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2019-nCoV N protein linear epitope peptide, monoclonal antibody and application

A monoclonal antibody, 2019-ncov technology, applied in the field of immunology, can solve the problem that there is no specific treatment for the disease

Active Publication Date: 2021-06-15
INST OF MICROBIOLOGY - CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] There is currently no specific treatment for the disease caused by the new coronavirus

Method used

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  • 2019-nCoV N protein linear epitope peptide, monoclonal antibody and application
  • 2019-nCoV N protein linear epitope peptide, monoclonal antibody and application
  • 2019-nCoV N protein linear epitope peptide, monoclonal antibody and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0046] Embodiment 1, establishment of hybridoma cell line

[0047] 1. Experimental materials

[0048] 1. Immunogen: In this example, polypeptides 1-5 were coupled to the carrier protein KLH, and the KLH-coupled polypeptides were used as immunogens to immunize animals.

[0049] 2. Experimental animals: BALB / c mice, 4-6 weeks old, female, SPF grade.

[0050] 3. Other materials: Freund's complete adjuvant and Freund's incomplete adjuvant were purchased from Sigma Company; HRP-labeled goat anti-mouse IgG antibody was purchased from JacksonImmune Company; other reagents were domestic analytically pure products.

[0051] 2. Establishment of hybridoma cell lines

[0052] 1. Animal immunization

[0053] 1) Basic immunization: the antigen was mixed with Freund's complete adjuvant in equal volumes and fully emulsified, and injected subcutaneously in points, each injection volume of each Balb / c mouse was 100 μg.

[0054] 2) Booster immunization: The emulsion of antigen and Freund's i...

Embodiment 2

[0063] Example 2 Preparation and evaluation of anti-new coronavirus mouse monoclonal antibody

[0064] 1. Antibody preparation

[0065] Adult BALB / c mice were selected and inoculated intraperitoneally with Freund's incomplete adjuvant, 0.5ml per mouse. After 7-10 days, the 16th generation hybridoma cells were inoculated intraperitoneally, 1×10 per mouse 6 -2×10 6 indivual. After an interval of 5 days, when the abdomen is obviously enlarged and the skin feels tense when touched with hands, the ascites can be collected with a No. 9 needle.

[0066] Centrifuge the ascitic fluid (13000r / min for 30 minutes), remove cell components and other precipitates, and collect the supernatant. Purify with Protein G~Sepharose CL-4B, the upper column liquid is 20mM PBS buffer, and the eluent of column chromatography is: pH 2.7, 20mM glycine buffer, respectively, to obtain the mouse monoclonal anti-new coronavirus N protein Cloned antibodies. Among them, the monoclonal antibody Clone 4G1 w...

Embodiment 3

[0084] Example 3, Application of Purified Antibody Clone 4G1 and Clone 11D5 to Prepare New Coronavirus N Protein Detection Reagent

[0085] Clone 4G1 and Clone 11D5 antibodies were used for paired detection experiments, and Clone4G1 was determined as the capture antibody, and HRP-labeled Clone 11D5 was used as the detection antibody. The ELISA detection method was determined, and the detection sensitivity reached 2.5ng / mL. See Figure 9 .

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Abstract

The invention provides a 2019-nCoV N protein linear epitope peptide, a monoclonal antibody and application. The amino acid sequence of the 2019-nCoV N protein linear epitope peptide is as shown in any one of SEQ ID NO.1-5. The polypeptide carrier protein containing the linear epitope peptide is coupled with an immunized mouse to prepare a plurality of hybridoma cells which secrete monoclonal antibodies which can specifically recognize2019-nCoV N protein and have biological activity.

Description

[0001] This application claims that the application date is February 4, 2020, the application number is 2020100799132, and the invention title is "2019 Novel Coronavirus N Protein Linear Epitope Peptide and Monoclonal Antibody and Its Application". technical field [0002] The present invention relates to the field of immunology and antibody preparation technology, in particular to 2019 novel coronavirus N protein linear epitope peptide, monoclonal antibody and application. Background technique [0003] Coronaviruses are a large family of viruses known to cause colds as well as more serious illnesses such as Middle East Respiratory Syndrome (MERS) and Severe Acute Respiratory Syndrome (SARS). The new coronavirus is a new strain of coronavirus that has never been found in humans before. The common signs of people infected with coronavirus include respiratory symptoms, fever, cough, shortness of breath and dyspnea. In more severe cases, infection can lead to pneumonia, severe ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/165C07K16/10C07K16/46C07K19/00G01N33/569G01N33/577C12N5/20A61K39/215A61K39/42A61P31/14A61P11/00
CPCC07K14/005C07K16/10G01N33/56983G01N33/577A61K39/12A61P31/14A61P11/00C12N2770/20022C12N2770/20034C07K2317/56C07K2317/31C07K2317/92C07K2319/00G01N2333/165G01N2469/10A61K2039/505
Inventor 高福孙乐宋豪李茂华
Owner INST OF MICROBIOLOGY - CHINESE ACAD OF SCI
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