CN111337669A - Novel coronavirus detection test paper and detection method for rapid detection - Google Patents
Novel coronavirus detection test paper and detection method for rapid detection Download PDFInfo
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Abstract
The invention discloses a novel test paper for rapidly detecting coronavirus and a detection method, wherein the novel test paper for rapidly detecting coronavirus comprises a sample pad, a colloidal gold pad, a reaction chromogenic region, a water absorption region and a support plate, wherein the sample pad and the water absorption pad are respectively arranged on the support plate and are positioned on two opposite sides of the support plate, and the sample pad is absorbent filter paper or a glass fiber membrane and is used for bearing an object to be detected; a nitrocellulose membrane is fixed on the supporting plate, and the absorbent pad is absorbent paper; through double-binding of a novel coronavirus antibody and a receptor ACE2 protein of a virus, a coronavirus protein antigen is specifically captured and bound by adopting double enzyme-linked immunosorbent assay; and then combining an ACE2 antibody and biotin in a test paper reaction color development area, connecting avidin with horseradish peroxidase, and rapidly judging through a biotin-avidin system through chemical color development or accurately detecting the virus content by using an enzyme-labeling instrument.
Description
Technical Field
The invention relates to the technical field of novel rapid detection reagents for coronavirus antigens, in particular to novel test paper and a detection method for rapidly detecting coronavirus, which can detect human whole blood, serum, plasma, saliva, throat swab, excrement, urine and other samples.
Background
Because of limited materials and detection means, qPCR method is needed for diagnosing the novel coronavirus, the operation difficulty is high, and the cost is high compared with other detection methods. Therefore, the number of the existing test paper cannot meet the unified detection of a large number of citizens, the suspected cases of the novel coronavirus are high at present, so that early and rapid diagnosis of the virus infection is urgent, and the invention of the novel test paper for detecting the coronavirus to improve the early diagnosis of the virus infection has important clinical value.
The method specifically captures and binds coronavirus protein antigens by double-binding novel coronavirus (2019-nCoV) IgM and IgG antibodies and a receptor ACE2 protein of the virus by adopting double enzyme-linked immunosorbent assay; and then the ACE2 antibody in the test paper reaction color development area is combined with biotin, the avidin is connected with horse radish peroxidase, the detection result can be rapidly judged through a biotin-avidin system and chemical color development, and the virus content can also be accurately detected by using an enzyme-labeling instrument. The antibody of the virus and the ACE2 protein of the receptor of the virus are combined in a double mode, the novel coronavirus is combined in a specific mode, positive and negative results are observed through a color development strip, and meanwhile, the protein can be extracted for quantitative analysis, and the method is the innovation point of the patent. Compared with the products of the existing companies, the technology can only detect novel coronavirus (2019-nCoV) antigens in samples such as human whole blood, serum and plasma, and the technology adopts double enzyme-linked immunosorbent assay to specifically capture and combine samples such as saliva, throat swabs and excrement, and can even capture viruses in free air in a closed environment, thereby being another advantage of the products.
Disclosure of Invention
The invention aims to provide a novel test paper for rapidly detecting coronavirus and a detection method, so as to solve the problems of higher operation difficulty, higher cost compared with other detection methods and the like in the background technology; the detection method detects the novel coronavirus through acquiring common ACE2 protein and novel coronavirus antibodies IgM and IgG and detecting the novel coronavirus through double enzyme-linked immunosorbent assay, the product uses the antibody and receptor protein which can react with the coronavirus specifically, the receptor protein is fixed on a nitrocellulose membrane, the other antibody is combined with colloidal gold, if the novel coronavirus CV exists in a sample to be detected, CV antigen firstly reacts with an anti-CV antibody colloidal gold conjugate, moves on the membrane through capillary phenomenon, and finally is combined with the receptor protein fixed on an NC membrane to form ACE2 protein-novel coronavirus antibody IgM and IgG conjugate, through combining with ACE2 antibody biotin and avidin horse radish peroxidase, further activates chromogenic peroxide and hydrogen donor to present color change, the combined product reacts with a substrate and a chromogenic solution of enzyme, the infection condition of the virus is judged through the strip, compared with the products of the existing companies, the technology can only detect the novel coronavirus 2019-nCoV antigen in human whole blood, serum, plasma and other samples, and the adopted 'double enzyme-linked immunosorbent' can specifically capture and combine saliva, throat swab, excrement and other samples, and even viruses in free air; the color can be displayed by using the simplest test paper, no instrument and equipment are needed, and the method is simple and quick and is suitable for field screening; reducing the risk of nosocomial infections in an individual; the waiting time of the patient is shortened, and the treatment is carried out in time; the workload of the epidemic situation fixed-point hospital is reduced.
In order to solve the technical problems, the invention provides the following technical scheme:
a novel test paper for rapidly detecting coronavirus comprises a sample pad, a colloidal gold pad, a reaction chromogenic region, a water absorption pad and a support plate, wherein the sample pad, the colloidal gold pad, the reaction chromogenic region and the water absorption pad are sequentially arranged on the support plate from left to right, and the sample pad and the water absorption pad are respectively arranged on two opposite sides of the support plate; the right side of the sample pad is provided with a gold rubber pad, and a connecting and overlapping area of 0.5cm is arranged on the right side of the sample pad; the right side of the colloidal gold pad is adjacent to a reaction color development area, the former covers 0.5cm of the adjacent side of the latter, the water absorption pad is positioned on the right side of the reaction color development area and covers 0.5cm of the reaction color development area, and the sample pad is used for bearing an object to be detected; the sample pad is suction filter paper or a glass fiber membrane; the water absorption pad is absorbent paper; the colloidal gold pad is attached with IgM antibody and IgG antibody of the novel coronavirus.
Further, the reaction color development area consists of an NC membrane, a quality control line C and a detection line T, wherein the quality control line C and the detection line T are respectively arranged on the surface of the NC membrane.
Furthermore, the quality control line C and the detection line T on the color development area are separated by 2-4 cm.
Further, the NC membrane of the reaction color development area consists of enzyme color development liquid, a virus receptor ACE2 protein connected with biotin, a substrate and horseradish peroxidase HRP connected with avidin, the enzyme color development liquid is TMB color development liquid or DAB horseradish peroxidase color development liquid, the substrate consists of peroxide capable of developing color and hydrogen donor, and the dosage ratio of the ACE2 protein to the novel coronavirus IgM and IgG antibodies is 1:2-1: 3; the molar ratio of the enzyme color development solution, the virus receptor ACE2 protein connected with biotin, the substrate and the horseradish peroxidase HRP connected with avidin is 3:1:2: 2; the substrate has a chromogenic peroxide and hydrogen donor molar ratio of 1:1, two different antibodies are respectively fixed on a detection line T and a quality control line C and respectively identify different antigenic determinants, wherein an antibody ACE2 protein on the detection line T identifies an antigen to be detected, an antibody anti-IgM on the quality control line C and IgM and IgG antibodies coupled on an anti-IgG identification colloidal gold pad, colloidal gold particles are distributed on the surface of the colloidal gold pad, the colloidal gold is a chromogenic marker and is coupled with the IgM antibody and the IgG antibody, and red bands displayed by the detection line T and the quality control line C are the result of mass aggregation of the colloidal gold.
Furthermore, the supporting plate is made of a non-water-absorbing material, such as PVC or other hard materials, and a nitrocellulose membrane is fixed on the supporting plate.
Furthermore, if the novel coronavirus exists in the sample to be detected, the novel coronavirus antigen firstly reacts with the gold-labeled antibody IgM antibody and IgG antibody on the gold pad to form an antigen-antibody complex, the antigen-antibody complex moves on the NC membrane through capillary phenomenon and continuously flows towards the water absorption pad, and finally is combined with ACE2 protein fixed on the NC membrane, an antibody aiming at the antigen, namely ACE2 protein, is arranged on the detection line T, the epitope recognized by the antibody is different from the gold-labeled antibody, so that the same antigen can be recognized by the two antibodies at the same time, the gold-labeled antibody is intercepted at the detection line to form ACE2 protein-new coronavirus antibody IgM, an IgG conjugate, and then is combined with ACE2 antibody biotin and avidin horse radish peroxidase to further activate peroxide and hydrogen donor to show color change, therefore, the color is developed on the detection line T, the amount of the gold-labeled antibody is surplus certainly, so that the gold-labeled antibody cannot be trapped only at the detection line T and can continuously flow towards the water absorption pad, a quality control line C exists at the moment, antibodies, namely anti-IgM and anti-IgG, which are specially used for the gold-labeled antibody exist on the quality control line C, the line can be developed certainly because the quality control line C has very strong capacity of recognizing the gold-labeled antibody, and if the line is not developed, the detection result is invalid, so that the infection condition of the virus can be judged through the strip; if the test sample does not contain the novel coronavirus, no conjugate is formed.
Furthermore, the novel coronavirus detection test paper body for rapid detection is square.
A detection test paper for rapidly detecting novel coronavirus and a detection method of the detection method comprise the following steps:
the first step, preparation: before use, the detection reagent, the sample and the RIPA lysate are restored to room temperature;
the second step is that: vertically dropping 11-13 drops of lysis solution to a first scale mark of the sample extraction tube by using a diluent bottle;
the third step: collecting throat swab or nasal cavity swab of patient with cotton swab, inserting the sample into the extraction tube, and standing for 1 min;
the fourth step: rotating the throat swab or the nasal swab in the extraction tube for 6 times to completely mix the sample with the lysate;
the fifth step: pinching and extracting the tube wall by fingers, squeezing the solution on the cotton swab as dry as possible and taking out the cotton swab;
and a sixth step: inserting a dripper of the extraction pipe into the extraction pipe;
and a sixth step: the novel coronavirus rapid detection test paper is horizontally placed on a table;
the seventh step: dripping 3 drops of liquid in the extraction tube into a sample pad of the rapid detection novel coronavirus detection test paper;
eighth step: the object to be detected enters the reaction color development area through the colloidal gold pad by the water absorption force of the water absorption pad, the result is observed after 10min, and the result is invalid after 10 min;
the ninth step: if the substance to be detected contains the novel coronavirus, the obtained antigen-antibody conjugate is coupled with enzyme, the conjugate product reacts with a corresponding substrate, a reaction color development area has a detection line T and a quality control line C, when the detection line T and the quality control line C both have strips for displaying, namely two strips are POSITIVE POSITIVE, and the detection line T has no strip and the quality control line C has a strip for displaying as NEGATIVE NEGATIVE;
the tenth step: the test is repeated for three times, a substrate on the novel coronavirus detection test paper is extracted for rapid detection, chemiluminescence is realized through a biotin-avidin system, the result is directly judged through color change, and further, the novel coronavirus with accurate quantification can be accurately detected through an enzyme-labeling instrument.
Further, the room temperature in the first step is 15-30 ℃.
Further, the first scale mark of the sample extraction tube in the second step is 0.5 mL.
Advantageous effects
Compared with the prior art, the invention has the following beneficial effects:
1. the result can be presented by using the simplest test paper color;
2. no instrument and equipment are needed, and the method is simple and quick;
3. is suitable for on-site screening;
4. reducing the risk of nosocomial infections in an individual;
5. the waiting time of the patient is shortened, and the treatment is carried out in time;
6. the workload of the epidemic situation fixed-point hospital is reduced;
7. compared with products of the existing companies, the technology can only detect the novel coronavirus 2019-nCoV antigen in samples such as human whole blood, serum, plasma and the like, and the adopted 'double enzyme-linked immunosorbent assay' can specifically capture and combine samples such as saliva, throat swabs, excrement and the like and even free viruses in the air;
8. the immunochromatographic test paper provided by the invention is added with a sample, a substance to be detected enters a reaction chromogenic region through the water absorption force of a water absorption pad, if the substance to be detected contains the novel coronavirus, an obtained conjugate is coupled with enzyme, the conjugate product reacts with a corresponding substrate, and the infection condition of the virus is judged through chromogenic reaction; if the object to be detected does not contain the novel coronavirus, no conjugate is formed;
9. the test system is made into a test paper form, whether the coronavirus is infected or not is judged primarily through primary color change, and the primarily screened test paper material can be used on a mask and helps to judge the service life of the mask and whether the mask is infected with the virus or not through color change; further, a substrate on the test paper can be extracted, chemiluminescence is realized through a biotin-avidin system, and accurate detection is realized through a microplate reader;
10. when the detection test paper is used for detection, professional detection personnel are not needed, the operation is very simple, the virus infection condition is intuitively sensed, and the result can be quickly obtained; the novel coronavirus detection test strip provided by the invention facilitates the detection of the novel coronavirus;
11. the invention utilizes the novel coronavirus which can be specifically and doubly combined with the novel coronavirus antibodies IgG, IgM and ACE2 in vitro so as to judge the infection condition of the virus through simple ELISA and immune colloidal gold technology, and also relates to the color change from biotin and avidin to reaction so as to react to the color change on test paper; and then, the chemiluminescence with the biotin and avidin amplification levels is used for more accurate detection by an enzyme-labeling instrument, the detection method is simple, the detection speed is high, the detection effect is obvious, and the kit can be used as an important means for primary screening.
Drawings
FIG. 1 is a schematic theoretical guidance diagram of a test strip for rapid detection of a novel coronavirus according to the present application;
FIG. 2 is a schematic structural diagram of the test paper for rapid detection of coronavirus according to the present application;
FIG. 3 is a schematic diagram showing the second to fifth steps of the rapid test method for coronavirus test strip of the present application;
FIG. 4 is a schematic diagram of the seventh operation of the detection method for rapidly detecting the novel coronavirus test strip according to the present application;
FIG. 5 is a schematic diagram of the test result of the test paper for rapidly detecting coronavirus of the present application:
FIG. 6 is a schematic diagram of the test strip for rapid detection of coronavirus according to the present invention.
Description of reference numerals: 1. A sample pad; 2. a gold rubber pad; 3. a reaction color development area; 4. A water absorbent pad; 5. and a support plate.
Detailed Description
The technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the drawings in the embodiments of the present invention, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
Example 1:
a novel test paper for rapidly detecting coronavirus is composed of a sample pad 1, a colloidal gold pad 2, a reaction chromogenic region 3, a water absorption pad 4 and a support plate 5, wherein the sample pad 1, the colloidal gold pad 2, the reaction chromogenic region 3 and the water absorption pad 4 are sequentially arranged on the support plate 5 from left to right, and the sample pad 1 and the water absorption pad 4 are respectively arranged on two opposite sides of the support plate 5; the right side of the sample pad 1 is provided with a gold rubber pad 2 with a joining and overlapping area of 0.5 cm; the right side of the colloidal gold pad 2 is adjacent to a reaction chromogenic region 3, the former covers 0.5cm of the adjacent side of the latter, the water absorption pad 4 is positioned on the right side of the reaction chromogenic region 3 and covers 0.5cm of the reaction chromogenic region, and the sample pad 1 is used for bearing an object to be detected; the sample pad 1 is suction filter paper or a glass fiber membrane; the absorbent pad 4 is absorbent paper; the colloidal gold pad 2 is attached with IgM and IgG antibodies of a novel coronavirus.
The reaction chromogenic region 3 consists of an NC membrane, a quality control line C and a detection line T, the quality control line C and the detection line T are respectively arranged on the surface of the NC membrane, the quality control line C and the detection line T are arranged on the chromogenic region 3 at intervals of 2-4 cm, the support plate 5 is made of a non-water-absorbing material such as PVC or other hard materials, a nitrocellulose membrane is fixed on the support plate 5, and the novel rapid coronavirus detection test paper body is square.
The NC membrane of the reaction chromogenic region 3 consists of enzyme color solution, a virus receptor ACE2 protein connected with biotin, a substrate and horseradish peroxidase HRP connected with avidin, the enzyme color solution is TMB color solution or DAB horseradish peroxidase color solution, the substrate consists of peroxide capable of developing colors and hydrogen donor, and the dosage ratio of the ACE2 protein to the novel coronavirus IgM and IgG antibodies is 1:2-1: 3; the molar ratio of the enzyme color development solution, the virus receptor ACE2 protein connected with biotin, the substrate and the horseradish peroxidase HRP connected with avidin is 3:1:2: 2; the substrate has a chromogenic peroxide and hydrogen donor molar ratio of 1:1, two different antibodies are respectively fixed on a detection line T and a quality control line C and respectively identify different antigenic determinants, wherein an ACE2 protein of the antibody of the detection line T identifies an antigen to be detected, an anti-IgM of the antibody on the quality control line C and IgM and IgG antibodies coupled on an anti-IgG identification colloidal gold pad 2, colloidal gold particles are distributed on the surface of the colloidal gold pad 2, the colloidal gold is a chromogenic marker and is coupled with the IgM antibody and the IgG, and red bands displayed by the detection line T and the quality control line C are the result of mass aggregation of the colloidal gold.
If the novel coronavirus exists in the sample to be detected, the novel coronavirus antigen firstly reacts with the gold-labeled antibody IgM antibody and IgG antibody on the colloidal gold pad 2 to form an antigen-antibody complex, moves on the NC membrane through capillary phenomenon, continuously flows towards the water absorption pad 4, and finally is combined with ACE2 protein fixed on the NC membrane, an antibody aiming at the antigen, namely ACE2 protein, is arranged on the detection line T, the epitope recognized by the antibody is different from the gold-labeled antibody, so that the same antigen can be recognized by the two antibodies at the same time, the gold-labeled antibody is intercepted partially at the detection line at the moment to form ACE2 protein-new coronavirus antibody IgM and IgG conjugate, and then is combined with ACE2 antibody biotin and avidin horse radish peroxidase to further activate chromogenic peroxide and hydrogen donor to show color change, so that the detection line T shows color, the quantity of the gold-labeled antibody is surplus certainly, so that the gold-labeled antibody cannot be trapped only at the detection line T and can continuously flow towards the water absorption pad, at this time, a quality control line C exists, antibodies, namely anti-IgM and anti-IgG, which are specially used for the gold-labeled antibody exist on the quality control line C, the gold-labeled antibody is very strong in capacity of recognizing the quality control line C, the line is colored certainly, if the line is not colored, the detection result is invalid, and therefore the infection condition of the virus can be judged through the strip; if the test sample does not contain the novel coronavirus, no conjugate is formed.
A detection test paper for rapidly detecting novel coronavirus and a detection method of the detection method comprise the following steps:
the first step, preparation: before use, the detection reagent, the sample and the RIPA lysate are recovered to 15-30 ℃;
the second step is that: vertically dropping 11-13 drops of lysis solution to a first scale mark of a sample extraction tube by using a diluent bottle, wherein the first scale mark of the sample extraction tube is 0.5 mL;
the third step: collecting throat swab or nasal cavity swab of patient with cotton swab, inserting the sample into the extraction tube, and standing for 1 min;
the fourth step: rotating the throat swab or the nasal swab in the extraction tube for 6 times to completely mix the sample with the lysate;
the fifth step: pinching and extracting the tube wall by fingers, squeezing the solution on the cotton swab as dry as possible and taking out the cotton swab;
and a sixth step: inserting a dripper of the extraction pipe into the extraction pipe;
and a sixth step: the novel coronavirus rapid detection test paper is horizontally placed on a table;
the seventh step: dripping 3 drops of liquid in the extraction tube into a sample pad 1 of the rapid detection novel coronavirus detection test paper;
eighth step: the substance to be detected enters the reaction color development area through the gold rubber pad 2 by the water absorption force of the water absorption pad 4, the result is observed after 10min, and the result is invalid after 10 min;
the ninth step: if the substance to be detected contains the novel coronavirus, the obtained antigen-antibody conjugate is coupled with enzyme, the conjugate product reacts with a corresponding substrate, a reaction color development area has a detection line T and a quality control line C, when the detection line T and the quality control line C both have strips for displaying, namely two strips are POSITIVE POSITIVE, and the detection line T has no strip and the quality control line C has a strip for displaying as NEGATIVE NEGATIVE;
the tenth step: the test is repeated for three times, a substrate on the novel coronavirus detection test paper is extracted for rapid detection, chemiluminescence is realized through a biotin-avidin system, the result is directly judged through color change, and further, the novel coronavirus with accurate quantification can be accurately detected through an enzyme-labeling instrument.
As shown in fig. 5, 1 test line positive, quality control line weak positive, 2 test line positive, quality control line positive, 1, 2 are positive results; 3, the test line is negative, the quality control line is positive, 4, the test line is weak positive, only the ACE2 protein is singly combined, the quality control line is positive, and 3 and 4 are negative results; 5 test line positive and quality control line negative, 6 test line negative and quality control line negative, 5 and 6 are all ineffective in detection.
As shown in fig. 6, the research team and the ministerial medical technology ltd of Jiangsu, according to the invention, created the prototype of the test paper, and developed the product verification by the scientific research cooperation of the subsidiary hospital of Nantong university. The patient is admitted to the hospital for treatment of the severe novel coronavirus pneumonia in 3 days 2 months in 2020, the number 001 is 5 days 2 months, the test result of the sputum sample of the patient is collected, and the result detection is positive; the number 002 is 2 months and 15 days, and after the same patient is hospitalized for 10 days, the test result of the sputum sample is collected, and the result detection is negative; and the results of the two detections are completely consistent with the results of the novel coronavirus nucleic acid.
The product uses an antibody and a receptor protein which can specifically react with coronavirus, the receptor protein is fixed on a nitrocellulose membrane, the other antibody is combined with colloidal gold, if a novel coronavirus CV exists in a sample to be detected, CV antigen firstly reacts with the anti-CV antibody colloidal gold conjugate, moves on the membrane through capillary phenomenon, and finally is combined with the receptor protein fixed on an NC membrane to form ACE2 protein-novel coronavirus antibody IgM and IgG conjugate, the ACE2 antibody biotin and avidin horse radish peroxidase are combined, so that chromogenic peroxide and hydrogen donor are activated to show color change, the combined product reacts with a substrate and a chromogenic solution of enzyme, if the novel coronavirus exists in the sample to be detected, the novel coronavirus antigen firstly reacts with a gold-labeled antibody IgM antibody and an IgG antibody on a colloidal gold pad 2, forming an antigen-antibody complex, moving on an NC membrane through capillary phenomenon, continuously flowing towards the direction of a water absorption pad 4, finally combining with ACE2 protein fixed on the NC membrane, wherein an antibody aiming at the antigen, namely ACE2 protein, exists on a detection line T, the epitope recognized by the antibody is different from a gold-labeled antibody, so that the same antigen can be recognized by the two antibodies simultaneously, at the moment, the gold-labeled antibody is partially intercepted at the detection line to form ACE2 protein-new coronavirus antibody IgM, IgG conjugate, then combining with ACE2 antibody biotin and avidin horse radish peroxidase, further activating superoxide and hydrogen donor to present color change, so that color is developed on the detection line T, the amount of the gold-labeled antibody is surplus, so that the gold-labeled antibody can not be intercepted only at the detection line T, but also continuously flowing towards the direction of the water absorption pad, at this time, a quality control line C is arranged, antibodies anti-IgM and anti-IgG which are specially used for the gold-labeled antibody are arranged on the quality control line C, the line can be colored because the capacity of the quality control line C for identifying the gold-labeled antibody is very strong, if the line is not colored, the detection result is invalid, and therefore the infection condition of the virus can be judged through the band; if the test substance does not contain the novel coronavirus, no conjugate is formed, and the negative result should exclude interference of whether the test method is wrong.
The judgment of the result based on the above colors further includes the following four cases:
1. the negative detection test result is a negative result, otherwise, the environment is unqualified;
2. the positive control test result is a positive result, otherwise, the test paper is invalid;
3. the three tests are negative results, and the test result is judged to be negative;
4. at least one test is positive, and the test result is judged to be positive;
the positive results should exclude interference from the following factors: other influenza virus infections.
Although the novel coronavirus detection test paper is convenient for patient self-detection and can obtain a result by a simple detection method, the novel coronavirus detection test paper can only be used as a novel coronavirus infection screening means and still cannot completely eliminate false positive and false negative caused by various factors. Therefore, the test paper results and clinical symptoms are needed, and the physician experience is combined to finally judge the results, so that the patients can see the doctor timely when the test paper is positive.
Finally, it should be noted that: although the present invention has been described in detail with reference to the foregoing embodiments, it will be apparent to those skilled in the art that changes may be made in the embodiments and/or equivalents thereof without departing from the spirit and scope of the invention. Any modification, equivalent replacement, or improvement made within the spirit and principle of the present invention should be included in the protection scope of the present invention.
Claims (10)
1. A novel coronavirus detection test paper for rapid detection is characterized in that: the novel rapid coronavirus detection test paper consists of a sample pad (1), a colloidal gold pad (2), a reaction chromogenic region (3), a water absorption pad (4) and a support plate (5), wherein the sample pad (1), the colloidal gold pad (2), the reaction chromogenic region (3) and the water absorption pad (4) are sequentially arranged on the support plate (5) from left to right, and the sample pad (1) and the water absorption pad (4) are respectively arranged on two opposite sides of the support plate (5); the right side of the sample pad (1) is provided with a gold rubber pad (2) with a joining and overlapping area of 0.5 cm; the right side of the colloidal gold pad (2) is closely adjacent to a reaction chromogenic region (3), the former covers 0.5cm of the adjacent side of the latter, the water absorption pad (4) is positioned on the right side of the reaction chromogenic region (3) and covers 0.5cm of the reaction chromogenic region, and the sample pad (1) is used for bearing an object to be detected; the sample pad (1) is a suction filter paper or a glass fiber membrane; the water absorption pad (4) is absorbent paper; the colloidal gold pad (2) is attached with IgM antibody and IgG antibody of novel coronavirus.
2. The test strip of claim 1, wherein the test strip for rapid detection of coronavirus comprises: the reaction color development area (3) consists of an NC membrane, a quality control line C and a detection line T, wherein the quality control line C and the detection line T are respectively arranged on the surface of the NC membrane.
3. The test strip according to claim 2, wherein the test strip for rapid detection of coronavirus comprises: the quality control line C and the detection line T on the color development area (3) are spaced by 2-4 cm.
4. The test strip according to claim 2, wherein the test strip for rapid detection of coronavirus comprises: the NC membrane of the reaction chromogenic region (3) consists of enzyme color development liquid, a virus receptor ACE2 protein connected with biotin, a substrate and horseradish peroxidase HRP connected with avidin, the enzyme color development liquid is TMB color development liquid or DAB horseradish peroxidase color development liquid, the substrate consists of peroxide capable of developing colors and hydrogen donor, and the dosage ratio of the ACE2 protein to the novel coronavirus IgM and IgG antibodies is 1:2-1: 3; the molar ratio of the enzyme color development solution, the virus receptor ACE2 protein connected with biotin, the substrate and the horseradish peroxidase HRP connected with avidin is 3:1:2: 2; the substrate has a chromogenic peroxide and hydrogen donor molar ratio of 1:1, two different antibodies are respectively fixed on a detection line T and a quality control line C and respectively identify different antigenic determinants, wherein an ACE2 protein of the antibody of the detection line T identifies an antigen to be detected, an anti-IgM of the antibody on the quality control line C and IgM and IgG antibodies coupled on an anti-IgG identification colloidal gold pad (2), colloidal gold particles are distributed on the surface of the colloidal gold pad (2), the colloidal gold is a chromogenic marker and is simultaneously coupled with the IgM antibody and the IgG, and red bands displayed by the detection line T and the quality control line C are the result of mass aggregation of the colloidal gold.
5. The test strip of claim 1, wherein the test strip for rapid detection of coronavirus comprises: the supporting plate (5) is made of a non-water-absorbing material, such as PVC or other hard materials, and a nitrocellulose membrane is fixed on the supporting plate (5).
6. The test strip of claim 4, wherein the test strip for rapid detection of coronavirus comprises: if the novel coronavirus exists in the sample to be detected, the novel coronavirus antigen firstly reacts with a gold-labeled antibody IgM antibody and an IgG antibody on a colloidal gold pad (2) to form an antigen-antibody complex, moves on an NC membrane through capillary phenomenon, continuously flows towards a water absorption pad (4), and finally is combined with ACE2 protein fixed on the NC membrane, an antibody aiming at the antigen, namely ACE2 protein, is arranged on a detection line T, an epitope recognized by the antibody is different from the gold-labeled antibody, so that the same antigen can be recognized by the two antibodies simultaneously, at the moment, the gold-labeled antibody is intercepted partially at the detection line to form ACE2 protein-new coronavirus antibody IgM, an IgG conjugate, and then is combined with ACE2 antibody biotin and avidin horse radish peroxidase to further activate peroxide and hydrogen donor to show color change, therefore, the color is developed on the detection line T, the amount of the gold-labeled antibody is surplus certainly, so that the gold-labeled antibody cannot be trapped only at the detection line T and can continuously flow towards the water absorption pad, a quality control line C exists at the moment, antibodies, namely anti-IgM and anti-IgG, which are specially used for the gold-labeled antibody exist on the quality control line C, the line can be developed certainly because the quality control line C has very strong capacity of recognizing the gold-labeled antibody, and if the line is not developed, the detection result is invalid, so that the infection condition of the virus can be judged through the strip; if the test sample does not contain the novel coronavirus, no conjugate is formed.
7. The test strip of claim 1, wherein the test strip for rapid detection of coronavirus comprises: the novel coronavirus rapid detection test paper body is square.
8. A detection method for rapidly detecting novel coronavirus detection test paper is characterized by comprising the following steps:
the first step, preparation: before use, the detection reagent, the sample and the RIPA lysate are restored to room temperature;
the second step is that: vertically dropping 11-13 drops of lysis solution to a first scale mark of the sample extraction tube by using a diluent bottle;
the third step: collecting throat swab or nasal cavity swab of patient with cotton swab, inserting the sample into the extraction tube, and standing for 1 min;
the fourth step: rotating the throat swab or the nasal swab in the extraction tube for 6 times to completely mix the sample with the lysate;
the fifth step: pinching and extracting the tube wall by fingers, squeezing the solution on the cotton swab as dry as possible and taking out the cotton swab;
and a sixth step: inserting a dripper of the extraction pipe into the extraction pipe;
and a sixth step: the novel coronavirus rapid detection test paper is horizontally placed on a table;
the seventh step: dripping 3 drops of liquid in the extraction tube into a sample pad (1) of the rapid detection novel coronavirus detection test paper;
eighth step: the object to be detected enters the reaction color development area (3) through the colloidal gold pad (2) by the water absorption force of the water absorption pad (4), the result is observed after 10min, and the result is invalid after 10 min;
the ninth step: if the substance to be detected contains the novel coronavirus, the obtained antigen-antibody conjugate is coupled with enzyme, the conjugate product reacts with a corresponding substrate, a reaction color development area has a detection line T and a quality control line C, when the detection line T and the quality control line C both have strips for displaying, namely two strips are POSITIVE POSITIVE, and the detection line T has no strip and the quality control line C has a strip for displaying as NEGATIVE NEGATIVE;
the tenth step: the test is repeated for three times, a substrate on the novel coronavirus detection test paper is extracted for rapid detection, chemiluminescence is realized through a biotin-avidin system, the result is directly judged through color change, and further, the novel coronavirus with accurate quantification can be accurately detected through an enzyme-labeling instrument.
9. The method according to claim 8, wherein the reagent strip for rapid detection of coronavirus is characterized in that: the room temperature in the first step is 15-30 ℃.
10. The method according to claim 8, wherein the reagent strip for rapid detection of coronavirus is characterized in that: the first scale line of the sample extraction tube in the second step was 0.5 mL.
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