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Intracellular fluorescence labeling method of food-derived probiotics and in-vivo application thereof

A labeling method and probiotic technology, applied in the research field of food-derived probiotics, can solve the problems of low sensitivity and short emission wavelength, and achieve the effects of simple steps, low cost, good stability and labeling reproducibility

Inactive Publication Date: 2019-06-28
NANKAI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, these two methods still have problems such as low sensitivity and short emission wavelength, which have certain limitations when applied to in vivo imaging.

Method used

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  • Intracellular fluorescence labeling method of food-derived probiotics and in-vivo application thereof
  • Intracellular fluorescence labeling method of food-derived probiotics and in-vivo application thereof
  • Intracellular fluorescence labeling method of food-derived probiotics and in-vivo application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0055] A method for intracellular fluorescent labeling of food-derived probiotics

[0056] (1) Preparation of DNA-coated long-lasting fluorescent nanoprobes.

[0057] Mix 40mg DNA with 10mL PBS and gently stir, add 40mg EDC and 40mgNHS, stir well, react at room temperature for 30 minutes, weigh accurately, add 10mg long persistence nanoprobes (ZGGO PLNPs), react at room temperature for 30 minutes, and finally wash with PBS Wash and purify, and store away from light.

[0058] (2) Preparation of competent probiotic bacteria that are easy to ingest and accommodate foreign DNA.

[0059] Cultivate probiotics overnight at 37°C, transfer 25mL to 100mL MRS medium containing 1% glycine, culture to logarithmic phase (OD600=0.6), 4°C, use 3000g centrifugal force, centrifuge for 5min, collect the bacteria, and place on ice 10min, with 100mL pre-cooled 10mM MgCl 2 Wash once, then wash with pre-cooled 3.5mM MgCl 2 (Containing sucrose at a concentration of 925mM) solution was washed twic...

Embodiment 2

[0064] A method for intracellular fluorescent labeling of food-derived probiotics. The steps and method are basically the same as in Example 1, except that the probiotics are Bifidobacterium lactis V9, and correspondingly fluorescently labeled Bifidobacterium lactis V9 is obtained.

Embodiment 3

[0066] A method for intracellular fluorescent labeling of food-derived probiotics, the steps and method are basically the same as in Example 1, the difference is that the probiotics are Lactobacillus reuteri, and fluorescently labeled Lactobacillus reuteri is obtained accordingly.

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Abstract

The invention relates to an intracellular fluorescence labeling method of food-derived probiotics. The method comprises the following steps that a, a long-afterglow nano-luminescent material is modified by using a DNA; b, a probiotic competence is prepared; c, a DNA coated nano fluorescent probe is electrically transferred to the probiotics; d, the nano fluorescent probe is orally taken into a mouse body for imaging. The method has the advantages that the method for using the nano-luminescent material for intracellular fluorescence labeling is achieved for the first time, complex surface functionalization and antibody modification of a fluorescent material are not needed, the steps are simple and convenient, the cost is low, and compared with surface marking, the method has better stability and marking reproducibility. According to a near-infrared fluorescence labeling probiotic body in-vivo biological imaging technology developed through the method, the long-afterglow nano-luminescentproperty inside thalli can be used, metabolic behaviors and distribution conditions of probiotic bodies in a biological body can be monitored losslessly in situ in real time, and an innovative research means is provided for functional sites and nutriology of the food-derived probiotics.

Description

technical field [0001] The invention belongs to the research field of food-derived probiotics, and relates to a method for intracellular fluorescent labeling of food-derived probiotics and its application in vivo. Background technique [0002] The Food and Agriculture Organization of the United Nations and the World Health Organization point out that probiotics are a type of living microorganisms that, when ingested in a certain amount, have beneficial effects on the health of the host. Studies have shown that probiotics can resist the invasion of pathogenic bacteria, inhibit the reproduction of pathogenic bacteria, improve the barrier function of intestinal epithelial cells, maintain the homeostasis of the gastrointestinal tract, and rebuild the stability of intestinal flora. Probiotics play a beneficial role mainly through the production of dominant flora, biological oxygen capture and biological antagonism, and through the production of beneficial substances and antibacte...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K49/00A61B5/00C09K11/02
Inventor 刘敬民王硕
Owner NANKAI UNIV
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