Method for detecting escherichia coli O157:H7

A technology of O157 and Escherichia coli, applied in the field of microbial detection, can solve problems such as low precision, and achieve the effect of saving cost and improving detection sensitivity

Inactive Publication Date: 2016-07-13
NANCHANG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0005] The present invention aims at the technical defects of the prior art, and provides a detection method for Es...

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  • Method for detecting escherichia coli O157:H7
  • Method for detecting escherichia coli O157:H7
  • Method for detecting escherichia coli O157:H7

Examples

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Embodiment 1

[0050] Example 1 (Application of the novel fluorescent ELISA detection method for detection of Escherichia coli O157:H7 in lettuce in the present invention)

[0051] When the novel fluorescent ELISA detection method of the present invention is used to detect the content of Escherichia coli O157:H7 in lettuce, it is implemented through the following steps: sample pretreatment, detection by the detection method of the present invention, and analysis of 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 Escherichia coli O157:H7 content in the above-mentioned sample with detection method of the present invention

[0055] Take the enzyme label coated with polyclonal antibody against Escherichia coli O157:H7 ...

Embodiment 2

[0067] Example 2 (Escherichia coli O157: H7 ELISA detection method using horseradish peroxidase as antibody labeling enzyme and TMB as chromogenic substrate)

[0068] When the traditional ELISA detection method is used to detect the content of E. coli O157:H7 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.

[0069] (1) Sample pretreatment

[0070] 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.

[0071] (2) Use the traditional ELISA detection method to detect the content of E. coli O157:H7 in the above samples

[0072] Take the enzyme label coated with polyclonal antibody against Escherichia coli O157:H7 content, add standard substance / sample 1...

Embodiment 3

[0078] The invention relates to a detection method for Escherichia coli O157:H7, which belongs to the double-antibody sandwich enzyme-linked immunosorbent method, and the method is for the detection of Escherichia coli O157:H7 antigen, and the enzyme used for labeling the antibody in the method is catalase C100.

[0079] On the basis of the above technical solutions, the following conditions are met:

[0080] The substrate in the method includes hydrogen peroxide and mercaptopropionic acid modified cadmium telluride quantum dots.

[0081] The specific detection method includes the following steps:

[0082] 1) Escherichia coli O157:H7 antibody coated;

[0083] 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;

[0084] 3) Then add biotinylated Escherichia coli O157:H7 monoclonal antibody to mix, react at 35°C in a dark environment for 40min, and wash;

[0085] 4) Then add streptavi...

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Abstract

The invention provides a method for detecting escherichia coli O157:H7. According to the method, firstly, escherichia coli O157:H7 and coated polyclonal antibodies are combined, then connected with biotinylated monoclonal antibodies and connected with catalase C100 labeled with streptavidin, fluorescence quenching of cadmium telluride quantum dots modified with mercaptopropionic acid is reduced through decomposition of hydrogen peroxide under the catalytic function of catalase, and the concentration of escherichia coli O157:H7 in a sample is judged according to the fluorescence intensity. The method is based on a double-antibody sandwich enzyme immunoassay technique and adopts a biotin-avidin system for amplifying a reaction. More importantly, a novel antibody labeling enzyme (catalase C100) and a more sensitive fluorogenic substrate (cadmium telluride quantum dots) are adopted and matched with an effective reaction condition, accordingly, the detection sensitivity is remarkably improved, the cost is reduced, the detection efficiency is improved, and the method has good popularization prospect.

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 Escherichia coli O157:H7. Background technique [0002] Escherichia coli O157:H7 (Escherichiacoli O157:H7) belongs to the genus of Enterobacteriaceae and is the main serotype of enterohaemorrhagic Escherichia coli; cattle are its main host, and can cause hemorrhagic enteritis, diarrhea, Hemolytic uremia and other symptoms are characterized by the production of a large amount of Shiga-LikeToxin (SLT), which causes microvascular changes in the body; some studies have found that the pathogenic dose of this bacteria is extremely low, only 10 live bacteria are needed That's it. Escherichia coli O157:H7 is easy to culture, reproduces rapidly, has strong infectivity, and has a wide range of infection routes. Therefore, establishing a sensitive and rapid detection method is an impo...

Claims

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Application Information

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IPC IPC(8): G01N33/569G01N33/535
CPCG01N33/535G01N33/56916G01N2333/245
Inventor 许恒毅熊勇华黄小林赖卫华陈锐湛胜楠
Owner NANCHANG UNIV
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