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Detection method for listeria monocytogenes

A technology for the detection of Listeria monocytogenes, which is applied in the direction of measuring devices, instruments, scientific instruments, etc., can solve the problems of low precision of the detection method, and achieve the effects of improving detection sensitivity, saving costs, and detecting sensitivity

Inactive Publication Date: 2016-08-10
NANCHANG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The present invention aims to provide a detection method for Listeria monocytogenes aimed at the technical defects of the prior art, so as to solve the problem of the low accuracy of the ELISA detection method for Listeria monocytogenes in the prior art.

Method used

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  • Detection method for listeria monocytogenes
  • Detection method for listeria monocytogenes
  • Detection method for listeria monocytogenes

Examples

Experimental program
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Effect test

Embodiment 1

[0050] Embodiment 1 (the present invention detects the application of the novel fluorescence ELISA detection method of LM in the LM content in detection lettuce)

[0051] When the novel fluorescent ELISA detection method of the present invention is used to detect the LM content in the lettuce, it is implemented through the following steps: sample pretreatment, detection by the detection method of the present invention, and analysis of the results.

[0052] (1) Sample pretreatment

[0053] Take 1g of the processed lettuce sample, add it to 9ml of sterile PBS, then add 1ml of bacterial solution, shake vigorously for 30 minutes; take the supernatant and add immunomagnetic beads to enrich and concentrate to 1mL liquid, take it out for later use.

[0054] (2) detect LM content in the above-mentioned sample with detection method of the present invention

[0055] Take the microtiter plate coated with anti-LM monoclonal antibody, add 100 μL / well of standard / sample to the correspondin...

Embodiment 2

[0068] Embodiment 2 (using horseradish peroxidase as antibody labeling enzyme, using TMB as the LM ELISA detection method of chromogenic substrate)

[0069] When the traditional ELISA detection method is used to detect the LM content in lettuce and beef products, it is implemented through the following steps: sample pretreatment, detection by traditional ELISA detection method, and analysis of the results.

[0070] (1) Sample pretreatment

[0071] Take 1g of the processed lettuce sample, add it to 9ml of sterile PBS, then add 1ml of bacterial solution, shake vigorously for 30 minutes; take the supernatant and add immunomagnetic beads to enrich and concentrate to 1mL liquid, take it out for later use.

[0072] (2) Detect the content of LM in the above samples by traditional ELISA detection method

[0073] Take the microtiter plate coated with anti-LM monoclonal antibody, add 100 μL / well of standard / sample to the corresponding microwell; take the microtiter plate coated with an...

Embodiment 3

[0079] A detection method for LM, the method belongs to double-antibody sandwich ELISA, the method is for the detection of LM antigen, and the enzyme used to label the antibody in the method is catalase C100.

[0080] On the basis of the above technical solutions, the following conditions are met:

[0081] The substrate in the method includes hydrogen peroxide and mercaptopropionic acid modified cadmium telluride quantum dots.

[0082] The specific detection method includes the following steps:

[0083] 1) coated with LM antibody;

[0084] 2) Take the coated antibody in step 1), mix it with the sample to be tested, react in a light-proof environment at 35°C for 40 minutes, and wash;

[0085] 3) Then add biotinylated LM polyclonal antibody to mix, react at 35°C in a dark environment for 40min, and wash;

[0086] 4) Then add streptavidin-labeled catalase C100 to mix, react at 35°C in a dark environment for 40min, and wash;

[0087] 5) Then add hydrogen peroxide solution with...

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Abstract

The invention provides a detection method for listeria monocytogenes (LM). The method comprises the steps of firstly combining the LM with a coated monoclonal antibody, then connecting with a biotinylated polyclonal antibody, next connecting with a streptavidin labeled catalase C100, catalyzing decomposition of hydrogen peroxide by the catalase, reducing fluorescence quenching of mercaptopropionic acid modified cadmium telluride quantum dots, and determining the concentration of the LM in a sample according to the level of the fluorescence intensity. The method is based on the double-antibody sandwich enzyme-linked immunosorbent assay, and a biotin-avidin system is used for amplification of the reaction. More importantly, due to use of the new antibody labeled enzyme (catalase C100) and the more sensitive fluorescent substrate (cadmium telluride quantum dots), and matching of the effective reaction conditions, the detection sensitivity is significantly improved, at the same time, the cost is reduced, and the detection efficiency is improved; therefore, the detection method has good prospects for promotion.

Description

technical field [0001] The invention relates to the technical field of microorganism detection, and further relates to an ELISA-based antigen detection technology, in particular to a detection method for Listeria monocytogenes. Background technique [0002] Listeria monocytogenes (Listeria monocytogenes, LM) is one of the most common food-borne pathogens, belonging to aerobic or facultative anaerobic Gram-positive bacteria, short bacilli with blunt ends, single Arranged in a V shape, the size is about (0.4-0.5μm)×(0.5-2.0μm); LM is the most pathogenic bacteria in the Listeria genus, and it is a zoonotic pathogen , is also a common foodborne pathogen. It can parasitize and proliferate in animal and human cells, mainly through Listeria lysin (LLO), ActA protein and internalization to achieve infection. The main targets of LM infection are newborns, the elderly, pregnant women, and people with immunodeficiency. Generally, mild poisoning mainly manifests as enteritis symptoms,...

Claims

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Application Information

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IPC IPC(8): G01N33/569
CPCG01N33/56911G01N2333/32
Inventor 许恒毅熊勇华黄小林赖卫华段宏裴可
Owner NANCHANG UNIV
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