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.
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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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