Method and kit for detecting acephate

A technology of acephate and molecular acephate, applied in the field of detection of acephate and kits, can solve the problems of poor stability, low anti-interference ability, lack of specificity and the like

Inactive Publication Date: 2014-06-04
HUAZHONG UNIV OF SCI & TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0007] In order to overcome the technical problems of traditional enzyme inhibition methods such as lack of specificity, low anti-interference ability, and poor stability, the present invention provides a MIP-enzyme inhibition method for detecting acephate, which can be specific, rapid, sensitive, and high-throughput Reliable Detection of Trace Acephate in Vegetable and Environmental Samples

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  • Method and kit for detecting acephate
  • Method and kit for detecting acephate
  • Method and kit for detecting acephate

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0047] Embodiment 1: the preparation of acephate MIP

[0048] The preparation of acephate MIP uses dopamine as a functional monomer and acephate as a template molecule, and obtains acephate MIP through the oxidative polymerization reaction of dopamine and the elution process of template molecules. The specific method includes the following steps:

[0049] Step 1: Polymerization:

[0050] a) dissolving the template molecule acephate and dopamine in a Tris-HCL buffer solution with a molar ratio of 1:10-50, and mixing;

[0051] b) Adding carrier materials: carrier materials such as microspheres and filter membranes are directly added to the mixture, or the mixture is added to the wells of the microplate.

[0052] c) Polymerization: Expose to air for 48-72 hours at room temperature to carry out polymerization.

[0053] d) The synthesis method of blank imprinted polymer (NIP) is similar to that of MIP, except that template molecules are not added during the synthesis process.

[...

Embodiment 2

[0059] Example 2: Sensitivity analysis of the MIP-enzyme inhibition kit (the carrier material is a 96-well microtiter plate as an example)

[0060] Add 300 μL of acephate standard aqueous solution (0, 2.5, 5, 25, 50, 100, 200 ng / mL) in a series of concentration gradients to the wells of the microplate containing MIP, act at 37°C for 1-6 hours, then rinse with pure water Wash 2-3 times, then add 120 μL triple distilled water and 10-20 μL cholinesterase (AChE) (500 mU / mL) to each well, incubate at 37 ° C for 10-20 min, add 105 μL containing 0.1 μmol / mL Lumin to each well Mixture of luminol, 0.5mg / mL choline oxidase (ChOX) and 10U / mL horseradish peroxidase (HRP), add 10-20μL (20μmol / mL) of acetylcholine (ATCI) to each well before detection , and immediately use a multi-functional microplate reader to detect and measure the change of the cumulative luminescence value within 5 minutes, the reading interval is 10s, and the luminescence value reaches equilibrium within 5 minutes. Th...

Embodiment 3

[0061] Example 3: Specificity analysis of MIP-enzyme inhibition kit (carrier material is 96-well ELISA plate as an example)

[0062] (1) Add 300 μL of trichlorfon, dimethoate, and glufosinate-ammonium (standard aqueous solutions (0, 2.5, 5, 25, 50, 100, 200 ng / mL)) to the wells of the microplate containing MIP, and the different concentrations are similar The luminescence value of the substance is basically consistent with the luminescence value when the concentration is 0ng / mL. According to the method in Example 2, the inhibition rate obtained by using the balance value of the chemiluminescence intensity has no linear relationship with the concentrations of trichlorfon, dimethoate, and glufosinate-ammonium. It shows that the kit prepared by the present invention cannot be combined with trichlorfon, dimethoate, and glufosinate-ammonium, but can only be specifically combined with acephate, and can be conveniently and accurately used for the detection of acephate in the environme...

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Abstract

The invention relates to a method for detecting a trace amount of acephate in a sample and provides a kit. According to a dopamine oxidation polymerization method, an artificial antibody namely an MIP (molecularly imprinted polymer) which can be specifically bonded with the acephate is synthesized on a carrier material; after a trace amount of acephate enriched in different samples is specifically separated by using the MIP, the content of the enriched acephate is determined according to an enzyme inhibition-chemiluminescence method. The MIP is high in temperature resistance and organic solvent resistance, the trace amount of acephate enriched in the different samples can be directly and specifically separated through the MIP, and the influence of various interference matters such as other organophosphorus pesticides and chlorophyll is removed. An MIP specific sample pretreatment method is organically combined with an enzyme inhibition method into an MIP-enzyme inhibition method, so that a certain pesticide can be specifically and quickly detected by the enzyme inhibition method, the detection sensitivity and the detection specificity can be improved, and a trace amount of target pesticides in environment and food can be directly, quickly and sensitively detected with high throughput.

Description

technical field [0001] The invention belongs to the technical field of molecular imprinting technology and rapid detection of pesticide residues, and specifically relates to the preparation of an acephate artificial antibody (MIP), and the set-specific sample established by organically combining MIP with enzyme inhibition-chemiluminescence method Pretreatment and enzyme inhibition-chemiluminescence detection in one, a new method for specific, rapid, sensitive and high-throughput detection of trace acephate in samples: MIP-enzyme inhibition method, and provide corresponding detection kits . technical background [0002] Organophosphorus pesticides are the most widely used pesticides in my country. Since January 1, 2007, methamidophos and other five kinds of medium and high-toxic organophosphorus pesticides have been completely banned in my country. Acephate has been used for its low toxicity, safety, and broad spectrum. , long-lasting effect, good field application effect and...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N21/76G01N33/531
Inventor 吕斌王雪娟刘燕婕石云
Owner HUAZHONG UNIV OF SCI & TECH
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