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A method for quantitative detection of β-lactoglobulin in milk powder

A technology for quantitative detection of lactoglobulin, applied in the field of fluorescent sensing technology, biological analysis and detection, and nanomaterials, to achieve good water solubility, high sensitivity, and good selectivity

Active Publication Date: 2019-07-26
大庆市绿叶乳品有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In recent years there have been many aptamer-based assays for the detection of various target molecules, but to the best of our knowledge, there is currently no carbon dot-based fluorescent aptamer sensing platform for the detection of β-lactoglobulin

Method used

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  • A method for quantitative detection of β-lactoglobulin in milk powder
  • A method for quantitative detection of β-lactoglobulin in milk powder
  • A method for quantitative detection of β-lactoglobulin in milk powder

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0037] Example 1 Preparation of carboxylated ferroferric oxide magnetic nanoparticles, the steps are as follows:

[0038] 0.54g of ferric chloride hexahydrate and 1.44g of sodium acetate were dissolved in 10mL of ethylene glycol. Subsequently, the sodium acetate solution was slowly added dropwise to the ferric chloride solution under stirring. After stirring for 30 minutes, it was placed in a high-pressure reactor, reacted at 200°C for 4h, cooled to room temperature, washed several times with ethanol and deionized water, and dried in vacuum at 40°C for 12h. For the Fe 3 o 4 To modify the carboxyl group on the surface, ultrasonically dissolve 10.5 g of citric acid monohydrate in 100 mL of ultrapure water, and add 0.5 g of Fe prepared above 3 o 4 , and stirred mechanically at room temperature for 4h. Finally, the resulting product was washed several times with deionized water and ethanol, and dried under vacuum at 40°C to obtain the carboxyl-modified Fe 3 o 4 magnetic nan...

Embodiment 2

[0039] Example 2 The preparation of carbon dot CDs, the steps are as follows:

[0040] Take 1.2g of citric acid monohydrate, 0.6mL of diethylenetriamine and 20mL of ultrapure water in a 30mL autoclave, ultrasonically dissolve, and react at 200°C for 4h; take out the autoclave and place it at room temperature to obtain a brown solution, which is CDs solution; in order to purify CDs, the resulting CDs solution was concentrated, added acetone for purification and then dried in vacuum at 50°C; the obtained CDs were light yellow powder. Its transmission electron microscope picture, ultraviolet absorption and fluorescence excitation and emission picture are as follows figure 2 shown.

Embodiment 3

[0041] Embodiment 3 prepares Fe 3 o 4 -Aptamer complex, the steps are as follows:

[0042] Carboxy-modified Fe 3 o 4 Magnetic nanoparticles were dissolved in acidic PBS and prepared into 0.1, 0.2, 0.5, 1.0 mg / mL solutions, EDC and NHS were added to the above solution, and activated at room temperature for 30 min; the pH was adjusted to weak alkalinity with 1M NaOH, and appropriate Ligand, incubate overnight at room temperature. Finally, magnetic separation, washed with PBS to remove excess aptamers and impurities, redissolved in PBS for later use, and investigated carboxy-modified Fe 3 o 4 The effect of the concentration of magnetic nanoparticles on the response of the target, the results are as follows image 3 As shown in A.

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Abstract

The invention discloses a method for quantitatively detecting beta-lactoglobulin in milk powder. The method comprises: conjugating carboxylated Fe3O4 magnetic nanoparticles and a beta-LG aptamer to form a Fe3O4-aptamer; synthesizing carbon quantum dots (CDs) by using citric acid as a carbon source and using diethylenetriamine as a passivator, and conjugating the carbon quantum dots and the complementary strand of the beta-LG aptamer through an amide bond to form a fluorescent labeled probe CDs-cDNA; and carrying out hybridization on the Fe3O4-aptamer and the CDs-cDNA to form a complex Fe3O4-aptamer-cDNA-CDs, adding beta-LG to make the beta-LG and the aptamer in the Fe3O4-aptamer-cDNA-CDs complex be combined so as to separate and release part of the CDs-cDNA into the supernatant, and quantitatively determining the beta-LG content by detecting the fluorescence intensity of the supernatant. The method of the present invention has advantages of high sensitivity, good selectivity, and easydetection.

Description

technical field [0001] The invention belongs to the fields of nanomaterials, fluorescent sensing technology and biological analysis and detection, in particular to a method for quantitatively measuring β-lactoglobulin (β-LG) in milk powder with a fluorescent aptamer sensing platform based on magnetic nanoparticles and carbon dots. Background technique [0002] In today's society, food allergy is one of the food safety issues that people are concerned about. Among them, milk protein allergy is the most common allergic reaction in early childhood. According to statistics, in the first few years after birth, the milk protein allergy rate will reach 2-3%. β-lactoglobulin (β-LG) is considered the main allergenic protein in milk, even at low concentrations, it can cause allergic reactions, symptoms include: gastrointestinal lesions, atopic dermatitis, urticaria , allergic rhinitis, asthma, angioedema, chronic cough, etc. In order to reduce the incidence of milk allergy, relevan...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N33/68G01N33/543G01N33/533
Inventor 胡琴时梦岚许贯虹魏芳弟岑瑶徐晓曼程霞柴煜莹
Owner 大庆市绿叶乳品有限公司
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