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Nitrofurantoin residue enzyme-linked immunoassay kit

An enzyme-linked immunosorbent assay and nitrofurantoin technology, which can be used in measurement devices, instruments, scientific instruments, etc., can solve the problems of unsuitability for on-site detection and promotion, and high detection costs

Inactive Publication Date: 2009-10-21
FOOD INSPECTION CENT OF CIQ SHENZHEN
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although high performance liquid chromatography and tandem mass spectrometry have the advantages of high sensitivity and qualitative and quantitative identification; however, high performance liquid chromatography and tandem mass spectrometry require expensive equipment and high detection costs
Not suitable for on-site detection and promotion, and detection of a large number of samples

Method used

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  • Nitrofurantoin residue enzyme-linked immunoassay kit
  • Nitrofurantoin residue enzyme-linked immunoassay kit
  • Nitrofurantoin residue enzyme-linked immunoassay kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0045] Embodiment 1: the preparation of antigen and antibody

[0046] 1 Preparation of immunogen

[0047] Add 50mg of m-carboxybenzaldehyde, 151mg of Hitein hydrochloride, and 7mL of pyridine into a 25mL round bottom flask, reflux overnight, evaporate the pyridine to dryness, add 6mL of water, adjust the pH to 1~2 with 1M HCl, collect the precipitate by filtration, and wash with water , and dried to obtain 72 mg of off-white solid.

[0048] Using the mixed anhydride method, 18.6 mg of the Hitein derivative A prepared above was dissolved in 1.8 mL of dimethylformamide, and 21 uL of N-methylmorpholine and 18 uL of isobutyl chloroformate were added under ice-cooling. The reaction was carried out under ice water cooling for 30 minutes.

[0049] Weigh 60mg bovine serum albumin, dissolve it in 3mL 0.1M, pH9.0 boric acid buffer, add 1.6mL dimethylformamide, add dropwise 1.2mL of the above mixed anhydride solution, react overnight at 4°C, dialyze against PBS for two days , change t...

Embodiment 2

[0062] Embodiment 2: Nitrofurantoin detection kit performance

[0063] 1 sensitivity

[0064] The minimum detection limit of the zero standard of this kit is 0.1ng / mL. Due to matrix effects, the lowest detection limit of blank fish and shrimp samples was 0.4ng / mL.

[0065] Statistical table of test results of blank fish meat ng / g

[0066]

[0067] Statistical table of determination results of blank shrimp meat ng / g

[0068]

[0069] 2 Cross-reactivity

[0070] To determine the cross-reactivity of monoclonal antibodies with the metabolite AOZ of furazolidone, the metabolite of furaltadone AMOZ, and the metabolite of nitrofurazone SEM, the metabolites are first derivatized with o-nitrobenzaldehyde, and then the half inhibitory concentration IC is determined 50 , the cross-reactivity was found to be less than 0.1%.

[0071] Cross Reaction Results

[0072] product name

IC 50 (ng / mL)

CR (cross-reactive

sex,%)

Heitin

0.9

...

Embodiment 3

[0088] Embodiment 3: the detection of nitrofurantoin in the sample

[0089] 1. Preparation of ELISA plate

[0090] Dilute the coated antigen to a certain concentration with coating buffer, add 100 μL to each well, incubate at 37°C for 2 hours or overnight, pour off the coating solution, wash three times with washing solution, and pat dry; then add 200 μL of blocking solution (containing 3% skimmed milk powder), incubated at 37°C for 2 hours, poured off the blocking solution, dried and stored in a sealed and dry place.

[0091] 2. Solution preparation

[0092] 1M NaOH: Weigh 4 grams of sodium hydroxide and dissolve in distilled water, add distilled water to make up to 100mL

[0093] 2M HCl: Take 17.2mL of concentrated hydrochloric acid and add distilled water to make up to 100mL

[0094] 0.1M K 2 HPO 4 : 2.28 g K 2 HPO 4 ·3H 2 O, add distilled water to 100mL

[0095] Derivatization reagent 10mM o-nitrobenzaldehyde: weigh 151mg o-nitrobenzaldehyde, add 100mL methanol to...

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Abstract

The invention discloses a nitrofurantoin residue enzyme-linked immunoassay kit which comprises a) an ELIAS plate, b) a standard solution, c) an antibody working solution, d) a cleaning solution, e) an enzyme marker, f) a substrate developing solution and g) a stopping solution, wherein the ELIAS plate is coated with nitrofurantoin metabolite AHD or coupling antigen of a derivative of the nitrofurantoin metabolite and a protein; and the standard solution is prepared by o-nitrobenzaldehyde which is dissolved in the dimethyl sulfoxide and then reacts with hydantoin derivative. The invention is mainly provided in a form of working solution, is characterized by convenient use, high specificity, high sensitivity, high precision, high accuracy and the like, and is applied to the detection of nitrofurantoin residues in animal food such as aquatic products and the like.

Description

technical field [0001] The invention belongs to the technical field of ELISA, and in particular relates to an ELISA detection kit for nitrofurantoin residues. Background technique [0002] Nitrofuran is a synthetic broad-spectrum antibiotic. Due to its low price and good effect, it has been widely used in the disinfection and sterilization of aquatic products, animals and poultry. In the 1990s, nitrofuran and its metabolites were found to be carcinogenic and teratogenic, and countries around the world began to ban the use of nitrofuran. At present, the standards adopted by food supervision agencies in various countries in the world are that nitrofuran metabolites cannot be detected, and the detection limit is 1ppb. [0003] After entering the body, nitrofuran is rapidly metabolized and difficult to detect, but the formed metabolites are stable in combination with proteins and can be stored for several weeks. The current international method is to detect residues of nitrofu...

Claims

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Application Information

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IPC IPC(8): G01N33/577G01N33/543G01N33/531
Inventor 朱海岳振峰叶卫翔范放汤慕瑾张恒郑晓燕
Owner FOOD INSPECTION CENT OF CIQ SHENZHEN
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