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Antibody pointed at SARS coronavirus N protein antigen and its use in detecting SARS coronavirus or its antigen

A coronavirus, protein antigen technology, applied in the field of antibodies against SARS coronavirus N protein antigen and its application in SARS coronavirus or its antigen detection

Inactive Publication Date: 2005-03-09
马杰
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

There is no report on using N antigen as the detection antigen

Method used

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  • Antibody pointed at SARS coronavirus N protein antigen and its use in detecting SARS coronavirus or its antigen
  • Antibody pointed at SARS coronavirus N protein antigen and its use in detecting SARS coronavirus or its antigen

Examples

Experimental program
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Embodiment 1

[0063] Embodiment 1, N protein gene synthesis:

[0064] The full-length nucleic acid sequence of the N protein is 1269bp, and the full-length protein sequence has a total of 422 amino acids. Because the SARS virus is the pathogen of a serious life-threatening infectious disease transmitted through the air, it is safe and simple to artificially synthesize it from safety considerations. According to the SARS coronavirus nucleocapsid protein (NP) gene complete coding region nucleotide sequence (Locus: AY307165 Protein_id=" AAP49024.1 " published in GenBank on June 19, 2003) synthetic whole coding region sequence, this sequence is shown in SEQ ID NO: 1, the deduced amino acid sequence of which is shown in SEQ ID No: 26.

[0065] The strategy for the synthesis of N protein-coding genes is as follows:

[0066] 1 3 5 7 9 11 13 15 17 19

[0067] 5' - - - - - - - - - 3'

[0068] 3' - - - - - - - - - 5'

[0069] 2 4 6 8 10 12 14 16 18 20

[0070] To complete the synthesis of the c...

Embodiment 2

[0105] Embodiment 2. Utilize goat anti-SARS coronavirus N protein polyclonal antibody to carry out double antibody sandwich analysis

[0106] (1) Preparation of polyclonal antibodies

[0107] According to conventional methods, goats were immunized with the full-length SARS virus N protein prepared in Example 1, and boosted. Polyclonal antibodies were isolated from immunized goat sera and purified with ammonium sulfate and dialyzed against PBS. precipitation.

[0108] (2) Conjugate polyclonal antibodies to colloidal gold particles

[0109] Using the colloidal gold labeling kit of Arista Biologicals Inc., the goat polyclonal antibody against SARS virus N protein was conjugated to 40nm colloidal gold particles for use.

[0110] (3) Prepare test strips in a conventional manner, wherein the goat anti-SARS coronavirus N protein polyclonal antibody conjugated with colloidal gold is the first antibody, and the simple goat anti-SARS virus N protein polyclonal antibody is used as "ca...

Embodiment 3

[0111] The preparation of embodiment 3 mouse anti-SARS-N protein monoclonal antibody:

[0112] I. Antigen preparation: The full-length SARS-N recombinant protein antigen prepared in Example 1 was dissolved in 1×PBS to prepare a 0.5 mg / ml working solution.

[0113] II. Mice: Balb / c female mice, 5-6 weeks old. Immunization begins at 8-9 weeks of age.

[0114] III. Preparation of hybridoma:

[0115] 1). Mix 1.5ml of full-length SARS-N protein with a concentration of 1.0mg / ml and an equal volume of complete Freund's adjuvant.

[0116] 2). Primary immunization of mice: each mouse was injected with 0.5ml, and a total of 6 mice were immunized.

[0117] 3). Booster immunization: start booster immunization on the 14th day after the initial immunization, and then booster immunization every 14 days. The booster immunization was carried out with a mixture of antigen and incomplete Freund's adjuvant, and a total of 3 booster immunizations were performed. On the 10th day after the thir...

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Abstract

An antibody against the nucleocapsidprotein of SARS coronavirus, the method for using said antibody to detect the SARS coronavirus or its antigen, and the reagent kit and relative products for said method are disclosed.

Description

technical field [0001] The present invention relates to an antibody against SARS coronavirus nucleocapsid protein (nucleocapsid protein), and a method for detecting the presence of SARS coronavirus or its antigen in a sample by using the antibody. The invention also relates to kits for these methods and corresponding products. Background technique [0002] Severe Acute Respiratory Syndrome (SARS) is an acute and fatal viral infectious disease. From the first infection reported in Guangdong Province, China, it spread to 32 countries in the world in just a few months. As of June 11, 2003, there were 8,400 infected persons and 789 deaths in the world (WHO statistics). It is a viral infection that threatens the world due to its rapid spread, high mortality rate, and lack of effective treatment. This disease is highly contagious, seriously affects the health of patients, and has a high fatality rate. [0003] There is definite evidence that SARS is caused by a previously unkn...

Claims

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Application Information

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IPC IPC(8): C07K16/10G01N33/563G01N33/569G01N33/577
Inventor 马杰
Owner 马杰
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