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PBP2a monoclonal antibody as well as preparation method and application thereof

A monoclonal antibody, 1B4 technology, applied in the direction of antibodies, chemical instruments and methods, antibacterial drugs, etc., can solve the problems of low purity titer, poor specificity, long time consumption, etc., and achieve high sensitivity, strong specificity and high purity. Effect

Active Publication Date: 2021-07-23
深圳市龙华区疾病预防控制中心
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] However, the existing PBP2a detection method has poor specificity and takes a long time, which delays the diagnosis and treatment of patients; the monoclonal antibody of PBP2a protein in the prior art has low purity and low titer and poor specificity

Method used

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  • PBP2a monoclonal antibody as well as preparation method and application thereof
  • PBP2a monoclonal antibody as well as preparation method and application thereof
  • PBP2a monoclonal antibody as well as preparation method and application thereof

Examples

Experimental program
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Embodiment

[0044] Experimental steps and methods:

[0045] 1. Acquisition of PBP2a gene

[0046] Obtain the amino acid sequence of the PBP2a gene from the GenBank database, optimize the codon of the base sequence of the PBP2a gene according to the codon preference of Escherichia coli, and entrust Sangon Bioengineering (Shanghai) Co., Ltd. to synthesize the optimized codon gene sequence. PBP2a encodes 668 amino acids.

[0047] PBP2a amino acid sequence:

[0048] MKKIKIVPLILIVVVVGFGIYFYASKDKEINNTIDAIEDKNFKQVYKDSSYISKSDNGEVEMTERPIKIYNSLGVKDINIQDRKIKKVSKNKKRVDAQYKIKTNYGNIDRNVQFNFVKEDGMWKLDWDHSVIIPGMQKDQSIHIENLKSERGKILDRNNVELANTGTAYEIGIVPKNVSKKDYKAIAKELSISEDYIKQQMDQNWVQDDTFVPLKTVKKMDEYLSDFAKKFHLTTNETESRNYPLGKATSHLLGYVGPINSEELKQKEYKGYKDDAVIGKKGLEKLYDKKLQHEDGYRVTIVDDNSNTIAHTLIEKKKKDGKDIQLTIDAKVQKSIYNNMKNDYGSGTAIHPQTGELLALVSTPSYDVYPFMYGMSNEEYNKLTEDKKEPLLNKFQITTSPGSTQKILTAMIGLNNKTLDDKTSYKIDGKGWQKDKSWGGYNVTRYEVVNGNIDLKQAIESSDNIFFARVALELGSKKFEKGMKKLGVGEDIPSDYPFYNAQISNKNLDNEILLADSGYGQGEILINPVQILSI...

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Abstract

The invention discloses a PBP2a monoclonal antibody as well as a preparation method and application thereof. The preparation process of the PBP2a monoclonal antibody comprises the following steps: synthesizing a staphylococcus aureus PBP2a gene, expressing and purifying PBP2a protein, immunizing a Balb / c mouse by taking the recombinant PBP2a protein as an immunogen, fusing splenocytes of the immunized mouse with myeloma NS-1 cells of the mouse, screening hybridoma cells secreting specific McAb through an indirect ELISA method, inducing the mouse to generate ascites by using a hybridoma cell strain, purifying the antibody by using a protein G affinity chromatography method, and finally obtaining the PBP2a protein monoclonal antibody. The PBP2a monoclonal antibody disclosed by the invention can be combined with PBP2a. The obtained PBP2a monoclonal antibody has a titer of more than 2 million, and the monoclonal antibody of the PBP2a protein has the advantages of high purity, strong specificity and high sensitivity, and can be used for preparing a diagnostic reagent for detecting the PBP2a gene.

Description

technical field [0001] The invention relates to the field of immunotherapy biomedicine, in particular to a PBP2a monoclonal antibody and its preparation method and application. Background technique [0002] Due to the widespread use of broad-spectrum antibacterial agents, the proportion of methicillin-resistant Staphylococcus aureus (MRSA) in clinically isolated Staphylococci has been increasing, and it has developed into one of the main pathogens of serious infections in hospitals. Since MRSA was first discovered in early 1961, the infection of MRSA is spreading and becoming popular all over the world at an alarming speed. Therefore, the rapid detection of MRSA in clinical samples is of great significance. Current studies have shown that the main cause of MRSA is the production of a penicillin-binding protein PBP2a by Staphylococcus aureus. This protein is present on the cell surface by drug-resistant genes and has extremely low affinity for β-lactam antibacterial agents....

Claims

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Application Information

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IPC IPC(8): C07K16/12A61K39/40A61P31/04G01N33/577G01N33/569
CPCC07K16/1271A61P31/04G01N33/577G01N33/56938C07K2317/56C07K2317/76Y02A50/30
Inventor 邬玉兰汪涛刘丽陈钰羊林启辉任燕许少坚
Owner 深圳市龙华区疾病预防控制中心
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