Method for rapidly detecting and screening staphylococcus aureus

A staphylococcus, golden yellow technology, applied in the direction of measuring devices, instruments, scientific instruments, etc., can solve the problems of difficult simultaneous analysis of multiple components, molecular weight analysis, shortened detection time, etc., to achieve low requirements for supporting equipment, high efficiency and rich Set, good effect of reliability

Inactive Publication Date: 2012-07-18
SHANGHAI NORMAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although these methods have improved the sensitivity of detection and greatly shortened the detection time, they all have certain defects.
For example, ELISA has high selectivity to reagents, and it is difficult to analyze multiple components at the same time; there is a certain degree of cross-reaction to compounds with similar structures; it is difficult to analyze compounds with small molecular weight or very unstable compounds

Method used

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  • Method for rapidly detecting and screening staphylococcus aureus
  • Method for rapidly detecting and screening staphylococcus aureus
  • Method for rapidly detecting and screening staphylococcus aureus

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0032] Example 1 Preparation of Immunized Superparamagnetic Nanomagnetic Beads

[0033] 1. Preparation of aldylated superparamagnetic nano-magnetic beads

[0034]Mix 0.06 g of magnetic beads with ferric oxide as the magnetic core and 6.0 g of polyethylene glycol, add to 6 mL of double distilled water, and disperse for 30 minutes. The mixture was transferred to a shaker flask at room temperature with continuous stirring. 115 mL of dispersion medium ethanol / double distilled water (V:V=3:2) was added to the bottle. 8mL (0.68g) of acrolein, 15mL (13.5g) of styrene, 0.3mL (0.28g) of divinylbenzene, and 6.0g of benzoyl peroxide were added in sequence, and stirred for 30min. The temperature was raised to 65°C, and the reaction was continued for 10 hours to obtain a light brown emulsion. Wash several times with 0.1M hydrochloric acid and double distilled water, and separate by magnetic field. Aldehyde functionalized magnetic beads were obtained.

[0035] 2. Linkage of aldylated m...

Embodiment 2

[0037] Example 2 Connection of fluorescent quantum dots to human immunoglobulin IgG

[0038] 1-Ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC·HCL) was used as a cross-linking agent, N-hydroxysuccinimide (NHS) was used as a protective agent, and the Quantum dots modified with carboxyl groups on the surface (QDs-COOH) cross-link with the terminal amino groups on the antibody (Ab) to generate amide bonds, and then immune quantum dots are obtained. The molar ratio of antibody to quantum dot is 1:10 to 1:30.

[0039] The specific method is: 60ul (7.8*10 -6 mol) QDs (red light) stock solution was added to the mixture of phosphate buffer solution PBS (0.01mol / L, pH 7.2) and 6mg EDC·HCL (final concentration 20mg / mL), and the mixture was vortexed at room temperature for 5min. React for another 15-20min to fully activate the free carboxyl groups on the QDs; similarly, 1.5mg NHS (final concentration: 5mg / mL) was added to the above mixture, vortexed at room temperature fo...

Embodiment 3

[0040] The detection of embodiment 3 samples

[0041] The principle of detection is as figure 1 shown. The immunized superparamagnetic nano-magnetic beads prepared in Example 1 were dispersed in phosphate buffer (0.01 mol / L, pH=7.2) (concentration is mg / ml, and 50 μL of immuno-magnetic beads were used dropwise into 3mL different concentrations (10 5 cfu / mL, 10 4 cfu / mL, 10 3 cfu / mL, 10 2 cfu / mL, 10cfu / mL) in the sample liquid of Staphylococcus aureus. Shake at room temperature to fully react for 0.5h. Under the action of an external magnetic field, wash with phosphate buffer (0.01mol / L, pH 7.2) for 2 to 3 times, and dilute to 0.2mL to obtain "magnetic beads-Salmonella" Complex.

[0042] Then, 50uL of the immunized quantum dots prepared in Example 2 were added dropwise to the washed system, and shaken for 0.5h under a shaker at room temperature, so that the antigen on the surface of Staphylococcus aureus and the IgG antibody connected to the quantum dots were fully For ...

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Abstract

The invention relates to the detection field of microorganism foodborne pathogens and discloses a method for rapidly detecting and screening staphylococcus aureus, which adopts bio-functionalized superparamagnetic nano particles and immunized quantum dots to immunologically recognize target microorganisms to realize to specificity detection on the foodborne pathogens (staphylococcus aureus). According to the method, target bacteria can be rapidly separated from various samples, and enrichment is also performed efficiently; and besides, bacteria separated by the enrichment can be marked rapidly by fluorescence, so that the bacteria can be qualitatively identified by a fluorescence microscope and can be quantitatively detected by a fluorescence photometer. The method is convenient and simple to operate, and has high reliability and a low requirement on corollary equipment.

Description

technical field [0001] The invention relates to the field of detection of microbial food-borne pathogenic bacteria, and discloses a method of using biologically functionalized superparamagnetic nanoparticles and immunized fluorescent quantum dots to immunize and identify target microorganisms to achieve specific detection of food-borne pathogenic bacteria golden yellow Staphylococcus method. Background technique [0002] With the improvement of living standards, people pay more and more attention to the issue of food safety. Statistics from 2003 to 2004 show that the pathogenic factors of food poisoning in my country are microbial, chemical, and toxic animal and plant pathogens. Microbial pathogens are the main factor leading to food poisoning, and the number of people poisoned is the largest. Therefore, rapid and accurate detection and identification of pathogenic bacteria in food is an important prerequisite for timely and effective control and prevention of pathogenic b...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/569
Inventor 赵渝陈艳顾鸣
Owner SHANGHAI NORMAL UNIVERSITY
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