Production method and use for imidacloprid artificial hapten, artificial antigen and specific antibody

An artificial hapten and imidacloprid technology, applied in chemical instruments and methods, specific peptides, material inspection products, etc., can solve the problems of cumbersome process, great influence, unsuitable for large-scale sample detection and analysis, etc., and achieve high sensitivity, Strong specificity and easy on-site monitoring effect

Inactive Publication Date: 2005-01-26
ZHEJIANG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, the sensitivity of these methods is greatly affected by steps such as sample purification and concentration; moreover, these methods require compl...

Method used

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  • Production method and use for imidacloprid artificial hapten, artificial antigen and specific antibody
  • Production method and use for imidacloprid artificial hapten, artificial antigen and specific antibody
  • Production method and use for imidacloprid artificial hapten, artificial antigen and specific antibody

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preparation example Construction

[0037] 3 Antibody preparation and imidacloprid enzyme-linked immunoassay method establishment

[0038] The steps of the preparation method of imidacloprid-specific antibody are as follows:

[0039] 1) The experiment selects healthy male rabbits or mice about half a year old and weighing 2-3 kg. The experimental immunization dose is 0.25-2.0 mg / kg for basic immunization, and 0.5-2.0 mg / kg for booster immunization. Dilute appropriate amount of artificial antigen complexes with normal saline respectively, add an equal volume of Freund's complete adjuvant, fully emulsify until instilled Emulsion droplets do not disperse in water. A combination of subcutaneous multi-point injection on the back and intramuscular injection on the thigh was used. Back subcutaneous immunization at 4 to 6 points, thigh intramuscular injection at 2 to 4 points, booster immunization after 3 to 4 weeks, and then booster immunization every 2 weeks, using incomplete Freund's adjuvant for booster immunizati...

Embodiment 1

[0043] Example 1: Synthesis of Hapten IM

[0044] According to the feeding ratio of imidacloprid: 3-mercaptopropionic acid as 1: 1, 5.15g (20mmol) of imidacloprid (20mmol) was dissolved in 20mL dimethyl sulfoxide, put into a three-necked flask, and then 2.25g (40mmol) of KOH was added, and magnetically stirred to dissolve it, weighing Get 2.10g (20mmol) of β-mercaptopropionic acid and dissolve it in 10mL dimethyl sulfoxide and transfer it to a constant pressure dropping funnel, slowly drop the β-mercaptopropionic acid solution into a three-necked flask, and slowly heat the oil bath to 100°C. After keeping warm for 2 hours, remove the oil bath. After the reaction solution is naturally cooled to room temperature, add 20 mL of water and adjust the pH value to 3.0-5.0 with dilute hydrochloric acid; extract with 3×30 ml of dichloromethane, collect the organic phase, and wash the organic phase with 3×15 ml of water. phase, dried over anhydrous sodium sulfate, and concentrated to ob...

Embodiment 2

[0048] Example 2: Synthesis of Hapten IM

[0049] According to the feeding ratio of imidacloprid: 3-mercaptopropionic acid as 1:2, 5.15g (20mmol) of imidacloprid (20mmol) was dissolved in 20mL dimethyl sulfoxide, put into a three-necked flask, and then 4.5g (80mmol) of KOH was added, and magnetically stirred to dissolve it, weighing Get 4.2g (40mmol) of β-mercaptopropionic acid and dissolve it in 20mL dimethyl sulfoxide and transfer it to a constant pressure dropping funnel, slowly drop the β-mercaptopropionic acid solution into the three-necked flask, and slowly heat the oil bath to 100°C. After keeping warm for 2 hours, remove the oil bath. After the reaction solution is naturally cooled to room temperature, add 40mL of water and adjust the pH value to 3.0-5.0 with dilute hydrochloric acid; extract with 3×30ml of dichloromethane, collect the organic phase, and wash the organic phase with 3×15ml of water. phase, dried over anhydrous sodium sulfate, and concentrated to obtain...

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Abstract

The invention discloses the production method and use for imidacloprid artificial hapten, artificial antigen and specific antibody, wherein the production method comprises, using imidacloprid (1-(6-chlorine-3-picolyl)-N-nitro-2-imidazoline imine) as raw material for reaction with 3-mercaptopropionic acid under alkaline condition, thus synthesizing hapten 1-(6-(2-carboxyethyl) sulfo-3-picolyl)-N-nitro-2-imidazoline imines (IM), then coupling with proteins through carbodiimide method and mixed anhydride method to prepare artificial antigens (immunogens and peridium antigens).

Description

technical field [0001] The invention relates to a method for preparing imidacloprid artificial hapten, artificial antigen and specific antibody and its application. The invention belongs to the technical field of pesticide immunochemistry. Background technique [0002] The invention belongs to the technical field of immunochemistry and residue analysis of small-molecule pesticide compounds (molecular weight less than 1000 Daltons), relates to organic synthesis, immunochemistry and biochemistry, and relies on basic principles of immunology and immunochemistry and biotechnological means to design and synthesize The small molecule target analyte hapten is coupled with the carrier protein to prepare an effective artificial antigen, and the specific antibody for the small molecule analyte is prepared by immunizing animals, using the specific immunological reaction of the antigen antibody and the marker that is easily detected and identified It can quantitatively detect ultra-tra...

Claims

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Application Information

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IPC IPC(8): C07D233/30C07K16/00G01N33/02G01N33/563
Inventor 魏方林程敬丽郑尊涛吴慧明朱国念
Owner ZHEJIANG UNIV
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