A hybridoma cell line js secreting anti-phenolphthalein monoclonal antibody and its application
A technology of hybridoma cell lines and monoclonal antibodies, which can be used in hybrid peptides, analytical materials, and methods based on microorganisms. It can solve the problems of high detection cost, poor accuracy, and quantitative analysis, and achieve good detection sensitivity. , good specific effect
Active Publication Date: 2022-04-19
JIANGNAN UNIV
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Problems solved by technology
These detection methods all have the disadvantages of high detection cost, long time-consuming and high professional quality requirements for detection personnel.
In addition, it is also reported that phenolphthalein is colorless in acidic and neutral solutions and pink in alkaline environments with pH > 10 to prepare a phenolphthalein detection card for detection. Although this method is simple to operate, it is convenient Carry but has the disadvantage of poor accuracy and cannot be quantitatively analyzed
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Embodiment 1
[0021] Embodiment 1 Preparation of hybridoma cell line JS
[0022] (1) Synthesis of complete antigen: Weigh 3.55mg of phenolphthalein (the molar ratio of phenolphthalein to keyhole limpet hemocyanin KLH is 9000:1), 17.6mg of N,N'-carbonyldiimidazole (CDI), dissolve in 300μL N,N - In dimethylformamide (DMF), stir and react at 37°C for 1 h (referred to as liquid A); take 6 mg of KLH and dissolve it in 2 mL of 0.01M borate buffer solution (BB, pH=8.6) (referred to as liquid B ), then slowly add liquid A to liquid B drop by drop, and react at room temperature for 36 hours; then dialyze with 0.01M PBS solution to remove unreacted small molecule haptens, and obtain the complete antigen phenolphthalein-KLH, which is carried out by ultraviolet absorption scanning method Identification.
[0023] (2) Animal immunization: After mixing and emulsifying the complete phenolphthalein antigen with an equal amount of Freund's adjuvant, BALB / c mice were immunized with multiple subcutaneous inje...
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Abstract
A hybridoma cell line JS secreting anti-phenolphthalein monoclonal antibody and its application belong to the field of food safety immunoassay. A hybridoma cell line JS that secretes an anti-phenolphthalein monoclonal antibody of the present invention has been deposited in the General Microbiology Center CGMCC of the China Microbiological Culture Collection Management Committee. The preservation date is March 7, 2019, and the preservation number is CGMCC No.17394. The invention mixes and emulsifies the complete antigen of phenolphthalein with the same amount of Freund's adjuvant, and immunizes BALB / c mice through multi-point subcutaneous injection on the back of the neck. The cells were then screened by indirect competitive ELISA and subcloned three times, and finally a monoclonal antibody hybridoma cell line was obtained. The monoclonal antibody secreted by this cell line has good specificity and detection sensitivity to phenolphthalein (IC 50 The value is 15.4 ng / mL), which can realize the detection of phenolphthalein residues in health food, and provides raw materials for the immunological detection of phenolphthalein residues in food, which has practical application value.
Description
technical field [0001] The invention relates to a hybridoma cell line JS that secretes anti-phenolphthalein monoclonal antibody and its application, which belongs to the field of food safety immune detection. Background technique [0002] Phenolphthalein (Phenolphthalein) is a weak organic acid, which is white or slightly yellowish fine crystals at room temperature. It has multiple uses. Phenolphthalein is colorless in acidic solution and red in lye or alkali metal carbonate solution. It is often used as an indicator of acid-base titration in chemical experiments. In organic synthesis, it is mainly used to synthesize plastics, especially phthalazinone polyaryletherketone polymers. [0003] Phenolphthalein is used as a contact laxative in medicine, which stimulates the colonic mucosa to strengthen and organize the absorption of intestinal fluid, and accumulates water and electrolytes in the colon to produce a laxative effect and promote defecation. It is clinically used as ...
Claims
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IPC IPC(8): C12N5/20C07K14/795C07K16/44G01N33/577C12R1/93
CPCC07K16/44C07K14/795C07K19/00G01N33/577
Inventor 胥传来陈雨汐匡华徐丽广马伟刘丽强吴晓玲宋珊珊胡拥明
Owner JIANGNAN UNIV