Pesticide and veterinary drug multi-residue detection method based on microarray detection chip

A technology of microarray chips and detection chips, which is applied in the direction of biological testing, material inspection products, etc., can solve the problems of expensive instruments, complicated and difficult operations, false positives, etc., to ensure accuracy and sensitivity, simple operation process, and preparation The effect of cost reduction

Active Publication Date: 2013-12-11
内蒙古敖敦食品股份有限公司
View PDF10 Cites 15 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

(3) Chromatography-mass spectrometry (GC-MS, HPLC-MS): It can achieve qualitative and quantitative detection purposes at the same time, especially suitable for the detection of pesticide metabolites, degradation products and multi-residue detection, etc., but this method requires expensive instruments And the operation is complicated and difficult, not suitable for frequent detection
(4) Supercritical Fluid Chromatography (SFC): It not only has the characteristics of fast, efficient, and sensitive gas spectroscopy, but also has the characteristics of liquid chromatography that can detect thermally unstable and macromolecular compounds, but requires expensive instruments and complicated operations Difficulty, etc.
(5) Capillary electrophoresis (CE): It is suitable for the separation and analysis of some ionized samples that are difficult to separate by traditional chromatography. It has 10-1000 times higher analytical capacity than HPLC. It requires expensive instruments and is complicated and difficult to operate. It is not suitable for for regular testing
However, due to the influence of antibodies such as dependence on foreign imports, at present, the ELISA finished kits in the Chinese market rely on imports from abroad.
(3) Enzyme inhibition method is the most mature and widely used rapid pesticide residue detection technology. This method can only detect organophosphorus and carbamate pesticides, and the inhibition rate of different pesticides of the same kind varies greatly. , so using a uniform inhibition rate to determine whether pesticide residues exceed the standard will inevitably lead to false positive or false negative missed detection, which can be said to be the current "rapid detection method adopted as a last resort", which is only applicable to the initial inspection at the grassroots level and plays a warning role , when the phenomenon of exceeding the standard is found, it must be retested and confirmed by standard methods, negative reactions should also be sampled in proportion, and retested and confirmed by reliable methods
[0015] Existing research at home and abroad is mainly focused on establishing a specific immunoassay method for a single pesticide and veterinary drug, but in fact there are often many kinds of pesticide and veterinary drug residues in agricultural products, Immunoassay methods and corresponding detection kits for single agricultural and veterinary drug components have been difficult to meet the needs of actual detection

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Pesticide and veterinary drug multi-residue detection method based on microarray detection chip
  • Pesticide and veterinary drug multi-residue detection method based on microarray detection chip
  • Pesticide and veterinary drug multi-residue detection method based on microarray detection chip

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0071] Part I: Synthesis of Pefloxacin (PEFloxacion, PEF) Immunization Antigen and Detection Antigen

[0072] (1) Accurately weigh 45 mg of N-hydroxysuccinimide (NHS), 210 mg of carbodiimide (EDC), 36 mg of pefloxacin (PEF), and add 5 mL of N,N-dimethyl Formamide (DMF). And incubate at room temperature for 24 h with shading and stirring, and this solution is called A.

[0073] (2) Accurately weigh 105 mg bovine serum albumin (BSA), add 0.01 mol / L (pH 7.4) phosphate buffer saline (PBS) 15 mL, this solution is called B.

[0074] (3) Add the incubated solution A to solution B dropwise, stir while adding, and dialyze with 0.01 mol / L (pH value 7.4) PBS magnetic stirring for 6 days, and change the solution several times during the period to Remove unreacted small molecule species. Centrifuge at 12,000 r / min for 30 min, collect the supernatant to obtain the PEF-BSA conjugate, and store the obtained PEF-BSA conjugate in a -20°C refrigerator for future use.

[0075] After replaci...

Embodiment 2

[0146] The first part of melamine (Melamine) immune antigen and detection antigen synthesis

[0147] The immune antigen melamine-BSA and the detection antigen melamine-OVA were synthesized by combining polybasic anhydrides and mixed anhydrides.

[0148] (1) Synthesis of melamine glutaric anhydride semialdehyde by polybasic anhydride method (where pyridine is used as solvent and catalyst). Weigh 125 mg of melamine (about 1 mmol), add it into 4 mL of pyridine solution containing 114 mg (about 1 mmol) of glutaric anhydride, and stir the reaction with a magnetic stirrer for 22 h at room temperature. After the reaction was complete, the pyridine was blown dry with nitrogen.

[0149] (2) Use the above product to react with isobutyl chloroformate to obtain mixed anhydrides. Dissolve melamine-glutaric anhydride semialdehyde in 40 mL solvent (DMF and 1,4-dioxane 1:1 mixture), add 262 μL (about 1 mmol) n-tributylamine, stir in ice for 10 min, add chloroformic acid 144 μL (about 1 m...

Embodiment 3

[0158] The first part of the synthesis of methamidophos immune antigen and detection antigen

[0159] Weigh 100 mg of methamidophos and dissolve in 10 mL of 0.1 mol / L HCl, add pre-cooled 1 % NaNO 2 Continue to stir for 30 min until the starch potassium iodide test paper turns blue; take another 10 mg BSA and dissolve it in 2 mL of borate buffer solution with pH 8.7, take 2 mL of the above diazotization product and add it dropwise to the protein solution, and stir in an ice bath After 2 hours, place overnight at 4°C and put the reaction solution into a dialysis bag. Dialyze with PBS at 4°C for 3 days, freeze-dry the dialysate to obtain a white powder, store at -20°C for use. The coated antigen methamidophos-OVA was prepared in the same way.

[0160] The second part is the preparation of monoclonal antibodies to agricultural and veterinary drugs, and the third part is the preparation of microarrays of monoclonal antibodies to agricultural and veterinary drugs.

[0161] U...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

PUM

No PUM Login to view more

Abstract

The invention relates to a pesticide and veterinary drug multi-residue detection method based on a microarray detection chip. The method comprises the steps that a micromolecular pesticide and veterinary drug hapten is coupled with bovine serum albumin to prepare an immunizing antigen; a mouse is immunized by the immunizing antigen to prepare a corresponding pesticide and veterinary drug monoclonal antibody; a biochip sample application instrument for a monoclonal antibody acquisition probe performs sample application on an agarose modified slide solid phase carrier to prepare a multi-repeat monoclonal antibody acquisition probe microarray; the pesticide and veterinary drug hapten is coupled with ovalbumin to prepare a hapten-OVA (ovalbumin) couplet; a fluorescent molecule Cy3 is then used for labeling; sample liquid to be detected is incubated with the chip after mixed with a labeling detection antigen according to a certain concentration; a to-be-detected antigen in a sample and the corresponding labeling detection antigen are directly and competitively bound with the corresponding monoclonal antibody acquisition probe fixed on the chip; elution is performed under a certain condition; and a result is detected by a biochip scanner. The method can detect multiple pesticide and veterinary drug residues in the subsidiary agricultural product sample simultaneously, and is applicable to high-throughput, quick and accurate detection of drug residues in planting and culture production.

Description

[0001] technical field [0002] The invention relates to a method for preparing a multi-residue detection microarray chip of agricultural and veterinary drugs in the field of food safety. [0003] Background technique [0004] Agricultural and veterinary drugs are effective means to prevent and control crop diseases and insect pests and aquaculture diseases in countries around the world. The use of a large number of chemical pesticides and veterinary drugs, especially the highly toxic and high-residue pesticides and veterinary drugs, has become the main "killer" of food safety and endangering people's health. [0005] At present, the main chemical pesticides used are mainly divided into three categories: insecticides, herbicides and fungicides according to their different functions. According to different chemical structures, they can be divided into organochlorines, organophosphorus, pyrethroids, carbamates, and inorganic pesticides. The common characteristics of the...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

Application Information

Patent Timeline
no application Login to view more
Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/68
Inventor 李同祥孙会刚黄天姿候进慧
Owner 内蒙古敖敦食品股份有限公司
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Try Eureka
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap