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Immunochromatographic test paper for detecting novel coronavirus

A technology of immunochromatographic test paper and coronavirus, which is applied in the field of immune detection, can solve the problems of lack of sensitivity and achieve the effect of improving sensitivity, speed and accurate identification

Active Publication Date: 2020-11-03
广州德成生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] Although there are immunochromatographic devices for the detection of new coronaviruses, they all use conventional markers, and the sensitivity is still lacking.

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0018] Example 1: Preparation of recombinant antigen

[0019] Referring to the CN202010105749.8 patent application, S protein, N protein, and M protein were synthesized. The amino acid sequence of the N protein is shown in SEQ ID NO.1, and the amino acid sequence of the S protein is shown in SEQ ID NO.12. The amino acid sequence of the M protein is shown in SEQ ID NO.13, and the amino acid sequence of the E protein is shown in SEQ ID NO.14.

[0020] The sequence of SEQID NO.1 is as follows:

[0021] MSDNGPQNQRNAPRITFGGPSDSTGSNQNGERSGARSKQRRPQGLPNNTASWFTALTQHGKEDLKFPRGQGVPINTNSSPDDQIGYYRRATRRIRGGDGKMKDLSPRWYFYYLGTGPEAGLPYGANKDGIIWVATEGALNTPKDHIGTRNPANNAAIVLQLPQGTTLPKGFYAEGSRGGSQASSRSSSRSRNSSRNSTPGSSRGTSPARMAGNGGDAALALLLLDRLNQLESKMSGKGQQQQGQTVTKKSAAEASKKPRQKRTATKAYNVTQAFGRRGPEQTQGNFGDQELIRQGTDYKHWPQIAQFAPSASAFFGMSRIGMEVTPSGTWLTYTAAIKLDDKDPNFKDQVILLNKHIDAYKTFPPTEPKKDKKKKADETQALPQRQKKQQTVTLLPAADLDDFSKQLQQSMSSADSTQA。

[0022] The sequence of SEQID NO.12 is as follows:

[0023]MGS...

Embodiment 2

[0028] Example 2: Preparation of monoclonal antibodies

[0029] Referring to the CN202010105749.8 patent application, selection of immunized animals, cell fusion, selection of specific hybridoma cells, mass preparation of monoclonal antibodies (preparation of ascites), and use of ammonium sulfate precipitation, Protein A affinity chromatography purification to obtain antibodies against S Protein, N protein, M protein, E protein monoclonal antibody.

Embodiment 3

[0030] Example 3: Preparation of recombinant ACE2 protein

[0031] RNA extraction of ACE2: Take about 100 mg of lung tissue, add 1 ml of Trizol reagent, and homogenize with a homogenizer. Transferred to a 1.5 ml EP tube, placed at room temperature for 5 min, centrifuged at 12 000 g at 4 °C for 10 min. Add 0.2 ml of chloroform, mix well, place at room temperature for 3 min, centrifuge at 4°C, 15 000 g for 15 min. Take the supernatant, add 0.5 ml of isopropanol, put it at room temperature for 10 min, then centrifuge at 12000 g at 4°C for 10 min. Discard the supernatant, add 1 ml of 75% ethanol, mix by shaking, centrifuge at 7800g at 4°C for 5 min. Discard the supernatant, and after air-drying, add 30 μL of 1‰DEPC-treated water to dissolve.

[0032] Reverse transcription nested polymerase chain reaction primers were designed according to GenBank ACE-2 sequence information (NM021804).

[0033] The outer primer pair is: ACE2-OutF: 5'-CCCAACCCAAGTTCAAAG-3', ACE2-OutR: 5'-GCATGCC...

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Abstract

The invention relates to immunochromatographic test paper for detecting novel coronavirus SARS-CoV-2. The test paper comprises a substrate, and a sample pad, a combination pad, a nitrocellulose membrane and a water absorption pad which are arranged on the substrate and are sequentially and fixedly adhered to the substrate along the flowing direction of a to-be-detected liquid sample, the substrateis a PVC plate. The sample pad is coated with a biotin labeled anti-SARS-CoV-2N protein monoclonal antibody and biotin labeled angiotensin converting enzyme 2 (ACE2), the combination pad is coated with avidin crosslinked colloidal gold, the nitrocellulose membrane is provided with a detection line and a quality control line, the detection line is coated with an anti-SARS-CoV-2M protein monoclonalantibody, and the quality control line is coated with goat anti-mouse IgG. The immunochromatographic test paper provided by the invention achieves high-sensitivity, high-specificity, high-speed and convenient-to-operate detection, and can be applied to large-scale rapid screening of people in primary hospitals and communities.

Description

technical field [0001] The invention relates to the technical field of immunoassay, in particular to an immunochromatographic test paper for detecting novel coronavirus. Background technique [0002] The novel coronavirus pneumonia that broke out at the end of 2019 has seriously affected global public health security. On January 12, 2020, the World Health Organization (WHO) officially named the new coronavirus "2019-nCov". On February 11, 2020, WHO named the pneumonia caused by the new coronavirus as "COVID-19". Meanwhile, the Coronavirus Study Group of the International Committee on Taxonomy of Viruses (ICVT-CSG) named the virus "Severe Acute Respiratory Syndrome Coronavirus 2" (SARS-CoV-2). [0003] So far, although more and more candidate drugs have entered the field of vision of researchers, and a large number of clinical trials are being carried out around the world, the real new crown specific drugs have not yet appeared. At the same time, many new developments have...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/577G01N33/569G01N33/558G01N33/543G01N33/532
CPCG01N33/577G01N33/56983G01N33/558G01N33/54346G01N33/532Y02A50/30
Inventor 刘伟芳
Owner 广州德成生物科技有限公司
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