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Immunochromatographic kit for detecting novel coronavirus SARS-CoV-2

A technology of coronavirus and immunochromatography, applied in the direction of immunoglobulin, analytical materials, measuring devices, etc., can solve the problems of only 60.5%, detection lag, etc.

Inactive Publication Date: 2020-07-03
CHONGQING TANSHENG TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, whether it is IgM or IgG, there is a window period from virus infection to antibody production, so its detection often lags behind nucleic acid detection. It is generally believed that the sensitivity of specific antibody detection is lower than nucleic acid detection, but Zheng et al. The study of 129 cases of SARS coronavirus infection found that the positive rate of nucleic acid fluorescent PCR detection was only 60.5% (78 / 129), while the positive rate of specific antibody detection was 100% (129 / 129). For coronavirus infection, research on specific antibody detection should be actively carried out to make up for the deficiency of nucleic acid detection

Method used

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  • Immunochromatographic kit for detecting novel coronavirus SARS-CoV-2
  • Immunochromatographic kit for detecting novel coronavirus SARS-CoV-2
  • Immunochromatographic kit for detecting novel coronavirus SARS-CoV-2

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0043] The preparation of embodiment 1 neopterin antigen and novel coronavirus N antigen

[0044] (1) Preparation of neopterin antigen

[0045] Cross-linking buffer preparation: 0.1mol / L PBS, 0.15mol / L NaCl, pH7.2;

[0046] Weigh 20 mg of neopterin, add 5 ml of dimethyl sulfoxide, adjust the pH to 7.5 with 1 mol / L sodium hydroxide solution, and stir at 37°C until completely dissolved. Weigh 20mg of SMCC, dissolve it in 5ml dimethyl sulfoxide, add it to the completely dissolved neopterin solution, stir at 37°C for 30min, then add 8ml BSA solution dissolved in cross-linking buffer with a concentration of 5mg / ml Stir at 37°C for 30 minutes, then dialyze 4 times with 1L of 10mM PBS with pH 7.4 and concentrate to a volume of about 8ml. The obtained concentrate is the neopterin antigen (Neo-BSA) of cross-linked BSA protein. Add 0.1% preservative and freeze for later use.

[0047] In the same way, the neopterin antigen (Neo-KLH) of the cross-linked KLH protein can be obtained;

...

Embodiment 2

[0063] Example 2 Preparation and identification of neopterin monoclonal antibody

[0064] (1) Preparation of neopterin monoclonal antibody

[0065] The Neo-BSA prepared in Example 1 was emulsified with an equivalent amount of Freund's complete adjuvant, and the emulsification of the antigen was carried out by double syringes. After the emulsification was completed, 4 Balb / c mice aged about 8 weeks were basally immunized (100 μg / ml, 200 μl per mouse) by multi-point subcutaneous injection in limbs and intraperitoneal injection. After 2 weeks, Neo-BSA was emulsified with the same amount of Freund's incomplete adjuvant, and the mice were boosted by multi-point subcutaneous injection of limbs plus intraperitoneal injection (100 μg / ml, 200 μl per mouse); 2 weeks Afterwards, an additional immunization was carried out in the same way. After 7 days, blood was collected from the tail vein of the mice to detect the antiserum titer by indirect ELISA. The ELISA antiserum titer reached 1:5...

Embodiment 3

[0084] Example 3 Preparation of Colloidal Gold Immunochromatography Detection Kit for Detecting New Coronavirus Infection

[0085] 1) buffer solution

[0086] Sample pad treatment solution (50mM Tris, 3.6mM EDTA-Na2, 3% trehalose, 0.1% BSA, 0.4% Tween-20, pH8.6);

[0087] Conjugate pad treatment solution (50mM Tris-HCl, 3% trehalose, 0.5% BSA, pH9.0);

[0088] Colloidal gold diluent (50mM Tris-HCl, 0.5% casein, 3% trehalose, pH8.0);

[0089] Coating buffer (10mM PBS, 3% trehalose, 0.5% methanol, pH7.4);

[0090] Sample diluent (10 mM PBS, 0.1% Triton, pH 7.4).

[0091] 2) Colloidal gold preparation

[0092] Measure 200ml of ultrapure water and add it to a 500ml Erlenmeyer flask, place the Erlenmeyer flask on the heating plate of a magnetic heating stirrer, put a magnetic stirrer, turn on the stirring knob to an appropriate speed, stir slowly and heat until the ultrapure water boils. When the ultrapure water boils, use a pipette to draw 2mL of 1% tetrachloroauric acid solu...

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Abstract

The invention discloses an immunochromatographic kit used for detecting novel coronavirus SARS-CoV-2 infection. The kit comprises a test strip. The test strip comprises a sample pad, a combination pad, a chromatography membrane and an absorption pad which are correspondingly pasted and laminated, and the combination pad is coated with a colloidal gold labeled neopterin antibody conjugate, a novelcoronavirus N protein conjugate, a novel coronavirus S1 protein conjugate and a DNP-BSA conjugate. A quality control line C and a detection line are arranged on the NC membrane. The C line is coated with a rabbit anti-DNP antibody, the detection line comprises a T1 line, a T2 line and a T3 line. The T1 line and the T2 line are respectively coated with one of an anti-human IgG antibody and an anti-human IgM antibody. The T3 line is coated with a Neo-BSA antigen, and the C line, the T1 line and the T2 line are three parallel bands perpendicular to the long side of the test strip. The kit provided by the invention is beneficial to early diagnosis and rapid confirmation of novel coronavirus infection, and has high sensitivity.

Description

technical field [0001] The application relates to the technical field of biological detection, in particular to an immunochromatographic kit for detecting the novel coronavirus SARS-CoV-2. Background technique [0002] The new coronavirus, namely "SARS-CoV-2", was named by the World Health Organization on January 12, 2020. It belongs to the same category of coronaviruses as the SARS coronavirus in 2002 and the Middle East respiratory syndrome coronavirus (MERS) in 2012. Family β genus. Comparing SARS-CoV-2 with the 2002 "SARS" SARS-CoV, it has about 70% sequence similarity and 40% sequence similarity with MERS-CoV. WHO Director-General Tedros Adhanom Ghebreyesus announced on February 11, 2020 that the novel coronavirus-infected pneumonia was named "COVID-19" (Corona Virus Disease 2019). At the same time, the International Virus Taxonomy Committee stated that the new coronavirus was named "SARS-CoV-2" (Severe Acute Respiratory Syndrome Coronavirus2); and that this virus is ...

Claims

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Application Information

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IPC IPC(8): G01N33/577G01N33/569G01N33/558C07K16/44C07K1/22
CPCC07K16/44G01N33/558G01N33/56983G01N33/577G01N2333/165
Inventor 胡川闽王江明范舒李斌徐淞
Owner CHONGQING TANSHENG TECH
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