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Metanephrine derivative, immunogen, anti-metanephrine specific antibody and preparation method and application thereof

A technology of metanephrine and epinephrine, which is applied in the field of biomedical detection, can solve the problems of poor accuracy, long detection time, and difficulty in batch detection.

Active Publication Date: 2021-01-26
长沙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

Enzyme-linked immunosorbent assay has high sensitivity and good specificity, but the detection time is long and the accuracy is poor, so it is difficult to use in batch detection

Method used

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  • Metanephrine derivative, immunogen, anti-metanephrine specific antibody and preparation method and application thereof
  • Metanephrine derivative, immunogen, anti-metanephrine specific antibody and preparation method and application thereof
  • Metanephrine derivative, immunogen, anti-metanephrine specific antibody and preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0083] Example 1: Synthesis of Methoxyephrine Derivatives

[0084] Methoxyephrine derivatives were synthesized via the following synthetic route:

[0085]

[0086] Concrete synthetic steps are as follows:

[0087] (1) Synthesis of compound 3:

[0088]

[0089] Weigh 50.0g of methoxyepinephrine and 25.0g of compound 2, dissolve them in 250mL N,N-dimethylformamide (DMF), and then add 20.0g of K 2 CO 3 , to make a reaction mixture; the reaction mixture was stirred at room temperature for 12 hours; after the reaction, 250 mL of purified water was added to the reaction mixture, and then 200 mL of dichloromethane (DCM) was used for extraction, and the extraction step was repeated 3 times; The combined organic phase obtained by the extraction was passed through Na 2 SO 4 Drying was carried out, followed by concentration; the residue obtained after concentration was purified by a silica gel drying column (PE:EA=5:1) to obtain 26.0 g of compound 3 with a yield of 52%.

[00...

Embodiment 2

[0093] Example 2: Preparation of Methoxyephrine Immunogen

[0094] The specific steps of the preparation method of methoxyepinephrine immunogen are as follows:

[0095] (1) Preparation of carrier solution: Dissolve the carrier protein in 0.35mol / L potassium phosphate buffer (pH=8.5), and the final concentration of the carrier protein is 5.0mg / mL to obtain the carrier protein solution;

[0096] (2) Preparation of methoxyephrine derivative solution: 250.0 mg of the above methoxyephrine derivative, 7.5 mL of dimethylformamide, 7.5 mL of ethanol, 15.0 mL of potassium phosphate buffer (10.0 mmol / L, pH=8.0), 150.0mg 1-ethyl-3-(-3-dimethylaminopropyl) carbodiimide, 90.0mg N-hydroxysulfosuccinimide were mixed, stirred and dissolved for 3 hours to obtain Methoxyepinephrine derivative solution;

[0097] (3) Synthesis of methoxyepinephrine immunogen: Add the methoxyephrine derivative solution obtained in step (2) to the carrier protein solution obtained in step (1) dropwise, and stir ...

Embodiment 3

[0098] Example 3: Preparation of anti-methoxyadrenaline specific antibody

[0099] The specific steps of the preparation method of anti-methoxyepinephrine specific antibody are as follows:

[0100] (1) Dilute the above-mentioned methoxyepinephrine immunogen with 0.15mol / L sodium phosphate buffer (pH=7.0) to a final concentration of 3.5mg / mL to obtain an artificial antigen solution, and then mix 3.0mL artificial antigen solution with The same amount of Freund's complete adjuvant was mixed, and the experimental animal rabbit was injected at multiple points;

[0101] (2) After 4 weeks, use 3.0mL of the same artificial antigen solution and the same amount of Freund's incomplete adjuvant to inject the above-mentioned experimental animal rabbits at multiple points, and then inject once every 5 weeks, for a total of 6 injections;

[0102] (3) Blood was collected from rabbits that had been injected in step (2), separated and purified to obtain anti-methoxyadrenaline-specific antibodi...

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Abstract

The invention discloses a metanephrine derivative, an immunogen, an anti-metanephrine specific antibody as well as a preparation method and application thereof. The preparation method comprises the following steps: firstly, coupling a novel metanephrine derivative with recombinant bovine serum albumin obtained by genetic engineering modification to prepare a metanephrine artificial antigen, and then immunizing an experimental animal with the metanephrine artificial antigen to obtain an anti-metanephrine specific antibody. ELISA detection shows that the specific antibody is strong in specificity and high in sensitivity, and an interference experiment shows that the specific antibody has no cross reaction with 32 common hormones; and the anti-metanephrine specific antibody is applied to preparation of metanephrine detection reagents, including a metanephrine homogeneous enzyme immunoassay reagent and a metanephrine latex enhanced turbidimetric immunoassay reagent. The detection reagent can realize high-throughput and rapid detection of metanephrine on a full-automatic biochemical analyzer.

Description

technical field [0001] The invention relates to a methoxyephrine derivative, an immunogen, an anti-methoxyephrine specific antibody, a preparation method and application thereof, and belongs to the technical field of biomedical detection. Background technique [0002] Methoxyepinephrine (Metanephrine, MN) is 3-methoxyephrine, which is mainly an intermediate metabolite of adrenaline. Catecholamine (CA) is a hormone secreted by the adrenal medulla, adrenal neurons and extra-adrenal chromaffin bodies, mainly including epinephrine (epinephrine, E) and norepinephrine (Norepinephrine, NE). Adrenaline is mainly converted into MN through the adrenal medulla and chromaffin cells under the action of catechol-O-methyltransferase (COMT), and norepinephrine is mainly converted into MN through tissues other than neurons and chromaffin cells Under the action of COMT, it is converted into methoxynorepinephrine (Normetanephrine, NMN). The determination of methoxyepinephrine in urine can in...

Claims

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Application Information

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IPC IPC(8): C07C229/14C07C227/20C07K14/765C07K16/44G01N33/94G01N33/539G01N33/543
CPCC07C229/14C07C227/20C07C227/06C07K14/765C07K16/44G01N33/9433G01N33/539G01N33/54313C07B2200/07
Inventor 张小可王英高慧李冬
Owner 长沙博源医疗科技有限公司
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