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Novel coronavirus nucleocapsid protein antibody or antigen binding fragment for in vitro diagnosis

A technology of nucleocapsid protein and coronavirus, which is applied in the direction of antiviral immunoglobulin, antibody, antiviral agent, etc., can solve the problems of low sensitivity, prone to missed detection, prone to false positive results, etc., to achieve high sensitivity and specific effect

Active Publication Date: 2022-02-01
北京健乃喜生物技术有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The main manifestations are: 1) The binding affinity between the antibody and the antigen is not high enough, resulting in low detection sensitivity, which is prone to missed detection in practical applications
2) Antibodies are easily interfered by other substances in the sample, resulting in poor specificity of detection and prone to false positive results in practical applications

Method used

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  • Novel coronavirus nucleocapsid protein antibody or antigen binding fragment for in vitro diagnosis
  • Novel coronavirus nucleocapsid protein antibody or antigen binding fragment for in vitro diagnosis
  • Novel coronavirus nucleocapsid protein antibody or antigen binding fragment for in vitro diagnosis

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Embodiment 1

[0065] Antibodies were obtained by the hybridoma method. Specific steps are as follows:

[0066] The new coronavirus nucleocapsid protein was used as an antigen to immunize mice. Mouse spleens were collected and splenocytes were prepared. Mouse splenocytes were fused with Myeloma cells and stable hybridoma cells were screened. Antigen-specific antibodies were then screened by ELISA.

[0067] The obtained highly specific antibodies were sequenced. One of the antibody numbers is mAb#9, its heavy chain variable region sequence is shown in SEQ ID NO:17, and its light chain variable region sequence is shown in SEQ ID NO:18.

[0068] Then the coding sequence of the antibody was introduced into pCMV_HC and pCMV_LC antibody expression vectors. The pCMV_HC and pCMV_LC vectors were transfected into CHO cell lines by liposome method for expression of recombinant antibodies. The antibody was separated and purified with Protein A affinity column, and the purity reached more than 95%....

Embodiment 2

[0073] Embodiment 2 provides a kind of test strip, such as figure 1 shown. figure 1 The middle mark 1 is the sample pad (glass fiber film), 2 is the colloidal gold pad, 3 is the nitrocellulose membrane, 4 is the absorbent paper, 5 is the bottom plate, 6 is the detection line (also known as the T line), 7 is the quality Control line (also known as C line). The preparation method of each part in the test strip is as follows:

[0074] 1. Preparation of nitrocellulose membrane

[0075] Preparation of coating buffer: 0.05M pH8.5 PBS buffer is used as coating buffer, filtered through 0.22 μm membrane, and stored at 4°C for later use. The buffer formula: NaCl 40g, KCl 1g, NaCl 2 HPO 4 12H 2 O 14.5g, KH 2 PO 4 1g, dilute to 1000mL with double-distilled deionized water.

[0076] Preparation of nitrocellulose membrane: Dilute the recombinant antibody (mAb#4) of the new coronavirus nucleocapsid protein obtained in Example 1 to 0.5-2 mg / mL with coating buffer, adjust the machine...

Embodiment 3

[0096] Embodiment 3 adopts the test strip of embodiment 2 to verify the detection performance of novel coronavirus:

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Abstract

The present invention provides a novel coronavirus nucleocapsid protein antibody or antigen binding fragment for in vitro diagnosis. The antibody comprises a heavy chain variable region and a light chain variable region, the heavy chain variable region comprises CDR sequences as shown in SEQ ID NO: 11, 12 and 13, and the light chain variable region comprises CDR sequences as shown in SEQ ID NO: 14, 15 and 16. The antibody provided by the invention is used for in-vitro detection of novel coronavirus, and has extremely high sensitivity and specificity.

Description

technical field [0001] The invention relates to the field of medical detection, in particular to a novel coronavirus nucleocapsid protein antibody or antigen-binding fragment for in vitro diagnosis. Background technique [0002] Since the outbreak of the novel coronavirus (SARS-COV-2) in December 2019, it has spread widely around the world. After it infects the human body, it will lead to the new type of coronavirus pneumonia (COVID-19) with a high probability. It is highly contagious and pathogenic. Impact. [0003] After the outbreak of the new coronavirus, in order to realize the diagnosis of the disease and block the chain of virus transmission, a variety of new coronavirus in vitro diagnostic technologies have come out. The main methods include new coronavirus nucleic acid detection, new coronavirus antibody detection and new coronavirus antigen Several types are detected. Antibody detection is the main detection method in the early stage of the epidemic, but it is a...

Claims

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Application Information

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IPC IPC(8): C07K16/10C12N15/13A61K39/42A61P31/14G01N33/569G01N33/577G01N33/558G01N33/58
CPCC07K16/10A61P31/14G01N33/56983G01N33/577G01N33/558G01N33/587C07K2317/565C07K2317/56C07K2317/76G01N2333/165G01N2469/10
Inventor 陈亚萍尹午山王美娜张磊贾晨晨石云敬赵雷张丛
Owner 北京健乃喜生物技术有限公司
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