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Hybridoma cell strain capable of secreting tigecycline monoclonal antibody and application of hybridoma cell strain

A technology of hybridoma cell line and monoclonal antibody, which is applied in the field of immunoassay, can solve the problems of long detection time, inapplicability of rapid detection, complicated processing, etc., and achieve good detection sensitivity and specificity

Active Publication Date: 2021-02-19
JIANGNAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, these methods have shortcomings such as complicated sample pretreatment and long detection time, and are not suitable for rapid detection. Therefore, it is necessary to establish an efficient and rapid detection method for tigecycline

Method used

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  • Hybridoma cell strain capable of secreting tigecycline monoclonal antibody and application of hybridoma cell strain

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0046] Example 1: Tigecycline Haptens

[0047] Since the small molecule of tigecycline is not immunogenic and cannot stimulate the mice to produce an immune response and then produce antibodies, it is necessary to couple tigecycline to the protein through protein linkage technology to obtain immunogenicity;9 -Aminominocycline is similar in structure to tigecycline and contains a reactive group NH 2 , so it is the tigecycline hapten.

[0048] The tigecycline hapten structure of the present invention is as follows:

[0049] ;

Embodiment 2

[0050] Example 2: Synthesis of Tigecycline Complete Antigen

[0051] Weigh 11.42mg tigecycline hapten TGC-NH 2 Dissolve in anhydrous N,N-dimethylformamide DMF, add 57 μL 1M HCl in an ice bath and avoid light, add excess NaNO after 30 min 2 React for 30 minutes to obtain solution A; dilute 6 mg of keyhole limpet hemocyanin KLH with 0.01M carbonate buffer solution CBS to 3 mg / mL to obtain solution B; add solution A to solution B and react for 2 hours to obtain a reaction solution; Dialyze the reaction solution with 0.01M phosphate buffered saline PBS to obtain the complete antigen TGC-NH 2 -KLH, and identified by UV absorption scanning method.

Embodiment 3

[0052] Embodiment 3: the synthesis of tigecycline coating former

[0053] Weigh 8.72mg tigecycline hapten TGC-NH 2 Dissolve in anhydrous N,N-dimethylformamide DMF, add 32 μL 1M HCl in ice bath and avoid light, add excess NaNO after 30 min 2 React for 30 minutes to obtain liquid A; dilute 6 mg of chicken ovalbumin (OVA) with 0.01M carbonate buffer solution CBS to 3 mg / mL to obtain liquid B; add liquid A to liquid B and react for 2 hours to obtain a reaction liquid; The reaction solution was dialyzed with 0.01M phosphate buffered saline PBS to obtain the coated original TGC-NH 2 -OVA, and identified by UV absorption scanning method.

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Abstract

The invention discloses a hybridoma cell strain capable of secreting a tigecycline monoclonal antibody and application of the hybridoma cell strain, and belongs to the technical field of immunodetection. The hybridoma cell strain ABC03 capable of secreting the tigecycline monoclonal antibody is preserved in the China General Microbiological Culture Collection Center (CGMCC), the preservation dateis November 28, 2019, and the preservation number is CGMCC No.19176. The tigecycline monoclonal antibody secreted by the strain is used for analyzing and detecting the blood concentration of tigecycline. The tigecycline monoclonal antibody cell strain obtained in the invention can be used for immunoassay detection, has good detection sensitivity and specificity (IC50 value is 2.3 ng / mL) on tigecycline, and the cross rate on tigecycline analogues (cross rate = (IC50 of tigecycline / IC50 of analogues) * 100% is less than 10%.

Description

technical field [0001] The invention relates to a hybridoma cell strain secreting tigecycline monoclonal antibody and application thereof, belonging to the technical field of immunoassay. Background technique [0002] Tigecycline (tigecycline) is a broad-spectrum glycyl cyclic peptide antibiotic administered intravenously, belonging to the third-generation tetracycline antibiotics. In June 2005, the US FDA approved the drug for complex skin and soft tissue infections in adults. and complicated intra-abdominal infections in adults. Tigecycline is a new generation of tetracycline antibiotics developed after doxycycline, minocycline and metacycline. Compared with tetracycline, on the side chain of the central skeleton, the N-alkyl-glycylamino group is replaced by the D-ring glycyl ring at the 9th position, which makes the antibacterial spectrum wider and the antibacterial activity stronger. It can overcome the mechanism of resistance of most bacteria to tetracycline. However...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/20C07K16/44C07K14/795C07K14/77C07C237/48G01N33/577C12R1/91
CPCC07K16/44C07K14/795C07K14/77C07C237/48G01N33/577G01N33/9446C07C2603/46
Inventor 胥传来路倩倩匡华徐丽广孙茂忠刘丽强宋珊珊吴晓玲郝昌龙胡拥明
Owner JIANGNAN UNIV
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