Recombinant protein, test strip, preparation method and application for detecting 2019 novel coronavirus antibody by double antigen sandwich method

A double-antigen sandwich and coronavirus technology, applied in the direction of virus/bacteriophage, biochemical equipment and methods, viruses, etc., can solve the problems of large influence, unsatisfactory sensitivity, poor specificity, etc., and achieve high sensitivity, good stability, and easy operation easy effect

Active Publication Date: 2021-08-31
GENERAL HOSPITAL OF PLA +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

The reported methodologies of immunological diagnosis are mainly colloidal gold method, ELISA, etc., to detect virus-specific IgM and IgG antibodies in the blood. The impact is relatively large, the specificity is poor, and the sensitivity is not ideal
The double-antigen sandwich method has high sensitivity and strong specificity, but there is no report on the ideal double-antigen sandwich raw materials for 2019-nCoV on the market and in the literature

Method used

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  • Recombinant protein, test strip, preparation method and application for detecting 2019 novel coronavirus antibody by double antigen sandwich method
  • Recombinant protein, test strip, preparation method and application for detecting 2019 novel coronavirus antibody by double antigen sandwich method

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preparation example Construction

[0034] The present invention also provides a preparation method for the test strip described in the above technical scheme, comprising the following steps:

[0035] 1) Mix and couple the activated fluorescent microspheres with the recombinant protein described in the above technical scheme to obtain the multi-dominant epitope fusion protein of 2019 novel coronavirus labeled with fluorescent microspheres;

[0036] 2) spraying the 2019 novel coronavirus multi-dominant epitope fusion protein labeled with fluorescent microspheres onto the glass fiber membrane to obtain a fluorescent microsphere pad;

[0037] 3) Spray the 2019 novel coronavirus multi-dominant epitope fusion protein and the anti-N protein antibody on the detection area and the quality control area of ​​the nitrocellulose membrane respectively to obtain a nitrocellulose membrane with the detection area and the quality control area fixed;

[0038] 4) Paste the sample absorbent pad, the fluorescent microsphere pad, the...

Embodiment 1

[0048] Preparation of reagent strips for detection of novel coronavirus 2019-nCoV antibody by double-antigen sandwich method.

[0049] 1. Preparation of novel coronavirus 2019-nCoV multi-dominant epitope fusion recombinant protein:

[0050] The dominant antigenic epitopes of the N protein, S protein, and M protein of the new coronavirus 2019-nCoV were screened by computational prediction, and linked together by a flexible polypeptide (GGGS) to form a 2019-nCoV virus multi-dominant epitope fusion protein, amino acid sequence As shown in the sequence SEQ ID NO.1. Entrusted Beijing Deaoping Biotechnology Co., Ltd. to produce and prepare, the purity is greater than 95%.

[0051] DQVILLNKHIDAYKTFPPTEPKKDKKKKADETQALPQRQKKQQTVTLLPAADLDDFSKQ GGGS DISTEIYQAGSTPCNGVEGFNCYFPLQSYGFQPTNGVGYQPYRVVVLSFELLHAPATVCGPKKSTNLVKNKCVNFNFNGLTGTGVLTESNKKFLPFQQFGRDIADTTDAVRDPQTLEILDITPCSFGGVSVITPGNTSNQVAVLYQDVNCTEVPVAIHA GGGS TLAILTALRLCAYCCNIVNVSLVKPSFYVYSRVKNLNSSRVPDLLV

[0052] 2. Preparation ...

Embodiment 2

[0064] Application of double-antigen sandwich method for detection of novel coronavirus 2019-nCoV antibody test strips

[0065] 1. Sample pretreatment

[0066] Aspirate 20 μL whole blood sample and add to 200 μL sample diluent, mix thoroughly.

[0067] 2. Test with test strips

[0068] Use a micropipette to accurately draw 80 μL of the sample solution to be tested into the sample hole of the test strip, and act for 15 minutes at room temperature (20-25°C); insert the test paper card into the carrier of the fluorescence detector, and select the sample to be tested through the touch screen. item, press the "Start Detection" button, the fluorescence detector will automatically scan the test paper card; read or print the test results on the display screen of the instrument.

[0069] 3. Analysis of test results

[0070] Quantitative detection

[0071] After the test is completed, the instrument obtains the ratio of the time-resolved fluorescence intensity of the detection area ...

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Abstract

The invention relates to a recombinant protein for detecting 2019 novel coronavirus antibody by a double-antigen sandwich method, a test strip, a preparation method and application thereof, and belongs to the technical field of virus detection. The amino acid sequence of the recombinant protein of the 2019 novel coronavirus antibody detected by the double-antigen sandwich method of the present invention is shown in SEQ ID NO.1. The recombinant protein of the present invention is a fusion protein of multiple dominant epitopes of 2019-nCoV, which can be used to prepare reagents for detection of 2019-nCoV virus antibodies by antigen double-sandwich method, can be stored at room temperature, and can be fast, high-sensitivity, high-throughput, Low instrument cost, single-person detection at any time, easy operation, can greatly improve the convenience of clinical use.

Description

technical field [0001] The invention relates to the technical field of virus detection, in particular to a recombinant protein, a test strip, a preparation method and an application for detecting 2019 novel coronavirus (2019-nCoV) antibodies by a double-antigen sandwich method. Background technique [0002] Coronaviruses are a large family of viruses known to cause more serious illnesses such as colds, 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, and 2019-nCoV is the seventh type of coronavirus isolated from humans. According to relevant disease monitoring, many cases of viral pneumonia were found, all of which were diagnosed as viral pneumonia / lung infection. [0003] After the new coronavirus infects the human body, there is generally an incubation period of 3 to 14 days (in some cases, up to 24 days). During the incubation period, there ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K19/00G01N33/533G01N33/543G01N33/558G01N33/569
CPCC07K14/005C07K2319/00C12N2770/20022G01N33/533G01N33/54313G01N33/558G01N33/56983G01N2333/165
Inventor 何昆仑田亚平张思兵周建平周裕军
Owner GENERAL HOSPITAL OF PLA
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