Test paper for detecting ethinyl estradiol and preparation method thereof

Abstract

The application discloses a kind of detection ethinyl estradiol test paper and preparation method thereof, comprising the following steps: (1) preparation Sb2WO6@NiS complex, surface adsorption ethinyl estradiol antibody, i.e. first antibody obtains photo-thermal signal probe;(2) assembly test strip, ethinyl estradiol antigen and second antibody are respectively fixed on detection area and quality control area;(3) after photo-thermal signal probe is mixed with sample liquid to be measured, test strip sample pad end is inserted into the mixed solution and carries out chromatography;(4) detection photo-thermal signal: with near infrared LED illumination detection area and with thermal imaging or temperature measuring equipment acquisition temperature, according to the linear relationship between detection area temperature and sample concentration to be measured quantitative analysis.The application detection mode is flexible, linear range is wide, sensitivity is high, and specificity is good.

Description

Technical Field

[0001] This invention relates to the field of small molecule analysis and detection technology, and in particular to a test strip for detecting ethinylestradiol and its preparation method. Background Technology

[0002] Lateral flow immunochromatographic assay (LFIA) strips are widely used in the design of portable analytical sensors due to their advantages of rapid analysis, good selectivity, low cost, small sample volume requirements, ease of large-scale production, and stable long-term storage. Photothermal LFIA strips, which utilize photothermal signal probes, offer sensitivity improvements of tens to hundreds of times compared to colorimetric methods. The signal from this type of detection is generated by temperature changes caused by the photothermal effect, resulting in low background signal, high sensitivity, and the ability to operate on various substrates of different colors, demonstrating strong application potential. Currently, there is still a need for new nanomaterials with higher photothermal conversion efficiency to serve as photothermal signal probes, forming a more pronounced thermal contrast to further improve the performance of photothermal LFIA strips.

[0003] Sb₂WO₆ is a compound with unique photoelectric properties, showing promising applications in photocatalysis and photothermal conversion. For example, studies have constructed a three-dimensional biomimetic bird's nest-shaped nanocomposite material by in-situ combining Sb₂WO₆ and D-fructose (D-Fru), which exhibits excellent photothermal conversion performance, enabling efficient solar photothermal conversion and water evaporation. Furthermore, NiS, a sulfide semiconductor material with a narrow band gap and high light absorption coefficient, also shows potential for applications in photocatalysis and photothermal conversion. However, compared to Sb₂WO₆, direct research on the photothermal conversion applications of NiS is relatively limited. Summary of the Invention

[0004] To address the aforementioned problems in existing technologies, this invention provides a test strip for detecting ethinylestradiol and its preparation method. The Sb2WO6@NiS composite material is used to prepare the photothermal test strip for ethinylestradiol detection. The test strip of this invention is used to detect ethinylestradiol with a detection limit as low as 10. -1 pg / mL.

[0005] The technical solution of the present invention is as follows:

[0006] The first objective of this invention is to provide a Sb2WO6@NiS composite material, the preparation method of which includes the following steps:

[0007] (1) Add Sb2WO6 nanoparticles to a polyethylene glycol aqueous solution, disperse them ultrasonically, and stir to prepare a mixture;

[0008] (2) The mixture obtained in step (1) is added to NiS aqueous solution, stirred and reacted, and centrifuged and dried after the reaction is completed to obtain the composite material.

[0009] In one embodiment of the present invention, in step (1), one or more of the following conditions are met:

[0010] The weight-average molecular weight of polyethylene glycol is 5000-10000;

[0011] The concentration of the polyethylene glycol aqueous solution is 1-4 mg / mL;

[0012] The mass ratio of Sb2WO6 nanoparticles to polyethylene glycol is 1:10-1:5;

[0013] The ultrasonic dispersion conditions are: ultrasonic power of 120-600W and ultrasonic time of 20-30min;

[0014] The reaction conditions with stirring are: room temperature, 500-1000 rpm, 10-15 h.

[0015] In step (2), one or more of the following conditions must be met:

[0016] The concentration of the NiS aqueous solution is 0.1-1% (containing 5% nitric acid);

[0017] The volume ratio of the mixed solution to the NiS aqueous solution is 1:5-5:1.

[0018] The conditions for the stirred reaction are: 80-95℃, 10-15h.

[0019] In one embodiment of the present invention, 1-2 mg of Sb2WO6 nanoparticles are added to 5-10 mL of deionized water containing 10-20 mg of polyethylene glycol (MW = 5000-10000), sonicated for 20-30 minutes, and stirred overnight; 1 mL of the above mixture is added to 1 mL of NiS aqueous solution and stirred overnight; the dark gray powder is collected by centrifugation and dried for further use.

[0020] In one embodiment of the present invention, Sb2WO6 nanoparticles are synthesized by a hydrothermal method, and the particle size is 10-100 nm.

[0021] The second objective of this invention is to provide an application of the above-mentioned Sb2WO6@NiS composite material for the preparation of photothermal signal probes.

[0022] The third objective of this invention is to provide a photothermal signal probe prepared from the above-mentioned Sb2WO6@NiS composite material, containing Sb2WO6@NiS composite material, ethinylestradiol antibody, and bovine serum albumin.

[0023] In one embodiment of the present invention, the photothermal signal probe preparation method is as follows: Sb2WO6@NiS composite material is mixed with analyte antibody and bovine serum albumin to obtain Sb2WO6@NiS-mAb mixture.

[0024] In one embodiment of the present invention, Sb2WO6@NiS composite material is placed in a centrifuge tube, a weak alkaline solution is added to adjust the pH of the system to 6-8, the mixture is shaken and mixed, ethinylestradiol antibody is added, and the mixture is shaken and reacted at room temperature for 30-60 min. Then, bovine serum albumin (BSA) is added for blocking, and the mixture is shaken and reacted at room temperature for 1-1.5 h. After centrifugation, the supernatant is removed, and the remaining substance is reconstituted in buffer solution to obtain Sb2WO6@NiS-mAb mixture, which is then refrigerated for later use.

[0025] In one embodiment of the present invention, the buffer solution is a phosphate buffer, a borate buffer, or a carbonate buffer.

[0026] A fourth objective of this invention is to provide a test strip containing the aforementioned photothermal signal probe, the test strip comprising a photothermal signal probe and a test strip body;

[0027] The photothermal signal probe is prepared by modifying an ethinylestradiol polyclonal antibody with an Sb2WO6@NiS composite nanomaterial, and is used either in a form independent of the main structure of the photothermal test strip or in a form dried on the conjugate pad.

[0028] The main body of the test strip includes a PVC base plate, a sample pad, a nitrocellulose membrane (NC membrane), and an absorbent pad.

[0029] In one embodiment of the present invention, the test strip body comprises: a sample pad, a nitrocellulose membrane (NC membrane), and an absorbent pad, which are sequentially and overlappingly pasted from top to bottom on a PVC base plate in a horizontal direction; wherein, the nitrocellulose membrane is used to separate and detect analytes from other substances in the sample, the sample pad is used for sample loading, the absorbent pad is used to absorb excess liquid, and the PVC base plate provides physical support for the test strip; the nitrocellulose membrane includes a detection zone (T zone) and a control zone (C zone); wherein, ethinylestradiol antigen (ethinylestradiol-protein conjugate) is immobilized on the T zone; and a secondary antibody is immobilized on the C zone, the secondary antibody being a secondary antibody against the primary antibody (i.e., anti-primary antibody);

[0030] The photothermal signal probe can be used independently of the main structure of the photothermal test strip, or the photothermal signal probe can be dried on the conjugate pad and the conjugate pad can be inserted between the sample pad and the NC membrane.

[0031] In one embodiment of the present invention, the ethinylestradiol antibody, i.e., the primary antibody, is derived from at least one of mice, rats, and rabbits; the secondary antibody is derived from at least one of goat anti-mouse secondary antibody, rabbit anti-mouse secondary antibody, goat anti-rabbit secondary antibody, and donkey anti-rabbit secondary antibody.

[0032] The fifth object of the present invention is to provide a method for preparing the above-mentioned test strip, comprising the following steps:

[0033] Preparation of Sb2WO6@NiS composite material;

[0034] Preparation of Sb2WO6@NiS composite material, ethinylestradiol antibody, and bovine serum albumin Sb2WO6@NiS-mAb mixture;

[0035] Constructing the test strip: Add or spray ethinylestradiol antigen to the T zone, and add or spray secondary antibody against the primary antibody (i.e., anti-primary antibody) to the C zone. After drying, store in a vacuum bag for later use.

[0036] The photothermal signal probe, namely the Sb2WO6@NiS-mAb mixture, is stored in a sealed container as a solution or lyophilized powder; or the photothermal signal probe, namely the Sb2WO6@NiS-mAb mixture, is dried on the binding pad and the binding pad is inserted between the sample pad and the NC membrane for use.

[0037] In one embodiment of the present invention, the addition or spraying of ethinylestradiol antigen to the T region specifically involves adding or spraying a 5-20 mM PBS solution containing 0.5-10 mg / mL ethinylestradiol antigen to the T region; the addition or spraying of a secondary antibody against the primary antibody, i.e., anti-primary antibody, to the C region specifically involves adding or spraying a 5-20 mM PBS solution containing 0.05-10 mg / mL secondary antibody to the C region.

[0038] The sixth objective of this invention is to provide an application of the above-mentioned test strip for detecting ethinylestradiol; the photothermal temperature of the detection area is negatively correlated with the content of ethinylestradiol to be tested, and the color intensity is negatively correlated with the content of ethinylestradiol in the sample, specifically:

[0039] When the sample does not contain ethinylestradiol, the T region appears purple and the temperature is high.

[0040] When the sample contains ethinylestradiol, the T region appears light purple or even disappears, and the temperature decreases.

[0041] The quality control area, which serves as a reference for verifying the validity of the test strip results, always appears purple.

[0042] The sixth objective of this invention is to provide a test strip for detecting ethinylestradiol, comprising a photothermal signal probe and a test strip body;

[0043] The photothermal signal probe was prepared by mixing the Sb2WO6@NiS composite material with the analyte antibody and bovine serum albumin to obtain the Sb2WO6@NiS-mAb mixture.

[0044] The main body of the test strip comprises: a sample pad, a nitrocellulose membrane (NC membrane), and an absorbent pad, which are horizontally overlapped and pasted from top to bottom on a PVC base plate. The nitrocellulose membrane is used to separate and detect analytes from other substances in the sample; the sample pad is used for sample loading; the absorbent pad is used to absorb excess liquid; and the PVC base plate provides physical support for the test strip. The nitrocellulose membrane includes a detection zone (T zone) and a control zone (C zone). The T zone contains immobilized ethinylestradiol antigen, i.e., ethinylestradiol-protein conjugate; the C zone contains immobilized secondary antibody, which is a secondary antibody derived from the primary antibody, i.e., anti-primary antibody; the primary antibody is a murine ethinylestradiol polyclonal antibody; and the secondary antibody is a goat anti-mouse antibody.

[0045] The photothermal signal probe can be used independently of the main structure of the photothermal test strip, or the photothermal signal probe can be dried on the conjugate pad and the conjugate pad can be inserted between the sample pad and the NC membrane.

[0046] In one embodiment of the present invention, the method for detecting ethinylestradiol using test strips includes the following steps:

[0047] When the photothermal signal probe is used independently of the main structure of the photothermal test strip, it specifically includes:

[0048] The sample to be tested was mixed with Sb2WO6@NiS-mAb solution in the running buffer, and then the test strip was placed in it. After 10-15 minutes, the test strip was removed and dried. The photothermal analysis results were then read and quantitative analysis was performed.

[0049] When the photothermal signal probe is dried on the conjugate pad and the conjugate pad is inserted between the sample pad and the NC membrane in the main body structure of the test strip, the specific steps include:

[0050] Mix the sample to be tested with the running buffer, then place it into the test strip. After 10-15 minutes, remove the test strip and let it dry. Read the photothermal analysis results and perform quantitative analysis.

[0051] The method for reading photothermal analysis results after the test strip is dried is as follows: The test strip is placed under a near-infrared LED light source for excitation, and the temperature result is obtained using a thermal imaging or temperature measuring device and a smart display terminal. The thermal imaging or temperature measuring device includes at least one of a mobile phone infrared thermal imaging analysis accessory, an infrared thermal imager, and a temperature measuring gun. The smart display terminal includes at least one of a computer and a smartphone. The thermal imaging device acquires a photothermal image and outputs it for display through the connected smart display terminal.

[0052] In one embodiment of the present invention, the running buffer is a PBS solution with pH 6.5 to 8.0 and a total concentration of 10 mM, and the following components are in mass percentage, specifically including: 0 to 15% sucrose, 1 to 10% BSA, and 0.15 to 1% Tween-20.

[0053] In one embodiment of the present invention, the wavelength of the near-infrared LED light source is 980nm.

[0054] The detection principle of the photothermal test strip for ethinylestradiol in this invention is as follows: The control zone (zone C) serves as a reference for verifying the validity of the test strip results and always appears purple. After the sample solution and Sb2WO6@NiS-mAb are premixed, they move towards the absorbent paper under capillary action. When there is no ethinylestradiol in the sample, Sb2WO6@NiS-mAb is captured by ethinylestradiol-BSA in zone T, and zone T appears purple to the naked eye. Under laser excitation, Sb2WO6@NiS-mAb exhibits an LSPR effect, and the temperature rises. Excess Sb2WO6@NiS-mAb is captured by the secondary antibody in zone C, so zone C also appears purple. When the sample contains ethinylestradiol, ethinylestradiol binds to some of the Sb2WO6@NiS-mAb, thereby reducing the total amount of Sb2WO6@NiS-mAb captured in zone T. The LSPR effect of Sb2WO6@NiS-mAb in zone T decreases or even disappears, and the temperature drops. As the concentration of ethinylestradiol in the sample increases, less and less Sb2WO6@NiS-mAb is trapped in the T region, and the temperature of the T region gradually decreases, which is inversely proportional to the analyte concentration.

[0055] The beneficial technical effects of this invention are as follows:

[0056] The Sb2WO6@NiS composite material provided by this invention can be used in the preparation and application of photothermal test strips without changing the conventional structure of the test strips. It can be used as a photothermal signal probe, and can be used independently of the aforementioned photothermal test strip form for easy storage, or it can be fixed to the bonding pad of the test strip for easy portability.

[0057] The present invention provides a detection method including a photothermal test strip for detecting ethinylestradiol. The photothermal test strip is based on the plasma resonance effect, using a high photothermal conversion efficiency and a small size Sb2WO6@NiS composite material as a T-zone fixed photothermal signal probe, excited by a near-infrared LED light source, and combined with a smart terminal and infrared thermal imaging accessories to realize the acquisition of photothermal signals, effectively removing the sample liquid base color and test strip fluorescence background interference, thereby improving the signal-to-noise ratio.

[0058] The detection method provided by this invention has a detection limit as low as 10 for ethinylestradiol. -1 pg / mL. Attached Figure Description

[0059] Figure 1 This is a schematic diagram of the photothermal test strip of the present invention;

[0060] Figure 2The images show (A) scanning electron microscope (SEM) image, (B) transmission electron microscope (TEM) image, and (C) X-ray diffraction pattern of the Sb2WO6@NiS composite material in the photothermal signal probe of the test strip of this invention.

[0061] Figure 3 The photothermal heating curves of Sb2WO6@NiS composite material, Sb2WO6, and NiS in the photothermal signal probe of the test strip of this invention are shown.

[0062] Figure 4 The test strip of this invention exhibits its response to water samples of different concentrations of ethinylestradiol under photothermal mode and its operating curves.

[0063] Figure 5 This is a diagram showing the response of the test strip of the present invention to ethinylestradiol and four other estrogens. Detailed Implementation

[0064] The present invention will now be described in detail with reference to the accompanying drawings and embodiments.

[0065] Example 1: Preparation of raw materials

[0066] 1.1 Preparation of Sb₂WO₆ nanocrystals

[0067] Sb₂WO₆ nanoparticles were synthesized via a hydrothermal method. 0.5 g of Na₂WO₄·2H₂O and 1.5 g of Bi(NO₃)₃·5H₂O were dissolved separately in 5 mL of ethylene glycol. After mixing, 35 mL of ethanol was added. After stirring for 0.5 h, the solution was transferred to a 50 mL autoclave and heated at 180 °C for 10 h. The precipitate after centrifugation was washed with deionized water and ethanol and dried at 80 °C for 10 h. Then, the resulting powder was calcined at 450 °C (heating rate of 2 °C / min) for 2 h to obtain Sb₂WO₆ nanocrystals.

[0068] 1.2 Preparation of Sb2WO6@NiS composite material and characterization of its structural properties

[0069] Add 1 mg of Sb2WO6 nanoparticles to 5 mL of deionized water containing 10 mg of polyethylene glycol (MW = 7000), sonicate at 200 W for 200 minutes, and stir for 10 h; add 1 mL of the above mixture to 1 mL of NiS aqueous solution and stir overnight; collect the dark gray powder by centrifugation and dry it for further use.

[0070] The scanning electron microscope, transmission electron microscope and XRD of Sb2WO6@NiS composite material were measured as follows: Figure 2 As shown; the photothermal heating curve is as follows Figure 3 As shown, the Sb2WO6@NiS composite material exhibits the strongest heating capacity, superior to both Sb2WO6 and NiS.

[0071] 1.3 Adsorption of Sb2WO6@NiS with Antibody: A mixture of Sb2WO6@NiS composite material, ethinylestradiol antibody, and bovine serum albumin was prepared, namely the Sb2WO6@NiS-mAb mixture.

[0072] Take 1 mL of Sb2WO6@NiS into a centrifuge tube, add 4 μL of 0.2 W / L K2CO3 solution to adjust the pH of the system, vortex to mix, then add 5 μL of 1 mg / mL ethinylestradiol antibody, mix well, and incubate at room temperature with shaking for 45 min. After the reaction is complete, add 100 μL of BSA (m / m = 5%) for blocking for 1 h, centrifuge at 12000 r / min for 30 min, remove the supernatant, and then redissolve in 100 μL of buffer (20 mol / L Na3PO4, 5% BSA, 0.25% Tween-20, 10% sucrose) to obtain the Sb2WO6@NiS-mAb mixture, and store at 4℃ for later use.

[0073] 1.4 Preparation of the solution for the detection zone (T zone)

[0074] Ethinyl estradiol-BSA was diluted to 0.6 mg / mL with 10 mM PBS solution at pH 7.4.

[0075] 1.5 Preparation of solutions for the quality control zone (Zone C)

[0076] The goat anti-mouse secondary antibody was diluted to 0.4 mg / mL with 10 mM PBS solution at pH 7.4.

[0077] Example 2: Preparation of test strips

[0078] according to Figure 1 The NC membrane is attached to the middle of the PVC base plate using a modular assembly method. The sample pad and absorbent pad are overlapped at the left and right ends of the NC membrane, respectively, covering the NC membrane by about 2 mm. The assembled card is cut into strips with a width of 3 mm. Detection lines (T zone) and control lines (C zone) are drawn on the NC membrane, with a 4 mm interval between the two lines, thus obtaining a blank test strip. 0.5 μL of T zone solution and 0.5 μL of C zone solution are added to the T zone and C zone respectively. The sampled test strips are placed in an oven and dried at 37°C for 60 min, then stored in a vacuum bag for later use.

[0079] Example 3: Preparation of test strips containing binding pads

[0080] test strips according to Figure 1The binding method consists of a sample pad, a nitrocellulose membrane (NC), a conjugation pad, and a PVC base plate. The NC membrane is adhered to the center of the PVC base plate, and a detection line (T zone) and a control line (C zone) are drawn on the NC membrane, with a 4mm gap between the two lines. Then, the right end of the conjugation pad overlaps the left end of the NC membrane, covering it by about 2mm. The sample pad overlaps the left end of the conjugation pad, covering it by about 2mm. The absorbent pad overlaps the right end of the NC membrane, covering it by about 2mm. The assembled test strip is cut into 3mm wide strips. A mixture of 10μL Sb2WO6@NiS composite material, ethinylestradiol antibody, and bovine serum albumin (BSA) solution, i.e., Sb2WO6@NiS-mAb mixture, is fixed on the conjugation pad. 0.5μL of T zone solution and 0.5μL of C zone solution are added to the T zone and C zone, respectively. The sampled test strips are placed in an oven and dried at 37℃ for 60min, and then stored in a vacuum bag for later use.

[0081] Example 4: Working Curve Plotting

[0082] Prepare 10 mL of 1 mg / mL ethinylestradiol standard solution using acetonitrile, and dilute to a concentration of 10 mM PBS. -2 pg / mL, 10 -1 pg / mL, 10 0 pg / mL, 10 1 pg / mL, 10 2 pg / mL, 10 3 pg / mL, 10 4 pg / mL, 10 5 pg / mL, prepared as the test solution. Mix 80 μL of the test solution with 10 μL of Sb2WO6@NiS-mAb and 10 μL of running buffer (10 mM PBS solution containing 5% sucrose, 1% BSA, 1% Tween-20, pH 7.4) in a centrifuge tube. Insert the test strip into the centrifuge tube, remove the test strip after 10 min, and allow it to dry. Then, use a near-infrared LED (power 1.36 W / cm²) to test. 2 Irradiation for 3 minutes was performed, and temperature changes were monitored using a mobile phone and infrared thermal imaging accessories. The results of the photothermal mode are as follows: Figure 4 As shown; the working curve is y = 1.983x + 0.377 (R²). 2 =0.9934).

[0083] The results showed that under photothermal mode, the negative T region had the highest temperature and the ethinylestradiol concentration was 10. 5 The temperature in region T drops to its lowest point at a concentration of pg / mL; this concentration is used as the detection limit for the photothermal mode. Therefore, the detection limit of the constructed dual-mode test strip can reach 10. -1 pg / mL.

[0084] Example 5: Detection of ethinylestradiol

[0085] The specificity of the ethinylestradiol Sb2WO6@NiS complex photothermal quantitative test strip was verified, including the following steps:

[0086] 1. Preparation of test paper materials

[0087] Same as Example 1.

[0088] 2. Preparation of test strips

[0089] Same as Example 3.

[0090] 3. Drawing working curves

[0091] Same as Example 4.

[0092] 4. Specificity

[0093] Prepare 10 mL of 1 mg / mL ethinylestradiol, estradiol, estrone, estriol, and diethylstilbestrol standard solutions using acetonitrile. Dilute these solutions with 10 mM PBS to a concentration of 120 ng / mL and use them as test solutions.

[0094] 5. Sample testing

[0095] Mix 80 μL of the test solution with 10 μL of Sb2WO6@NiS-mAb and 10 μL of running buffer (10 mM PBS solution containing 5% sucrose, 1% BSA, 1% Tween-20, pH 7.4) in a centrifuge tube for 10 min. Then insert the test strip into the centrifuge tube and, after the test strip has dried, use a near-infrared LED (power 1.36 W / cm²) to measure the mixture. 2 Irradiation for 3 minutes was performed, and temperature changes were monitored using a mobile phone and infrared thermal imaging accessories. The results of the photothermal mode are shown as follows: Figure 5 Compared to the blank, estradiol, estrone, estriol, and diethylstilbestrol had very weak effects on the T-zone temperature of the test strip, while ethinylestradiol had a very significant effect on the T-zone temperature of the test strip. This indicates that the test strip has a weak response to the four substances, meaning that the test strip has good specificity.

[0096] As can be seen from the above embodiments, the Sb2WO6@NiS composite material provided in this application, when used in the preparation of photothermal test strips, does not require alteration of the conventional structure of the test strip. It can be used as a photothermal signal probe, either independently of the aforementioned photothermal test strip form for convenient storage, or fixed to the bonding pad of the test strip for easy portability. The detection method provided by this invention, which includes a photothermal test strip for detecting ethinylestradiol, is based on the plasma resonance effect. It uses the high photothermal conversion efficiency and small size of the Sb2WO6@NiS composite material as a T-zone fixed photothermal signal probe, excited by a near-infrared laser source. Combined with a smart terminal and infrared thermal imaging accessories, it achieves photothermal signal acquisition, effectively removing sample liquid base color and test strip fluorescence background interference, thereby improving the signal-to-noise ratio. The detection method has a lower detection limit, a wider detection range, and better specificity, exhibiting higher sensitivity than other photothermal test strips.

[0097] The embodiments provided above are not intended to limit the scope of the invention, nor are the described steps intended to limit the order of execution. Any obvious modifications made to the invention by those skilled in the art based on existing common knowledge also fall within the scope of protection defined by the claims.

Claims

1. A test strip for detecting ethinylestradiol, characterized in that, The test strip includes a photothermal signal probe and a test strip body; The photothermal signal probe is prepared by modifying an ethinylestradiol polyclonal antibody with an Sb2WO6@NiS composite material, and is used either in a form independent of the main structure of the test strip or dried on the conjugate pad. The main body of the test strip includes a PVC base plate, a sample pad, a nitrocellulose membrane, and an absorbent pad; The preparation method of the Sb2WO6@NiS composite material includes the following steps: (1) Add 1 mg of Sb2WO6 nanoparticles to 5 mL of deionized water containing 10 mg of polyethylene glycol, sonicate at 200 W for 200 minutes, and stir for 10 h to obtain a mixture; (2) Add 1 mL of the above mixture to 1 mL of NiS aqueous solution and stir overnight; collect the dark gray powder by centrifugation, dry it, and obtain the composite material; In step (1), the weight-average molecular weight of polyethylene glycol is 7000.

2. A photothermal signal probe prepared from the Sb2WO6@NiS composite material as described in claim 1, characterized in that, It contains Sb2WO6@NiS composite material, ethinylestradiol antibody and bovine serum albumin.

3. The photothermal signal probe according to claim 2, characterized in that, Sb2WO6@NiS composite material was mixed with ethinylestradiol antibody and bovine serum albumin to obtain Sb2WO6@NiS-mAb mixture.

4. A test strip containing the photothermal signal probe of claim 2, characterized in that, The test strip includes a photothermal signal probe and a test strip body; The main body of the test strip comprises: a sample pad, a nitrocellulose membrane, and an absorbent pad, which are sequentially and horizontally overlapped from top to bottom on a PVC base plate; wherein, the nitrocellulose membrane is used to separate and detect analytes from other substances in the sample, the sample pad is used for sample loading, the absorbent pad is used to absorb excess liquid, and the PVC base plate provides physical support for the test strip; the nitrocellulose membrane includes a detection area and a quality control area; wherein, ethinylestradiol antigen is immobilized on the detection area, and the ethinylestradiol antigen is an ethinylestradiol-protein conjugate; a secondary antibody is immobilized on the quality control area, and the secondary antibody is a secondary antibody derived from the primary antibody; The photothermal signal probe can be used independently of the main structure of the test strip, or the photothermal signal probe can be dried on the conjugate pad and the conjugate pad can be inserted between the sample pad and the nitrocellulose membrane.

5. The test strip according to claim 4, characterized in that, Ethinyl estradiol antibodies, or primary antibodies, can be derived from at least one of the following sources: mice, rats, and rabbits.

6. The test strip according to claim 4, characterized in that, The secondary antibody includes at least one of goat anti-mouse secondary antibody, rabbit anti-mouse secondary antibody, goat anti-rabbit secondary antibody, and donkey anti-rabbit secondary antibody.

7. An application of the test strip according to claim 4, characterized in that, Used for the detection of ethinylestradiol; the photothermal temperature of the detection zone is negatively correlated with the content of ethinylestradiol to be measured, and the color intensity is negatively correlated with the content of ethinylestradiol in the sample, specifically: When the sample does not contain ethinylestradiol, the detection area appears purple and the temperature is high; When the sample contains ethinylestradiol, the detection area appears light purple or even disappears, and the temperature decreases. The quality control area, which serves as a reference for verifying the validity of the test strip results, always appears purple.

8. The application according to claim 7, characterized in that, The method for detecting ethinylestradiol using test strips includes the following steps: When the photothermal signal probe is used independently of the main structure of the test strip, it specifically includes: The sample to be tested was mixed with Sb2WO6@NiS-mAb solution in the running buffer, and then the test strip was placed in it. After 10-15 min, the test strip was removed and dried. The photothermal analysis results were then read and quantitative analysis was performed. When the photothermal signal probe is dried on the conjugate pad and the conjugate pad is inserted between the sample pad of the test paper body structure and the nitrocellulose membrane, the specific steps include: Mix the sample to be tested with the running buffer, then place it into the test strip. After 10-15 minutes, remove the strip and let it dry. Read the photothermal analysis results and perform quantitative analysis. The method for reading photothermal analysis results after the test strip is dried is as follows: The test strip is placed under a near-infrared LED light source for excitation, and the temperature result is obtained using a thermal imaging device or temperature measuring device and a smart display terminal. The thermal imaging device or temperature measuring device includes at least one of a mobile phone infrared thermal imaging analysis accessory, an infrared thermal imager, and a temperature gun. The smart display terminal includes at least one of a computer and a smartphone. The thermal imaging device acquires photothermal imaging photos and outputs and displays them through the connected smart display terminal.

Images