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Amikacin and kanamycin two-in-one fast detection enzyme linked immunosorbent assay kit and application thereof

A technology of amikacin and enzyme-linked immunosorbent reagent, which is applied in the field of enzyme-linked immunosorbent assay, can solve the problems of complicated pretreatment, complicated sequence, high cost, high cost and the like, and achieves a detection method with high efficiency, convenient portability and simple structure. Effect

Active Publication Date: 2017-06-20
四川精卫食品检测科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Traditional detection methods include microbiological assay and high performance liquid chromatography. These methods are used for the detection of amikacin or kanamycin in eggs or chicken samples. The pretreatment is tedious and complicated, and the determination cycle is long. Requires expensive instruments and high cost, which cannot meet the real-time and rapid detection of amikacin and kanamycin in eggs and chicken samples

Method used

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  • Amikacin and kanamycin two-in-one fast detection enzyme linked immunosorbent assay kit and application thereof
  • Amikacin and kanamycin two-in-one fast detection enzyme linked immunosorbent assay kit and application thereof
  • Amikacin and kanamycin two-in-one fast detection enzyme linked immunosorbent assay kit and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0017] Example 1 Amikacin and Kanamycin Two-in-One Rapid Detection ELISA Kit

[0018] Amikacin and kanamycin two-in-one rapid detection ELISA kit, which includes:

[0019] 1) A 96-well or 48-well microtiter plate coated with an amikacin hapten-ovalbumin conjugate;

[0020] 2) The enzyme-labeled anti-antibody is a goat anti-mouse anti-antibody labeled with horseradish peroxidase;

[0021] 3) 6 bottles of Kanamycin standard solution, the concentrations are 0μg / L, 0.5μg / L, 1.5μg / L, 4.5μg / L, 13.5μg / L, 40.5μg / L;

[0022] 4) The substrate chromogenic solution is composed of a substrate chromogenic solution A and a substrate chromogenic B solution, the substrate chromogenic solution A is carbamide peroxide, and the substrate chromogenic solution B is tetramethylbenzidine;

[0023] 5) The stop solution is 2mol / L hydrochloric acid;

[0024] 6) The washing solution is a Tris buffer solution containing 0.5-1.5‰ Triton X-100 and 0.5‰ Proclin300, wherein the concentration of the Tris bu...

Embodiment 2

[0026] Example 2 Preparation of Main Components of Amikacin and Kanamycin Two-in-One Rapid Detection ELISA Kit

[0027] 1. Synthesis of immunogen and coater

[0028] 1) Synthesis of immunogen

[0029] Weigh 20 mg of BSA to fully dissolve it in 3 mL of 0.05 mol / L MES (pH 6.0) buffer solution, then quickly add 50 mg of EDC and 50 mg of NHS, and react in a shaker for 12 hours at 4°C in the dark to obtain solution Ⅰ; Add 50mg of amikacin standard substance into 1.5mL 0.05mol / L MES (pH6.0) buffer solution, fully dissolve to make amikacin solution; add amikacin dropwise to solution Ⅰ Stir the kanamycin solution while adding, avoid light, and stir overnight at 4°C; pass through the G-25 desalting column once, put the desalted product into the treated dialysis bag, and place it in a 0.1mol / L In the PBS dialysate, the dialysate was changed every 4 hours for 2 days, and the dialyzed product was divided into 1.5 mL centrifuge tubes.

[0030] 2) Coating original synthesis

[0031] Wei...

Embodiment 3

[0052] The detection of amikacin and kanamycin in embodiment 3 eggs, chicken sample

[0053] 1. Sample pretreatment method

[0054] egg sample

[0055] 1) Stir the egg white and egg yolk thoroughly to make them evenly mixed;

[0056] 2) Weigh 1.0±0.05g sample into a 10mL polystyrene centrifuge tube, add 1mL 3% trichloroacetic acid solution, 1mL chloroform and 2mL ethyl acetate respectively, vortex for 3min, 3000g at room temperature (20-25℃) Centrifuge for 5min;

[0057] 3) Pipette 400 μL of the supernatant into a 2 mL polystyrene centrifuge tube, add 600 μL of 0.05 mol / L citrate buffer (pH 6.0), vortex for 1 min, and mix well; take 50 μL for analysis.

[0058] chicken sample

[0059] 1) Weigh 1.0±0.05g of the homogenized chicken sample into a 10mL polystyrene centrifuge tube, add 4ml of 0.05mol / L citric acid buffer (pH6.0), vortex for 3min with a vortexer, 3000g at room temperature (20 -25°C) centrifuge for 5min;

[0060] 2) Pipette 100 μL of the supernatant into a 2 mL...

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Abstract

The invention provides an amikacin and kanamycin two-in-one fast detection enzyme linked immunosorbent assay kit, which comprises an enzyme labeling plate, an enzyme labeling antibody, a amikacin standard product, substrate color developing liquid, terminating liquid, washing liquid and an amikacin antibody, wherein amikacin hapten-protein conjugates are coated on the enzyme labeling plate. The kit disclosed by the invention can be used for qualitative and quantitative detection of residue of amikacin and kanamycin in egg and chicken samples. According to the method for detecting amikacin and kanamycin in the egg and chicken sample, firstly, the egg and chicken sample to be tested is treated; then, the enzyme linked immunosorbent assay kit is used for detection; finally, a detection result is analyzed. The enzyme linked immunosorbent assay kit provided by the invention has the advantages that the structure is simple; the use is convenient; the carrying is convenient; the detection method is efficient, accurate, simple and convenient; the kit is suitable for the qualitative and quantitative detection of mass samples.

Description

technical field [0001] The invention relates to enzyme-linked immunosorbent detection technology, in particular to a two-in-one rapid detection enzyme-linked immunosorbent kit for amikacin and kanamycin, which is especially suitable for amikacin and kanamycin in eggs and chicken Detection of kanamycin drug residues. [0002] technical background [0003] Amikacin (Amikin) is also known as Amikacin, see figure 1 , is the sulfate salt of a semisynthetic derivative of kanapenicillin a. Kanamycin is a protein biosynthesis inhibitor, which can bind to the ribosomal 30s subunit in bacterial cells, resulting in misreading of mRNA, to inhibit the synthesis of bacterial proteins and inhibit bacterial growth. It is mainly used clinically for Qingda Various infections caused by kanamycin, kanamycin-resistant Gram-negative bacilli such as Escherichia coli, Proteus and Pseudomonas aeruginosa. Long-term or repeated large-scale intake of amikacin can lead to vestibular nerve and auditory...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/53G01N33/531G01N33/543
CPCG01N33/53G01N33/531G01N33/54306
Inventor 杨绍元杨少琼
Owner 四川精卫食品检测科技有限公司
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