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Enzyme linked immunosorbent assay kit for detecting omethoate, and application thereof

A technology of omethoate and enzyme-linked immunosorbent assay, which is applied in measurement devices, instruments, scientific instruments, etc., can solve the problems of unseen application and low accuracy, and achieves simple sample pretreatment process, high accuracy, and high accuracy. Handling less demanding effects

Inactive Publication Date: 2017-04-26
JIANGSU WISE SCI & TECH DEV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The advantage of the enzyme inhibition method is that it is easy to operate, fast, does not require expensive instruments, and is suitable for on-site detection and screening of a large number of samples. The disadvantage is that the accuracy is low, and the repeatability and recovery rate need to be improved. Qualitative detection of organophosphorus pesticide residues in vegetables, fruits and food, rapid and quantitative detection has not yet been applied

Method used

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  • Enzyme linked immunosorbent assay kit for detecting omethoate, and application thereof
  • Enzyme linked immunosorbent assay kit for detecting omethoate, and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0035] Embodiment 1 Preparation of kit components

[0036] 1. Preparation of omethoate hapten

[0037] 1) Get 20mg omethoate, 10ul, 1,3-propanediamine, and dissolve the catalytic amount of 4-dimethylaminopyridine in 2ml N,N'-dimethylformamide to obtain liquid I;

[0038] 2) Dissolve 20mg of N,N'-dicyclohexylcarbodiimide in 0.5ml of DMF to obtain liquid II;

[0039] 3) Slowly add solution II to solution I dropwise at 0°C, and continue to react for 20 h after returning to room temperature;

[0040] 4) evaporate solvent, column chromatography (eluent: dichloromethane / methanol, volume ratio 20:1), obtain omethoate hapten, such as figure 1 .

[0041] 2. Antigen preparation

[0042] Immunogen preparation—the omethoate hapten was coupled with bovine serum albumin (BSA) to obtain the immunogen.

[0043] 1) Take 5.3 mg of omethoate hapten and dissolve it with 0.5 ml DMF to obtain liquid I;

[0044] 2) Dissolve 30mg of BSA in 2.0ml, pH7.0, 0.1mol / L phosphate buffer solution to obt...

Embodiment 2

[0065] Example 2 The formation of the ELISA kit for detecting omethoate

[0066] An enzyme-linked immunosorbent assay kit for detecting omethoate is set up to include the following components:

[0067] (1) a microtiter plate coated with omethoate-coupled antigen;

[0068] (2) 6 bottles of omethoate standard solution, the concentration is respectively 0ug / L, 1ug / L, 3ug / L, 9ug / L, 27ug / L, 81ug / L.

[0069] (3) concentrated enzyme conjugate;

[0070] (4) enzyme conjugate dilution;

[0071] (5) Substrate chromogenic solution is made up of substrate liquid A liquid and substrate liquid B liquid, and substrate liquid A liquid is carbamide peroxide, and substrate liquid B liquid is tetramethylbenzidine;

[0072] (6) The stop solution is 2mol / L sulfuric acid.

Embodiment 3

[0073] The detection of omethoate in the sample of embodiment 3

[0074] 1. Sample pretreatment

[0075] Homogenize the vegetable sample with a homogenizer; weigh 5.0g±0.05g of the homogenized vegetable sample into a 50ml polystyrene centrifuge tube, add 25ml of 50% methanol, shake vigorously with an oscillator for 5min, over 3000r, at room temperature ( Centrifuge at 20-25℃ / 68-77℉) for 5min; take 500ul supernatant to a 2ml polystyrene centrifuge tube, add 500ul10% sodium chloride aqueous solution and shake for 1min with a shaker, mix well; take 20ul for analyze.

[0076] 2. Detection with kit

[0077] Add 20 μl of ochratoxin A standard solution / sample to the microwell of the enzyme plate coated with omethoate-coupled antigen, and then add 100 μl of enzyme conjugate working solution (use enzyme conjugate for the concentrated enzyme conjugate Dilute the diluent according to the volume of 1:20), seal the plate with a cover film, and react in the dark at 25°C for 10 minutes,...

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Abstract

The invention provides an enzyme linked immunosorbent assay kit for detecting omethoate. The kit comprises an enzyme label plate coated with an omethoate coupled antigen, an omethoate standard substance solution, a concentrated enzyme conjugate, an enzyme conjugate operating fluid, a substrate coloring solution and a stopping solution. The invention also discloses a method using the enzyme linked immunosorbent assay kit to detect omethoate in a sample. The method comprises the following steps: preprocessing the sample, detecting omethoate by using the kit, and analyzing a detection result. The enzyme linked immunosorbent assay kit can be used for detecting the residual quantity of omethoate in vegetable (raw material, batch and concentrate) samples, and has the advantages of simplicity in operation, low cost, high sensitivity, realization of onsite monitoring, and suitableness for screening a lot of samples.

Description

technical field [0001] The invention relates to an enzyme-linked immunosorbent detection technology, in particular to an enzyme-linked immunosorbent assay kit for detecting omethoate, which is suitable for the determination of omethoate in vegetables. [0002] technical background [0003] In recent years, organophosphorus pesticides (Ops) have played an extremely important role in world agriculture due to their high efficiency and low cost, and are one of the main pesticides. Organophosphorus pesticides are medium- and high-toxic pesticides with systemic toxicity, large residues in the environment, and most of them are difficult to degrade. While protecting crops, they also cause serious environmental pollution and directly or indirectly endanger human health. Omethoate is one of the most widely used and most used organophosphorus pesticides. Like other organophosphorus pesticides, its advantages and harms exist simultaneously: Omethoate uses high-concentration root-irrigati...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/577
CPCG01N33/543
Inventor 洪霞刘静张淑雅
Owner JIANGSU WISE SCI & TECH DEV
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