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Reagent of multi-residue simultaneous rapid fluorescence detection of aminoglycoside antibiotics and application thereof

An aminoglycoside and fluorescence detection technology, which is applied in the field of nanomaterial preparation, can solve the problems of high cost, stability easily affected by temperature, and difficulty in detection with large batches of samples

Active Publication Date: 2019-10-08
JIANGSU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although the immunological method can realize the rapid detection of antibiotic residues, most of them are single residue detection, and the acquisition of antibodies requires animal experiments or cell culture, which is costly, and the stability is easily affected by temperature. detection

Method used

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  • Reagent of multi-residue simultaneous rapid fluorescence detection of aminoglycoside antibiotics and application thereof
  • Reagent of multi-residue simultaneous rapid fluorescence detection of aminoglycoside antibiotics and application thereof

Examples

Experimental program
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Effect test

Embodiment 1

[0033] Measure 5 μL each of QD1, QD2 and QD3 with concentrations of 21.11, 13.09 and 6.01 μmol / L, and add 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide at a concentration of 10 g / L respectively 10 μL hydrochloride, dilute to 40 μL with deionized water, and react QD1, QD2 and QD3 with 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride under ultrasonic vibration 4, 10 and 15 min. Then add KAPT, NAPT and TAPT which have been denatured at 95°C according to the ratio of KAPT:QD1 molar ratio 1:4, NAPT:QD2 molar ratio 1:5 and TAPT:QD3 molar ratio 1:5, and use phosphoric acid with pH 7.4 The volume of salt buffer solution was adjusted to 400 μL, and KAPT, NAPT, and TAPT were respectively reacted with activated QD1, QD2, and QD3 in the dark for 2 h. After the reaction, the solutions were filtered with 50kD ultrafiltration tubes, and the remaining solutions in the tubes were adjusted to 100 μL with phosphate buffer solution. In the KAPT-QD1, NAPT-QD2 and TAPT-QD3 conjugate sol...

Embodiment 2

[0036] Measure 10mL blank milk, add kanamycin, netilmicin and tobramycin mixed standard solution, then add 2mL 15% trichloroacetic acid solution, then ultrasonically shake the sample for 20min, centrifuge at 12000r / min for 10min, collect The supernatant was adjusted to 5 mL with deionized water, and the concentrations of kanamycin, netilmicin and tobramycin in it were 1, 1 and 10 μg / L, respectively. The concentrations of kanamycin, netilmicin and tobramycin were 1, 1, 10, 5, 5, 50, 10, 10, 100, 15, 15, 200, 20, 20, 500 μg / L 1 μL of the mixed standard solution series and the spiked milk sample were respectively drawn into 100 μL of the fluorescence detection reagent of the present invention, and reacted for 35 minutes. With 370nm as the excitation wavelength, measure the fluorescence intensity F at 555 and 607nm, 583 and 526nm, and 673 and 736nm for each solution after the reaction. 555nm and F 607nm , F 583nm and F 526nm and F 673nm and F 736nm . ΔF of each mixed standa...

Embodiment 3

[0038] Measure 10mL blank milk, add kanamycin, netilmicin and tobramycin mixed standard solution, then add 2mL 15% trichloroacetic acid solution, then ultrasonically shake the sample for 20min, centrifuge at 12000r / min for 10min, collect The supernatant was adjusted to 5 mL with deionized water, and the concentrations of kanamycin, netilmicin and tobramycin in it were 10, 10 and 100 μg / L, respectively. The concentrations of kanamycin, netilmicin and tobramycin were 1, 1, 10, 5, 5, 50, 10, 10, 100, 15, 15, 200, 20, 20, 500 μg / L 1 μL of the mixed standard solution series and the spiked milk sample were respectively drawn into 100 μL of the fluorescence detection reagent of the present invention, and reacted for 35 minutes. With 370nm as the excitation wavelength, measure the fluorescence intensity F at 555 and 607nm, 583 and 526nm, and 673 and 736nm for each solution after the reaction. 555nm and F 607nm , F 583nm and F526nm and F 673nm and F 736nm . Take the ΔF of each mi...

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Abstract

The invention relates to a preparation and using method of a reagent of multi-residue simultaneous rapid fluorescence detection of aminoglycoside antibiotics. Three kinds of quantum dots with the samematerial core and different emission wavelengths are coupled with an aptamer having the high specific recognition ability for kanamycin, netilmicin and tobramycin; the three kinds of coupling mattersand graphene oxide to form a fluorescence resonance energy transfer system; and a reagent capable of realizing simultaneous rapid fluorescence detection of kanamycin, netilmicin and tobramycin residues is prepared. With the provided reagent, rapid, accurate and sensitive detection of kanamycin, netilmicin and tobramycin residues in milk samples can be realized without depending on the separationanalysis technology; and the powerful technical guarantee is provided for the g food safety supervision enhancement.

Description

technical field [0001] The invention relates to the technical field of preparation of nanomaterials and the technical field of veterinary drug residue detection, in particular to a combination of high specificity of aptamers and excellent fluorescence properties of quantum dots, which can selectively label kanamycin, netilmicin and tobu A fluorescent detection reagent for mycin, and it was applied to the simultaneous and rapid fluorescent detection of multiple residues of kanamycin, netilmicin and tobramycin. Background technique [0002] Kanamycin, netilmicin and tobramycin are three aminoglycoside antibiotics that are widely used in the aquaculture industry. After long-term consumption of animal-derived foods containing their drug residues, human beings will lead to aminoglycoside antibiotics. Adverse effects such as ototoxicity, nephrotoxicity, and allergic reactions shared by antibiotics. In order to prevent food containing its residues from entering the market, it is n...

Claims

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Application Information

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IPC IPC(8): G01N33/94G01N33/58G01N21/64
CPCG01N33/9446G01N33/582G01N21/6428G01N2021/6432G01N2021/6439
Inventor 郭欣陈冠华宋尚红高志飞
Owner JIANGSU UNIV
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