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Murine monoclonal antibody against Zika virus envelope protein

A monoclonal antibody, envelope protein technology, applied in the direction of antiviral immunoglobulin, antiviral agent, immunoglobulin, etc., to achieve the effect of strong neutralization activity, good specificity and enhanced anti-aggregation

Active Publication Date: 2021-09-28
WUHAN INST OF VIROLOGY CHINESE ACADEMY OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, there are no approved vaccines and specific drugs. Here we use the Zika virus envelope protein constructed by ourselves to immunize mice, and obtain a mouse-derived monoclonal antibody with strong neutralizing activity, which can be used in the future for Zika virus. prevention

Method used

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  • Murine monoclonal antibody against Zika virus envelope protein
  • Murine monoclonal antibody against Zika virus envelope protein
  • Murine monoclonal antibody against Zika virus envelope protein

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0015] Example 1: Obtaining of D10 antibody sequence

[0016] Based on the constructed immunized mouse Fab phage display library (the acquisition method of the library comes from the fusion protein of the envelope protein of Flaviviridae virus disclosed in Chinese patent application 201811613477.1 and its preparation method and application, the fusion protein is used as the immunogen for immunization After obtaining the cDNA of the mouse, we used it as the construction material of the phage display library), we carried out three rounds of screening, and through the monoclonal phage ELISA, the positive clones were sequenced, and the base sequence of the D10 candidate clone was obtained , and the amino acid sequence was obtained by amino acid translation software, the amino acid sequence of the light chain is shown in SEQ No.1, and the amino acid sequence of the heavy chain is shown in SEQ No.2.

Embodiment 2

[0017] Example 2: Using Tango software to optimize the obtained D10 amino acid sequence

[0018] Use Tango software (http: / / tango.crg.es / ) to predict the aggregation-prone region of the amino acid sequence of the D10 candidate clone, and find that there is a strong aggregation-prone region, such as Picture 1-1 , so point mutation is carried out in this region to obtain D11 antibody, the amino acid sequence of its light chain is shown in SEQ No.1, the amino acid sequence of its heavy chain is shown in SEQ No.3, the corresponding base sequence, and the light chain is shown in SEQ No. 4, the heavy chain is shown in SEQ No.5. The mutated amino acid sequence was re-predicted by Tango software, and it was found that the aggregation-prone region was significantly improved, as shown in Figure 1-2 , 1-3.

Embodiment 3

[0019] Example 3: 293F cells express D11 antibody and detect it by SDS-PAGE

[0020] The mutated base sequence was cloned into the dual promoter vector pVitro2-neo-mcs (InvivoGen), and the 293F cells were transfected with PEI to express the protein. After 5-7 days of expression, the culture supernatant of the 293F cells was treated with ProteinA (GE) was purified, and after the purified protein was concentrated and replaced, the D11 antibody dissolved in PBS buffer was obtained. After SDS-PAGE electrophoresis detection, it was found that it presented two bands of light chain and heavy chain, such as figure 2 shown.

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PUM

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Abstract

The invention discloses a mouse-derived monoclonal antibody directed at Zika virus envelope protein. After screening and genetic modification, the obtained antibody has good specificity, strong neutralizing activity, and increased anti-aggregation property, and is suitable for Zika virus prevention and treatment.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a mouse-derived monoclonal antibody against Zika virus envelope protein. Background technique [0002] Zika virus (ZIKV) is a virus of the family Flaviviridae. Infection with this virus can cause microcephaly in newborns and affect the neurodevelopment of infants. Guillain-Barré syndrome (GBS) may occur after infection in adults and lead to neurological disorders. damage. At present, there are no approved vaccines and specific drugs. Here we use the Zika virus envelope protein constructed by ourselves to immunize mice, and obtain a mouse-derived monoclonal antibody with strong neutralizing activity, which can be used in the future for Zika virus. Prevention and treatment. Contents of the invention [0003] The purpose of the present invention is to fill the gap in the prior art and provide a mouse-derived monoclonal antibody with good specificity against Zika virus envelope prote...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K16/10C12N15/13A61P31/14
CPCA61P31/14C07K16/1081C07K2317/76C07K2317/92
Inventor 龚睿
Owner WUHAN INST OF VIROLOGY CHINESE ACADEMY OF SCI
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