A kind of preparation method and application of dexamethasone hapten
A technology of dexamethasone and hapten, which is applied in the field of food safety detection, can solve the problems that the accuracy, sensitivity, and specificity cannot be fully achieved in detection, and achieve the effect of simple and feasible preparation method, high yield and low cost
- Summary
- Abstract
- Description
- Claims
- Application Information
AI Technical Summary
Problems solved by technology
Method used
Image
Examples
Embodiment 1
[0029] Embodiment 1, the preparation of dexamethasone hapten
[0030] 1. Preparation of dexamethasone hapten
[0031] Add 500 mg of dexamethasone raw material into a 50 ml round bottom flask, add 110 mg of succinic anhydride after dissolving 5 ml of pyridine F, react at 80 degrees for 3 hours, TLC detects that after the reaction of the raw materials is completed, remove pyridine under reduced pressure, add 10 ml of ice water to the residue, A white solid was precipitated, filtered, washed with water and dried to obtain 350 mg of the product, which was the hapten.
[0032] The reaction equation is as follows:
[0033]
[0034] 2. Identification of dexamethasone hapten
[0035] The resulting product was identified by mass spectrometry, -MS: M-1=491.2, +MS: M+K=531.7 ( figure 1 ).
[0036] The results showed that its chemical structure was as shown in Formula I (MW=492.53), which was the hapten of dexamethasone.
[0037] Formula 1.
Embodiment 2
[0038] Embodiment 2, preparation and identification of dexamethasone artificial antigen
[0039] 1. Synthesis of dexamethasone immune antigen (DSMS-BSA)
[0040] (1) Dissolve 22 mg of hapten in 1.5 mL of DMF, after complete dissolution, add 25 mg of EDC and 25 mg of NHS in turn, and stir for 3 hours at room temperature;
[0041] (2) Weigh 50mg of BSA, dissolve in 3.5mL 0.1M carbonic acid buffer pH=9.6, stir at 400rpm for 10min, fully dissolve;
[0042] (3) Take the activation solution in the above step 1, add it dropwise to the protein solution in an ice-water bath environment, stir while adding, and react with magnetic stirring at room temperature for 24 hours;
[0043] (4) Put the reaction product into a dialysis bag (15cm) rinsed with distilled water, dialyze with 1LPBS for 3 days, stir and dialyze at room temperature, change the dialysate 3 times a day, centrifuge the dialyzed product at 4500rpm for 6min, dispense 0.5ml / tube, and put Antigen number, stored at -20°C for f...
Embodiment 3
[0051] Example 3, Preparation and Specific Identification of Enzyme-labeled Monoclonal Antibody
[0052] 1. Preparation of dexamethasone monoclonal antibody
[0053] 1. Use the immunogen prepared above at 100 μg / mouse, dissolve the immunogen in normal saline and mix it with Freund’s complete adjuvant in an equal volume, and immunize 6-8 week-old Balb / c female mice subcutaneously on the back of the neck. After the initial immunization, On the 7th, 14th, and 28th days, mix the immunogen with incomplete Freund's adjuvant in equal volumes, and give each additional immunization once. Three days before the fusion, use 100 μg of the immune complex per mouse, without Freund's adjuvant, and then add another immunization.
[0054] 2. Proceed according to the conventional method, take the splenocytes of the immunized mice and mix them with the mouse myeloma cells (SP2 / 0) in the logarithmic growth phase, and then slowly add the preheated fusion agent (PEG4000) within 45s for fusion, Suspen...
PUM
Login to View More Abstract
Description
Claims
Application Information
Login to View More