Paclitaxel immunogen, anti-paclitaxel specific antibody and paclitaxel detection reagent
A technology of paclitaxel and immunogen, which is applied in the field of paclitaxel immunogen and paclitaxel detection reagents, can solve the problems of not being suitable for large-scale clinical application and failing to meet the needs of clinical detection, so as to facilitate clinical promotion and use, reduce detection costs, and immunogen sex high effect
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Embodiment 1
[0068] Example 1: Synthesis and quantitative detection of paclitaxel derivatives
[0069] The chemical structure of paclitaxel derivatives is shown in formula (IV):
[0070]
[0071] The synthetic route and preparation steps of the above-mentioned taxol derivatives are as follows:
[0072]
[0073] 1. Weigh 1g paclitaxel and 2g succinic anhydride into the reaction system, and then incubate them with 20ml pyridine or dry benzene in a glass flask with continuous reflux;
[0074] 2. Continue heating the reaction mixture at a reflux temperature of 70-80°C for 6 hours, then slowly cool the reaction mixture to room temperature, and gently pour off excess pyridine or benzene;
[0075] 3. The remaining organic components are continuously blown into a nitrogen stream under negative pressure to evaporate and dry, and the dried residual product obtained is the paclitaxel derivative;
[0076] 4. Wash the product obtained above with 60% ethanol prepared with distilled water for more than 10 times to ...
Embodiment 2
[0078] Example 2 Synthesis of Taxol Immunogen
[0079] Paclitaxel immunogen consists of bovine serum albumin (Bovine Serum Albumin, BSA) and the paclitaxel derivative represented by formula (II) -CO-(CH 2 ) n -COO- group is connected. In this embodiment, n=2 is taken as an example to illustrate the method for synthesizing the immunogen in detail. The specific steps are as follows:
[0080] 1. Dissolve 200mg of bovine serum albumin in 50ml of 0.2M, pH 8.5 phosphate buffer;
[0081] 2. Add the following chemicals to a small beaker and stir to dissolve: 200mg synthetic paclitaxel derivative, 3.5ml dimethylformamide, 3.5ml ethanol, 7.0ml 10mM potassium phosphate buffer pH 5.0, 200mg 1-ethyl -3-(-3-Dimethylaminopropyl)carbodiimide, 50mg N-hydroxysulfosuccinimide, stir and dissolve these chemicals at room temperature for 30min;
[0082] 3. Add the dissolved solution dropwise to the BSA solution and stir overnight at 2-8°C to obtain the antigen; the synthesized antigen is purified by dialysi...
Embodiment 3
[0083] Example 3: Preparation of specific antibodies against paclitaxel
[0084] The paclitaxel immunogen prepared in Example 2 was inoculated with experimental animal rabbits by conventional methods, and antiserum was obtained after booster immunization. The specific steps are as follows:
[0085] 1. Dilute the synthesized paclitaxel 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 to inject the experimental animal rabbit.
[0086] After 2.2 to 3 weeks, 1.0 ml of the same antigen solution and Freund's incomplete adjuvant were used to inject the above-mentioned experimental animal rabbits once, and then once every four weeks, for a total of 4 injections.
[0087] 3. Take blood from the experimental animal rabbit in step 2, and separate and purify to obtain the anti-paclitaxel specific antibody with a titer of 1:30000-1:50000.
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