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Magnetic granule chemiluminescence kit for detecting nitrofurantoin metabolite and application of magnetic granule chemiluminescence kit

A technology of nitrofurantoin metabolites and chemiluminescence reagents, which is applied in the field of magnetic particle competition direct chemiluminescence detection kits, can solve the problems of reaction time and temperature, poor stability of reagents, etc., and achieve low detection time, high sensitivity and fast detection Effect

Inactive Publication Date: 2012-07-11
BEIJING KWINBON BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The reaction time and temperature have a great influence on the enzyme activity, and the stability of the reagent is poor

Method used

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  • Magnetic granule chemiluminescence kit for detecting nitrofurantoin metabolite and application of magnetic granule chemiluminescence kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] The preparation of the concrete component of embodiment 1 kit

[0037] 1. Preparation of luminescent markers

[0038] a) Synthesis of haptens

[0039] The nitrofurantoin metabolite and p-carboxybenzaldehyde are derivatized by reaction in water to obtain the nitrofurantoin metabolite derivative hapten.

[0040] Take nitrofurantoin metabolite 1000mg and dissolve in 12ml pure water. Dissolve 1000 mg of p-carboxybenzaldehyde in 26 ml of water, add it to the nitrofurantoin metabolite solution and react at room temperature for 48 hours, and obtain a pale yellow precipitate which is the nitrofurantoin metabolite derivative hapten. Repeatedly washing with 50ml of water for 5-6 times, drying to obtain the hapten of the derivative of nitrofurantoin metabolite.

[0041] b) Preparation of luminescent markers

[0042] Take 4.5mmol / L ABEI, dissolve it in 4ml distilled water, dissolve 5.0mmol / L N-hydroxysuccinimide in 0.5ml N,N-dimethylformamide, mix the two thoroughly and react a...

Embodiment 2

[0058] The formation of embodiment two kits

[0059] A magnetic particle chemiluminescent detection kit for detecting nitrofurantoin metabolites was constructed so that it contained the following components:

[0060] Fluorescent marker of FITC-labeled nitrofurantoin metabolite monoclonal antibody

[0061] Luminescent marker of ABEI-labeled nitrofurantoin metabolite hapten

[0062] Separation reagent of paramagnetic nanobeads coated with goat anti-FITC monoclonal antibody

[0063] Nitrofurantoin metabolite standard solution (0ng / ml, 0.01ng / ml, 0.03ng / ml, 0.09ng / ml, 0.27ng / ml, 0.81ng / ml), the standard dilution is pH7.4, containing 0.05% thimerosal Preservative, 0.1mol / LTRIS buffer. The percentage content is a mass percentage content.

[0064] The concentrations of the nitrofurantoin metabolite control solution were 0.02ng / ml and 0.5ng / ml respectively, and the quality control substance was diluted to pH 7.4, containing 0.05% thimerosal preservative, and 0.1mol / LTRIS buffer. ...

Embodiment 3

[0066] Detection of Nitrofurantoin Metabolites in Example Three Actual Samples

[0067] 1. Sample pretreatment

[0068] (1) Pretreatment methods for muscle, liver, and other tissues

[0069] Homogenize the sample with a homogenizer; take 1.0±0.05g of the homogenate into a 50ml polystyrene centrifuge tube; add 5ml of methanol-water solution, shake with a shaker for 5min, and centrifuge at room temperature (20-25°C) for 10min at 3000g or more , remove all supernatant; (this step can reduce sample matrix interference) add 4ml deionized water, vortex to dissolve with vortex, add 0.5ml 1M hydrochloric acid solution and 100μl derivatization reagent, shake fully with a shaker for 2min; Incubate overnight at 37°C (about 16h); add 5ml 0.1M dipotassium hydrogen phosphate solution, 0.4ml 1M sodium hydroxide solution and 5ml ethyl acetate respectively, shake vigorously with a shaker for 30s; over 3000g, room temperature (20-25°C) ) and centrifuge for 10 min; put 2.5 ml of ethyl acetate ...

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Abstract

The invention relates to a magnetic granule chemiluminescence kit for detecting nitrofurantoin metabolite, which comprises the following reagents: a luminous marker, a fluorescein marker, a standard substance, a quality control substance and a separation reagent. The luminous marker is nitrofurantoin metabolite hapten marked by an isoluminol luminous marker; the fluorescein marker is a nitrofurantoin metabolite monoclonal antibody marked by fluorescein or a fluorescein derivative; and the separation reagent is paramagnetic nano micro-beads enveloped in an anti-goat FITC (fluorescein isothiocyanate) monoclonal antibody. The invention further relates to a method adopting the kit to detecting the nitrofurantoin metabolite in animal derived food; and the method has higher sensitivity, specificity and detecting speed for nitrofurantoin metabolite.

Description

technical field [0001] The invention relates to a direct chemiluminescence detection kit and a testing method thereof, in particular to a magnetic particle competition direct chemiluminescence detection kit for detecting residual furaltadone metabolites in samples such as honey, aquatic products and tissues. technical background [0002] Nitrofuran drugs are synthetic broad-spectrum antibiotics, which have very good antibacterial effects and pharmacokinetic properties, and have been widely used as growth-promoting additives for livestock, poultry, and artificially farmed aquatic products. However, long-term experimental studies have found that both nitrofuran drugs and metabolites can cause cancer and gene mutations in experimental animals, and it can be inferred that humans have long-term use of such drugs or long-term consumption of poultry, Livestock and artificially raised aquatic products can also undergo canceration and gene mutations. Therefore, such drugs are prohib...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N21/76G01N33/577G01N33/533
Inventor 何方洋万宇平冯才伟刘平赵正苗张丽杰杨秀贤
Owner BEIJING KWINBON BIOTECH
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