Dengue fever detection kit

Abstract

The invention discloses a dengue fever detection kit, and belongs to the technical field of detection kits, the dengue fever detection kit comprises a sample pad, a colloidal gold pad, a nitrocellulose membrane, a detection test strip, a PVC bottom plate and a water absorption pad, the nitrocellulose membrane is coated with an IgG test line, an IgM test line and a control line; the preparation method of the colloidal gold pad comprises the following steps: S1, preparing a dengue fever (I + III) type labeled antigen colloidal gold compound; preparing a dengue (II + IV) type labeled antigen colloidal gold compound; preparing a chicken IgY colloidal gold compound; and S2, uniformly mixing the dengue (I + III) type labeled antigen-colloidal gold compound, the dengue (II + IV) type labeled antigen-colloidal gold compound and the chicken IgY colloidal gold compound according to a volume ratio, then spraying the mixture on a glass cellulose membrane, and drying to obtain the product. The dengue detection kit has the characteristics of accurate detection and high sensitivity, is wide in application range, and provides powerful assistance for detection of dengue diseases.

Description

Technical Field

[0001] The invention belongs to the technical field of detection kits, and in particular relates to a dengue fever detection kit. Background Art

[0002] Dengue fever is a febrile disease. The dengue virus includes 4 serotypes. In nature, it is mainly transmitted by the bites of Aedes aegypti and Aedes albopictus mosquitoes, and generally occurs in tropical and subtropical areas. Symptoms generally appear 3 to 14 days after the infected bite. The diseases caused include: asymptomatic infection, general fever, mild dengue fever, and severe dengue hemorrhagic fever and dengue shock syndrome, which affect infants, young children and adults. Signs and symptoms of dengue fever may include fever, headache, rash, body pain and bleeding manifestations. Symptoms can be mild or severe. Severe dengue fever usually requires hospitalization. Dengue fever occasionally presents with symptoms and signs of aseptic meningitis or encephalitis.

[0003] The acute phase of dengue virus infection lasts about 5 to 7 days, followed by an immune response. After the first infection, protection against the other three types of viruses is only temporary and partial. After a short time, the immune protection weakens and the patient may be at risk of a second or subsequent infection.

[0004] Therefore, the present invention provides a colloidal gold immunochromatography detection method for dengue fever IgG / IgM antibody detection, which is a non-invasive test, has a fast and simple operation process, and is easily accepted by patients, so as to solve the problems raised in the above background technology. Summary of the invention

[0005] The present invention provides a dengue fever detection kit, which overcomes the shortcomings of the prior art and realizes the rapid detection of dengue fever markers-IgG / IgM antibodies. It has the advantages of being non-invasive, fast and simple to operate, accurate in detection, high in sensitivity, low in cost, etc., has a wide range of applications, is suitable for wide promotion, and provides powerful assistance for the detection of dengue fever.

[0006] The purpose of the present invention can be achieved by the following technical solutions:

[0007] A dengue fever detection kit comprises a sample pad, a colloidal gold pad, a nitrocellulose membrane (NC membrane), a test paper strip, a PVC bottom plate, and a water-absorbing pad, wherein the nitrocellulose membrane is coated with an IgG test line (G line), an IgM test line (M line), and a control line (C line);

[0008] Wherein, the preparation of the colloidal gold pad is as follows:

[0009] S1. preparing colloidal gold complex of dengue fever (I+III) type labeled antigen; preparing colloidal gold complex of dengue fever (II+IV) type labeled antigen; preparing colloidal gold complex of chicken IgY;

[0010] S2. Evenly mix the dengue fever (I+III) type labeled antigen-colloidal gold complex, dengue fever (II+IV) type labeled antigen-colloidal gold complex, and chicken IgY colloidal gold complex in a volume ratio of 1:1:1, then evenly spray them on the glass cellulose membrane KB50, and dry them at 37-45°C for 4-24h to obtain.

[0011] Further, the preparation of dengue fever (I+III) type labeled antigen colloidal gold complex:

[0012] A colloidal gold solution with a particle size of 530-533 nm and an OD value of 4.0-5.0A is prepared by a citric acid reduction method; 12.5 μL of 0.2M K2CO3 is added to 1 mL of the colloidal gold solution, mixed, 5 μg of dengue fever (Ⅰ+Ⅲ) type labeled antigen is added, mixed, and reacted at 18-25°C for 30-60 minutes by standing or stirring, BSA is added until the final concentration of BSA is 1%, and blocked, and reacted at 18-25°C for 30-60 minutes by standing or stirring, and then centrifuged at 4-25°C at 10000 rpm for 15-45 minutes, the supernatant is removed by suction to obtain a precipitate, and a gold labeling solution is added for resuspending to obtain a dengue fever (Ⅰ+Ⅲ) type labeled antigen-colloidal gold complex.

[0013] Further, the preparation of dengue fever (Ⅱ+Ⅳ) type labeled antigen colloidal gold complex:

[0014] A colloidal gold solution with a particle size of 530-533 nm and an OD value of 4.0-5.0A is prepared by a citric acid reduction method; 12.5 μL of 0.2M K2CO3 is added to 1 mL of the colloidal gold solution, mixed to obtain a mixed solution, 5 μg of dengue fever (II+IV) type labeled antigen is added, mixed, and reacted at 18-25°C for 30-60 minutes by standing or stirring, BSA is added until the final concentration of BSA is 1%, and blocked, and reacted at 18-25°C for 30-60 minutes by standing or stirring, and then centrifuged at 4-25°C at 10000 rpm for 15-45 minutes, the supernatant is removed by suction to obtain a precipitate, and a gold labeling solution is added for re-suspending to obtain a dengue fever (II+IV) type labeled antigen-colloidal gold complex.

[0015] Further, the preparation of chicken IgY colloidal gold complex:

[0016] A colloidal gold solution with a particle size of 530-535 nm and an OD value of 4.0-5.0A is prepared by a citric acid reduction method; 15-25 μl of 0.2M K2CO3 is added to 1 mL of the colloidal gold solution, mixed, 0.7-1.2 μg of chicken IgY antibody is added, mixed, reacted at 18-25°C for 30-60 minutes by standing or stirring, BSA is added until the final concentration of BSA is 1%, reacted at 18-25°C for 30-60 minutes by standing or stirring, then centrifuged at 4-25°C for 15-45 minutes at 10000 rpm, the supernatant is removed by suction to obtain a precipitate, a gold labeling solution is added for re-suspending, and a chicken IgY colloidal gold complex is obtained.

[0017] Furthermore, the sample pad is prepared by:

[0018] Pour the sample pad treatment solution into a container, slowly place the glass cellulose membrane 8965 into the solution, and keep tapping to drive out the bubbles. Rotate and drain until no obvious water drops fall. Place it on a drying rack, keep it flat, and dry it at 37-45℃ until it is completely dry.

[0019] Furthermore, the sample pad treatment solution used is: 50mM Tris + 0.2% S9 + 0.6% PVPK-30 + 0.8% NaCl + 0.3% C-Na + 0.2% EDTA-Na2 + 0.03mg / mL blocker-H + 0.2mg / mL RBC antibody, pH = 8.6. Among them, Tris is trishydroxymethylaminomethane; S9 is a surfactant, which belongs to propylene oxide-ethylene oxide-ethylene diamine copolymer and has good water solubility; PVPK-30 is polyvinyl pyrrolidone; C-Na is Casein-Na, that is, sodium caseinate.

[0020] Furthermore, the coating method of the detection line is:

[0021] Dengue mouse anti-human IgG antibody and dengue mouse anti-human IgM antibody were diluted to 1.0 mg / mL with coating buffer, coated on the NC membrane, and dried at 45°C for 24 to 48 hours;

[0022] The control line is coated by:

[0023] The goat anti-chicken IgY monoclonal antibody was diluted to 1.0 mg / mL with coating buffer and then coated on the NC membrane, and then dried at 45° C. to 55° C. for 24 to 72 hours.

[0024] Furthermore, the coating buffer used was: 20 mM Tris, 100 mM NaCl, and trehalose accounting for 3% of the total mass of the coating buffer.

[0025] Furthermore, the assembly method of the test strip is:

[0026] After the humidity of the environment drops below 40%, take out the required materials and assemble them as required, tear off the plastic sticker at the absorbent paper position, and stick the absorbent paper of the specified size with the upper end aligned with the plastic sticker; tear off the plastic sticker at the sample pad and gold label pad positions, first stick the gold label pad with the upper end covering 1-2mm on the NC film, then stick the lower end of the sample pad aligned with the plastic sticker, and the upper end covers the gold label pad by 1-2mm.

[0027] Furthermore, the gold label complex solution is: Tris+pvpk-30+TX-100+C-Na+trehalose+BSA; Tris is trishydroxymethylaminomethane; PVPK-30 is polyvinylpyrrolidone; C-Na is sodium caseinate; TX-100 is triton.

[0028] Beneficial effects of the present invention:

[0029] The dengue fever IgG / IgM antibody detection kit (colloidal gold method) of the present invention adopts a highly specific antigen-antibody reaction principle and an immunochromatographic analysis technique to detect dengue fever IgG / IgM antibodies in a blood sample through an antibody capture immunochromatographic detection method.

[0030] Furthermore, colloidal gold solution was prepared by citric acid reduction method, colloidal gold labeled dengue (I+III) type labeled antigen, dengue (II+IV) type labeled antigen and chicken IgY antibody were pre-coated on glass fiber membrane, dengue mouse anti-human IgG antibody, dengue mouse anti-human IgM antibody and goat anti-chicken IgY antibody were coated on nitrocellulose membrane test line (G / M) and control line (C) respectively. Sample pad, gold label pad, nitrocellulose membrane and absorbent paper were attached to the PVC bottom plate in the chromatography direction.

[0031] The present invention adopts colloidal gold chromatography method, can realize the rapid detection of dengue fever IgG / IgM antibody in blood, and according to the different color development of the detection line of dengue fever IgG / IgM antibody of different concentrations, a standard colorimetric card is established, and the detection of dengue fever symptoms can be further realized by comparison. The dengue fever detection kit provided by the present invention has the characteristics of accurate detection and high sensitivity, wide application range, suitable for wide promotion, and provides powerful assistance for the detection of dengue fever disease. BRIEF DESCRIPTION OF THE DRAWINGS

[0032] The present invention will be further described below in conjunction with the accompanying drawings.

[0033] Figure 1 It is a schematic diagram of the structure of the dengue fever detection kit of the present invention;

[0034] Figure 2 This is the color development diagram obtained when the raw material dosage in the dengue fever detection kit in Comparative Example 1 is 4 μg;

[0035] Figure 3 This is the color development diagram obtained when the raw material dosage in the dengue fever detection kit in Comparative Example 1 is 6 μg;

[0036] Figure 4 This is the color development diagram obtained when the raw material dosage in the test kit for detecting fever in Example 1 is 5 μg;

[0037] Figure 5 This is a sample sensitivity test diagram of the fever detection kit in Example 1;

[0038] Figure 6 This is a sample sensitivity test diagram after the NC membrane of the leukemia detection kit in Comparative Example 2 is replaced;

[0039] Figure 7 This is a sample sensitivity test diagram after the NC membrane of the leukemia detection kit in Comparative Example 2 is replaced;

[0040] Figure 8 This is a sample sensitivity test diagram of the test kit for detecting fever in Example 1 when different sample buffers are replaced;

[0041] Figure 1 Middle: 1. PVC base plate; 2. Sample pad; 3. Colloidal gold pad; 4. Nitrocellulose membrane; 5. Absorbent pad; 6. IgG test line (G line); 7. Control line (C line); 8. IgM test line (M line). DETAILED DESCRIPTION

[0042] The following will be combined with the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0043] Example 1

[0044] This embodiment provides a dengue fever detection kit, including a sample pad, a colloidal gold pad, a nitrocellulose membrane (NC membrane), a test paper strip, a PVC bottom plate, and an absorbent pad. The nitrocellulose membrane is coated with an IgG test line (G line), an IgM test line (M line) and a control line (C line); (the specific structural schematic diagram is shown in Figure 1 (shown)

[0045] Wherein, the preparation of the colloidal gold pad is as follows:

[0046] S1. preparing colloidal gold complex of dengue fever (I+III) type labeled antigen; preparing colloidal gold complex of dengue fever (II+IV) type labeled antigen; preparing colloidal gold complex of chicken IgY;

[0047] S2. Evenly mix the dengue fever (I+III) type labeled antigen-colloidal gold complex, dengue fever (II+IV) type labeled antigen-colloidal gold complex, and chicken IgY colloidal gold complex in a volume ratio of 1:1:1, then evenly spray them on a glass cellulose membrane KB50, and dry them at 45°C for 24 hours.

[0048] Among them, the preparation of dengue fever (Ⅰ+Ⅲ) type labeled antigen colloidal gold complex:

[0049] A colloidal gold solution with a particle size of 530-533 nm and an OD value of 4.0-5.0A was prepared by a citric acid reduction method; 12.5 μL of 0.2M K2CO3 was added to 1 mL of the colloidal gold solution and mixed, 5 μg of dengue fever (Ⅰ+Ⅲ) type labeled antigen was added and mixed, and the mixture was allowed to stand or stir for 60 minutes at 25°C, BSA was added until the final concentration of BSA was 1%, and the mixture was blocked and allowed to stand or stir for 60 minutes at 25°C, and then the mixture was centrifuged at 10,000 rpm for 45 minutes at 25°C, the supernatant was removed and the precipitate was obtained, and the gold labeling solution was added for re-suspending to obtain a dengue fever (Ⅰ+Ⅲ) type labeled antigen-colloidal gold complex.

[0050] Among them, the preparation of dengue fever (Ⅱ+Ⅳ) type labeled antigen colloidal gold complex:

[0051] A colloidal gold solution with a particle size of 530-533 nm and an OD value of 4.0-5.0A was prepared by a citric acid reduction method; 12.5 μL of 0.2M K2CO3 was added to 1 mL of the colloidal gold solution, mixed to obtain a mixed solution, 5 μg of dengue fever (II+IV) type labeled antigen was added, mixed, and reacted at 18-25°C for 60 minutes with standing or stirring, BSA was added until the final concentration of BSA was 1%, and blocked, and reacted at 25°C for 60 minutes with standing or stirring, and then centrifuged at 25°C for 45 minutes at 10,000 rpm, the supernatant was removed by aspiration to obtain a precipitate, and a gold labeling solution was added for re-suspending to obtain a dengue fever (II+IV) type labeled antigen-colloidal gold complex.

[0052] Among them, the preparation of chicken IgY colloidal gold complex:

[0053] A colloidal gold solution with a particle size of 530-535 nm and an OD value of 4.0-5.0A is prepared by a citric acid reduction method; 15-25 μ 0.2 M K2CO3 is added to 1 mL of the colloidal gold solution, mixed, 1.2 μ g of chicken IgY antibody is added, mixed, reacted at 25° C. for 60 minutes by standing or stirring, BSA is added until the final concentration of BSA is 1%, reacted at 25° C. for 60 minutes by standing or stirring, then centrifuged at 25° C. for 45 minutes at 10,000 rpm, the supernatant is removed by suction to obtain a precipitate, a gold labeling solution is added for resuspending, and a chicken IgY colloidal gold complex is obtained.

[0054] Wherein, the preparation method of the sample pad is:

[0055] Pour the sample pad treatment solution into a container, slowly place the glass cellulose membrane 8965 into the solution, tap it continuously to remove the bubbles, and rotate it to drain until no obvious water drops fall. Place it on a drying rack, keep it flat, and dry it at 45°C until it is completely dry.

[0056] The sample pad treatment solution used is: 50mM Tris + 0.2% S9 + 0.6% PVPK-30 + 0.8% NaCl + 0.3% C-Na + 0.2% EDTA-Na2 + 0.03mg / mL blocker-H + 0.2mg / mL RBC antibody, pH = 8.6. (Tris is trishydroxymethylaminomethane; S9 is a surfactant, which belongs to propylene oxide-ethylene oxide-ethylene diamine copolymer and has good water solubility; PVPK-30 is polyvinyl pyrrolidone; C-Na is Casein-Na, i.e. sodium caseinate).

[0057] Wherein, the coating method of the detection line is:

[0058] Dengue mouse anti-human IgG antibody and dengue mouse anti-human IgM antibody were diluted to 1.0 mg / mL with coating buffer, coated on the NC membrane, and dried at 45°C for 48 h;

[0059] The control line is coated by:

[0060] Sheep anti-chicken IgY monoclonal antibody was diluted to 1.0 mg / mL with coating buffer and coated on the NC membrane, and then dried at 45°C for 48 h;

[0061] The coating buffer used above is: 20mM Tris, 100mM NaCl, and trehalose accounting for 3% of the total mass of the coating buffer.

[0062] Wherein, the assembly method of the test strip is:

[0063] After the humidity of the environment drops below 40%, take out the required materials and assemble them as required, tear off the plastic sticker at the absorbent paper position, and stick the absorbent paper of the specified size with the upper end aligned with the plastic sticker; tear off the plastic sticker at the sample pad and gold label pad positions, first stick the gold label pad with the upper end covering 1-2mm on the NC film, then stick the lower end of the sample pad aligned with the plastic sticker, and the upper end covers the gold label pad by 1-2mm.

[0064] The gold label complex solution is: Tris+PVPK-30+TX-100+C-Na+trehalose+BSA; Tris is trishydroxymethylaminomethane; PVPK-30 is polyvinylpyrrolidone; C-Na is sodium caseinate; TX-100 is Triton.

[0065] Comparative Example 1

[0066] Compared with Example 1 ( Figure 4 ) The only difference is: adjust the amount of raw materials used in the G line marking process ( Figure 2 The dosage of the raw material is 4 μg. Figure 3 The dosage of raw materials was 6 μg) to verify at which dosage a higher sensitivity could be obtained.

[0067] Comparative Example 2

[0068] Compared with Example 1 ( Figure 5 )The only difference is: replace the different NC membrane (nitrocellulose membrane) ( Figures 6-7 ) to verify the effect of different NC membranes on the kit.

[0069] Comparative Example 3

[0070] The only difference from Example 1 is that different sample buffers are replaced to verify the influence of sample buffers with different formulations on the kit.

[0071] 1. The sensitivity of the dengue fever detection kits obtained in Example 1 and Comparative Example 1 was tested:

[0072] The dengue fever humanized recombinant antibody was subjected to 10-fold and 10-fold -2 times, 10 -3 times, 10 -4 times, 10 -5 times dilution, and then tested. The test results are as follows Figures 2 to 4 shown.

[0073] Depend on Figures 2 to 4 It can be seen that the dengue fever detection kit in Example 1 is diluted to a concentration of 10 -2 The color is clearer when 10 -3 It can also be observed that in Comparative Example 1, after adjusting the amount of raw materials, whether it is reducing the amount of raw materials ( Figure 2), or increase the amount of raw materials ( Figure 3 ), after detecting 10 -2 When diluted 100 times, the color development did not reach that of Example 1 ( Figure 4 ) to a certain extent.

[0074] It can be seen that the dengue fever detection kit provided in the present invention selects the raw material dosage in Example 1, so that the kit has excellent detection sensitivity.

[0075] 2. The dengue fever detection kits obtained in Example 1 and Comparative Example 2 were tested in terms of performance:

[0076] The dengue fever humanized recombinant antibody was subjected to 10-fold and 10-fold -2 times, 10 -3 times, 10 -4 times, 10 -5 times dilution, and then tested. The test results are as follows Figures 2 to 4 shown.

[0077] Depend on Figures 5 to 7 It can be seen that the NC membrane selected in Example 1 ( Figure 5 ), compared with Comparative Example 2 ( Figures 6-7 ), no false positives, high sensitivity, and non-diffuse lines.

[0078] It can be seen that the dengue fever detection kit provided in the present invention selects the NC membrane in Example 1, which can better improve the performance of the kit and make the kit have excellent performance.

[0079] 3. Testing the dengue fever detection kit obtained in Example 1 for false positives:

[0080] The dengue fever detection kit obtained in Example 1 was used to screen the sample buffer to screen out the most suitable sample buffer. The test results are as follows: Figure 8 shown.

[0081] Depend on Figure 8 It can be seen that PB buffer cannot block blood, PBS buffer and isotonic PBS buffer have false positives, and normal saline can block blood without obvious false positives, and the background color is beautiful.

[0082] It can be seen that the use of physiological saline as the sample buffer in this kit can better improve the performance of the kit and make the kit have excellent performance.

[0083] It should be noted that, in this article, relational terms such as first and second, etc. are only used to distinguish one entity or operation from another entity or operation, and do not necessarily require or imply any such actual relationship or order between these entities or operations. Moreover, the terms "include", "comprise" or any other variants thereof are intended to cover non-exclusive inclusion, so that a process, method, article or device including a series of elements includes not only those elements, but also other elements not explicitly listed, or also includes elements inherent to such process, method, article or device.

[0084] Although embodiments of the present invention have been shown and described, it will be appreciated by those skilled in the art that various changes, modifications, substitutions and variations may be made to the embodiments without departing from the principles and spirit of the present invention, and that the scope of the present invention is defined by the appended claims and their equivalents.

Claims

1. A dengue fever detection kit, characterized in that, It includes a sample pad, a colloidal gold pad, a nitrocellulose membrane, a test paper strip, a PVC bottom plate, and a water-absorbing pad, wherein the nitrocellulose membrane is coated with an IgG test line, an IgM test line, and a control line; Wherein, the preparation of the colloidal gold pad is as follows: S1. preparing colloidal gold complex of dengue fever (I+III) type labeled antigen; preparing colloidal gold complex of dengue fever (II+IV) type labeled antigen; preparing colloidal gold complex of chicken IgY; S2. Evenly mix the dengue fever (I+III) type labeled antigen-colloidal gold complex, dengue fever (II+IV) type labeled antigen-colloidal gold complex, and chicken IgY colloidal gold complex in a volume ratio of 1:1:1, then evenly spray them on the glass cellulose membrane KB50, and dry them at 37-45°C for 4-24h to obtain.

2. A dengue fever detection kit according to claim 1, characterized in that: Preparation of dengue fever (Ⅰ+Ⅲ) type labeled antigen colloidal gold complex: A colloidal gold solution with a particle size of 530-533 nm and an OD value of 4.0-5.0A is prepared by a citric acid reduction method; 12.5 μL of 0.2M K2CO3 is added to 1 mL of the colloidal gold solution, mixed, 5 μg of dengue fever (Ⅰ+Ⅲ) type labeled antigen is added, mixed, and reacted at 18-25°C for 30-60 minutes by standing or stirring, BSA is added until the final concentration of BSA is 1%, and blocked, and reacted at 18-25°C for 30-60 minutes by standing or stirring, and then centrifuged at 4-25°C at 10000 rpm for 15-45 minutes, the supernatant is removed by suction to obtain a precipitate, and a gold labeling solution is added for re-suspending to obtain a dengue fever (Ⅰ+Ⅲ) type labeled antigen-colloidal gold complex.

3. A dengue fever detection kit according to claim 1, characterized in that: Preparation of dengue fever (Ⅱ+Ⅳ) type labeled antigen colloidal gold complex: A colloidal gold solution with a particle size of 530-533 nm and an OD value of 4.0-5.0A is prepared by a citric acid reduction method; 12.5 μL of 0.2M K2CO3 is added to 1 mL of the colloidal gold solution, mixed to obtain a mixed solution, 5 μg of dengue fever (II+IV) type labeled antigen is added, mixed, and reacted at 18-25°C for 30-60 minutes by standing or stirring, BSA is added until the final concentration of BSA is 1%, and blocked, and reacted at 18-25°C for 30-60 minutes by standing or stirring, and then centrifuged at 4-25°C at 10000 rpm for 15-45 minutes, the supernatant is removed by suction to obtain a precipitate, and a gold labeling solution is added for resuspending to obtain a dengue fever (II+IV) type labeled antigen-colloidal gold complex.

4. A dengue fever detection kit according to claim 1, characterized in that: Preparation of Chicken IgY Colloidal Gold Complex: A colloidal gold solution with a particle size of 530-535 nm and an OD value of 4.0-5.0A is prepared by a citric acid reduction method; 15-25 μl of 0.2M K2CO3 is added to 1 mL of the colloidal gold solution, mixed, 0.7-1.2 μg of chicken IgY antibody is added, mixed, reacted at 18-25°C for 30-60 minutes by standing or stirring, BSA is added until the final concentration of BSA is 1%, reacted at 18-25°C for 30-60 minutes by standing or stirring, then centrifuged at 4-25°C for 15-45 minutes at 10000 rpm, the supernatant is removed by suction to obtain a precipitate, a gold labeling solution is added for re-suspending, and a chicken IgY colloidal gold complex is obtained.

5. A dengue fever detection kit according to claim 1, characterized in that: The preparation method of the sample pad is as follows: Pour the sample pad treatment solution into a container, slowly place the glass cellulose membrane 8965 into the solution, and keep tapping to drive out the bubbles. Rotate and drain until no obvious water drops fall. Place it on a drying rack, keep it flat, and dry it at 37-45℃ until it is completely dry.

6. A dengue fever detection kit according to claim 5, characterized in that: The sample pad treatment solution used was: 50 mM Tris + 0.2% S9 + 0.6% PVPK-30 + 0.8% NaCl + 0.3% C-Na + 0.2% EDTA-Na2 + 0.03 mg / mL Blocker-H + 0.2 mg / mL RBC antibody, pH = 8.

6.

7. A dengue fever detection kit according to claim 1, characterized in that: The coating method of the test line is: Dengue mouse anti-human IgG antibody and dengue mouse anti-human IgM antibody were diluted to 1.0 mg / mL with coating buffer, coated on the NC membrane, and dried at 45°C for 24 to 48 hours; The control line is coated by: Dilute goat anti-chicken IgY monoclonal antibody to 1.0 mg / mL with coating buffer and coat it on the NC membrane, then dry it at 45°C to 55°C for 24 to 72 hours.

8. A dengue fever detection kit according to claim 7, characterized in that: The coating buffer used was: 20 mM Tris, 100 mM NaCl, and trehalose accounting for 3% of the total mass of the coating buffer.

9. A dengue fever detection kit according to claim 1, characterized in that: The assembly method of the test strip is as follows: After the humidity of the environment drops below 40%, take out the required materials and assemble them as required, tear off the plastic sticker at the absorbent paper position, and stick the absorbent paper of the specified size with the upper end aligned with the plastic sticker; tear off the plastic sticker at the sample pad and gold label pad positions, first stick the gold label pad with the upper end covering 1-2mm on the NC film, then stick the lower end of the sample pad aligned with the plastic sticker, and the upper end covers the gold label pad by 1-2mm.

10. A dengue fever detection kit according to any one of claims 2 to 4, characterized in that: The gold label complex solution is: Tris+pvpk-30+TX-100+C-Na+trehalose+BSA.

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