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Method for detecting the colon bacillus by combining magnetic nanoparticle enrichment with bi-color flow cytometry

A technology of magnetic nanoparticles and Escherichia coli, which is applied in the direction of individual particle analysis, chemical instruments and methods, particle and sedimentation analysis, etc., can solve problems such as difficulties and false positive signals, and achieve improved accuracy, high detection sensitivity, and reduced false positives. The effect of positive signals

Inactive Publication Date: 2013-02-27
HUNAN UNIV
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Problems solved by technology

However, in the case of low bacterial concentration, it is still difficult to overcome problems such as false positive signals, so as to meet the requirements of rapid and accurate detection

Method used

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  • Method for detecting the colon bacillus by combining magnetic nanoparticle enrichment with bi-color flow cytometry
  • Method for detecting the colon bacillus by combining magnetic nanoparticle enrichment with bi-color flow cytometry
  • Method for detecting the colon bacillus by combining magnetic nanoparticle enrichment with bi-color flow cytometry

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Embodiment

[0043] Application of the detection method of the present invention in the detection of Escherichia coli O157:H7

[0044] 1. Capture and enrichment of E.coli O157:H7 by magnetic nanoparticles

[0045] The preparation concentration is 2.6×10 6 cells / mL, 2.6×10 5 cells / mL, 2.6×10 4 cells / mL, 2.6×10 3 cells / mL, 1.3×10 3 cells / mL of E.coli O157:H7 bacterial suspension, incubate each bacterial suspension with concanavalin A at 35°C-37°C for 1 hour, centrifuge and wash twice; then take 6 centrifuge tubes with a volume of 10mL (the numbers are respectively a, b, c, d, e, f), respectively add the following bacterial suspension and PB buffer solution to each tube to prepare the samples to be tested with gradient concentration, and add 100 μL to each centrifuge tube at the same time Silica-coated aminated magnetic nanoparticles with surface-modified mannose (the transmission electron microscope image of the magnetic nanoparticles is shown in Figure 4 shown), the material composit...

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Abstract

The invention relates to a detection method of colon bacillus, particularly discloses a method for detecting the colon bacillus by combining magnetic nanoparticle enrichment with bi-color flow cytometry. The method comprises the following steps of: firstly co-culturing nanoparticles and samples to be measured and enriching composites formed by the magnetic nanoparticles and the samples to be measured under the action of an externally applied magnetic field; then carrying out immune co-culture and secondary enrichment on colon bacillus antibodies labelled by fluorescent nanoparticles and the enriched samples to be measured; then dyeing the secondarily-enriched samples to be measured by using nucleic acid dyes; finally detecting and viewing the dyed samples to be measured by adopting the multi-parameter flow cytometry and a laser confocal microscope, and analyzing and judging detection results of the colon bacillus contained in the samples to be measured according to the number of detected objects with fluorescent double-positive signals. The detection method has the advantages of rapidness, accuracy, sensitivity, low false positive rate, and the like.

Description

technical field [0001] The invention relates to a method for detecting Escherichia coli, in particular to a method for detecting Escherichia coli using a biological immunoassay technique. Background technique [0002] Escherichia coli (E.coli), commonly known as Escherichia coli, was discovered by Escherich in 1885. For a long period of time, it has been regarded as a component of normal intestinal flora and considered to be a non-pathogenic bacterium . It was not until the middle of the 20th century that people realized that some special serotypes of Escherichia coli were pathogenic to humans and animals, especially infants and young animals (poultry), often causing severe diarrhea and sepsis. Escherichia coli is a common prokaryotic organism that belongs to bacteria. According to different biological characteristics, we can divide pathogenic Escherichia coli into five categories: pathogenic Escherichia coli (EPEC), enterotoxigenic Escherichia coli ( ETEC), enteroinvasive...

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N15/10C09K11/59
Inventor 何晓晓王柯敏周丽霞何定庚曹杰
Owner HUNAN UNIV
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