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ELISA Kit for Detecting Triazophos and Its Application

An enzyme-linked immunosorbent reagent, triazophos technology, applied in the direction of measuring devices, instruments, scientific instruments, etc., can solve the problems of high capital and personnel investment costs, and achieve the effect of simple sample pretreatment process, high accuracy, and simple structure

Active Publication Date: 2016-09-21
BEIJING KWINBON BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The instrument method has the advantages of high sensitivity and accurate results, but the investment cost of capital and personnel is high

Method used

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  • ELISA Kit for Detecting Triazophos and Its Application
  • ELISA Kit for Detecting Triazophos and Its Application
  • ELISA Kit for Detecting Triazophos and Its Application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] Embodiment 1 Preparation of kit components

[0029] 1. Preparation of triazophos hapten

[0030] Add 10mL of dichloromethane and 0.5g of triazophos into a 100mL three-necked flask, cool down to -5°C, add 1.01 equivalents of acetyl chloride under stirring, add 2 equivalents of aluminum trichloride under temperature control, and react at 5°C for 6 hours , add dilute hydrochloric acid and ice water, extract with ethyl acetate, wash with water, dry the organic phase with anhydrous magnesium sulfate, evaporate the solvent under reduced pressure, and recrystallize from petroleum ether-ethyl acetate system to obtain acetylated product.

[0031] Add the acetylated product from the previous step into a 100 mL three-neck flask, dissolve 10 mL of pyridine, add 1.2 equivalents of carboxymethyl hydroxylamine, react at 65°C for 5 hours, extract with ethyl acetate, wash with water, dry the organic phase with anhydrous magnesium sulfate, and distill under reduced pressure to obtain hal...

Embodiment 2

[0047] Example 2 Establishment of an enzyme-linked immunosorbent assay kit for detecting triazophos

[0048] An ELISA kit for detecting triazophos was set up to include the following components:

[0049] (1) a microtiter plate coated with a triazophos-coupled antigen;

[0050] (2) 6 bottles of triazophos standard solution, the concentrations are 0 μg / L, 1 μg / L, 3 μg / L, 9 μg / L, 27 μg / L, 81 μg / L;

[0051] (3) Goat anti-mouse anti-antibody labeled with horseradish peroxidase;

[0052] (4) Triazophos-specific antibody;

[0053] (5) Substrate chromogenic solution is made up of A liquid and B liquid, and A liquid is carbamide peroxide, and B liquid is tetramethylbenzidine;

[0054] (6) The stop solution is 2mol / L sulfuric acid;

[0055] (7) The washing solution has a pH value of 7.4, contains 0.5% to 1.0% Tween-20, 0.01‰ to 0.03‰ sodium azide preservative, and 0.1 to 0.3mol / L phosphate buffer, and the percentage is weight volume percentage;

[0056] (8) The complex solution is...

Embodiment 3

[0057] Example 3 Detection of triazophos in vegetables

[0058] 1. Sample pretreatment

[0059] Homogenize the tissue sample with a homogenizer; weigh 2.0±0.05g of the homogenate into a 10ml polystyrene centrifuge tube, add 2ml of 0.1M sodium hydroxide solution, 10ml of ethyl acetate, shake with an oscillator for 5min, and mix thoroughly ; Above 3000g, centrifuge at room temperature (20-25℃ / 68-77℉) for 5min; pipette 1ml of the upper organic phase into a 10ml clean and dry glass test tube; blow dry with nitrogen in a water bath at 50-60℃ (122-140℉); Add 1ml reconstitution working solution, vortex for 1min. Take 200μl and add it to 1800μl reconstituted working solution, mix well; take 50μl for analysis.

[0060] 2. Detection with kit

[0061] Add 50 μl of standard / sample to the corresponding microwell, then add 50 μl / well of antibody working solution, shake and mix gently, cover the plate with a cover film and place it in a dark environment at 25°C for 30 minutes. Carefully ...

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Abstract

The invention provides an enzyme-linked immunosorbent assay kit for detecting triazophos. The enzyme-linked immunosorbent assay kit comprises an ELISA plate containing a coating antigen, a triazophos standard solution, an ELIAS secondary antibody, a triazophos specificity antibody, a substrate color development solution, a stopping solution, a cleaning solution and a re-dissolving solution, wherein the coating antigen is a triazophos coupling antigen; the ELIAS secondary antibody is an enzyme-labeled goat-anti-mouse IgG antibody. The invention further discloses a method for applying the enzyme-linked immunosorbent assay kit to detection of triazophos. The method comprises the following steps: first, carrying out sample pretreatment; then, carrying out the detection with the enzyme-linked immunosorbent assay kit; finally, analyzing the detection result. The enzyme-linked immunosorbent assay kit provided by the invention can be utilized for detecting the content of triazophos in a vegetable sample, has an on-site monitoring function, and is simple and convenient to operate, low in cost, high in sensitivity, and suitable for screening of lots 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 triazophos, which can qualitatively and quantitatively detect the residual amount of triazophos in vegetables. Background technique [0002] Triazophos, also known as triazolphos, English common name Triazophos, chemical name O, O-diethyl-O-(1-phenyl-1,2,4-triazol-3-yl) phosphorothioate ester. The pure product is light brown yellow liquid, the solubility in water is 30-40mg / L at 20°C, soluble in most organic solvents; stable to light, hydrolyzed in acid and alkali medium, and decomposed at 140°C. Triazophos has strong contact and stomach poisoning effects, good insecticidal effect, obvious ovicidal effect, strong permeability, and no systemic effect. It is a moderately toxic, broad-spectrum organophosphorus insecticide and acaricide. It also has a certain nematicide effect and has been registered on rice...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N33/535
CPCG01N33/535
Inventor 冯才伟罗晓琴吴鹏何方洋冯静杨烁宋茹王然
Owner BEIJING KWINBON BIOTECH
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