阅读说明：本技术 一种抗新型冠状病毒n蛋白的中和抗体 (Neutralizing antibody for resisting novel coronavirus N protein ) 是由 吴信忠 蔡小辉 陈泓霖 于 2020-07-03 设计创作，主要内容包括：本发明公开了一种抗新型冠状病毒N蛋白的中和抗体,它由轻链和重链组成,所述轻链可变区的氨基酸序列如SEQ ID NO：1所示,所述重链可变区的氨基酸序列如SEQ ID NO：2所示。本发明所述抗体能够特异性识别和结合SARS-CoV-2 N蛋白,实验结果显示,其与2019-NCOV Nucleocapsid的亲和常数KD值为111nM,EC50值为3.975μg/mL；其与NCOV-SARI-N-his的EC50值为4.752μg/mL。(The invention discloses a neutralizing antibody for resisting novel coronavirus N protein, which consists of a light chain and a heavy chain, wherein the amino acid sequence of a variable region of the light chain is shown as SEQ ID NO: 1, and the amino acid sequence of the heavy chain variable region is shown as SEQ ID NO: 2, respectively. The antibody can specifically recognize and combine SARS-CoV-2N protein, and the experimental result shows that the affinity constant KD value of the antibody and 2019-NCOV Nucleocapsid is 111nM, and the EC50 value is 3.975 mug/mL; it has an EC50 value of 4.752. mu.g/mL with NCOV-SARI-N-his.)

1. A neutralizing antibody against N protein of novel coronavirus, which consists of a light chain and a heavy chain, and is characterized in that the amino acid sequence of the variable region of the light chain is shown as SEQ ID NO: 1, and the amino acid sequence of the heavy chain variable region is shown as SEQ ID NO: 2, respectively.

2. The neutralizing antibody against the N protein of novel coronavirus according to claim 1, wherein the amino acid sequence of the light chain of said antibody is as shown in SEQ ID NO: 3, the amino acid sequence of the heavy chain is shown as SEQ ID NO: 4, respectively.

3. Use of the antibody of claim 1 for the preparation of a reagent for the diagnosis of a novel coronavirus, or for the preparation of a medicament for the treatment of a novel coronavirus.

4. A product for the diagnosis and/or treatment and/or prevention of diseases caused by a novel coronavirus, wherein the antibody of claim 1 is the active ingredient in the product.

Technical Field

The invention relates to a virus from RNA, in particular to a neutralizing antibody for resisting novel coronavirus N protein.

Background

2019 the pathogen of the novel coronavirus is a novel coronavirus (2019-nCoV, COVID-19 or SARS-CoV-2). SARS-CoV-2 genus Coro-naviridae (Coronavir) genus Coronavirus (Coronavir) is an enveloped single-stranded positive-strand RNA virus. The novel coronavirus gene codes a plurality of structural proteins, such as N protein, M protein, E protein, S protein and the like, wherein the S protein can be specifically combined with a host receptor and is a key protein for virus invasion to host susceptible cells, the M protein and the E protein refer to the formation of virus cell membranes, and the N protein is involved in the assembly of the virus. The proteins comprise a plurality of antigen epitopes, and the existence of the antigen can be detected through the antibody by utilizing the principle that the antigen is specifically combined with the antibody, so that the fact that the sample contains the novel coronavirus is directly proved.

At present, various research organizations are dedicated to research and develop antibody products aiming at the S protein of SARS-CoV-2, such as the invention patent with publication number CN111333722A, the invention patent with publication number CN111303280A adopts phage display technology to construct a high-capacity human immune phage antibody library, and SARS-CoV-2-S protein is taken as a target, a human antibody single-chain antibody fragment is screened and an antibody with strong neutralizing effect on SARS-CoV-2 virus is obtained, for example, the invention patent with publication number CN111303280A discloses a fully human monoclonal antibody for resisting SARS-CoV-2, the affinity constant KD of the monoclonal antibody and the S antigen is 1. 1.0 nM., the antibody products aiming at the N protein of the novel coronavirus are few, 111269313A is published as the invention patent with publication number CN 8625, and the result of indirect measurement of the monoclonal antibody is obtained by adopting ELISA method for measuring the relative affinity of the antibody and the S protein as the result of the invention antibody with publication number 1.835 × 10^-10mol/L。

Disclosure of Invention

The invention aims to provide a neutralizing antibody for resisting a novel coronavirus N protein, which has ideal affinity with the N protein.

The neutralizing antibody for resisting the novel coronavirus N protein consists of a light chain and a heavy chain, wherein the amino acid sequence of a variable region of the light chain is shown as SEQ ID NO: 1, the amino acid sequence of the heavy chain variable region is shown as SEQ ID NO: 2, respectively.

The amino acid sequence of the light chain of the neutralizing antibody of the anti-novel coronavirus N protein is shown as SEQ ID NO: 3, the amino acid sequence of the heavy chain is shown as SEQ ID NO: 4, respectively. The antibody is an antibody product aiming at novel coronavirus N protein (SARS-CoV-2N protein), and the amino acid sequence of the novel coronavirus N protein is shown as SEQ ID NO: 5, respectively.

The applicant finds that the antibody of the invention can specifically recognize and bind to SARS-CoV-2N antigen protein, therefore, the application also includes the application of the antibody in preparing reagent for diagnosing new type coronavirus or preparing medicine for treating new type coronavirus.

The present application further includes a product for diagnosing and/or treating and/or preventing diseases caused by the novel coronavirus, which contains the antibody of the present invention as an active ingredient.

Compared with the prior art, the antibody can specifically recognize and bind SARS-CoV-2N protein, and the experimental result shows that the affinity constant KD value of the antibody and 2019-NCOV Nucleocapsid (prokaryotic expression N protein) is 111nM, and the EC50 value is 3.975 mug/mL; the EC50 value of the protein was 4.752. mu.g/mL compared with NCOV-SARI-N-his (eukaryotic expression N protein).

Drawings

FIG. 1 is a SDS-PAGE detection profile of neutralizing antibodies against the N protein of the novel coronavirus after purification in an affinity column, wherein M represents the molecular weight of the standard protein; me represents the combined media; FT represents flow-through; w represents a washing solution; e represents the purified neutralizing antibody against the N protein of the novel coronavirus.

FIG. 2 is an electrophoresis chart of protein expression of a neutralizing antibody of Western Blot for verifying N protein of the novel coronavirus, wherein M represents the molecular weight of a standard protein; e represents the purified neutralizing antibody against the N protein of the novel coronavirus.

Detailed Description

The present invention will be better understood from the following detailed description of specific examples, which should not be construed as limiting the scope of the present invention.

The "Fab 20 antibody" or "Fab 20" appearing in the following is short for the neutralizing antibody against the novel coronavirus N protein of the present invention.