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Anti-quinolone antibiotic class specific monoclonal antibody hybridoma cell strain YH6 and application thereof

A hybridoma cell line and monoclonal antibody technology, applied in the field of immunochemistry, can solve the problems of inapplicable daily rapid detection of quinolone antibiotics

Active Publication Date: 2017-03-22
JIANGNAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, these methods require expensive instruments, professional operators and complicated sample pretreatment, and are not suitable for daily rapid detection of quinolone antibiotics in food

Method used

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  • Anti-quinolone antibiotic class specific monoclonal antibody hybridoma cell strain YH6 and application thereof
  • Anti-quinolone antibiotic class specific monoclonal antibody hybridoma cell strain YH6 and application thereof
  • Anti-quinolone antibiotic class specific monoclonal antibody hybridoma cell strain YH6 and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0018] Example 1 Preparation of hybridoma cell line YH6

[0019] 1. Animal immunity and serum screening

[0020] The conjugate of hapten 1 (H1) and hemocyanin (KLH) (H1-KLH) was selected as the immunogen, and the complete adjuvant was mixed with the immunogen at a volume of 1:1 for the first dose, and BALB / c was immunized by subcutaneous immunization For mice, the interval between immunizations was 4 weeks, and 7-10 days after five immunizations, the titer and inhibition of serum were measured by indirect competitive enzyme-linked immunosorbent assay, and mice with high titer and good inhibition were selected for fusion.

[0021] 2. Cell Fusion

[0022] 7 to 10 days before fusion, in RPMI-1640 medium containing 10% fetal bovine serum in 5% CO 2 Tumor cells were cultured in an incubator. Three days before the fusion, the selected mice were immunized by abdominal sprinting. On the day of fusion, the eyeballs were removed to collect blood. After the mice were killed by cervic...

Embodiment 2

[0027] Embodiment 2 Preparation and purification of the anti-quinolone antibiotic monoclonal antibody secreted by hybridoma cell line YH6

[0028] 1. Preparation of monoclonal antibodies

[0029] Select healthy BALB / c mice and inject sterile paraffin oil at a volume of 0.6 mL / mouse. After 10 days, each mouse was intraperitoneally injected with 1×10 6 hybridoma cells. After the 6th day, observe the state of the mouse every day. If the abdomen of the mouse is obviously enlarged, feels tight when touched by hands, and is unwilling to move, collect ascites; then centrifuge (6000 rpm, 12 min) to remove red blood cells Wait for impurities, aliquot, and freeze at -20°C.

[0030] 2. Monoclonal antibody purification

[0031] Ascitic fluid was purified by octanoic acid-ammonium sulfate precipitation. Under acidic conditions, n-octanoic acid can precipitate other miscellaneous proteins in ascites except IgG immunoglobulin, then centrifuge, and discard the precipitate; then use a cer...

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Abstract

An anti-quinolone antibiotic class specific monoclonal antibody hybridoma cell strain YH6 and an application thereof belong to the technical field of immunochemistry. The monoclonal cell strain YH6 is preserved in China General Microbiological Culture Collection Center with the preservation number of CGMCC No.12024. A monoclonal antibody secreted by the YH6 is detected by indirect competitive enzyme-linked immunosorbent assay, and has cross reaction with the following 21 pyrethroids: norfloxacin, ofloxacin, enrofloxacin, ciprofloxacin, flumequine, nafloxacin, enoxacin, lomefloxacin, levofloxacin, pefloxacin, nalidixic acid, danofloxacin, pyridine acid, cinoxacin, oxolinic acid, marbofloxacin, pazufloxacin, sparfloxacin, gatifloxacin, orbifloxacin and fleroxacin, and the IC50 value of the monoclonal antibody is 0.1-50 ng / mL. The class specific monoclonal antibody can be used for developing colloidal gold immunochromatographic test strips and immunosensors, provides a raw material for the immunodetection of quinolone antibiotic residues in foods, and has practical application values.

Description

technical field [0001] The invention relates to a mouse-derived hybridoma cell strain YH6 and the group-selective monoclonal antibody produced by it that can simultaneously recognize 21 kinds of quinolone antibiotics, which can be used in the development of colloidal gold immunochromatographic test strips and immunosensors. It belongs to the technical field of immunochemistry. Background technique [0002] Quinolones are a class of synthetic antibacterial drugs with 4-quinolone as the basic structure. They act on bacterial deoxyribonucleic acid (DNA), hinder DNA gyrase, and cause irreversible damage to bacterial DNA to achieve antibacterial effects. After more than forty years of development, quinolones are divided into four generations. Quinolones antibiotics are widely used in clinical and animal anti-infection because of their broad antibacterial spectrum, high bioavailability, and few adverse reactions. However, many quinolone antibiotics have been confirmed to have a ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/20C07K16/44G01N33/577C12R1/91
CPCC07K16/44G01N33/577G01N2430/00
Inventor 刘丽强彭娟胥传来匡华徐丽广宋珊珊吴晓玲胡拥明
Owner JIANGNAN UNIV
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