Catecholamine immunogen, derivative and synthesis method, specific antibody and detection reagent and preparation method

A catecholamine, immunogenic technology, applied in the fields of specific antibodies and detection reagents and preparation, catecholamine immunogens, derivatives and synthesis, can solve problems such as the limitations of clinical large-scale application, reduce detection costs, immunogenicity High, specific effect

Active Publication Date: 2018-04-20
苏州博源医疗科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

These methods have their own advantages and disadvantages, but they all have certain limitations in large-scale clinical application.
At present, there is a lack of catecholamine detection reagents with good stability, high sensitivity, and strong specificity in the market, especially good-quality automated detection reagents. Therefore, the quality of research and development and production meets clinical requirements, strong practicability, and high cost performance. It can be applied to automatic biochemistry The catecholamine determination reagent of the analyzer has become a hot spot in the in vitro diagnostic reagent industry at home and abroad

Method used

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  • Catecholamine immunogen, derivative and synthesis method, specific antibody and detection reagent and preparation method
  • Catecholamine immunogen, derivative and synthesis method, specific antibody and detection reagent and preparation method
  • Catecholamine immunogen, derivative and synthesis method, specific antibody and detection reagent and preparation method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0067] Example 1 Synthesis and Quantitative Detection of Catecholamine Derivatives

[0068] The chemical structure of catecholamine derivatives is shown in formula (Ⅲ):

[0069]

[0070] The synthetic route and preparation steps of the above-mentioned catecholamine derivatives are as follows:

[0071]

[0072] Concrete synthetic steps are as follows:

[0073] Synthesis of compound 2

[0074]

[0075] 1) Weigh 20.41g (147.9mmol) of compound 1, 25.29g (147.9mmol) of benzyl bromide and 30.62g (221.9mmol) of K 2 CO 3 , were dissolved together in 200 mL of acetone, and stirred overnight at room temperature. Thin-layer chromatography (TLC) detection showed that the reactant had reacted completely. The reacted synthetic solution was filtered, the filtrate was concentrated in vacuo, and the residue obtained after concentration was diluted with 300 mL of EA. Rinse the organic layer with 200mL purified water and 200mL brine, pass Na 2 SO 4 Drying was carried out, and th...

Embodiment 2

[0109] The synthesis of embodiment dicatecholamine immunogen

[0110] The catecholamine immunogen is composed of bovine serum albumin (Bovine Serum Albumin, BSA) and the catecholamine derivative -(CH 2 ) n The -COO- group is connected. In this embodiment, the synthesis method of the immunogen is described in detail by taking n=3 as an example. The specific steps are as follows:

[0111] 1. Dissolve 200mg bovine serum albumin in 50ml 0.2M, pH 8.5 phosphate buffer;

[0112] 2. Add the following chemicals into a small beaker and stir to dissolve: 200mg synthetic catecholamine derivatives, 3.5ml dimethylformamide, 3.5ml ethanol, 7.0ml 10mM, pH 5.0 potassium phosphate buffer, 200mg 1-ethyl -3-(-3-dimethylaminopropyl) carbodiimide, 50mg N-hydroxyl sulfosuccinimide, these chemicals were stirred and dissolved at room temperature for 30 minutes;

[0113] 3. Add the dissolved solution dropwise to the BSA solution, and stir overnight at 2-8° C. to obtain the antigen; purify the synthe...

Embodiment 3

[0114] Example 3: Preparation of anti-catecholamine specific antibody

[0115] The catecholamine immunogen prepared in Example 2 was inoculated into experimental animal rabbits by conventional methods, and the antiserum was taken after booster immunization. The specific steps were as follows:

[0116] 1. Dilute the synthesized catecholamine immunogen to 1.0 mg / ml with PBS to obtain an antigen solution, then mix 1.0 ml of the antigen solution with Freund's complete adjuvant, and inject the experimental animal rabbit.

[0117] 2. After 2-3 weeks, inject 1.0ml of the same antigen solution and incomplete Freund's adjuvant to the above-mentioned experimental animal rabbit once, and then inject once every four weeks, a total of 4 injections.

[0118] 3. Taking blood from the experimental animal rabbit in step 2, separating and purifying to obtain specific anti-catecholamine antibodies with a titer of 1:30000-1:50000.

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Abstract

The invention discloses a catecholamine immunogen, a ramification and a synthetic method, a specific antibody and a detection reagent and a preparation method. The prepared catecholamine immunogen is high in immunogenicity, a high-titer anti-catecholamine specific antibody can be obtained by inducing of the prepared catecholamine immunogen, and no cross reaction of the prepared catecholamine immunogen with 62 common medicines occurs; catecholamine detection reagent is obtained by preparing the antibody, and catecholamine content of a sample can be determined precisely and rapidly. Compared with an existing detection reagent on the market, the detection reagent has the advantages that the operation is simple, the sensitivity is high, the specificity is strong, and the result is precise, the catecholamine detection cost can also be lowered, and the detection reagent can be easily popularized and used at clinic in a large scale.

Description

technical field [0001] The invention belongs to the field of biotechnology, and relates to catecholamine immunogens, derivatives and synthesis methods, specific antibodies, detection reagents and preparation methods. Background technique [0002] Catecholamine is a general term for a class of small molecular substances derived from tyrosine and containing catechol and amine groups. Common catecholamines mainly include epinephrine (E), norepinephrine , NE) and dopamine (dopamine, DA) etc. (the structural formula is shown in formula 1). In the human body, catecholamines are mainly secreted by the chromaffin cells of the adrenal medulla and sympathetic neurons. , endocrine glands, kidneys, smooth muscles and other tissues and organs have a wide range of regulatory effects. Among them, dopamine plays a very important role in the brain and nerve signal transmission, which can improve the response speed of the nervous system and enhance the body's ability to adapt to the environ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K14/765C07K14/795C07K14/435C07K16/44C07C217/60C07C213/02G01N33/53
CPCC07C213/08C07C217/60C07K14/435C07K14/765C07K14/795C07K16/44C07K19/00G01N33/535
Inventor 虞留明颜光涛洪天配邱玲
Owner 苏州博源医疗科技有限公司
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