CN112611863A - Colloidal tellurium test paper for detecting 2019 novel coronavirus and preparation method - Google Patents
Colloidal tellurium test paper for detecting 2019 novel coronavirus and preparation method Download PDFInfo
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Abstract
The invention discloses a colloidal tellurium test paper for detecting 2019 novel coronavirus and a preparation method thereof, belonging to the technical field of immunodiagnosis. The test paper comprises a PVC rubber plate, and a sample absorption pad, a colloidal tellurium binding pad, a chromatographic membrane and a water absorption material which are sequentially lapped on the PVC rubber plate; the colloidal tellurium binding pad is a chicken IgY compound containing colloidal tellurium markers and a novel coronavirus S1 antibody 1 compound containing colloidal tellurium markers; the chromatography membrane is provided with a detection line and a quality control line at intervals, wherein the detection line and the quality control line are sprayed with a novel coronavirus S1 antibody 2 and a goat anti-chicken IgY polyclonal antibody; the novel coronavirus S1 antibody 1 and the antibody 2 are respectively antibodies with the product numbers of 40592-R001 and 40591-T62 of Beijing Yiqiao Shenzhou science and technology Co. The invention adopts a double-antibody sandwich method, uses a specific antibody aiming at the key S protein of the novel coronavirus, has convenient operation and high sensitivity, and realizes noninvasive detection by taking saliva as a detection sample.
Description
Technical Field
The invention relates to the technical field of immunodiagnosis, in particular to test paper for detecting 2019 novel coronavirus and a preparation method thereof.
Background
The pandemic caused by the novel coronavirus SARS-CoV-2 is a sudden public health event of international concern. At present, the epidemic situation in most countries has not reached a peak, and future cases and death rates are expected to continue to rise. The diagnosis of suspected cases of COVID-19 and asymptomatic infectors is very important for timely receiving and treating patients and taking effective isolation measures as soon as possible.
The current widely used etiology detection is based on real-time PCR nucleic acid detection (RT-PCR) of collected nasopharyngeal swab samples. However, in practical application, because the RT-PCR detection operation is complicated and long in time consumption, the detection result is easily influenced by a plurality of factors such as sample quality, virus infection parts, sampling time and the like, the RT-PCR positive detection rate of suspected cases in COVID-19 clinical and epidemiology is about 50%, a large number of false negative results appear in the detection process, and the difficulty of epidemic situation prevention and control is further increased.
The Chinese patent publication No. CN111273003A discloses a novel immunochromatographic test strip for detecting coronavirus, which is formed by bonding a sample pad, a bonding pad, a nitrocellulose membrane and a water absorption plate on a bottom plate in a way of mutually lapping in sequence, and can quickly detect specific antibody IgM or IgG of a patient; however, the test paper still has high false negative rate, and simultaneously the antibody detection has the problem of window period, and the condition of missed detection exists in the initial infection stage. Chinese patent publication No. CN111337669A discloses a novel test paper for rapid detection of coronavirus and a detection method thereof, wherein a coronavirus protein antigen is specifically captured and combined by double enzyme-linked immunosorbent assay, and then judgment is performed by chemical color development; however, the test paper is complex to operate and poor in accuracy, detection is completed by matching with an enzyme-labeling instrument, results cannot be obtained quickly, and meanwhile, the test paper is easily interfered by other viruses or biological macromolecules.
In the face of increasingly severe epidemic situations, many suspected cases still cannot be diagnosed due to insufficient detection capability, so how to rapidly screen out the suspected cases on a large scale, improve detection accuracy and safety, provide rapid diagnosis basis for clinicians, and is a problem to be solved urgently at present.
Disclosure of Invention
The invention aims to solve the defects in the prior art and provides a colloidal tellurium test paper for detecting 2019 novel coronavirus in saliva and a preparation method thereof. The test paper can directly detect the novel coronavirus antigen, the detection sample is saliva, the source is convenient, the sensitivity is high, large-scale accurate and quick screening is convenient to realize, and noninvasive detection of the novel coronavirus is realized.
The purpose of the invention is realized by the following technical scheme:
the utility model provides a colloid tellurium test paper for saliva detects 2019 novel coronavirus, includes the PVC offset plate and overlap joint in proper order inhale a kind pad, colloid tellurium combination pad, chromatographic column, water absorption material on the PVC offset plate.
The sample sucking pad is a glass fiber membrane which is impregnated by a first treatment solution containing a nonionic surfactant. Preferably, the nonionic surfactant is triton-100.
The colloidal tellurium binding pad is a glass fiber film or a polyester film which is impregnated by a second treatment solution containing a colloidal tellurium-labeled chicken IgY compound and a colloidal tellurium-labeled novel coronavirus S1 antibody 1 compound, wherein the novel coronavirus S1 antibody 1 is a rabbit monoclonal antibody with the product number of 40592-R001 of Beijing Yi Qiao Shenzhou science and technology Limited. Furthermore, the mass ratio of the chicken IgY to the novel coronavirus S1 antibody 1 in the second treatment solution is 3: 1. Furthermore, the colloidal tellurium is prepared by taking hydroxypropyl methyl cellulose as a template and tellurite and vitamin C as reaction raw materials. Preferably, the mass ratio of the hydroxypropyl methyl cellulose to the tellurite to the vitamin C is 1-5:2: 10-30.
The chromatographic membrane is a nitrocellulose membrane, detection lines and quality control lines are arranged on the chromatographic membrane at intervals, novel coronavirus S1 antibody 2 is sprayed on the detection lines, and goat anti-chicken IgY polyclonal antibodies are sprayed on the quality control lines. Wherein, the novel coronavirus S1 antibody 2 is a rabbit polyclonal antibody with the product number of 40591-T62 of Beijing Yizhushanhua science and technology GmbH; the relative positions of the quality control line and the detection line are adjustable. Preferably, the distance between the detection line and the quality control line is 4 mm.
The colloidal tellurium test paper has the advantages of simple structure, easily obtained materials, low cost and convenient production.
Compared with the prior art, the colloidal tellurium test paper for saliva detection 2019 novel coronavirus has the following advantages:
(1) the invention uses a specific antibody aiming at 2019 novel coronavirus key S protein (spike protein), and the sensitivity can reach 1 ng/mL.
(2) The sample used by the invention is saliva, has wide sources, can be collected at any time and any place, and can really realize the noninvasive detection of the novel coronavirus.
(3) The operation process is simple, and the summary is as follows: SUCK & CHECK, the detection result is rapidly obtained in 15 minutes.
(4) The antigen detection can be used as a secondary diagnosis method for nucleic acid detection of discharged cases, so that new transmission risks caused by discharge of false negative patients are avoided.
(5) Compared with the conventional colloidal gold test paper, the colloidal tellurium test paper has higher sensitivity and lower price.
The preparation method of the colloidal tellurium test paper for detecting 2019 novel coronavirus by saliva comprises the following steps:
s1, preparing a sample suction pad: and (3) dipping the first membrane material by using a first treatment solution, and drying to obtain the sample pad. The first membrane material is a glass fiber membrane. The first treatment liquid comprises the following components in percentage by mass: 0.02-0.1% of boric acid, 0.05-0.25% of borax, 0.1-1.5% of triton-1000.5, 0.1-1% of casein and the balance of water. Preferably, the first treatment liquid comprises the following components in percentage by mass: 0.06% of boric acid, 0.15% of borax, 0.5% of triton-1001%, 0.5% of casein and the balance of water. Wherein, triton-100 is a commonly used nonionic surfactant and is the prior art.
S2, preparing a colloidal tellurium bonding pad: soaking a second treatment solution prepared by mixing colloidal tellurium-labeled chicken IgY and colloidal tellurium-labeled novel coronavirus S1 antibody 1 on a second membrane material, and drying to obtain a colloidal tellurium binding pad; as an example of the present invention, the second film material may be a glass fiber film or a polyester film.
S3, preparation of chromatographic membranes: respectively spraying a novel coronavirus S1 antibody 2 with the concentration of 1.0-2.0 mg/mL and goat anti-chicken IgY with the concentration of 1.0-2.0 mg/mL on a third membrane material at intervals to form a detection line and a quality control line, wherein the interval between the detection line and the quality control line is 1-5 mm; preferably, the detection line and the quality control line are separated by 4 mm. Preferably, the third membrane material is a nitrocellulose membrane, such as a commercially available NC membrane of sartorius CN 140.
S4, assembling: and (3) sticking the water absorbing material, the prepared sample absorbing pad, the colloidal tellurium binding pad and the chromatographic membrane at fixed positions in sequence, and cutting to obtain the product. The test strip may have a width of 3mm, 3.5mm or 4.0 mm.
Further, in step S2, the method for preparing the second processing liquid includes:
s21, preparing colloidal tellurium: adding 1g of hydroxypropyl methyl cellulose into 100mL of purified water, and uniformly stirring to obtain a template; respectively adding 10g of vitamin C and 2g of tellurite, and uniformly stirring; and continuing to react for 30 minutes to obtain the product.
S22, preparing a chicken IgY-colloidal tellurium compound: taking 1mL of the colloidal tellurium prepared in the step S21, adding 5-50 mu L0.1M of potassium carbonate, uniformly mixing, and adjusting the pH value to 8.0-10.0; then adding 5-30 mug of chicken IgY and 50-100 mug L of 10% bovine serum albumin in sequence, and mixing evenly.
Preparation of S23, novel coronavirus S1 antibody 1-colloidal tellurium complex: adding 10-50 mu L of 0.1M potassium carbonate into 1mL of the colloidal tellurium prepared in the step S21, uniformly mixing, adjusting the pH to 8.0-10.0, sequentially adding 5-20 mu g of novel coronavirus S1 antibody 1 and 50-100 mu L of 10% bovine serum albumin, and uniformly mixing; the pH may be 8.0, 8.5, 9.5, or 10.0.
S24, purification of colloidal tellurium complexes: and (4) respectively centrifuging the chicken IgY-colloidal tellurium compound prepared in the step S22 and the novel coronavirus S1 antibody 1-colloidal tellurium compound prepared in the step S23, discarding the supernatant, and adding an isovolumetric re-dissolving solution for re-dissolving to obtain the compound. Preferably, the re-solution comprises the following components in percentage by mass: 0.06% boric acid, 0.15% borax, 0.5% casein, 0.5% surfactant S9 (Tetronic 1307), 0.1% bovine serum albumin, 0.5% polyvinylpyrrolidone, 20% sucrose and 5% trehalose, and the balance water.
S25, mixing the re-dissolved chicken IgY-colloidal tellurium compound and the novel coronavirus S1 antibody 1-colloidal tellurium compound according to the volume ratio of 2:1 to obtain a second treatment solution.
Further, the drying conditions of step S1 are: drying for 10-24 h at 30-60 ℃; for example, drying at 30 ℃ for 24h or at 40 ℃ for 18h, at 50 ℃ for 12h or at 60 ℃ for 10 h; the drying conditions of step S2 are: drying for 10-24 h at 30-55 ℃; the drying conditions of step S3 are: drying for 4-12 h at 30-50 ℃.
The preparation method of the colloidal tellurium test paper for detecting 2019 novel coronavirus has the same beneficial effects as the test paper, and the description is omitted here.
Drawings
FIG. 1 is a schematic structural diagram of the colloidal tellurium test paper for saliva detection of COVID-19 coronavirus according to the present invention.
Detailed Description
It should be noted that the embodiments and features of the embodiments may be combined with each other without conflict.
Experimental methods without specific conditions noted in the following examples, generally according to conventional conditions; the various chemicals used in the examples are commercially available. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. The terminology used in the description of the invention herein is for the purpose of describing particular embodiments only and is not intended to be limiting of the invention.
Unless otherwise specified, the units% in the present invention are mass percentages.
Example 1:
the embodiment provides a colloidal tellurium test paper for saliva detects 2019 novel coronavirus, including the PVC offset plate and overlap joint in proper order inhale a kind pad, colloidal tellurium combines pad, chromatographic column, water absorption material on the PVC offset plate, its structural diagram is shown in fig. 1, wherein:
the sample absorbing pad is made of glass fiber membrane. Soaking the glass fiber membrane in the first treating solution for 5-10min, wherein the soaking treatment amount is 800g/m2(i.e., each square meter of the glass fiber membrane is soaked by 800g of the first treatment solution), the glass fiber membrane is dried in an air-blast drying oven at 55 ℃ for 16h, and the glass fiber membrane is cut into the size of 25mm by 300mm for standby after drying. The first treatment liquid comprises the following components in percentage by mass: 0.06% of boric acid, 0.15% of borax, 0.5% of casein, triton-1000.5% and the balance of purified water. Adjusting the pH of the first treatment solution to 7.5 +/-0.05 by using 6M NaOH, and then supplementing purified water to 100 percent.
Colloidal tellurium bonding pads: adding 1g of hydroxypropyl methyl cellulose into 100mL of purified water, and uniformly stirring to obtain a template; respectively adding 10g of vitamin C and 2g of tellurite, and uniformly stirring; the reaction was continued for 30 minutes to obtain colloidal tellurium. Adding 20 mu L of 0.1M potassium carbonate into 1mL of colloidal tellurium, uniformly mixing, adjusting the pH to 9.0 by using 6M NaOH, adding 15 mu g of chicken IgY, fully and uniformly mixing, adding 100 mu L of 10% bovine serum albumin, and uniformly mixing to obtain a chicken IgY-colloidal tellurium compound; adding 25 mu L of 0.1M potassium carbonate into 1mL of colloidal tellurium, uniformly mixing, adjusting the pH to 9.2 by using 6M NaOH, adding 10 mu g of novel coronavirus S1 antibody 1 (purchased from Beijing Yizhao Shenzhou science and technology Co., Ltd., product name: novel coronavirus spike protein neutralizing antibody, rabbit monoclonal antibody, product number: 40592-R001), fully mixing, adding 100 mu L of 10% bovine serum albumin, and uniformly mixing to obtain a novel coronavirus S1 antibody 1-colloidal tellurium compound; centrifuging the chicken IgY-colloidal tellurium compound and the novel coronavirus S1 antibody 1-colloidal tellurium compound respectively, removing the supernatant, and adding a re-dissolving solution containing 20% of sucrose and 5% of trehalose for re-dissolving in equal volume; mixing the re-dissolved chicken IgY-colloidal tellurium compound and the novel coronavirus S1 antibody 1-colloidal tellurium compound according to the volume ratio of 2:1 to obtain a second treatment solution, wherein the concentration of the chicken IgY in the second treatment solution is 10 mu g/mL, and the concentration of the novel coronavirus S1 antibody 1 is 3.3 mu g/mL. Immersing the glass fiber membrane in a second treatment solution, wherein the immersion treatment amount is 650g/m2(i.e., each square meter of the glass fiber membrane is soaked by 650g of the second treatment solution), the glass fiber membrane is placed into a blast drying oven to be dried, the drying temperature is 37 ℃, the drying time is 20 hours, and the glass fiber membrane is cut into the size of 5mm x 300mm for standby after drying. Wherein the redissolution is prepared from the following components in percentage by massThe components by percentage are as follows: 0.06% of boric acid, 0.15% of borax, 0.5% of casein, 0.5% of surfactant S9 (Tetronic 1307) (purchased from Beijing Baiolai Boke technology Co., Ltd.), 0.1% of bovine serum albumin, 0.5% of polyvinylpyrrolidone, 20% of sucrose, 5% of trehalose and the balance of purified water.
Chromatography membrane: nitrocellulose membrane, a commercially available NC membrane of sartorius CN140, was selected. Goat anti-chicken IgY polyclonal antibody with the concentration of 1.0mg/mL, novel coronavirus S1 antibody 2 with the concentration of 1.5mg/mL (purchased from Beijing Yiqiao Shenzhou science and technology Co., Ltd., product name: New coronavirus (SARS-CoV-2) Spike antibody, rabbit polyclonal antibody, product number: 40591-T62) are respectively sprayed on an NC membrane to prepare a quality control line C and a detection line T, and the quality control line C and the detection line T are placed into a blast drying oven to be dried for 12 hours at 37 ℃. The size of the nitrocellulose membrane was 25mm by 300 mm.
Water-absorbing material: the absorbent paper is cut into the size of 20mm 300mm for standby.
PVC rubber plate: PVC plates of 75mm by 300mm were selected.
And sequentially sticking the prepared sample sucking pad, the colloidal tellurium binding pad, the chromatographic membrane and the water absorbing material at fixed positions on a bottom plate, and cutting into pieces of 75mm x 4mm to obtain the colloidal tellurium test paper for detecting 2019 novel coronavirus.
Comparative example 1:
the procedure was as in example 1 except that the label used was gold colloid.
Example 2:
(1) detection of a sample
Collecting saliva with a sampling cup, inserting a sample sucking pad of the detection test paper into the saliva for at least 30 seconds, then flatly placing the detection test paper on a table, waiting for 15 minutes, and observing a detection result. When the detection line T and the quality control line C have strips, the display result is positive; when no strip appears in the detection line T, the display result is negative when a strip appears in the quality control line C.
(2) Sensitivity testing
A novel coronavirus S protein recombinant antigen (purchased from Beijing Yiqiao Shenzhou science and technology Co., Ltd., product name: SARS-CoV-2 (2019-nCoV) Spike S1-Fc recombinant protein, product number: 40591-V02H) is selected as a quality control product, the quality control product is respectively diluted to 10ng/mL, 5ng/mL, 1ng/mL and 0.1ng/mL for dilution, test is carried out by using the test paper prepared in example 1 and comparative example 1, each quality control product is tested repeatedly for 3 times, and the test results are shown in Table 1.
TABLE 1 results of sensitivity detection
Note: the negative test results are represented by "-", and the positive test results are represented by "+" (weak), "+" (middle), "+" (strong) respectively according to the intensity of color development.
As can be seen from the test results in Table 1, the test paper prepared in example 1 of the present invention shows positive test results at the concentrations of the quality control substances of 10ng/mL, 5ng/mL and 1ng/mL, and shows weak positive test results at the concentration of 0.1 ng/mL. The test paper prepared in comparative example 1 showed positive results at the concentrations of the quality control substances of 10ng/mL and 5ng/mL, but showed negative results at the concentration of the quality control substance of 1 ng/mL. The result shows that the test paper prepared in the example 1 has higher detection sensitivity and can reach 1 ng/mL.
(3) Negative and positive compliance rate testing of clinical specimens
30 parts of the novel coronavirus sample with a positive nucleic acid detection result and 30 parts of the novel coronavirus sample with a negative nucleic acid detection result were selected and tested using the test paper prepared in example 1 and comparative example 1, and the test results are shown in tables 2 and 3.
TABLE 2 Positive sample test results
| Sample numbering | Example 1 | Comparative example 1 | PCR detection results |
| 1 | + | + | Positive for |
| 2 | + | + | Positive for |
| 3 | + | + | Positive for |
| 4 | + | + | Positive for |
| 5 | + | + | Positive for |
| 6 | + | - | Positive for |
| 7 | + | + | Positive for |
| 8 | + | + | Positive for |
| 9 | + | + | Positive for |
| 10 | + | + | Positive for |
| 11 | + | + | Positive for |
| 12 | + | + | Positive for |
| 13 | + | + | Positive for |
| 14 | + | + | Positive for |
| 15 | + | - | Positive for |
| 16 | + | + | Positive for |
| 17 | + | + | Positive for |
| 18 | + | - | Positive for |
| 19 | + | + | Positive for |
| 20 | + | + | Positive for |
| 21 | + | + | Positive for |
| 22 | + | + | Positive for |
| 23 | + | - | Positive for |
| 24 | - | + | Positive for |
| 25 | + | + | Positive for |
| 26 | + | - | Positive for |
| 27 | + | + | Positive for |
| 28 | + | + | Positive for |
| 29 | + | + | Positive for |
| 30 | + | + | Positive for |
TABLE 3 negative sample test results
| Sample numbering | Example 1 | Comparative example 1 | PCR detection results |
| 1 | - | - | Positive for |
| 2 | - | - | Positive for |
| 3 | - | + | Positive for |
| 4 | - | - | Positive for |
| 5 | - | - | Positive for |
| 6 | - | - | Positive for |
| 7 | - | - | Positive for |
| 8 | - | - | Positive for |
| 9 | - | - | Positive for |
| 10 | - | - | Positive for |
| 11 | - | - | Positive for |
| 12 | - | - | Positive for |
| 13 | - | - | Positive for |
| 14 | - | - | Positive for |
| 15 | - | - | Positive for |
| 16 | - | - | Positive for |
| 17 | - | - | Positive for |
| 18 | - | - | Positive for |
| 19 | - | - | Positive for |
| 20 | - | - | Positive for |
| 21 | - | - | Positive for |
| 22 | - | - | Positive for |
| 23 | - | + | Positive for |
| 24 | - | - | Positive for |
| 25 | - | - | Positive for |
| 26 | - | + | Positive for |
| 27 | - | - | Positive for |
| 28 | - | - | Positive for |
| 29 | - | - | Positive for |
| 30 | - | - | Positive for |
As can be seen from the test results in tables 2 and 3, the test strips prepared in example 1 had a positive coincidence rate of 96.7% (29/30) and negative coincidence rates of 100% (30/30), and the test strips prepared in comparative example 1 had a positive coincidence rate of 83.3% (25/30) and negative coincidence rates of 90% (27/30). The above results show that: the test paper prepared in example 1 has a significantly better negative and positive coincidence rate of clinical samples than the test paper prepared in comparative example 1.
(4) Cross-reaction and interference testing
Some other positive specimens of common infectious diseases were added to the novel coronavirus positive specimens and the novel coronavirus negative specimens, and the test results were shown in Table 4 using the test strips prepared in example 1 and comparative example 1.
TABLE 4 detection results of the Cross-reactivity
As can be seen from the test results in table 4, the test paper prepared in example 1 has no cross reaction with staphylococcus aureus, streptococcus pneumoniae, measles virus, mumps virus, adenovirus type 3, mycoplasma pneumoniae, parainfluenza type 2, metapneumovirus, coronavirus OC43, coronavirus 229E, parapertussis, influenza b virus (Victoria series) H1N1 (2009) influenza a virus, H3N2 influenza a virus, avian influenza virus H7N9, avian influenza virus H5N1, EB virus, enterovirus CA16, and rhinovirus, while the test paper prepared in comparative example 1 has cross reaction with adenovirus type 3 and mycoplasma pneumoniae, and therefore, the specificity of the test paper prepared in example 1 is superior to that of the test paper prepared in comparative example 1.
The above description is only for the purpose of illustrating the preferred embodiments of the present invention and is not to be construed as limiting the invention, and any modifications, equivalents, improvements and the like that fall within the spirit and principle of the present invention are intended to be included therein.
Claims (8)
1. A colloidal tellurium test paper for detecting 2019 novel coronavirus, which is characterized in that: comprises a PVC rubber plate, and a sample absorption pad, a colloidal tellurium binding pad, a chromatographic membrane and a water absorption material which are sequentially lapped on the PVC rubber plate;
the sample sucking pad is a glass fiber membrane containing a nonionic surfactant;
the colloidal tellurium binding pad is a glass fiber film or a polyester film containing a colloidal tellurium-labeled chicken IgY compound and a colloidal tellurium-labeled novel coronavirus S1 antibody 1 compound; wherein, the novel coronavirus S1 antibody 1 is a rabbit monoclonal antibody with the product number of 40592-R001 of Beijing Yizhushanhua science and technology Co., Ltd;
the chromatographic membrane is a nitrocellulose membrane, detection lines and quality control lines are arranged on the chromatographic membrane at intervals, a novel coronavirus S1 antibody 2 is sprayed on the detection lines, a goat anti-chicken IgY polyclonal antibody is sprayed on the quality control lines, and the novel coronavirus S1 antibody 2 is a rabbit polyclonal antibody with the product number of 40591-T62 of Beijing Yi Qianzhou science and technology Limited company.
2. The colloidal tellurium test strip of claim 1, for detecting 2019 a novel coronavirus, wherein: the non-ionic surfactant is triton-100.
3. The colloidal tellurium test strip of claim 1, for detecting 2019 a novel coronavirus, wherein: the mass ratio of the chicken IgY to the novel coronavirus S1 antibody 1 in the colloidal tellurium binding pad is 3: 1.
4. The colloidal tellurium test strip of claim 1, for detecting 2019 a novel coronavirus, wherein: the colloidal tellurium is prepared by taking hydroxypropyl methyl cellulose as a template and tellurite and vitamin C as reaction raw materials.
5. The colloidal tellurium test strip of claim 4, for detecting 2019 a novel coronavirus, wherein: the mass ratio of the hydroxypropyl methyl cellulose to the tellurite to the vitamin C is 1-5:2: 10-30.
6. The method of preparing the colloidal tellurium test paper for detecting 2019 novel coronaviruses as claimed in any one of claims 1 to 5, wherein: the method comprises the following steps:
s1, preparing a sample suction pad: dipping the first membrane material by using a first treatment solution, and drying to obtain a sample pad; the first membrane material is a glass fiber membrane; the first treatment liquid comprises the following components in percentage by mass: 0.02-0.1% of boric acid, 0.05-0.25% of borax, 0.1-1.5% of triton-1000.5, 0.1-1% of casein and the balance of water;
s2, preparing a colloidal tellurium bonding pad: soaking a second treatment solution prepared by mixing colloidal tellurium-labeled chicken IgY and colloidal tellurium-labeled novel coronavirus S1 antibody 1 on a second membrane material, and drying to obtain a colloidal tellurium binding pad; the second film material can be a glass fiber film or a polyester film;
s3, preparation of chromatographic membranes: spraying the novel coronavirus S1 monoclonal antibody 2 and goat anti-chicken IgY at intervals on a third membrane material to form a detection line and a quality control line; the third membrane material is a nitrocellulose membrane;
s4, assembling: and (3) sticking the water absorbing material, the prepared sample absorbing pad, the colloidal tellurium binding pad and the chromatographic film at the upper fixed positions in sequence, and slitting to obtain the colloidal tellurium test paper for detecting 2019 novel coronavirus in saliva.
7. The method for preparing the colloidal tellurium test paper for detecting 2019 novel coronavirus as claimed in claim 6, wherein the colloidal tellurium test paper is prepared by the following steps: in step S2, the method for preparing the second treatment liquid includes:
s21, preparing colloidal tellurium: adding 1g of hydroxypropyl methyl cellulose into 100mL of purified water, and uniformly stirring to obtain a template; respectively adding 10g of vitamin C and 2g of tellurite, and uniformly stirring; continuously reacting for 30 minutes to obtain colloidal tellurium;
s22, preparing a chicken IgY-colloidal tellurium compound: taking 1mL of the colloidal tellurium prepared in the step S21, adding 5-50 mu L of 0.1M potassium carbonate, uniformly mixing, and adjusting the pH value to 8.0-10.0; sequentially adding 5-30 μ g of chicken IgY and 50-100 μ L of 10% bovine serum albumin, and mixing;
preparation of S23, novel coronavirus S1 antibody 1-colloidal tellurium complex: adding 10-50 mu L of 0.1M potassium carbonate into 1mL of the colloidal tellurium prepared in the step S21, uniformly mixing, adjusting the pH to 8.0-10.0, sequentially adding 5-20 mu g of novel coronavirus S1 antibody 1 and 50-100 mu L of 10% bovine serum albumin, and uniformly mixing;
s24, purification of colloidal tellurium complexes: respectively centrifuging the chicken IgY-colloidal tellurium compound prepared in the step S22 and the novel coronavirus S1 antibody 1-colloidal tellurium compound prepared in the step S23, removing supernatant, and adding an isovolumetric re-dissolving solution for re-dissolving; the complex solution comprises the following components in percentage by mass: 0.06% of boric acid, 0.15% of borax, 0.5% of casein, 0.5% of S-9, 0.1% of bovine serum albumin, 0.5% of polyvinylpyrrolidone, 20% of sucrose and 5% of trehalose, and the balance of purified water;
s25, mixing the re-dissolved chicken IgY-colloidal tellurium compound and the novel coronavirus S1 monoclonal antibody 1-colloidal tellurium compound according to the volume ratio of 2:1 to obtain a second treatment solution.
8. The method for preparing the colloidal tellurium test paper for detecting 2019 novel coronavirus as claimed in claim 6, wherein the colloidal tellurium test paper is prepared by the following steps: the drying conditions of step S1 are: drying for 10-24 h at 30-60 ℃; the drying conditions of step S2 are: drying for 10-24 h at 30-55 ℃; the drying conditions of step S3 are: drying for 4-12 h at 30-50 ℃.
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113219174A (en) * | 2021-04-14 | 2021-08-06 | 中国科学院西北高原生物研究所 | Novel coronavirus S antigen detection kit and use method thereof |
Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP0321008A1 (en) * | 1987-11-19 | 1989-06-21 | H.B.T. Holland Biotechnology B.V. | Immunodetermination using non-metallic labels |
| US5348891A (en) * | 1988-11-15 | 1994-09-20 | H.B.T. Holland Biotechnology | Method for an increased visualization of the reaction product of a specifically binding substance and a corresponding bindable substance and test kit therefor |
| CN111273003A (en) * | 2020-03-05 | 2020-06-12 | 广州万孚生物技术股份有限公司 | 2019-nCoV novel coronavirus rapid detection immunochromatographic test strip |
| CN111366729A (en) * | 2020-03-26 | 2020-07-03 | 深圳市梓健生物科技有限公司 | Novel coronavirus COVID-19 antigen fluorescence detection kit and preparation method thereof |
| CN111521805A (en) * | 2020-05-12 | 2020-08-11 | 北京倍肯恒业科技发展股份有限公司 | 2019 novel coronavirus antigen detection reagent and preparation method thereof |
-
2020
- 2020-10-25 CN CN202011151402.3A patent/CN112611863A/en active Pending
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP0321008A1 (en) * | 1987-11-19 | 1989-06-21 | H.B.T. Holland Biotechnology B.V. | Immunodetermination using non-metallic labels |
| US5348891A (en) * | 1988-11-15 | 1994-09-20 | H.B.T. Holland Biotechnology | Method for an increased visualization of the reaction product of a specifically binding substance and a corresponding bindable substance and test kit therefor |
| CN111273003A (en) * | 2020-03-05 | 2020-06-12 | 广州万孚生物技术股份有限公司 | 2019-nCoV novel coronavirus rapid detection immunochromatographic test strip |
| CN111366729A (en) * | 2020-03-26 | 2020-07-03 | 深圳市梓健生物科技有限公司 | Novel coronavirus COVID-19 antigen fluorescence detection kit and preparation method thereof |
| CN111521805A (en) * | 2020-05-12 | 2020-08-11 | 北京倍肯恒业科技发展股份有限公司 | 2019 novel coronavirus antigen detection reagent and preparation method thereof |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113219174A (en) * | 2021-04-14 | 2021-08-06 | 中国科学院西北高原生物研究所 | Novel coronavirus S antigen detection kit and use method thereof |
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