A dual-mode immunochromatographic test strip and its application

Abstract

The present invention discloses a dual-modal immunochromatographic test strip and its application, which relates to the technical field of medical testing and food safety detection. The dual-modal immunochromatographic test strip uses AuNFs@TA / Zr as a probe, and the preparation method of AuNFs@TA / Zr includes: preheating deionized water, adding a chloroauric acid aqueous solution to obtain a chloroauric acid solution; dissolving tannic acid and a hydroxylamine hydrochloride aqueous solution in deionized water to obtain a mixed solution; reacting the chloroauric acid solution with the mixed solution to obtain an AuNFs@TA solution; adding zirconium chloride to the AuNFs@TA solution to obtain AuNFs@TA / Zr; the present invention also provides the application of the dual-modal immunochromatographic test strip in the detection of pesticides or pathogenic bacteria. The beneficial effect of the present invention is that AuNFs@TA / Zr is used as a beacon carrier to make an immunochromatographic test strip with colorimetric and photothermal dual-modal signals, which has the characteristics of high stability and high detection sensitivity.

Description

Technical Field

[0001] The present invention relates to the technical field of medical testing and food safety detection, and in particular to a dual-modal immunochromatographic test strip and applications thereof. Background Art

[0002] Immunochromatography is a rapid detection method based on the principle of antigen-antibody specific binding. With its advantages of rapid detection speed, strong specificity, ease of operation, and low cost, it has become the most mature on-site rapid detection technology and is widely used in clinical diagnosis, food safety testing, medical testing, and environmental pollutant monitoring. Among them, colloidal gold immunochromatographic test strips, as a classic application, offer the convenience of instant testing, but their low sensitivity and susceptibility to interfering substances in complex sample matrices limit their application in trace analysis.

[0003] In response to the above limitations, in recent years, immunochromatography technology has enriched the target through immunomagnetic separation to reduce matrix interference, and combined with signal amplification strategies such as biotin-streptavidin system and nanozyme to improve the sensitivity to the pg / mL level. At the same time, new labeling materials such as quantum dots and time-resolved fluorescent microspheres have been developed, and fluorescence enhancement or magnetic signal quantification modes have been used to break through the limitations of traditional colloidal gold, achieve synergistic optimization of sensitivity and anti-interference, and promote its application upgrade in trace biomarker detection and multi-target analysis.

[0004] For immunochromatographic detection, nanoprobe stability and biocompatibility are two key parameters. Stability determines the reliability of the detection method and the shelf life of the test product; biocompatibility affects the coupling of the nanoprobe with the recognition molecule, thus affecting the method's universality. Currently, the stability and biocompatibility of nanoprobes used in existing immunochromatographic techniques are mediocre, resulting in limited universality and low sensitivity for the detection of pesticides and pathogens. Summary of the Invention

[0005] The purpose of the present invention is to solve at least one of the technical problems existing in the prior art and provide a dual-modal immunochromatographic test strip and its application. Specifically, a dual-modal immunochromatographic test strip with metal polyphenol-coated flower-like gold nanoparticles (AuNFs@TA / Zr) and its application are designed. The present invention synthesizes a metal polyphenol-coated flower-like gold nanoparticle (AuNFs@TA / Zr) as a probe, which is applied to fields such as medical testing and rapid food safety detection, and improves the stability and detection sensitivity of the test strip.

[0006] On one hand, the present invention synthesized flower-like gold nanoparticles (AuNFs). Due to their unique, dendritic, hierarchical structure, AuNFs exhibit significant performance advantages over traditional colloidal gold, including a larger surface area (5-8 times that of spherical particles), enhanced localized surface plasmon resonance (LSPR), and peroxidase-like catalytic activity (3-5 times higher efficiency), significantly improving the signal transduction capability of sensor probes. On the other hand, the present invention also employed metal (Zr) and polyphenol (tannic acid, abbreviated as TA) coating of AuNFs to create dual-modal (colorimetric and photothermal) immunochromatographic test strips. The resulting AuNFs@TA / Zr exhibits excellent dispersibility, biocompatibility, colorimetric performance, and superior photothermal performance, making it suitable for use as a novel colorimetric / photothermal signal label in dual-modal immunochromatographic test strips. Compared with colloidal gold test strips, dual-modal immunochromatographic test strips prepared with metal polyphenol-coated flower-like gold nanoparticles (AuNFs@TA / Zr) as beacon carriers have the advantages of high sensitivity and short detection time.

[0007] The technical solutions of the present invention are as follows:

[0008] The first aspect of the present invention provides a dual-modal immunochromatographic test strip, wherein the dual-modal immunochromatographic test strip uses AuNFs@TA / Zr as a probe;

[0009] The preparation method of AuNFs@TA / Zr comprises the following steps:

[0010] S1. Preheat deionized water, add chloroauric acid aqueous solution and mix well to obtain chloroauric acid solution;

[0011] S2, dissolving tannic acid and hydroxylamine hydrochloride aqueous solution in deionized water, and mixing thoroughly to obtain a mixed solution;

[0012] S3, reacting the chloroauric acid solution with the mixed solution, cooling and centrifuging to obtain a precipitate after the reaction is completed; washing the precipitate and redissolving it in water to obtain an AuNFs@TA solution;

[0013] S4. Add zirconium chloride to the AuNFs@TA solution, stir and mix, centrifuge, and wash to obtain AuNFs@TA / Zr.

[0014] Preferably, in step S1, the temperature of the preheated deionized water is 65° C. to 75° C., the mass concentration of the chloroauric acid aqueous solution is 0.5% to 1.5%, and the added volume ratio of deionized water to chloroauric acid solution is 95 to 103:1.

[0015] Preferably, in step S2, the addition ratio of tannic acid and hydroxylamine hydrochloride aqueous solution is 18 mg-22 mg:0.5 mL, and the concentration of hydroxylamine hydrochloride aqueous solution is 0.05 mol / L-0.15 mol / L.

[0016] Preferably, in step S3, the cooling condition is room temperature, the cooling time is 35 min to 45 min, the centrifugal speed is 3000 r / min to 12000 r / min, the centrifugal time is 10 min to 30 min, the number of washing times is 1 to 3 times, and the volume of water added for re-dissolution is 7 mL to 9 mL.

[0017] Preferably, in step S4, the addition ratio of zirconium chloride and chloroauric acid solution is 4 mg~6 mg:1 mL, the stirring speed is 1000 r / min~1400 r / min, the stirring time is 20 h~28 h, the centrifugal speed is 3000 r / min~12000 r / min, the centrifugal time is 10 min~30 min, and the number of washing times is 1 time~3 times.

[0018] Preferably, the AuNFs@TA / Zr is loaded on the dual-modal immunochromatographic test strip in the form of an AuNFs@TA / Zr labeled antibody complex, wherein the preparation method of the AuNFs@TA / Zr labeled antibody complex comprises the following steps:

[0019] AuNFs@TA / Zr was dissolved in water to obtain AuNFs@TA / Zr solution.

[0020] The antibody to be labeled is added to the AuNFs@TA / Zr solution, and after thorough mixing, a blocking agent is added. After the reaction, the solution is centrifuged to obtain a precipitate, and the obtained precipitate is redissolved to prepare an AuNFs@TA / Zr labeled antibody complex.

[0021] Preferably, in the method for preparing the AuNFs@TA / Zr labeled antibody complex,

[0022] The final concentration of the antibody after adding the antibody to be labeled is 1 μg / mL to 100 μg / mL;

[0023] After the blocking agent is added, the final mass concentration of the blocking agent is 0.1% to 1%, and the blocking agent is selected from any one of casein, bovine serum albumin, ovalbumin, polyethylene glycol, and skim milk;

[0024] The centrifugal speed is 5000 r / min to 20000 r / min, and the centrifugal time is 10 min to 50 min;

[0025] The precipitate after centrifugation was re-dissolved with 0.01M-0.1M phosphate buffer, pH 6.0-9.0;

[0026] The antibody to be labeled includes any one of monoclonal antibodies, polyclonal antibodies, nanobodies, and phage-expressed antibodies.

[0027] Preferably, the dual-modal immunochromatographic test strip comprises:

[0028] base plate;

[0029] A nitrocellulose membrane is disposed on the bottom plate, and has detection lines and quality control lines spaced apart from each other;

[0030] A glass fiber mat is arranged on one end of the nitrocellulose membrane having the detection line, one end of the glass fiber mat is arranged on the bottom plate, and the other end is arranged on the nitrocellulose membrane, and the AuNFs@TA / Zr labeled antibody complex is arranged on the glass fiber mat;

[0031] absorbent paper, arranged at one end of the nitrocellulose membrane having the quality control line, one end of the absorbent paper being arranged on the bottom plate, and the other end being arranged on the nitrocellulose membrane;

[0032] A sample pad has one end arranged on the bottom plate and the other end arranged on the glass fiber pad.

[0033] Preferably, the method of placing the AuNFs@TA / Zr labeled antibody complex on the glass fiber mat comprises: spraying the AuNFs@TA / Zr labeled antibody complex onto the glass fiber mat at a volume of 7 μL / cm~9 μL / cm, and vacuum drying for 1.5 h~2.5 h.

[0034] Preferably, the test line is coated with an artificially coupled antigen or antibody to the analyte, and the quality control line is coated with a goat anti-mouse antibody.

[0035] The method for forming a detection line and a quality control line on the nitrocellulose membrane comprises the following steps:

[0036] Using 0.01 M to 0.5 M PBS solution (pH 6.0 to 8.0), adjust the concentration of the coated analyte artificially conjugated antigen or analyte antibody and goat anti-mouse antibody to 0.01 mg / mL to 10.0 mg / mL respectively; wherein the analyte antibody includes a monoclonal antibody, a polyclonal antibody, a nanobody, or a phage-expressed antibody;

[0037] The artificially conjugated antigen or antibody to be tested is sprayed on the upper part of the nitrocellulose membrane as a test line, and the goat anti-mouse antibody is sprayed on the lower part of the nitrocellulose membrane as a quality control line; wherein the spraying volume of the artificially conjugated antigen or antibody to be tested and the goat anti-mouse antibody is 0.25 μL / cm2~0.74 μL / cm2;

[0038] The nitrocellulose membrane sprayed with the test line and quality control line is dried.

[0039] Preferably, the assembly of the test strip comprises the following steps:

[0040] (1) The following materials are overlapped and pasted on the bottom plate: filter paper, sample pad, glass fiber pad sprayed with AuNFs@TA / Zr labeled antibody complex, nitrocellulose membrane sprayed with analyte-coupled artificial antigen or antibody as detection line and anti-mouse antibody / anti-rabbit antibody as quality control line, and absorbent paper, so as to assemble the dual-modal immunochromatographic test strip prepared using AuNFs@TA / Zr as beacon carrier of the present invention;

[0041] (2) The assembled test strip plate is cut into the required width by a cutter, which is the dual-modal immunochromatographic test strip prepared by the present invention using AuNFs@TA / Zr as a beacon carrier. The test strip can be used directly or can be placed in a plastic card shell for use.

[0042] A second aspect of the present invention provides a use of the aforementioned dual-modal immunochromatographic test strip in detecting pesticides or pathogens for non-disease diagnosis purposes. The method for detecting pesticides or pathogens using the dual-modal immunochromatographic test strip comprises the following steps:

[0043] The processed sample to be tested is added to the dual-modal immunochromatographic test strip with a sample volume of 50 μL to 200 μL and a reaction time of 3 minutes to 30 minutes. Quantitative detection is achieved by reading the grayscale data of the test strip and the imaging data of the infrared imager to calculate the concentration of the sample to be tested, or qualitative judgment of the sample to be tested is achieved by visually observing the presence or absence of colored strips on the test line and quality control line on the dual-modal immunochromatographic test strip.

[0044] The present invention has at least one of the following beneficial effects:

[0045] The present invention synthesizes flower-shaped gold nanoparticles (AuNFs), and uses metals and polyphenols to coat the flower-shaped gold nanoparticles to form AuNFs@TA / Zr. AuNFs@TA / Zr is then used as a beacon carrier to prepare a dual-modal immunochromatographic test strip for the detection of pathogens or pesticides. Compared with traditional colloidal gold test strips, the dual-modal immunochromatographic test strip prepared by the present invention has colorimetric and photothermal dual-modal signals, and the signals are strong. The prepared dual-modal immunochromatographic test strip has high detection sensitivity and can realize rapid quantitative or qualitative detection of various analytes. BRIEF DESCRIPTION OF THE DRAWINGS

[0046] Figure 1 This is a schematic diagram of the structure of a dual-modal immunochromatographic test strip, where 1 is the sample pad, 2 is the glass fiber pad, 3 is the nitrocellulose membrane, 4 is the test line, 5 is the quality control line, 6 is the base plate, and 7 is absorbent paper.

[0047] Figure 2 Figure 1 is a schematic diagram of the detection principle of the double-antibody sandwich method AuNFs@TA / Zr immunochromatographic test strip, where 8 is the analyte antibody on the test line, 9 is the anti-mouse antibody or anti-rabbit antibody on the quality control line, 10 is the macromolecular analyte, and 11 is the AuNFs@TA / Zr-antibody complex.

[0048] Figure 3 Figure 1 is a schematic diagram of the competitive AuNFs@TA / Zr immunochromatographic test strip detection principle, where 9 is the anti-mouse antibody or anti-rabbit antibody on the quality control line, 10 is the small molecule analyte, 11 is the AuNFs@TA / Zr-antibody complex, and 12 is the artificially coupled antigen of the analyte on the detection line.

[0049] Figure 4 Shown are transmission electron microscopy images of AuNFs@TA / Zr, where a in the figure is a transmission electron microscopy image with a scale of 0.2 μm, b in the figure is a transmission electron microscopy image with a scale of 100 nm, c in the figure is a transmission electron microscopy image with a scale of 50 nm, and d in the figure is a transmission electron microscopy image with a scale of 10 nm.

[0050] Figure 5 The figure shows the dual-modal immunochromatographic test strip with AuNFs@TA / Zr as the beacon carrier for the detection of Escherichia coli. E. coli O157:H7 Colorimetric diagram.

[0051] Figure 6 Shown is a colorimetric image of a traditional colloidal gold test strip detecting E. coli O157:H7.

[0052] Figure 7Shown is a photothermal image of a dual-modal immunochromatographic test strip with AuNFs@TA / Zr as a beacon carrier for detecting Escherichia coli E.coli O157:H7.

[0053] Figure 8 Shown is a colorimetric image of the dual-modal immunochromatographic test strip with AuNFs@TA / Zr as the beacon carrier for the detection of clothianidin.

[0054] Figure 9 Shown is a photothermal image of a dual-modal immunochromatographic test strip with AuNFs@TA / Zr as the beacon carrier for the detection of clothianidin.

[0055] Figure 10 Shown is a transmission electron micrograph of non-flower-shaped gold nanoparticles coated with metal polyphenols.

[0056] Figure 11 The figure shows the immunochromatographic test strip for detecting Escherichia coli using metal polyphenol-coated non-flower-shaped gold nanoparticles as beacon carriers. E. coli O157:H7 Physical picture.

[0057] Figure 12 Shown is the transmission electron microscopy image of AuNFs@polydopamine-Zr gold nanoparticles.

[0058] Figure 13 The image shows the immunochromatographic test strip using AuNFs@polydopamine-Zr gold nanoparticles as beacon carriers to detect Escherichia coli. E. coli O157:H7 Physical picture.

[0059] Figure 14 Shown is the transmission electron microscopy image of AuNPs@proanthocyanidin-Zr gold nanoparticles.

[0060] Figure 15 The figure shows the immunochromatographic test strip with AuNPs@proanthocyanidin-Zr gold nanoparticles as beacon carriers for detecting Escherichia coli. E. coli O157:H7 Physical picture.

[0061] Figure 16 Shown is the transmission electron microscopy image of AuNPs@epicatechin-Zr gold nanoparticles.

[0062] Figure 17 The figure shows the immunochromatographic test strip with AuNPs@epicatechin-Zr gold nanoparticles as beacon carriers for detecting Escherichia coli. E. coli O157:H7 Physical picture.

[0063] Figure 18 Shown is the transmission electron microscopy image of AuNFs@TA flower-like gold nanoparticles.

[0064] Figure 19The image shows the immunochromatographic test strip using AuNFs@TA flower-shaped gold nanoparticles as beacon carriers to detect Escherichia coli. E. coli O157:H7 Physical picture. DETAILED DESCRIPTION

[0065] In order to make the technical problems, technical solutions and beneficial effects solved by the present invention more clearly understood, the present invention is further described in detail below with reference to the accompanying drawings and embodiments. It should be understood that the specific embodiments described herein are only used to explain the present invention and are not intended to limit the present invention.

[0066] Example 1:

[0067] This embodiment provides a method for preparing AuNFs@TA / Zr, comprising the following steps:

[0068] (1) Preparation of chloroauric acid solution

[0069] 99 mL of deionized water was preheated to 70 °C in a microwave reactor. 1 mL of a 1% chloroauric acid (HAuCl4) aqueous solution was then quickly added to the preheated deionized water under continuous stirring and mixed evenly to obtain a chloroauric acid solution.

[0070] (2) Preparation of tannic acid-hydroxylamine hydrochloride-chloroauric acid mixed solution

[0071] Dissolve 20 mg of tannic acid (relative molecular mass 1701.2 g / mol) and 0.5 mL of 0.1 mol / L hydroxylamine hydrochloride aqueous solution in deionized water and mix thoroughly to form a homogeneous solution.

[0072] The tannic acid-hydroxylamine hydrochloride mixed solution was quickly added to the chloroauric acid solution obtained in step (1) and stirred continuously to allow the components to react fully. The mixed system was further heated at 70°C for 1 min to obtain a homogeneous mixed solution of tannic acid, hydroxylamine hydrochloride, and chloroauric acid.

[0073] (3) Collection of AuNFs@TA solution

[0074] The homogenized mixed solution was cooled at room temperature for 40 minutes and then centrifuged at 12,000 rpm for 10 minutes to collect the precipitate. The precipitate was washed three times with deionized water and finally redissolved in 8 mL of pure water to prepare the AuNFs@TA solution.

[0075] (4) Preparation of AuNFs@TA / Zr solution

[0076] Add 5 mg of zirconium chloride (ZrCl4) to the AuNFs@TA solution obtained in step 3 and stir vigorously (1200 r / min) at room temperature overnight; then centrifuge to separate the precipitate, wash it, and redissolve it in 1 mL of deionized water to obtain the AuNFs@TA / Zr solution, which was stored at 4°C.

[0077] The AuNFs@TA / Zr prepared in Example 1 was characterized. Figure 4 Shown is the transmission electron microscopy (TEM) image of AuNFs@TA / Zr. Figure 4 It can be seen that the AuNFs@TA / Zr prepared in Example 1 includes an inner layer of gold nanoparticles (AuNFs) and a metal polyphenol network structure formed by tannic acid (TA) and zirconium (Zr) loaded on the outer surface of the AuNFs. Among them, the AuNFs have a flower-like structure and have the characteristics of a large surface area; TA and Zr are evenly distributed on the surface of the AuNFs, which is conducive to improving biocompatibility.

[0078] The AuNFs@TA prepared in step (3) of Example 1 was characterized. Figure 18 Shown is the transmission electron microscopy image (TEM image) of AuNFs@TA. Figure 4 and Figure 18 It can be seen from the comparison that the AuNFs@TA / Zr prepared in Example 1 has a dense cross-linked structure compared to AuNFs@TA, indicating that the Zr 4+ The metal-polyphenol coordination with TA formed a stable three-dimensional cross-linked network, resulting in dense and orderly arrangement of the nanoparticles. Furthermore, the AuNFs in Example 1 exhibited a more uniform flower-like morphology, and the TEM images of Example 1 showed a more complete flower-like structure, indicating that the introduction of Zr enhanced the structural stability of the AuNFs.

[0079] Therefore, the introduction of Zr optimizes the morphology, chemical state, and optical properties of AuNFs@TA / Zr through metal-polyphenol coordination, which is manifested in the following ways: (1) Improved structural stability: The dense cross-linked network enhances mechanical strength and probe uniformity; (2) Functional site retention: Reduced shielding of Au-S bonds by TA improves the efficiency of antibody-directed coupling; (3) Enhanced signal amplification: High-sensitivity detection is achieved through the LSPR effect and enhanced T-line aggregation. However, AuNFs@TA lacks Zr cross-linking and has significant defects in structure, function, and signal intensity, resulting in decreased test strip sensitivity.

[0080] Example 2:

[0081] This embodiment provides a method for detecting Escherichia coli using AuNFs@TA / Zr as a beacon carrier. E. coli O157:H7 Preparation and detection method of sandwich immunochromatographic test strips, such as Figure 1As shown, the immunochromatographic test strip includes a base plate 6, and filter paper, a sample pad 1, a glass fiber pad 2, a nitrocellulose membrane 3 and an absorbent paper 7 sequentially overlapped and pasted on the base plate 6, and a detection line 4 and a quality control line 5 are provided on the nitrocellulose membrane 3.

[0082] 1. Preparation process of immunochromatographic test strips

[0083] 1. Preparation of nitrocellulose membrane;

[0084] Anti-E. coli E. coli O157:H7 Polyclonal and anti-mouse antibodies were coated onto nitrocellulose membranes: anti-E. coli was diluted in 0.01 M PBS (phosphate buffered saline) pH 7.5. E. coli O157:H7 The concentration of polyclonal antibody was 0.5 mg / mL, and the resulting solution was sprayed on the membrane as the test line; the concentration of diluted anti-mouse antibody was 0.5 mg / mL, and the resulting solution was sprayed on the membrane as the quality control line. The spray volume of both lines was 0.74 μL / cm. The test line was 10 mm away from the top edge of the membrane, and the distance between the two lines was 5 mm. The samples were dried at 30°C for 12 hours and stored in a drying cabinet for future use.

[0085] 2. Preparation of AuNFs@TA / Zr-antibody composite glass fiber mat:

[0086] 50 μL of the AuNFs@TA / Zr solution prepared in Example 1 was ultrasonicated for 3 minutes, mixed with 450 μL of 0.02 M borate buffer at pH 7.0, and then 10 μg of Escherichia coli was added. E. coli O157:H7 After thorough mixing, the monoclonal antibody was stirred for 2 h, and 50 μL of bovine serum albumin (0.5% final concentration) was added. The mixture was blocked at room temperature for 2 h and centrifuged at 8000 rpm for 10 min. The precipitate was reconstituted with 0.01 M phosphate buffered saline (PBS) (pH 7.4) to 1 / 10 of the initial volume and sprayed onto a glass fiber mat at a volume of 8 μL / cm. The mixture was vacuum dried for 2 h and stored in a drying cabinet for later use.

[0087] 3. Assemble the test strips:

[0088] (1) The filter paper and sample pad specifications are 1 cm × 30 cm;

[0089] ⑵ Glass fiber mat sprayed with AuNFs@TA / Zr-antibody complex, with a size of 0.8 cm × 30 cm;

[0090] (3) Nitrocellulose membrane sprayed with test lines and quality control lines, size 2.5 cm × 30 cm;

[0091] (4) Absorbent paper, size 1.2 cm × 30 cm;

[0092] ⑸ PVC baseboard, specifications are 5.5 cm × 30 cm.

[0093] Paste the above materials in sequence according to the positions of the components in the test strip structure diagram. After assembly, cut them into 4 mm × 55 mm test strips with a cutter, put them into a plastic card shell, press them tightly and put them into an aluminum foil bag. After adding desiccant, seal and store. The shelf life at room temperature is 12 months.

[0094] 2. Quantitative detection of E. coli in samples E. coli O157:H7

[0095] Use the above-mentioned immunochromatographic test strips to detect Escherichia coli in the sample E. coli O157:H7 The method comprises the following steps:

[0096] 1. Add 100 μL of sample to the sample well of the test strip and react for 15 minutes;

[0097] 2. Insert the test strip into the test window of the test strip reader. The intensity of the test line and the quality control line will be displayed on the display as a numerical value. According to the standard curve already recorded in the instrument, the E. coli content in the sample can be calculated. E. coli O157:H7 The content of Escherichia coli in the sample E. coli O157:H7 Quantitative detection of .

[0098] 3. Establish a standard curve: add the spike to the negative matrix and the E. coli in the standard curve E. coli O157:H7 Concentration: 0, 5×10 2 CFU / mL, 1×10 3 CFU / mL, 5×10 3 CFU / mL, 1×10 4 CFU / mL, 5×10 4 CFU / mL, 1×10 5 CFU / mL, 5×10 5 CFU / mL, 1×10 6 CFU / mL, 5×10 6 CFU / mL, the above samples were added to the sample pad respectively, and the reaction time was 15 minutes.

[0099] As a control, the above samples were added to traditional colloidal gold test strips, and the reaction time was 15 min.

[0100] Figure 2The figure shows the principle diagram of the double-antibody sandwich method AuNFs@TA / Zr immunochromatographic test strip detection, where 8 is the analyte antibody on the detection line, 9 is the anti-mouse antibody or anti-rabbit antibody on the quality control line, 10 is the macromolecular analyte, and 11 is the AuNFs@TA / Zr-antibody complex.

[0101] Figure 5 The immunochromatographic test strip prepared with AuNFs@TA / Zr as the beacon carrier is used to detect Escherichia coli. E. coli O157:H7 Schematic diagram of the actual results; the results show that as Escherichia coli E. coli O157:H7 As the colorimetric signal of the test strip detection line (T line) gradually increases, the lowest detection limit visible to the naked eye is 5×10 4 CFU / mL.

[0102] Figure 7 The immunochromatographic test strip prepared with AuNFs@TA / Zr as the beacon carrier was used to detect different concentrations (0 CFU / mL, 5×10 4 CFU / mL, 1×10 5 CFU / mL, 5×10 5 CFU / mL, 1×10 6 CFU / mL, 5×10 6 CFU / mL) Escherichia coli E. coli O157:H7 The results show that as Escherichia coli E. coli O157:H7 As the photothermal signal of the test strip detection line (T line) gradually increased, it showed that the immunochromatographic test strip prepared with AuNFs@TA / Zr as the beacon carrier was used to detect Escherichia coli. E. coli O157:H7 The colorimetric and photothermal effects are good.

[0103] In addition, as a control, Figure 6 Demonstrating the detection of different concentrations of E. coli using traditional colloidal gold test strips E. coli O157:H7 The results show that the lowest detection limit visible to the naked eye is 5×10 5 An obvious signal appeared when the CFU / mL was 0.1747 W / mL, which shows that compared with the traditional colloidal gold test strips, the detection sensitivity of the test strips based on the AuNFs@TA / Zr beacon carrier is 10 times higher than that of the traditional colloidal gold test strips, verifying the significant performance advantages of this new material in the immunochromatographic detection system.

[0104] Example 3:

[0105] This embodiment provides a preparation and detection method of a competitive immunochromatographic test strip for detecting clothianidin using AuNFs@TA / Zr as a beacon carrier, comprising the following steps:

[0106] 1. Preparation process of immunochromatographic test strips

[0107] The competitive immunochromatographic test strip for detecting clothianidin using the AuNFs@TA / Zr prepared in Example 1 as a beacon carrier comprises a PVC base plate, and a sample pad, a glass fiber pad, a nitrocellulose membrane, and absorbent paper that are overlapped and pasted on the base plate.

[0108] 1. Preparation of nitrocellulose membrane;

[0109] Clothianidin whole antigen and goat anti-mouse antibody were coated onto nitrocellulose membranes: the concentration of clothianidin whole antigen was diluted to 1 mg / mL in 0.01 M PBS (pH 7.4), and the resulting solution was sprayed on the membrane as a test line; the concentration of goat anti-mouse antibody was diluted to 0.5 mg / mL, and the resulting solution was sprayed on the membrane as a quality control line. The spray volume of both lines was 0.74 μL / cm. The test line was 10 mm away from the top edge of the membrane, and the distance between the two lines was 5 mm. The membranes were dried at 30°C for 12 h and stored in a drying cabinet until use.

[0110] 2. Preparation of AuNFs@TA / Zr-antibody composite glass fiber mat:

[0111] 50 μL of AuNFs@TA / Zr (prepared in Example 1) was sonicated for 5 minutes and mixed with 450 μL of 0.02 M borate buffer (pH 8.0) by oscillation. 2.5 μg of clothianidin monoclonal antibody was added and thoroughly mixed. The mixture was shaken for 3 hours, and 50 μL of bovine serum albumin (10%) was added. The mixture was blocked at room temperature for 1 hour and centrifuged at 8000 rpm for 20 minutes. The precipitate was reconstituted with 0.01 M phosphate buffered saline (PBS) (pH 7.4) to 1 / 10 of the initial volume. The mixture was sprayed onto a glass fiber mat at a volume of 8 μL / cm2, vacuum dried for 2 hours, and stored in a desiccator until ready for use.

[0112] 3. Assemble the test strips:

[0113] (1) The sample pad size is 1 cm × 30 cm;

[0114] ⑵ Glass fiber mat sprayed with AuNFs@TA / Zr-antibody complex, with a size of 0.8 cm × 30 cm;

[0115] (3) Nitrocellulose membrane sprayed with test lines and quality control lines, size 2.5 cm × 30 cm;

[0116] (4) Absorbent paper, size 1.2 cm × 30 cm;

[0117] ⑸ PVC baseboard, specifications are 5.5 cm × 30 cm.

[0118] Paste the above materials in sequence according to the positions of the components in the test strip structure diagram. After assembly, cut them into 4×55 mm test strips with a cutter, put them into a plastic card shell, press them tightly and put them into an aluminum foil bag. After adding desiccant, seal and store. The shelf life at room temperature is 12 months.

[0119] 2. Quantitative detection of clothianidin in samples

[0120] The method for detecting clothianidin in a sample using the above-mentioned immunochromatographic test strip comprises the following steps:

[0121] 1. Add 100 μL of sample to the sample well of the test strip and react for 15 minutes;

[0122] 2. Insert the test strip into the test strip reader's detection window. The intensity of the test and control lines will be indicated on the display as numerical values. Based on the standard curve already entered into the instrument, the clothianidin content in the sample can be calculated, enabling quantitative detection of positive samples.

[0123] 3. Prepare a standard curve for test strips: spike clothianidin into a negative matrix to concentrations of 0 ng / mL, 0.01 ng / mL, 0.05 ng / mL, 0.1 ng / mL, 0.5 ng / mL, 1 ng / mL, 5 ng / mL, 10 ng / mL, 50 ng / mL, 100 ng / mL, and 500 ng / mL, respectively. Apply each sample to the sample pad and allow the reaction to proceed for 15 minutes.

[0124] As a control, the above samples were added to traditional colloidal gold test strips, and the reaction time was 15 min.

[0125] Figure 3 Figure 1 is a schematic diagram of the competitive AuNFs@TA / Zr immunochromatographic test strip detection principle, where 9 is the anti-mouse antibody or anti-rabbit antibody on the quality control line, 10 is the small molecule analyte, 11 is the AuNFs@TA / Zr-antibody complex, and 12 is the artificially coupled antigen of the analyte on the detection line.

[0126] Figure 8 、 Figure 9Shown is a schematic diagram of the actual results of using an immunochromatographic test strip prepared using the AuNFs@TA / Zr beacon carrier to detect clothianidin. The experiment demonstrated that with increasing clothianidin concentration, both the colorimetric and photothermal signals of the test strip's detection line decreased in a concentration-dependent manner. Visible to the naked eye, a significant inhibitory effect was observed at 0.5 ng / mL, with complete disappearance at 50 ng / mL. These results confirm that the AuNFs@TA / Zr-based test strip can achieve semi-quantitative detection of clothianidin through dual-modal signaling (colorimetric / photothermal). Its naked-eye sensitivity (0.5 ng / mL) and dynamic response range (0.5 ng / mL-50 ng / mL) meet the requirements for trace analysis, further validating the practicality and reliability of this method.

[0127] Comparative Example 1

[0128] The difference from Example 1 is that: a TA and Zr coated non-flower-shaped gold nanoparticle is prepared, and the specific preparation method is as follows:

[0129] 1. Synthesis of Gold Nanoparticle-Tannic Acid Complex (AuNPs@TA)

[0130] 20 mg of tannic acid (TA) was dissolved in 100 mL of phosphate buffered saline (PBS, pH 8.5) and magnetically stirred to form a homogeneous solution. Under continuous stirring, 3 mL of a 1% aqueous solution of chloroauric acid (HAuCl4) was added, and the reaction continued for 10 minutes. The reaction system was centrifuged at 8000 rpm for 10 minutes to collect the product, which was then washed three times with deionized water to remove unreacted products. Finally, the purified AuNPs@TA complex was redispersed in 8 mL of deionized water to obtain a stable nanoparticle dispersion.

[0131] 2. Synthesis of Zirconium-Coordinated Gold Nanocomposites (AuNPs@TA-Zr)

[0132] To the AuNPs@TA dispersion, 5 mg of zirconium tetrachloride (ZrCl4) was added. The coordination reaction was carried out at room temperature (25 ± 2°C) with continuous stirring at 1200 rpm for 12 hours. After the reaction, the mixture was centrifuged at 8000 rpm for 10 minutes to collect the product, which was then washed three times with deionized water to remove free zirconium ions. The resulting AuNPs@TA-Zr composite nanoparticles can be used immediately for subsequent functionalization applications or stored refrigerated at 4°C for stability.

[0133] Figure 10 Shown is a transmission electron micrograph of non-flower-shaped gold nanoparticles coated with metal polyphenols, where spherical nanoparticles are clearly visible.

[0134] The present invention uses non-flower-shaped gold nanoparticles coated with metal polyphenols (AuNPs@TA-Zr) as signal probes, based on the sandwich immunochromatographic test strip construction method described in Example 2, to detect Escherichia coli. E. coli O157:H7 Quantitative detection is performed. Figure 11 As shown: When the concentration gradient of target bacteria is (0 CFU / mL, 5×10 4 CFU / mL, 1×10 5 CFU / mL, 5×10 5 CFU / mL, 1×10 6 CFU / mL, 5×10 6 CFU / mL), the colorimetric signal intensity of the test strip detection line (T line) is positively correlated with the bacterial concentration, and the lowest detection limit that can be identified by the naked eye is 1×10 5 CFU / mL.

[0135] Comparative experiments revealed that the detection sensitivity of the test strip using the metal polyphenol-coated flower-shaped gold nanoparticles (AuNFs@TA / Zr) as the signal carrier was 2 times higher than that of the non-flower-shaped structure system in comparative example 1. This difference is attributed to the following synergistic mechanisms: 1) the flower-shaped gold nanoparticles (AuNFs) have a higher specific surface area, which significantly enhances the loading efficiency and interfacial reaction activity of biomolecules such as antibodies; 2) the metal polyphenol coating layer is coated with Zr 4+ The coordination effect stabilizes the surface properties of the nanoparticles, effectively inhibiting nonspecific adsorption and improving biocompatibility. 3) Manipulating the nanoparticle morphology directly affects the efficiency of probe-target binding. The spatial topological properties of the flower-like structure are more conducive to the formation of highly sensitive immune complexes. These results demonstrate that the detection performance of nanoprobes can be significantly enhanced through the coordinated optimization of morphology design and surface coordination.

[0136] Comparative Example 2

[0137] The difference from Example 1 is that: a gold nanoparticle coated with Zr is prepared by replacing tannic acid with other polyphenols (such as polydopamine, proanthocyanidin, or epicatechin). The specific preparation method is as follows:

[0138] (1) Preparation of chloroauric acid solution

[0139] 99 mL of deionized water was preheated to 70 °C in a microwave reactor. 1 mL of a 1% chloroauric acid (HAuCl4) aqueous solution was then quickly added to the preheated deionized water under continuous stirring and mixed evenly to obtain a chloroauric acid solution.

[0140] (2) Preparation of a mixed solution of polydopamine, proanthocyanidins, or epicatechin-hydroxylamine hydrochloride-chloroauric acid

[0141] Dissolve 20 mg of polydopamine, proanthocyanidin, or epicatechin and 0.5 mL of 0.1 mol / L hydroxylamine hydrochloride aqueous solution in deionized water and mix thoroughly to form a uniform solution.

[0142] The polydopamine or proanthocyanidin or epicatechin-hydroxylamine hydrochloride mixed solution is quickly added to the chloroauric acid solution obtained in step (1), and stirring is continued to allow the components to fully react. The mixed system is further heated at 70°C for 1 minute to finally obtain a homogeneous mixed solution of polydopamine or proanthocyanidin or epicatechin, hydroxylamine hydrochloride and chloroauric acid.

[0143] (3) Collection of AuNFs@TA solution

[0144] The homogenized mixed solution was cooled at room temperature for 40 minutes and then centrifuged at 12,000 rpm for 10 minutes to collect the precipitate. The precipitate was washed three times with deionized water and finally redissolved in 8 mL of pure water to prepare the AuNFs@TA solution.

[0145] (4) Preparation of AuNFs@TA / Zr solution

[0146] Add 5 mg of zirconium chloride (ZrCl4) to the AuNFs@TA solution obtained in step 3 and stir vigorously (1200 r / min) at room temperature overnight; then centrifuge to separate the precipitate, wash it, and redissolve it in 1 mL of deionized water to obtain the AuNFs@TA / Zr solution, which was stored at 4°C.

[0147] The prepared polydopamine or proanthocyanidin or epicatechin and Zr-coated nano-gold particles were used to prepare sandwich immunochromatographic test strips according to the method in Example 2. E. coli O157:H7 Conduct testing.

[0148] Figure 12 The following is a transmission electron micrograph of polydopamine-Zr coated gold nanoparticles (AuNFs@polydopamine-Zr gold nanoparticles), showing flower-like nanoparticles. The target bacteria concentration gradient is 0 CFU / mL, 5×10 2 CFU / mL, 1×10 3 CFU / mL, 5×10 3 CFU / mL, 1×10 4 CFU / mL, 5×10 4 CFU / mL, 1×10 5 CFU / mL, 5×10 5 CFU / mL, 1×10 6 CFU / mL, 5×10 6 CFU / mL.

[0149] The test results of the test strip based on polydopamine and Zr-coated gold nanoparticles (AuNFs@polydopamine-Zr gold nanoparticles) as probes are shown in Figure 13 The colorimetric signal intensity of the detection line (T line) is positively correlated with the bacterial concentration, but the signal intensity of the T and C lines has decreased, the color is not dark enough, and the lowest detection limit that can be identified by the naked eye is 1×10 5 CFU / mL, a two-fold decrease in sensitivity compared to the AuNFs@TA / Zr-based dual-modal test strip of the present invention. Polydopamine-Zr-coated gold nanoparticles were prepared according to the synthesis method of AuNFs@TA / Zr, replacing tannic acid with polydopamine. All other preparation conditions and steps were identical. Although the synthesized polydopamine-Zr-coated gold nanoparticles exhibited a flower-like morphology, the detection sensitivity was somewhat reduced.

[0150] Figure 14 Shown is a transmission electron microscopy image of proanthocyanidin-Zr coated gold nanoparticles (AuNPs@proanthocyanidin-Zr gold nanoparticles), showing spherical nanoparticles.

[0151] The test results of the test strip based on proanthocyanidins and Zr-coated gold nanoparticles (AuNPs@proanthocyanidins-Zr gold nanoparticles) as probes are as follows Figure 15 As shown in the figure, the colorimetric signal intensity of the detection line (T line) is positively correlated with the bacterial concentration, but the signal intensity of the T and C lines has decreased, and the color has become wine red. The lowest detection limit that can be identified by the naked eye is 5×10 5 CFU / mL. The sensitivity of the dual-modal test strip based on AuNFs@TA / Zr of the present invention is reduced by 10 times. According to the synthesis method of AuNFs@TA / Zr, proanthocyanidin-Zr coated nano-gold particles were prepared, and tannic acid was replaced by proanthocyanidin. The other preparation conditions and steps were the same. The synthesized proanthocyanidin-Zr coated nano-gold particles did not show a flower-like shape, the morphology of the nanoparticles changed, and the detection sensitivity was significantly reduced.

[0152] Figure 16 Shown is a transmission electron micrograph of epicatechin-Zr coated gold nanoparticles (AuNPs@epicatechin-Zr gold nanoparticles), showing non-flower-shaped nanoparticles.

[0153] The test results of the test strip based on epicatechin and Zr-coated gold nanoparticles (AuNPs@epicatechin-Zr gold nanoparticles) as probes are as follows: Figure 17As shown, the test and control lines failed to develop color, indicating that the color is too light to be used for immunochromatographic test strips. Using the same preparation conditions and steps as AuNFs@TA / Zr, but replacing tannic acid with epicatechin, the synthesized epicatechin-Zr-coated gold nanoparticles changed morphology, no longer exhibiting a flower-like shape. The color was light gray and failed to develop on the test strip, making them unsuitable for immunochromatographic applications.

[0154] Comparative experiments showed that when tannic acid was replaced with polydopamine, proanthocyanidins, or epicatechin, the morphology, structure, and color of the nanoparticles changed significantly due to the change in polyphenol type, and the detection sensitivity decreased. This is because tannic acid's high density of catechol groups and stereochemical structure provide advantages in metal coordination and network stability, surpassing other polyphenols such as polydopamine, proanthocyanidins, and epicatechin.

[0155] Comparative Example 3

[0156] The difference from Example 1 is that a TA-coated flower-shaped gold nanoparticle is prepared. The specific preparation method is as follows:

[0157] (1) Preparation of chloroauric acid solution

[0158] 99 mL of deionized water was preheated to 70 °C in a microwave reactor. 1 mL of a 1% chloroauric acid (HAuCl4) aqueous solution was then quickly added to the preheated deionized water under continuous stirring and mixed evenly to obtain a chloroauric acid solution.

[0159] (2) Preparation of tannic acid-hydroxylamine hydrochloride-chloroauric acid mixed solution

[0160] Dissolve 20 mg of tannic acid (relative molecular mass 1701.2 g / mol) and 0.5 mL of 0.1 mol / L hydroxylamine hydrochloride aqueous solution in deionized water and mix thoroughly to form a homogeneous solution.

[0161] The tannic acid-hydroxylamine hydrochloride mixed solution was quickly added to the chloroauric acid solution obtained in step (1) and stirred continuously to allow the components to react fully. The mixed system was further heated at 70°C for 1 min to obtain a homogeneous mixed solution of tannic acid, hydroxylamine hydrochloride, and chloroauric acid.

[0162] (3) Collection of AuNFs@TA solution

[0163] The homogenized mixed solution was cooled at room temperature for 40 minutes and then centrifuged at 12,000 rpm for 10 minutes to collect the precipitate. The precipitate was washed three times with deionized water and finally redissolved in 8 mL of pure water to prepare the AuNFs@TA solution.

[0164] Figure 18 Shown is a transmission electron micrograph of TA-coated flower-shaped gold nanoparticles, showing flower-shaped nanoparticles.

[0165] The prepared TA-coated flower-like gold nanoparticles (AuNFs@TA flower-like gold nanoparticles) were used to prepare sandwich immunochromatographic test strips according to the method in Example 2. E. coli O157:H7 Test results are shown in Figure 19 As shown in the figure, the colorimetric signal intensity of the test strip detection line (T line) based on TA-coated flower-like gold nanoparticles (AuNFs@TA flower-like gold nanoparticles) as a probe is positively correlated with the bacterial concentration, but the signal intensity of the T and C lines has decreased, the color is lighter, and the lowest detection limit that can be identified by the naked eye is 1×10 5 CFU / mL. Compared with the sensitivity of the dual-modal test strip based on AuNFs@TA / Zr in Example 1, it decreased by 2 times.

[0166] The decrease in sensitivity of the test strip based on TA-coated flower-like gold nanoparticles (AuNFs@TA flower-like gold nanoparticles) is mainly caused by the following two key reasons:

[0167] (1) Reduced probe labeling efficiency: The dense TA coating covers the active sites on the gold surface, resulting in a decrease in antibody coupling density; at the same time, the nonspecific adsorption of TA causes non-directional antibody coupling, reducing the antigen binding efficiency;

[0168] (2) Insufficient signal amplification: The flower-like gold nanostructure formed by TA coordination alone is loose, the local surface plasmon resonance effect is weakened, and the lack of Zr 4+ The cross-linked dense polyphenol network resulted in a reduced aggregation of T-line gold particles and a significant weakening of the colorimetric signal.

[0169] The above description is only a preferred specific embodiment of the present invention, but the scope of protection of the present invention is not limited thereto. Any technician familiar with the technical field, within the technical scope disclosed by the present invention, who makes equivalent replacements or changes based on the technical solution and inventive concept of the present invention, should be covered by the scope of protection of the present invention.

Claims

1. A dual-modal immunochromatographic test strip, characterized in that: The dual-modal immunochromatographic test strip uses AuNFs@TA / Zr as a probe; The preparation method of AuNFs@TA / Zr comprises the following steps: S1. Preheat deionized water, add chloroauric acid aqueous solution and mix well to obtain chloroauric acid solution; S2, dissolving tannic acid and hydroxylamine hydrochloride aqueous solution in deionized water, and mixing thoroughly to obtain a mixed solution; S3, reacting the chloroauric acid solution with the mixed solution, cooling and centrifuging to obtain a precipitate after the reaction is completed; washing the precipitate and redissolving it in water to obtain an AuNFs@TA solution; S4, adding zirconium chloride to the AuNFs@TA solution, stirring and mixing, centrifuging, and washing to obtain AuNFs@TA / Zr; The AuNFs@TA / Zr is loaded on the dual-modal immunochromatographic test strip in the form of an AuNFs@TA / Zr labeled antibody complex, wherein the preparation method of the AuNFs@TA / Zr labeled antibody complex comprises the following steps: AuNFs@TA / Zr was dissolved in water to obtain AuNFs@TA / Zr solution. The antibody to be labeled is added to the AuNFs@TA / Zr solution, and after thorough mixing, a blocking agent is added. After the reaction, the solution is centrifuged to obtain a precipitate, and the obtained precipitate is redissolved to prepare an AuNFs@TA / Zr labeled antibody complex.

2. The dual-modal immunochromatographic test strip according to claim 1, characterized in that: In step S1, the temperature of the preheated deionized water is 65°C to 75°C, the mass concentration of the chloroauric acid aqueous solution is 0.5% to 1.5%, and the added volume ratio of deionized water to the chloroauric acid solution is 95 to 103:

1.

3. The dual-modal immunochromatographic test strip according to claim 1, characterized in that: In step S2, the addition ratio of tannic acid and hydroxylamine hydrochloride aqueous solution is 18 mg~22 mg:0.5 mL, and the concentration of hydroxylamine hydrochloride aqueous solution is 0.05 mol / L~0.15 mol / L.

4. The dual-modal immunochromatographic test strip according to claim 1, characterized in that: In step S3, the cooling condition is room temperature, the cooling time is 35 min to 45 min, the centrifugal speed is 3000 r / min to 12000 r / min, the centrifugal time is 10 min to 30 min, the number of washing times is 1 time to 3 times, and the volume of water added for re-dissolution is 7 mL to 9 mL.

5. The dual-modal immunochromatographic test strip according to claim 1, characterized in that: In step S4, the addition ratio of zirconium chloride and chloroauric acid solution is 4 mg~6 mg:1 mL, the stirring speed is 1000 r / min~1400 r / min, the stirring time is 20 h~28 h, the centrifugal speed is 3000 r / min~12000 r / min, the centrifugal time is 10 min~30 min, and the number of washing times is 1 time~3 times.

6. The dual-modal immunochromatographic test strip according to claim 1, characterized in that: In the preparation method of the AuNFs@TA / Zr labeled antibody complex, The final concentration of the antibody after adding the antibody to be labeled is 1 μg / mL to 100 μg / mL; After the blocking agent is added, the final mass concentration of the blocking agent is 0.1% to 1%, and the blocking agent is selected from any one of casein, bovine serum albumin, ovalbumin, polyethylene glycol, and skim milk; The centrifugal speed is 5000 r / min to 20000 r / min, and the centrifugal time is 10 min to 50 min; The precipitate after centrifugation was re-dissolved with 0.01M-0.1M phosphate buffer, pH 6.0-9.0; The antibody to be labeled includes any one of a monoclonal antibody and a polyclonal antibody.

7. The dual-modal immunochromatographic test strip according to claim 1, characterized in that: The antibody to be labeled includes any one of nanobodies and phage-expressed antibodies.

8. The dual-modal immunochromatographic test strip according to claim 6, characterized in that: The dual-modal immunochromatographic test strip comprises: base plate; A nitrocellulose membrane is disposed on the bottom plate, and has detection lines and quality control lines spaced apart from each other; A glass fiber mat is arranged on one end of the nitrocellulose membrane having the detection line, one end of the glass fiber mat is arranged on the bottom plate, and the other end is arranged on the nitrocellulose membrane, and the AuNFs@TA / Zr labeled antibody complex is arranged on the glass fiber mat; absorbent paper, arranged at one end of the nitrocellulose membrane having the quality control line, one end of the absorbent paper being arranged on the bottom plate, and the other end being arranged on the nitrocellulose membrane; A sample pad has one end arranged on the bottom plate and the other end arranged on the glass fiber pad.

9. The dual-modal immunochromatographic test strip according to claim 8, characterized in that: The method of placing the AuNFs@TA / Zr labeled antibody complex on the glass fiber mat comprises: spraying the AuNFs@TA / Zr labeled antibody complex onto the glass fiber mat at a volume of 7 μL / cm~9 μL / cm, and vacuum drying for 1.5 h~2.5 h; The detection line is coated with an artificially coupled antigen of the test object or an antibody to the test object, and the quality control line is coated with a goat anti-mouse antibody.

10. Use of the dual-modal immunochromatographic test strip according to any one of claims 1 to 9 in detecting pesticides or pathogenic bacteria for non-disease diagnosis purposes, characterized in that: The method for detecting pesticides or pathogenic bacteria using the dual-modal immunochromatographic test strip comprises the following steps: The processed sample to be tested is added to the dual-modal immunochromatographic test strip with a sample volume of 50 μL to 200 μL and a reaction time of 3 minutes to 30 minutes. Quantitative detection is achieved by reading the grayscale data of the test strip and the imaging data of the infrared imager to calculate the concentration of the sample to be tested, or qualitative judgment of the sample to be tested is achieved by visually observing the presence or absence of colored strips on the test line and quality control line on the dual-modal immunochromatographic test strip.

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