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Furantoin metabolite enzyme linked immunosorbent analytical reagent casing and uses thereof

A technology of nitrofurantoin metabolites and enzyme-linked immunological reagents, which is applied in the field of immunoassays, can solve problems such as cumbersome processes, unsuitability for on-site monitoring and screening of a large number of samples, complex instruments and equipment, etc., to overcome technical problems, reduce the lower limit of sample detection, Effect of Improving Detection Sensitivity

Active Publication Date: 2007-08-08
BEIJING WANGER BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The most common methods used to detect nitrofurantoin metabolites are LC-UV, LC-MS and LC-MS / MS, which are not suitable for on-site monitoring and screening of a large number of samples due to complex instruments and cumbersome processes

Method used

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  • Furantoin metabolite enzyme linked immunosorbent analytical reagent casing and uses thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0066] Embodiment 1 Preparation of kit components

[0067] 1. Antigen Synthesis

[0068] a. Synthesis of nitrofurantoin metabolite derivative hapten

[0069] The nitrofurantoin metabolite and p-carboxybenzaldehyde are derivatized by reaction in water to obtain the nitrofurantoin metabolite derivative hapten.

[0070] Preparation process of nitrofurantoin metabolite derivative hapten:

[0071] 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 light yellow precipitate which is the nitrofurantoin metabolite derivative hapten. The nitrofurantoin metabolite derivative hapten is obtained by washing repeatedly 5-6 times with 50 ml of water and drying.

[0072] b. Immunogen synthesis

[0073] The immunogen is obtained by coupling the nitrofurantoin metabolite derivative hapten and hemocyanin by ...

Embodiment 2

[0096] Example 2 Construction of an enzyme-linked immunosorbent assay kit for detecting nitrofurantoin metabolites

[0097] An enzyme-linked immunosorbent assay kit for detecting nitrofurantoin metabolites was constructed to include the following components:

[0098] (1) A microtiter plate coated with an anti-nitrofurantoin metabolite-coupled antigen;

[0099] (2) Goat anti-mouse anti-antibody labeled with horseradish peroxidase;

[0100] (3) Nitrofurantoin metabolite monoclonal antibody working solution;

[0101] (4) 6 bottles of nitrofurantoin standard solution, the concentrations were 0 μg / L, 0.1 μg / L, 0.3 μg / L, 0.9 μg / L, 2.7 μg / L, 8.1 μg / L;

[0102] (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;

[0103] (6) The stop solution is 2mol / L hydrochloric acid;

[0104] (7) The concentrated washing solution is 0....

Embodiment 3

[0106] Example 3 Detection of Nitrofurantoin Metabolite Residues in Samples

[0107] 1. Sample pretreatment

[0108] Animal tissues (chicken, pork, fish and shrimp)

[0109] Take 1±0.05g tissue sample homogenate, add 4ml distilled water, 0.5ml 1M HCl and 100μl 10mM 2-nitrobenzaldehyde, shake well; incubate overnight at 37°C (about 16h); add 5ml 0.1M K 2 HPO 4 , 0.4ml 1M NaOH and 5ml ethyl acetate, shake vigorously for 30s; centrifuge at room temperature (20-25℃ / 68-77) above 3000g for 10min; take out 2.5ml ethyl acetate into another container and blow with nitrogen at 50℃ Dry or evaporate to dryness with a rotary evaporator; dissolve the dry matter with 1ml of n-hexane, mix well with 1ml of the diluted complex solution; centrifuge at room temperature (20-25℃ / 68-77) above 3000g for 10min; use 50μl of the lower layer liquid for analysis.

[0110] 2. Detection with kit

[0111] Add 50 μl of a series of standard solution or sample solution to microwells of the microtiter pla...

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Abstract

The invention provides a detecting nitrofurantoin metabolite ELISA kit, which contains: an ELISA plate covered by the original coated, enzyme marker, nitrofurantoin metabolite specific antibody or nitrofurantoin metabolite derivative antibody, nitrofurantoin metabolite standard solution or nitrofurantoin metabolite derivative standard solution, the substrate color solution, the termination solution, the condensed washing solution, the condensed complex solution. The invention also provides a method to apply the above ELISA kit detecting nitrofurantoin metabolite, which includes steps: firstly, sample pre-treatment, and then using the kit for testing, and finally analyzing testing results. The invention is to provide the nitrofurantoin metabolite residues in the ELISA kit for detection of animal derived foods, such as chicken, pork, fish, shrimp, milk, honey, egg, and other samples, and the detection method is simple, low cost, high sensitivity, and it can monitor on the scene and suitable for screening large number of samples.

Description

technical field [0001] The invention relates to the field of immunoassay, in particular to an enzyme-linked immunoassay kit for detecting residues of nitrofurantoin metabolites in animal-derived foods and an application thereof. Background technique [0002] Nitrofuran drugs have been widely used as growth-promoting additives for poultry, aquatic products and pigs because of their excellent antibacterial and pharmacokinetic properties. However, in the course of long-term experimental research, it was found that both nitrofuran drugs and metabolites can cause cancer and gene mutation in experimental animals, so these drugs are prohibited from being used in treatment and feed. Nitrofurantoin was banned in 1995. [0003] Since nitrofuran drugs can be metabolized quickly in the body, and the metabolites combined in tissues can persist for a long period of time, it is often necessary to analyze the metabolized products when analyzing the residues of such drugs. The management d...

Claims

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Application Information

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IPC IPC(8): G01N33/543G01N33/577G01N33/566G01N33/535
Inventor 沈建忠冯才伟何方洋万宇平冯才茂吴小平赵正苗汪善良余厚美屈晓丽罗贵昆刘平陈炜琳丁双阳张素霞江海洋
Owner BEIJING WANGER BIOTECH
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