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Preparation method of monoclonal antibody to chloramphenicol

A technology of monoclonal antibody and chloramphenicol, applied in the direction of microorganism-based methods, biochemical equipment and methods, chemical instruments and methods, etc., can solve the problems of normal flora imbalance, susceptibility to infection, etc., reduce production costs, strengthen Controllable and reproducible effects

Inactive Publication Date: 2012-05-30
ZHEJIANG UNIV OF TECH +1
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

In addition, long-term micro-intake of chloramphenicol not only makes Escherichia coli, Salmonella, etc. produce drug resistance, but also causes the imbalance of normal flora in the body, making people susceptible to various diseases

Method used

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  • Preparation method of monoclonal antibody to chloramphenicol
  • Preparation method of monoclonal antibody to chloramphenicol

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Embodiment Construction

[0019] 1. Materials

[0020] 1.1 Animals, cell lines

[0021] BALB / c mice, Zhejiang Animal Experiment Center; F1 mice, Zhejiang University of Traditional Chinese Medicine Animal Experiment Center; SP2 / 0 myeloma cells, Nanjing KGI Biotechnology Development Co., Ltd.

[0022] 1.2 Main Instruments

[0023] Laminar flow clean bench SCV-4A1, Streamline Laboratory Products; trinocular inverted phase contrast microscope ELWD0.3T1-SNCP, Nikon; carbon dioxide constant temperature incubator 3111, Thermo Electron Corporation; HiTrap r-Protein A FF chromatography column (5mL), AKTA purifier, GE Healthcare; sterile culture plates, flasks, dishes, centrifuge tubes, Corning Company.

[0024] 1.3 Main Reagents

[0025] Immunogen CAP-BSA and detection antigen CAP-OVA, synthesized in our laboratory; chloramphenicol, Shanghai Jingchun Reagent Co., Ltd.; HAT culture medium, HT culture medium, PEG (1500D), Sigma Company; fetal bovine serum, Minhai Bioengineering Co., Ltd.; RPML 1640 culture me...

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Abstract

The invention discloses a preparation method of a monoclonal antibody to chloramphenicol (CAP). The method comprises the following steps of: (1) preparing an antigen of CAP-BSA by a diazotization method; (2) conducting lymph immunization to mice with the CAP-BSA antigen, leaving the antigen and a Freund's complete adjuvant to complete emulsification during the first time immunization, and during the second time and the third time immunization, leaving the antigen and a Freund's incomplete adjuvant to complete emulsification, and keeping an interval of 7 days between each time, and 3 days before fusion, carrying out direct antigen injection to the abdominal cavity to booster immunization; (3) measuring serum titers of the mice by an enzyme-linked immunosorbent assay (ELISA) method; (4) selecting a mouse with the highest serum titer, and taking a spleen cell and a myeloma cell for in vitro fusion; (5) culturing and screening a fusion cell in a selective medium, conducting positive cell detection and screening for cloning culture, further performing positive cell cloning and screening for positive cloning, expanding culture and cryopreservation; (6) injecting a cell strain undergoing expanded culture into the abdominal cavities of the mouse so as to produce a lot of ascites; (7) using a chromatographic column to purify the monoclonal antibody against chloramphenicol in the ascites. The prepared monoclonal antibody against CAP-BSA has the advantages of high sensitivity, strong specificity and good practicality.

Description

technical field [0001] The invention belongs to the technical field of monoclonal antibody preparation, in particular to a preparation method of chloramphenicol monoclonal antibody. Background technique [0002] Chloramphenicol (CAP) is a broad-spectrum antibiotic that has a good inhibitory effect on both Gram-negative and Gram-positive bacteria. Due to its excellent antibacterial properties, stable drug resistance and low price, it is used as a therapeutic drug for bacterial diseases in human and animal clinical applications, and is widely used in animal husbandry and artificial aquaculture production as a feed additive. However, chloramphenicol is toxic and can easily cause blood poisoning in humans, and long-term application can lead to aplastic anemia and agranulocytosis. Long-term consumption of milk contaminated by chloramphenicol may cause "grey baby syndrome". In addition, long-term micro-intake of chloramphenicol not only makes Escherichia coli and Salmonella resi...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K16/44C12N5/20C12R1/91
Inventor 应国清汪竹环易喻陈建澍梅建凤王鸿
Owner ZHEJIANG UNIV OF TECH
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