectrochemical methods and non-bioelectrochemical methods. The former includes impedance / conductivity method, potentiometric method, dye cell technology, cyclic / voltammetric method, amperometric method, etc. to detect microbial activity. A class of methods that induce a charge transfer signal between electrodes. The latter encompass...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N21/64
Inventor 王栋赵青华陆莹吴志红李沐芳
Owner WUHAN TEXTILE UNIV
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