Preparation method of an adaptive nano-integrated diagnosis and treatment platform

Abstract

The present invention is applicable to the field of nano-diagnosis and treatment platform technology, and provides a method for preparing an adaptive nano-integrated diagnosis and treatment platform, comprising the following steps: Step 1: preparing Apt-Ag + solution; Step 2: Synthesize GNRs by seed-mediated method; Step 3: Incubate GNRs with Apt-Ag + Solution mixing to prepare pure Apt‑Ag + The prepared adaptive nano-integrated diagnosis and treatment platform visualizes the degree of infection in real time through the concentration of inflammatory factor interferon-γ; it also triggers Ag + Release achieves immediate antibacterial effect; PTT synergistic treatment is activated based on the fluorescence intensity threshold to avoid overtreatment.

Description

Technical Field

[0001] The present invention belongs to the technical field of nano-diagnosis and treatment platforms, and in particular relates to a method for preparing an adaptive nano-integrated diagnosis and treatment platform. Background Art

[0002] Bacterial infections of the oral and maxillofacial area pose a major challenge to modern medicine due to their exposed location, proximity to vital organs, porous tissue, and rich blood supply. If left untreated, the infection can rapidly spread to adjacent vital structures, leading to life-threatening complications such as sepsis. Therefore, early, accurate diagnosis and timely intervention are crucial.

[0003] While existing methods for early diagnosis of maxillofacial infections (such as culture, molecular diagnostics like PCR, immunological tests like ELISA, and mass spectrometry) offer good sensitivity and specificity, their long diagnostic cycles, high costs, and complex procedures limit their application in early, real-time detection. Advances in nanotechnology and biomaterials have also led to the emergence of novel therapies such as chemodynamic therapy, photodynamic therapy, and photothermal therapy (PTT). These approaches, particularly those leveraging the PTT effect of nanomaterials for highly effective localized sterilization, not only demonstrate significant antibacterial efficacy but also reduce reliance on antibiotics and mitigate the risk of drug resistance.

[0004] However, current integrated nano-diagnosis and treatment platforms usually have a split in diagnosis and treatment logic: the diagnostic results require manual intervention to trigger treatment (such as manually turning on the laser after detection), resulting in delayed response and missing the golden treatment window. At the same time, there is a disadvantage of being unable to adjust the treatment method according to the disease condition. Summary of the Invention

[0005] The purpose of the embodiments of the present invention is to provide a method for preparing an adaptive nano-integrated diagnosis and treatment platform, aiming to solve the problems raised in the above-mentioned background technology.

[0006] The embodiment of the present invention is implemented as follows: a method for preparing an adaptive nano-integrated diagnosis and treatment platform comprises the following steps:

[0007] Step 1: Preparation of silver ion loaded aptamer sequence (Apt-Ag + ) solution;

[0008] Step 2: Synthesis of GNRs (gold nanorods) using a seed-mediated method;

[0009] Step 3: Incubate GNR with Apt-Ag + Solution mixing to prepare pure Apt-Ag + -GNR solution.

[0010] Further technical solution, said step 1 comprises the following specific steps:

[0011] Cytosine-rich (C) sequences were designed at both ends of the interferon-γ (IFN-γ) aptamer sequence, and a carboxyfluorescein (FAM) group was connected to one end, i.e., 5`-SH-C6-TTTTTCCCCCC GGGGTTGGTTGTGTTGGTGT CCCCCCCCCC-6-FAM-3' (the italicized part is the IFN-γ aptamer sequence). A 5-base polythymidine spacer was inserted after the 5' thiol to reduce the self-adhesion of the nucleic acid on the GNR surface. After purification by HPLC chromatography, it was dissolved in HPLC-grade water to construct the designed nucleic acid aptamer sequence (Apt) solution. The Apt solution was heated at 85°C for 5 minutes and then quickly cooled in ice. At the same time, Ag was added. + (500 uL, 6 μM) and stirred at room temperature in the dark for 2 hours to obtain Apt-Ag + solution.

[0012] Further technical solution, said step 2 includes the following specific steps:

[0013] Chloroauric acid (HAuCl4, 2.5 mL, 0.001 M) and cetyltrimethylammonium bromide (CTAB, 7.5 mL, 0.2 M) solutions were briefly mixed. Freshly prepared chilled sodium borohydride (NaBH4, 0.6 mL, 0.01 M) was then quickly added with vigorous stirring to form a brownish-yellow colloidal gold seed solution. The solution was stirred vigorously for an additional 2 minutes and allowed to stand at room temperature for 30 minutes before use. CTAB (200 mL, 0.2 M) solution was then gently mixed with HAuCl4 (200 mL, 0.001 M), silver nitrate (AgNO3, 8 mL, 0.004 M), and ascorbic acid (2.8 mL, 0.0778 M). 0.4 mL of the colloidal gold seed solution was then added, and the reaction mixture was left on the bench overnight. The resulting GNR solution was centrifuged (14,500 rpm, 12 min) and resuspended in 2 mL of distilled water.

[0014] Further technical solution, said step 3 includes the following specific steps:

[0015] The GNR solution was centrifuged three times (13000 rpm, 10 min) and then + The solution was mixed and ultrasonicated in a water bath for 30 seconds. It was then placed in a -4°C freezer for 120 minutes. After thawing at room temperature, unreacted oligonucleotides were removed by centrifugation (10,000 g, 10 minutes) and washed three times with PBS buffer (0.03 M, pH 7.4) containing 0.01% SDS. Pure Apt-Ag was finally obtained. + -GNR solution, store at room temperature until use.

[0016] The embodiment of the present invention provides a method for preparing an adaptive nano-integrated diagnosis and treatment platform, wherein the prepared adaptive nano-integrated diagnosis and treatment platform can visualize the infection degree in real time through the concentration of inflammatory factor interferon-γ; synchronously trigger Ag + Release achieves immediate antibacterial effect; PTT synergistic treatment is activated based on the fluorescence intensity threshold to avoid overtreatment. BRIEF DESCRIPTION OF THE DRAWINGS

[0017] Figure 1 A preparation flow chart of a method for preparing an adaptive nano-integrated diagnosis and treatment platform provided by an embodiment of the present invention;

[0018] Figure 2 Apt sequence and Apt-Ag + UV-visible and circular dichroism spectra;

[0019] Figure 3 For GNR and Apt-Ag + -Electron microscopy images, UV-visible spectra, surface potential, and dynamic particle size distribution of GNRs;

[0020] Figure 4 For GNR and Apt-Ag + - Fourier transform infrared spectroscopy and Raman spectroscopy of GNRs;

[0021] Figure 5 Apt-Ag + -Detection performance of the GNR nanoplatform;

[0022] Figure 6 Apt-Ag after LPS stimulation + -GNR detection of fluorescence intensity at different times corresponding to the confocal images, as well as cell level and serum level, Apt-Ag + -Comparison of IFN-g concentration calculated after GNR detection with that of ELISA kit and the corresponding fluorescence intensity;

[0023] Figure 7 The temperature changes, photothermal images, and heating-cooling curves after near-infrared irradiation of different powers are shown;

[0024] Figure 8 The structural change pattern of Apt under photothermal action and the Ag released under different IFN concentrations + concentration and corresponding fluorescence intensity;

[0025] Figure 9 After adding different concentrations of interferon, S. aureus and E. coli CFU statistical graph and corresponding CFU count graph;

[0026] Figure 10 is the OD value of bacterial biofilm after crystal violet staining after different treatments;

[0027] Figure 11 The CFU counts of bacterial biofilms after different treatments and photos of plates coated with CFU;

[0028] Figure 12 Scanning electron microscope images of bacterial biofilms after different treatments;

[0029] Figure 13 Live / dead staining of bacteria after different treatments Figure 3 D modeling images and live / dead bacteria ratio images, as well as changes in biofilm thickness and corresponding thickness statistics;

[0030] Figure 14 The relative fluorescence value statistics corresponding to the fluorescence photos of the backs of the two model animals after injection of AAG;

[0031] Figure 15 To assist with photothermal therapy for severe wounds, we took photos of back wounds at different irradiation times and the corresponding changes in wound temperature.

[0032] Figure 16 The wound change photos and fitting diagrams, the wound area fitting of the mild infection model, and the wound area fitting of the severe infection model are shown;

[0033] Figure 17 CFU statistics and corresponding CFU count chart for severe back wounds;

[0034] Figure 18 Analysis of the results of H&E and Masson staining of wound surfaces of different tissues;

[0035] Figure 19 Immunofluorescence analysis of inflammatory tissues in infected wounds after different treatments. DETAILED DESCRIPTION

[0036] In order to make the purpose, technical solutions and advantages of the present invention more clearly understood, the present invention will be 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.

[0037] The specific implementation of the present invention is described in detail below with reference to specific embodiments.

[0038] like Figure 1 As shown, a method for preparing an adaptive nano-integrated diagnosis and treatment platform provided by one embodiment of the present invention includes the following steps:

[0039] Step 1: Preparation of silver ion loaded aptamer sequence (Apt-Ag + ) solution;

[0040] Cytosine-rich (C) sequences were designed at both ends of the interferon-γ (IFN-γ) aptamer sequence, and a carboxyfluorescein (FAM) group was connected to one end, i.e., 5`-SH-C6-TTTTTCCCCCC GGGGTTGGTTGTGTTGGTGT CCCCCCCCCC-6-FAM-3' (the italicized part is the IFN-γ aptamer sequence, the nucleotide sequence is shown in Table 1). A 5-base polythymidine spacer was inserted after the 5' thiol to reduce the self-adhesion of the nucleic acid on the GNR surface. After purification by HPLC chromatography, it was dissolved in HPLC-grade water to construct the designed nucleic acid aptamer sequence (Apt) solution. The Apt solution was heated at 85°C for 5 minutes and then quickly cooled in ice. At the same time, Ag was added. + (500 uL, 6 μM) and stirred at room temperature in the dark for 2 hours to obtain Apt-Ag + solution.

[0041] Table 1 Nucleotide sequence listing

[0042]

[0043] Step 2: Synthesize GNRs using a seed-mediated method;

[0044] Chloroauric acid (HAuCl4, 2.5 mL, 0.001 M) and cetyltrimethylammonium bromide (CTAB, 7.5 mL, 0.2 M) solutions were briefly mixed. Freshly prepared chilled sodium borohydride (NaBH4, 0.6 mL, 0.01 M) was then quickly added with vigorous stirring to form a brownish-yellow colloidal gold seed solution. The solution was stirred vigorously for an additional 2 minutes and allowed to stand at room temperature for 30 minutes before use. CTAB (200 mL, 0.2 M) solution was then gently mixed with HAuCl4 (200 mL, 0.001 M), silver nitrate (AgNO3, 8 mL, 0.004 M), and ascorbic acid (2.8 mL, 0.0778 M). 0.4 mL of the colloidal gold seed solution was then added, and the reaction mixture was left on the bench overnight. The resulting GNR solution was centrifuged (14,500 rpm, 12 min) and resuspended in 2 mL of distilled water.

[0045] Step 3: After centrifuging the GNR solution for 3 times (13000 rpm, 10 min), +The solution was mixed and ultrasonicated in a water bath for 30 seconds. It was then placed in a -4°C freezer for 120 minutes. After thawing at room temperature, unreacted oligonucleotides were removed by centrifugation (10,000 g, 10 minutes) and washed three times with PBS buffer (0.03 M, pH 7.4) containing 0.01% SDS. Pure Apt-Ag was finally obtained. + -GNR solution, store at room temperature until use.

[0046] In the embodiment of the present invention, the Apt sequence and Apt-Ag + The UV-visible spectrum and circular dichroism spectrum of Figure 2 As shown, Figure 2 a is the Apt sequence and Apt-Ag + The UV-visible spectrum, Figure 2 b is the circular dichroism spectrum of Apt sequence and Apt-Ag+. + -Electron microscope images, UV-visible spectra, surface potential and dynamic particle size distribution of GNRs Figure 3 As shown, Figure 3 a is GNR and Apt-Ag + -Electron microscope images of GNRs (specifically, 1) electron microscope images of GNRs, 2) electron microscope images of Apt-Ag + -electron microscope image of GNRs), Figure 2 b is GNR and Apt-Ag + -UV-visible spectrum of GNR, Figure 3 c is GNR and Apt-Ag + -Surface potential of GNR, Figure 3 d is GNR and Apt-Ag + -Dynamic particle size distribution of GNR. GNR and Apt-Ag + -Fourier transform infrared and Raman spectra of GNRs Figure 4 As shown, Figure 4 a is GNR and Apt-Ag + - Fourier transform infrared spectroscopy of GNRs, Figure 4 b is GNR and Apt-Ag + -Raman spectrum of GNRs.

[0047] The following is the prepared Apt-Ag + -GNR conducted various performance tests, specifically:

[0048] (1) Detection performance:

[0049] The excitation wavelength / emission wavelength of the fluorescent group FAM is 417 nm / 521 nm. In order to determine the time required for fluorescence recovery, the fluorescence change was detected by a fluorescence spectrometer every 10 minutes. + After adding different concentrations of IFN-g to -GNR, the relationship between different concentrations and fluorescence intensity (FL) was explored, and a linear fitting curve was drawn based on the fluorescence value at 521 nm. Figure 5 As shown (where Figure 5 a is a diagram showing the structural changes after Apt recognizes IFN-g. Figure 5 b is the fluorescence recovery time after adding IFN-g, Figure 5 c is the different fluorescence intensities corresponding to different concentrations of IFN-g, Figure 5 d is the fitting diagram of the change of fluorescence intensity with IFN-g concentration at the cellular level). + - In the GNR nanoplatform, after the Apt structure recognizes IFN-g, due to local structural rearrangement and Ag + Upon release, the stem-loop structure undergoes conformational changes, and the FAM fluorophore at the 3' end recovers its fluorescence. + After adding IFN-g to the -GNR solution, fluorescence intensity was measured every 10 minutes. A significant fluorescence change was observed within 30 minutes, after which the rate of change slowed. Fluorescence intensity at 521 nm was measured after the same incubation time using different IFN-g concentrations. The results showed that fluorescence gradually recovered with increasing IFN-g concentration, following a linear relationship, as shown by the equation y = 0.90x + 244.91.

[0050] like Figure 6 As shown (where Figure 6 a is Apt-Ag after LPS stimulation + -Confocal images of fluorescence intensity corresponding to GNR detection at different times, Figure 6 b is the cellular level, Apt-Ag + - Comparison of IFN-g concentration calculated after GNR detection with that of ELISA kit and the corresponding fluorescence intensity, Figure 6 c is the serum level of Apt-Ag +-Comparison of IFN-g concentration calculated after GNR detection with ELISA kit and corresponding fluorescence intensity). At the cellular level, confocal images showed significant changes in fluorescence intensity after different stimulation times and changes in cell morphology after LPS stimulation. At the same time, the fluorescence intensity was measured, and the IFN-g concentration was calculated using the above linear equation, and the results were compared with the results of the IFN-g ELISA kit. The results showed that the fluorescence of the nanoplatform continued to increase, proving that the IFN-g concentration increased with the extension of the stimulation time, which was consistent with the results of ELISA. Further, the concentration of IFN-g in blood samples from inflamed mice was detected. Analysis showed that the IFN-g concentration was consistent with the detection results of ELISA. This demonstrates that the platform has the ability to detect pro-inflammatory signals in complex environments and the potential for subsequent therapeutic applications.

[0051] (2) Therapeutic properties:

[0052] The power density at 808 nm (1.0, 1.5, 2.0 W·cm -2 ) laser irradiation Apt-Ag + -GNR solution (50 μg mL -1 , 5 min), and the temperature changes were recorded by using an infrared thermal imager. In addition, at 1.0 W·cm -2 Apt-Ag was obtained under the following conditions + Finally, different 808 nm power densities (1.0, 1.5, 2.0 W·cm -2 ) After laser irradiation for 5 min, fluorescence changes were measured every 60 s using a fluorescence spectrometer. Figure 7 As shown (where Figure 7 a is the corresponding temperature change after irradiation with near-infrared rays of different powers, Figure 7 b is the photothermal image. Figure 7 c is the heating-cooling curve). As the laser power increases, the Apt-Ag + -GNRs showed a significant temperature rise in a short time. These results indicate that Apt-Ag + -GNRs exhibited good light-to-heat conversion performance. Moreover, the heating and cooling cycles showed stable temperature fluctuations over four consecutive on / off time intervals.

[0053] Rapid temperature fluctuations can also destroy C-Ag + -C structure, causing the change of Apt structure, thus changing the fluorescence signal, Ag + Release. At 50ug mL -1 Apt-Ag + 400, 600, 800, and 1000 pg·mL were added to the GNR solution.-1 IFN-g, and NIR (808 nm, 1.0 W·cm -2 After 30 minutes of incubation, the supernatant was taken for ICP analysis to measure Ag + At the same time, a fluorescence spectrometer is used to detect the corresponding fluorescence changes. Figure 8 As shown (where Figure 8 a is the structural change pattern of Apt under the action of light and heat, Figure 8 b Ag released under different IFN concentrations + concentration and corresponding fluorescence intensity), the results showed that: with the increase of IFN-g concentration, Ag + In contrast, when IFN-g was not added, the release of Ag + The concentration is still very low and almost undetectable. In addition, under NIR irradiation, Ag is observed + Released in large quantities, reaching 0.4 mg·L -1 Above, the fluorescence intensity increased significantly to 1824 au.

[0054] (3) In vitro antibacterial effect:

[0055] This nano-integrated diagnostic and therapeutic platform can release silver ions while recognizing interferon in vitro and produce a therapeutic effect instantly. However, severe bacterial infection can cause severe inflammation in the body. Therefore, in order to improve the efficiency of treatment, NIR is used in conjunction with PTT to achieve efficient sterilization when the inflammation intensity is obvious. The results show that AAG has an adaptive antibacterial effect. 1000pg·mL -1 The concentration of IFN-g was increased, and AAG reduced the concentration of bacterial solution to about 105, which proved that it had certain antibacterial ability, but it could be further enhanced. That is to say, in the case of severe infection, + On this basis, the PTT of GNR can be further coordinated to reduce the bacterial concentration to about 6 log, so as to enhance the antibacterial ability and shorten the antibacterial time.

[0056] The sterilization effect is shown in the figure, where Figure 9 After adding different concentrations of interferon, S. aureus and E. coli CFU statistics and corresponding CFU counts (where, Figure 9 a is S. aureus CFU statistics chart, Figure 9 b is the corresponding CFU count graph; Figure 9 c is E. coli CFU statistics chart, Figure 9 d is the corresponding CFU count graph). Figure 10OD values ​​of bacterial biofilms after crystal violet staining after different treatments (the vertical illustration on the right is the crystal violet staining image after 24 hours). Figure 11 The CFU counts of bacterial biofilms after different treatments and the photos of plates coated with CFU (where Figure 11 a and Figure 11 c is the CFU count of bacterial biofilm after different treatments, Figure 11 b and Figure 11 d is the corresponding photograph of the plate coated with CFU). Figure 12 Scanning electron microscope images of bacterial biofilms after different treatments. Figure 13 Live / dead staining of bacteria after different treatments Figure 3 3D modeling pictures and live / dead bacteria ratio pictures, as well as the changes in biofilm thickness and corresponding thickness statistics (wherein, Figure 13 a is the live / dead staining of bacteria after different treatments Figure 3 3D modeling pictures and live / dead bacteria ratio pictures, Figure 13 b is the change of biofilm thickness, Figure 13 c is the corresponding thickness statistical graph).

[0057] (4) Antibacterial effect in vivo:

[0058] Mouse back wound models with different infection degrees were established to compare the therapeutic effects. Figure 14 The relative fluorescence value statistics corresponding to the fluorescence photos of the backs of the two model animals after injection of AAG (where Figure 14 a is a fluorescent photo of the backs of the two model animals after injection of AAG. Figure 14 b is the corresponding relative fluorescence value statistics), Figure 15 In order to assist with photothermal therapy for severe wounds, the back wounds were treated with photothermal images at different irradiation times and the corresponding changes in wound temperature (among which, Figure 15 a is a photothermal treatment for severe wounds, with pictures of back wounds taken at different irradiation times. Figure 15 b is the light and heat change of the corresponding wound temperature). Figure 16 The wound change photos and fitting diagrams, the wound area fitting of the mild infection model and the wound area fitting of the severe infection model (wherein, Figure 16 a is the wound change photo and fitting diagram, Figure 16 b is the wound area fitting of the mild infection model, Figure 16 c is the wound area fitting of the severe infection model). Figure 17 The CFU statistics of the back wound of severe wound and the corresponding CFU count chart (wherein, Figure 17 a is the CFU statistics of the back wound of severe wound, Figure 17 b is the corresponding CFU count graph).

[0059] The results showed that after the model was established, the fluorescence intensity displayed by the material was used to distinguish between mild and severe cases, and different treatment methods were selected according to the diagnosis results. For mild models, the silver ions adaptively released by the AAG platform can produce a good antibacterial effect. For severe cases, Ag + Synergistic photothermal therapy can greatly improve treatment efficiency and shorten treatment time.

[0060] Furthermore, H&E and Masson staining were performed on the wound surfaces of different tissues to observe and compare the number of inflammatory cells and the recovery of collagen fibers. Figure 18 Analysis of the results of H&E and Masson staining of wound surfaces of different tissues (including Figure 18 a is H&E staining and Masson analysis of infected wounds after different treatments. Figure 18 b is the inflammatory cell counts in the mildly infected wounds in different treatment groups. Figure 18 c is the statistics of collagen content in different treatment groups in mildly infected wounds, Figure 18 d is the inflammatory cell counts in severely infected wounds of different treatment groups, Figure 18 e is the statistics of collagen content in different treatment groups in severely infected wounds).

[0061] Furthermore, CD31 was used as an anti-inflammatory factor and TNF-a as a pro-inflammatory factor to determine the recovery of inflammatory tissue at the wound. Figure 19 As shown (where Figure 19 a is the immunofluorescence confocal level image of inflammatory factors in infected wounds after different treatments. Figure 19 b is the CD31 fluorescence signal intensity of different treatment groups in mildly infected wounds. Figure 19 c is the CD31 fluorescence signal intensity of different treatment groups in severely infected wounds, Figure 19 d is the fluorescence signal intensity of TNF-a in different treatment groups in mildly infected wounds, Figure 19 e is the fluorescence signal intensity of TNF-a in different treatment groups in severely infected wounds).

[0062] The results showed that significant tissue healing and reduced inflammation were observed in the treatment groups of both models, demonstrating the good therapeutic effect of the platform.

[0063] The comparison of this method with other technologies is shown in Table 1 below.

[0064] Table 1

[0065]

[0066] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions and improvements made within the spirit and principles of the present invention should be included in the scope of protection of the present invention.

Claims

1. A method for preparing an adaptive nano-integrated diagnosis and treatment platform, characterized in that: The following steps are involved: Step 1: Preparation of Apt-Ag + solution; Step 2: Synthesize gold nanorods (GNRs) using a seed-mediated method; Step 3: Incubate GNR with Apt-Ag + Solution mixing to prepare pure Apt-Ag + -GNR solution; The step 1 includes the following specific steps: Cytosine-rich sequences were designed at both ends of the IFN-γ aptamer sequence, and a 6-carboxyfluorescein FAM group was connected to one end, i.e., 5′-SH-C6-TTTTTCCCCCC GGGGTTGGTTGTGTTGGTGT CCCCCCCCCC-6-FAM-3', a 5-base polythymidine spacer was inserted after the 5'thiol to reduce the self-adhesion of nucleic acids on the GNR surface; after purification by HPLC chromatography, it was dissolved in HPLC-grade water to construct the designed Apt solution, which was heated at 85°C for 5 minutes and then quickly cooled in ice. At the same time, 500 uL of 6 μM Ag was added. + , and stirred at room temperature in the dark for 2 hours to obtain Apt-Ag + solution.

2. The method for preparing the adaptive nano-integrated diagnosis and treatment platform according to claim 1, characterized in that: The step 2 includes the following specific steps: 2.5 mL of 0.001 M HAuCl4 was briefly mixed with 7.5 mL of 0.2 M cetyltrimethylammonium bromide (CTAB) solution, and then 0.6 mL of freshly prepared 0.01 M NaBH4 was quickly added under vigorous stirring to form a brown-yellow colloidal gold seed solution; The solution was stirred vigorously for another 2 minutes and allowed to stand at room temperature for 30 minutes before use. 200 mL of 0.2 M CTAB solution was gently mixed with 200 mL of 0.001 M HAuCl4, 8 mL of 0.004 M AgNO3, and 2.8 mL of 0.0778 M ascorbic acid. 0.4 mL of the above colloidal gold seed solution was then added, and the reaction mixture was left on the laboratory bench overnight. The resulting GNR solution was centrifuged at 14,500 rpm for 12 minutes and finally resuspended in 2 mL of distilled water.

3. The method for preparing the adaptive nano-integrated diagnosis and treatment platform according to claim 2, characterized in that: The step 3 includes the following specific steps: The GNR solution was centrifuged at 13000 rpm for 10 min, repeated 3 times, and then mixed with Apt-Ag + The solution was mixed and ultrasonicated in a water bath for 30 seconds; then it was placed in a -4°C refrigerator for 120 minutes. After thawing at room temperature, it was centrifuged at 10,000 g for 10 minutes to remove unreacted oligonucleotides and washed three times with 0.03 M PBS buffer containing 0.01% SDS to obtain pure Apt-Ag. + -GNR solution, store at room temperature until use.

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