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Detection kit for ciprofloxacin and detection method thereof

A technology of ciprofloxacin and kits, applied in the direction of measuring devices, instruments, scientific instruments, etc., can solve the problems of limited application, expensive, cumbersome sample processing, etc., and achieve the effects of high sensitivity, simple structure, and convenient use

Inactive Publication Date: 2010-05-26
JIANSGU INST OF MICROBIOLOGY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Chromatographic detection is sensitive and accurate, requires expensive instruments and specialized staff, and sample processing is cumbersome, so its application is limited, especially in large-scale screening and on-site testing at the grassroots level

Method used

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  • Detection kit for ciprofloxacin and detection method thereof
  • Detection kit for ciprofloxacin and detection method thereof
  • Detection kit for ciprofloxacin and detection method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0012] Example 1: Preparation of kit and detection of food of animal origin

[0013] Preparation of immune antigen:

[0014] The immunizing antigen of the present invention combines ciprofloxacin with a macromolecular carrier protein by a mixed acid anhydride method, and uses synthetic CIP-BSA as an immunizing antigen to immunize animals.

[0015] The carrier protein is bovine serum albumin (BSA). Weigh 30mg~300mg bovine serum albumin, dissolve in 6mL~10mL of 50% dioxane aqueous solution, adjust the pH value to 9~10, and make A liquid; weigh 10mg~40mg CIP, dissolve in 2mL dioxane Set it at 4°C for 10 minutes, add 20 μL of tri-n-butylamine and 10 μL of isobutyl chloroformate in sequence, stir at 4°C for 30 minutes, and use it as liquid B. Add liquid A dropwise to liquid B at 4°C, and stir overnight. The reaction solution was dialyzed against distilled water for 72 hours, during which the distilled water was changed 5 times. Or purified by Sephadex G50 column and eluted with...

Embodiment 2

[0055] Implementation example 2 preparation of kit

[0056] CIP-BSA antigen preparation:

[0057] Weigh 12mg of CIP 40mg, N-hydroxysuccinamide (NHS), 35mg of carbodiimide (EDC) and dissolve in N,N'-dimethylformamide (DMF), stir at room temperature for 24h to prepare A solution; Dissolve 50mg of BSA in 3mL of 0.01mol / mL pH7.4 PBS to prepare solution B. Add solution A dropwise to solution B, stir while adding, react at room temperature for 3 hours, dialyze the reaction solution in distilled water for 72 hours, and change distilled water 5 times during the period. Or purified by Sephadex G50 column and eluted with 0.05mol / L pH8.0 carbonate buffer solution. Collect the dialysis purified liquid or the column eluate of the first peak, aliquot and store at -20°C for antigen immunization.

[0058] Preparation of polyclonal ciprofloxacin antibody: same as in Example 1.

[0059] Eu 3+ - Preparation of goat anti-rabbit antibody: same as in Example 1.

[0060] Solid-phase antigen pr...

Embodiment 3

[0067] The reagent provided by the kit is the same as that in Example 1, and is used to detect honey samples.

[0068] Reagents that should be prepared by the laboratory:

[0069] (1) distilled water or deionized water;

[0070] (2) Dichloromethane;

[0071] (3) 0.02mol / L PB buffer solution: weigh Na 2 HPO 4 12H 2 O 5.16g, NaH 2 PO 4 0.87g into the volumetric flask, add water to dissolve and make the volume to 1L;

[0072] (4) Phosphate buffer working solution: pH7.2, take 1.45g disodium hydrogen phosphate (containing 12 crystal water), 0.1g potassium dihydrogen phosphate (anhydrous), 8.0g sodium chloride, dissolve in water and set Dissolve to 1000mL.

[0073] Matters needing attention before determination: same as Example 1.

[0074] The specific detection steps are as follows:

[0075] Accurately weigh 1.0g±0.01g honey sample into a 50mL centrifuge tube, add 2mL of 0.02mol / L PB buffer, shake until the honey is completely dissolved, add 8mL of dichloromethane, shake...

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Abstract

The invention discloses a detection kit for ciprofloxacin and a detection method thereof, which belong to the technical field of TRFIA and are used for detecting CIP in animal derived food, blood, urine and fodder. The prepared kit adopts the TRFIA to detect the CIP, and the measurement base is labeled immune response. A micro-pore plate is clad with CIP-carrier protein; and CIP standard or sample and CIP antibody are added into the micro-pore plate. The free CIP and the CIP-carrier protein on the micro-pore plate compete for the CIP antibody, the unconnected CIP antibody is removed by washing, then Eu3+-goat anti-rabbit antibody is added into the micro-pore plate, and the unconnected Eu3+-goat anti-rabbit antibody is removed by washing after the labeled immune response. After the enhancement liquid is added, the fluorescence intensity cps is measured by using a time-resolved fluorescence instrument, the fluorescence intensity is in inverse proportion to the concentration of the CIP in the sample, and the content of the CIP in the detected sample can be determined by referring to a stand curve. The kit provided by the invention for detecting the CIP has the advantages of simple structure, using convenience, low price and high sensitivity, and can reach over 0.01ng / mL.

Description

technical field [0001] The invention relates to a detection kit for detecting ciprofloxacin (CIP) and a detection method thereof, belonging to the technical field of time resolved fluoroisnmunoassay (TRFIA), and used for animal-derived food, blood and urine And detection of ciprofloxacin drug residues in feed. Background technique [0002] Ciprofloxacin (CIP), also known as ciprofloxacin, belongs to the third generation of quinolone antibacterial synthetic drugs. It has a broad antibacterial spectrum and strong antibacterial activity. It is one of the most powerful drugs and has been widely used in human clinical and animal husbandry. Because this type of drug is easy to induce bacterial drug resistance, long-term application will cause cumulative poisoning, which can cause articular cartilage, photosensitivity, liver and kidney toxicity, genotoxicity leading to lymphocyte chromosome mutation, and human tendon rupture. As a kind of shared medicine for humans and animals, d...

Claims

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Application Information

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IPC IPC(8): G01N33/543G01N33/533
Inventor 陆茂林宓晓黎李利东黄丽俊李英张六六储敏
Owner JIANSGU INST OF MICROBIOLOGY
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