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Bacteria detection kit

A gold nanorod and double-labeled technology, applied in the field of microbial detection, can solve the problems of high cost and time-consuming

Active Publication Date: 2019-08-16
JILIN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] The purpose of the present invention is to provide a convenient and economical bacterial detection kit to solve the problems of long time-consuming and high cost of the existing bacterial detection methods

Method used

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Examples

Experimental program
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Effect test

Embodiment 1

[0024] Example 1 Preparation of gold nanorods

[0025] Gold nanorods were prepared by the seed growth method, and the specific steps were as follows:

[0026] 1) Prepare the seed solution: add 96µl aqueous solution of tetrachloroauric acid trihydrate with a concentration of 25 mM to 7.5mL aqueous solution of cetyltrimethylammonium bromide with a concentration of 0.1M; then add 0.5-2mL aqueous solution with a concentration of 0.02 M sodium borohydride aqueous solution; when the mixed solution turns dark brown, stir at 30°C for 1-3 hours, and the resulting solution is the seed solution;

[0027] 2) Prepare growth solution: mix 100mL of 0.1M cetyltrimethylammonium bromide aqueous solution, 2mL of 0.5M sulfuric acid, 1.96mL of 25mM tetrachloroauric acid trihydrate aqueous solution, 0.5-2mL The silver nitrate aqueous solution with a concentration of 0.01M and the ascorbic acid aqueous solution with a concentration of 0.1M of 0.5-1mL are fully mixed, and the resulting mixed solutio...

Embodiment 2

[0030] Example 2 Preparation of double-labeled gold nanorod probes

[0031] Add 120 µL of urease solution with a concentration of 8-12 mg / mL dropwise into 1200 µL of the suspension containing gold nanorods described in Example 1, and vortex slowly for 5 minutes; 1 mL anti-Staphylococcus aureus egg yolk antibody solution, vortex slowly for 25 minutes; centrifuge at 8000 rpm for 10 minutes, discard the supernatant, and the precipitated part is the double-labeled gold nanorod probe;

[0032] Resuspend with 300-500µl ultrapure water, which is the suspension containing double-labeled gold nanorod probes; measured by using visible light-ultraviolet spectrophotometer, the absorption spectrum of double-labeled gold nanorod probes is better than that of unlabeled gold nanorod probes. The absorption spectrum of the rods was obviously red-shifted, which proved that the double-labeled gold nanorod probes were successfully prepared.

Embodiment 3

[0033] The preparation of embodiment 3 non-specific magnetic beads

[0034] The non-specific magnetic beads are Fe coated with carbon dots on the surface 3 o 4 Nanoparticles, synthesized by a two-step method;

[0035] Step 1: Preparation of Fe 3 o 4 Nanoparticle core: 0.5-1g ferrous chloride tetrahydrate and 1-2g ferric chloride hexahydrate are fully dissolved in 30mL ultrapure water; 35mL 28% (w / v) ammonia water is added dropwise; under the protection of nitrogen, 80 Stir at ℃ for 5 hours; use a permanent magnet to separate the black solid substance from the reaction solution, and wash it with ultrapure water, which is Fe 3 o 4 Nanoparticle cores, dried for later use;

[0036] Step 2: On Fe 3 o 4 Nanoparticle core surface modified carbon dots: add 0.05-0.2g of the above Fe 3 o 4 Mix nanoparticle core and 0.25-1g chitosan in 30mL 4% (w / v) glacial acetic acid, mix thoroughly; transfer the obtained liquid to a reaction kettle, and react at 180°C for 6-24 hours; Separa...

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Abstract

The invention discloses a double-labeled gold nanorod probe, which is obtained by adding a urease solution into a suspension of the gold nanorods dropwise for mixing and swirling, adding anti-staphylococcus aureus antibody solution dropwise for mixing and swirling, and finally centrifuging and supernatant abandoning. The double-labeled probe converts the quantity signal of SA into a pH change of the solution by the catalytic action of urease and the specific recognition of IgY. A detection kit for staphylococcus aureus is provided at the same time, comprising the double-labeled gold nanorod probe, non-specific magnetic beads, saturated urea and phenolphthalein test paper, wherein the double-labeled gold nanorod probe is labeled with urease and anti-staphylococcus aureus antibody; the result is rapidly detected by phenolphthalein test paper from white to magenta. The invention has the advantages that the kit is easy to operate; the test result can be interpreted by naked eyes under natural light without any instrument; comparison with a standard color chart can be carried out to obtain semi-quantitative detection results; and the detection is fast and can be completed in 20 minutes.

Description

technical field [0001] The invention belongs to the technical field of microorganism detection, and in particular relates to a bacteria detection kit. Background technique [0002] Staphylococcus aureus ( Staphylococcus aureus , SA) is an important zoonotic pathogen, one of the top five pathogens associated with foodborne diseases, and can cause a variety of serious infections. Because Staphylococcus aureus can tolerate a wide range of pH, temperature and humidity, it is widely distributed in nature, and there are many opportunities for food contamination. It is one of the main pathogenic bacteria that cause food poisoning. With the widespread use of antibiotics, a large number of drug-resistant Staphylococcus aureus have emerged. The diseases caused by Staphylococcus aureus have become a global public health problem, seriously endangering human safety and health. Studies have shown that 30%-50% of the general population are carriers of SA. Therefore, the product is easi...

Claims

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Application Information

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IPC IPC(8): G01N33/569G01N33/532G01N33/535
CPCG01N33/532G01N33/535G01N33/56938
Inventor 李娟庞博赵超徐坤王娟宋秀玲李金华
Owner JILIN UNIV
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