Method for rapidly detecting enterobacter sakazakii

A rapid technology of Enterobacter sakazakii, applied in the field of microbial detection, can solve the problems of limited optical signal of colloidal gold, limited application, low sensitivity of colloidal gold immunochromatography test strips, etc., to improve detection sensitivity, reduce workload, Uniformity of immunological response

Active Publication Date: 2016-06-08
NANCHANG UNIV
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Problems solved by technology

[0004] Colloidal gold immunochromatographic test strips have become an important tool for basic screening due to their simple operation, rapidity (10-15min), and accuracy. However, due to the limited optical signal of colloidal gold, the sensitivity of colloidal gold immunochromatographic test strips is not high High, the limit of detection for pathogenic bacteria is usually not higher than 10 5 CFU / mL, this defect limits its application in food and agricultural product testing

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  • Method for rapidly detecting enterobacter sakazakii

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Embodiment 1

[0039] Embodiment one: using Fe 3 o 4 / Ru(bqy) 3 2+ Detection of Enterobacter sakazakii in milk by nano-microsphere immunochromatographic test strip

[0040] 1. Preparation of Fc-conjugated mAbs 3 o 4 / Ru(bqy) 3 2+ Nanospheres

[0041] 1.1 Fe 3 o 4 / Ru(bqy) 3 2+ Preparation of nanospheres: adding 0.6mmolFeCl 3 ·6H 2 O and 0.3 mmol FeCl 2 4H 2 O into 100mL of deionized water, nitrogen gas was passed into the solution and heated to 90°C, and then 4.7mL of 25% NH 3 ·H 2 O was added to the mixture and reacted for 2h. Use a permanent magnet to separate the black solid matter from the reaction solution and wash it with high-purity water for 3 to 5 times to obtain Fe 3 o 4 Nanoparticles; take 12mg Fe 3 o 4 Nanoparticles were resuspended with a mixture of 3mL deionized water and 20mL ethanol, and 0.5mL of NH 4 OH solution, then dissolve 50uL tetraethyl orthosilicate in 50uL ethanol solution and add dropwise, react at room temperature for 12h, in Fe 3 o 4 A laye...

Embodiment 2

[0051] Embodiment two: using Fe 3 o 4 / Ru(bqy) 3 2+ Detection of Enterobacter sakazakii in beef by nano-microsphere immunochromatographic test strip

[0052] 1. Preparation of Fc-conjugated mAbs 3 o 4 / Ru(bqy) 3 2+ Nanospheres

[0053] 1.1Fe 3 o 4 / Ru(bqy) 3 2+ Preparation of nanospheres: adding 0.6mmolFeCl 3 ·6H 2 O and 0.3 mmol FeCl 2 4H 2 O into 100mL of deionized water, nitrogen gas was passed into the solution and heated to 90°C, and then 4.7mL of 25% NH 3 ·H 2 O was added to the mixture and reacted for 2h. Use a permanent magnet to separate the black solid matter from the reaction solution and wash it with high-purity water for 3 to 5 times to obtain Fe 3 o 4 Nanoparticles; take 12mg Fe 3 o 4 Nanoparticles were resuspended with a mixture of 3mL deionized water and 20mL ethanol, and 0.5mL of NH 4 OH solution, then dissolve 50uL tetraethyl orthosilicate in 50uL ethanol solution and add dropwise, react at room temperature for 12h, in Fe 3 o 4 A layer...

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Abstract

The invention provides a method for rapidly detecting enterobacter sakazakii. According to the scheme, the method comprises the steps of enriching Fe3O4 / Ru(bqy)3<2+> nanoparticles with bacteria, preparing test strips, and mounting a sample for detection. By means of the method, the step of eluting enterobacter sakazakii away from immunomagnetic beads is omitted, and capturing efficiency is improved; the step of spraying Ru (bqy) 3<2+> nanoparticles to a binding pad is omitted, an immunology reaction is more uniform, and the variation coefficient generated in the quantitative detection process is small; working load is reduced, and the probability of sundry bacteria contamination is lowered. The detection scheme is quite high in sensitivity and quite good in stability.

Description

technical field [0001] The invention relates to the field of microbial detection, specifically using Fe 3 o 4 / Ru(bqy) 3 2+ Rapid detection of Enterobacter sakazakii using nanosphere integrated immunomagnetic bead capture technology and immunochromatography. technical background [0002] Enterobacter sakazakii is a Gram-negative non-bacillus bacterium parasitic in the intestinal tract of humans and animals, and is an important food-borne bacterium. It infects mostly infants, especially premature, low birth weight or immunocompromised newborns. Formula milk powder is the main source of infection in infants and young children. Enterobacter sakazakii can cause severe bacteremia, necrotizing enterocolitis, and meningitis in newborns. Although the incidence of Enterobacter sakazakii is not high, the mortality rate can reach 50%-80%. Although most cases of E. sakazakii infection have been associated with infants, recent studies have found that the bacteria can also infect a...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/543G01N33/569
CPCG01N33/54326G01N33/56916
Inventor 赖卫华王景云刘道峰邓省亮山珊熊勇华魏华
Owner NANCHANG UNIV
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