Rapid pathogenic microorganism detection method

A technology of pathogenic microorganisms and detection methods, applied in the field of Raman detection, can solve the problems of difficult to meet the urgent needs of rapid detection, complicated and time-consuming operations, difficult to apply to complex samples, etc., to achieve excellent separation and enrichment capabilities, uniform distribution of hot spots, The effect of enhancing the effect is excellent

Active Publication Date: 2020-04-17
WUHAN ACADEMY OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Among them, the culture method is the gold standard method for microbial detection. Although the result is accurate, it usually needs to go through the steps of separation, identification, culture and counting. The operation is complicated and time-consuming, and generally takes 3-5 days.
However, although the detection time of PCR method and ELISA method is greatly shortened, they need to rely on expensive special instruments and professional technicians, and are difficult to apply to complex samples.
In summary, the above methods are difficult to meet the urgent needs of rapid on-site detection of pathogenic microorganisms

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0034] 1. Preparation of nano-gold@nitrogen-doped graphene quantum dot nanomaterials

[0035] (1) Synthesis of seed crystals: 50mL 1.0mM HAuCl 4 Add the solution into a 150mL flask; stir vigorously, and after heating to boiling, quickly add 5mL of sodium citrate solution (w / v 1%). When the color of the mixed solution turns wine red, stop the reaction immediately. Finally, the product was filtered through a 0.22 μm nitrocellulose membrane and stored in a refrigerator at 4°C until use. The particle size of the seed crystal is about 15nm.

[0036] (2) Synthesis of nano-gold: Add 48mL deionized water into a 150mL flask, stir vigorously, heat to boiling, then quickly add 2mL 0.1M Tris solution. After continuing to reflux for 5min, 0.5mL HAuCl 4 The solution (w / v of which is 1%) and 0.5 mL of the seed crystal synthesized in step (1) were rapidly injected into the flask. Within 3 minutes, the color of the solution changed from pink to purple to ruby ​​red immediately. After con...

Embodiment 2

[0047] Salmonella mock sample testing

[0048] First, the room temperature milk (3000rpm, 5min) purchased from the supermarket was centrifuged to remove impurities. Then it was treated by diluting 10 times with PBS solution. Add 1,000 and 10,000 CFU / mL Salmonella standard solutions respectively therein, and perform the Salmonella detection shown in Example 2 after thorough mixing. By measuring it at 2227cm -1 According to the working curve obtained in Example 2, it is estimated that the concentration of Salmonella in the simulated sample is 985 and 10224 CFU / mL, and then the recoveries of the two samples are 98.50% and 102.24 %. Therefore, it can be verified that the method has good feasibility and accuracy in the detection of actual samples.

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PUM

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Abstract

The invention belongs to the technical field of microorganism detection, and discloses a rapid pathogenic microorganism detection method which comprises the following specific steps: preparing a detection probe and a capture probe, carrying out incubating to obtain a sandwich product, drawing a standard working curve, and detecting a to-be-detected sample. The detection probe comprises a nanogold@ nitrogen doped graphene quantum dot nano material, and a sulfydryl-containing silent region Raman signal molecule and a broad-spectrum recognition molecule which are modified on the surface of the nano material; and the capture probe is an immunomagnetic bead modified with an antibody. Raman detection of pathogenic microorganism is realized by combining the detection probe with immunomagnetic beads, namely a mechanism of specific recognition coupled with broad-spectrum recognition. The sensor construction and detection process do not need large instruments, operation is simple and rapid, andthe sensitivity is high.

Description

technical field [0001] The invention relates to the technical field of pathogenic microorganism detection, in particular to a pathogenic microorganism composed of nano-gold@nitrogen-doped graphene quantum dot nanomaterials, Raman signal molecules in the silent zone, immune magnetic beads, and specific recognition coupled with broad-spectrum recognition Fast, accurate and highly sensitive Raman detection method. Background technique [0002] With the improvement of people's living standards and the changes of the main contradictions in our country, food safety has become an important issue of public health, and food-borne diseases are a serious global problem. It not only seriously affects people's life and health, but also affects the development of the global economy. According to the U.S. Centers for Disease Control and Prevention (CDC), it is estimated that about one-sixth of Americans get sick every year due to eating contaminated food or water, and 3,000 of them die as...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/543G01N21/65
CPCG01N33/54326G01N21/658Y02A50/30
Inventor 王利华白向茹吴文辉肖康飞王佳慧
Owner WUHAN ACADEMY OF AGRI SCI
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