Enzyme-linked immunalogical kit for detecting gentamicin medicine and method

An enzyme-linked immunosorbent reagent and gentamicin technology, applied in the field of immunological detection, can solve the problems of complex instruments and equipment, unsuitable for on-site monitoring and screening of a large number of samples, cumbersome process, etc., to overcome technical problems, preservation and good stability effect

Active Publication Date: 2007-08-22
BEIJING WANGER BIOTECH
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  • Abstract
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  • Claims
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Problems solved by technology

[0003] The current chemical method for detecting gentamicin residues is high-performance liquid chromatography, which is not...

Method used

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  • Enzyme-linked immunalogical kit for detecting gentamicin medicine and method

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0061] The preparation of embodiment 1 kit components

[0062] 1. Antigen Synthesis

[0063] a. Immunogen synthesis

[0064] Gentamicin and ovalbumin were coupled by the glutaraldehyde method to obtain the immunogen.

[0065] The preparation process of immunogen: get gentamicin 5mg and dissolve (I liquid) with 0.5ml water; Get 2.5% glutaraldehyde 0.1ml and add in the solution (I liquid), activate reaction 30min; Get ovalbumin 18mg with 2.4 ml of water was dissolved and added to the solution (II solution), stirred and reacted overnight to obtain the immunogen.

[0066] b. Preparation of coated original gentamicin-conjugated antigen

[0067] The gentamicin and hemocyanin were coupled by the glutaraldehyde method to obtain the coated antigen.

[0068] The preparation process of the original coating: take 5 mg of gentamicin and dissolve it in 0.5 ml of water (I solution); get 0.1 ml of 2.5% glutaraldehyde and add it into the solution (I solution), and activate the reaction for...

Embodiment 2

[0083] Embodiment 2 detects the formation of the ELISA kit of gentamicin

[0084] Set up the enzyme-linked immunosorbent assay kit for detecting gentamicin to include the following components:

[0085] (1) A microtiter plate coated with gentamicin-conjugated antigen;

[0086] (2) Goat anti-mouse anti-antibody labeled with horseradish peroxidase;

[0087] (3) gentamicin monoantibody concentrate;

[0088] (4) 6 bottles of gentamicin standard solution, the concentrations are 0 μg / L, 0.1 μg / L, 0.3 μg / L, 0.9 μg / L, 2.7 μg / L, 8.1 μg / L;

[0089] (5) The substrate chromogenic solution is composed of A liquid and B liquid, the substrate chromogenic liquid A liquid is carbamide peroxide, and the substrate chromogenic liquid B liquid is tetramethylbenzidine;

[0090] (6) The stop solution is 2mol / L hydrochloric acid;

[0091] (7) The concentrated washing solution is 0.01M, pH7.4, and contains 0.8%-1.2% Tween 20 and 0.5‰ sodium azide preservative phosphate buffer;

[0092] (8) The con...

Embodiment 3

[0093] The detection of gentamicin residue in the sample of embodiment 3

[0094] 1. Sample pretreatment

[0095] Animal tissue (chicken, chicken liver, pork, pork liver)

[0096] Weigh the fat-removed pulverized sample and mix it with 8ml 0.2M PB buffer for 5 minutes, put it in a water bath at 50°C for 30 minutes, centrifuge it at room temperature for 10 minutes above 3000g, take 50ml of the supernatant, add 450ml of the diluted reconstitution solution and mix, take 50μl for analysis.

[0097] 2. Detection with kit

[0098] Add 50 μl of gentamicin standard solution or sample solution to the microwells of the gentamicin-conjugated antigen-coated microtiter plate, then add 50 μl of gentamicin monoclonal antibody working solution, and seal the plate with a cover film. React in a 37°C incubator for 30 minutes, pour out the liquid in the wells, add 250 μl of washing solution to each well, pour out the liquid in the wells after 30 seconds, and repeat this operation to wash the p...

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Abstract

The invention provides an enzyme linked immune regent box to detect gentamycin. It includes: the enzyme labeled board which encrusts the encrusting substance, the enzyme marker, the special antibody for the gentamycin (when the antigen is on enzyme labeled board and the enzyme marker is the enzyme labeled antiantibody or the antiantibody is in the enzyme labeled board and the enzyme marker is the enzyme labeled antigen), the gentamycin standard solution, the substrate color solution, the stop solution, the condense washing solution, the condense recovery solution. The invention also discloses the method which includes the process: first to pre-treat the sample, then to detect by the regent box and last to analyze the detecting result. The invention is to detect the gentamycin residue weight in the animal food such as the chicken, chicken liver, pork, pork liver, milk powder, egg and feedstuff. The method is simple, low cost and has high sensitivity, also it is proper for detect the mass sample.

Description

technical field [0001] The invention relates to the field of immunological detection, in particular to an enzyme-linked immunoimmunoassay kit for detecting gentamicin drugs in animal-derived foods such as chicken, chicken liver, pork, pig liver, milk powder, eggs and feedstuffs and its detection method. Background technique [0002] Gentamicin (Gentamicin) is an aminoglycoside antibiotic, which is widely used in the treatment of animal diseases. Due to its neurotoxicity and nephrotoxicity, its residue in animal food will affect human health. European and American countries and my country require Its use is limited. [0003] The current chemical method for detecting gentamicin residues is high-performance liquid chromatography, which is not suitable for on-site monitoring and screening of a large number of samples due to complicated equipment and cumbersome process. Contents of the invention [0004] (1) Technical problems to be solved [0005] The purpose of the present ...

Claims

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Application Information

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IPC IPC(8): G01N33/569G01N33/543G01N33/577G01N33/535
Inventor 何方洋万宇平冯才伟吴小平郭勇朱亮赵正苗汪善良冯才茂刘平陈炜琳刘福林冯月君余厚美蒲小容罗贵昆刘玉梅
Owner BEIJING WANGER BIOTECH
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