A Raman spectroscopy-based detection technology for IDH mutations in gliomas

Abstract

This invention discloses a Raman spectroscopy-based detection technology for glioma IDH mutations, belonging to the field of tumor diagnosis. This invention prepares a core-shell structure based on gold nanoparticles and UiO-66, utilizing... N, N '-Di(2-aminoethyl)-1,3-propanediamine was used to modify the Raman signal of SERS-enhanced samples. Ratio analysis was then used to determine the presence of IDH mutations in gliomas, establishing a detection protocol for IDH mutations in glioma cells. This facilitates rapid intraoperative molecular pathology identification and provides effective guidance for surgical and subsequent treatment selection. The detection technology disclosed in this invention can obtain results within 2 minutes, greatly improving clinical diagnostic efficiency and enabling intraoperative molecular pathology diagnosis, which has significant clinical implications.

Description

Technical Field

[0001] This invention belongs to the field of tumor diagnosis and relates to a glioma IDH mutation detection technology based on Raman spectroscopy. It further relates to a method for rapidly detecting glioma homogenate samples using a surface-enhanced Raman scattering (SERS) substrate, combined with ratio analysis to determine whether they have IDH mutations. Background Technology

[0002] Gliomas are the most common malignant tumors of the central nervous system. High-grade gliomas have a median survival of only 15 months. While low-grade gliomas generally have a better prognosis, they are incurable and prone to malignant mutations after recurrence. IDH mutations are common in gliomas and significantly impact drug resistance and prognosis, thus becoming a crucial factor in treatment selection. The World Health Organization (WHO) classification of tumors of the central nervous system recommends using PCR, in situ hybridization, or NGS sequencing to detect molecular pathological markers in tumor samples, guiding surgical resection extent, precise intraoperative personalized antitumor therapy, and ultra-early postoperative adjuvant therapy. However, these techniques generally require completion 7-14 working days post-surgery, are costly, and complex, failing to meet the clinical needs of early perioperative treatment. With the advent of precision diagnostics, the field of molecular pathological diagnosis of gliomas urgently needs precise, rapid, and efficient detection technologies to guide the selection of comprehensive postoperative treatment strategies.

[0003] Surface-enhanced Raman spectroscopy (SERS) is a highly promising analytical tool with high sensitivity and non-destructive testing advantages, providing specific fingerprints of analytes. Currently, some studies have demonstrated the excellent performance of SERS technology in glioma detection. For example, Panda et al. fabricated a three-dimensional nested structure based on gold nanoparticles and L-DOPA microtubules, which can be used for in situ diagnosis of gliomas (ACS Appl. Nano Mater. 2019, 2, 5, 2663–2678). Sara Mahshid et al. reported a nanostructured microfluidic device that can use SERS technology to identify extracellular vesicles from different glioma cell subpopulations (Lab Chip, 2021, 21, 855-866). Although these technologies have made some contributions to glioma identification, they cannot provide molecular pathological information of gliomas, accurately diagnose gliomas, or guide postoperative comprehensive treatment. Furthermore, research using SERS technology to determine IDH mutations is very scarce, and no relevant literature has been found. Summary of the Invention

[0004] To address the aforementioned problems, this invention provides a Raman spectroscopy-based detection technology for glioma IDH mutations. By utilizing SERS-based substrate-enhanced Raman signals, a detection scheme for glioma cell IDH mutations is established, providing a reference for the selection of postoperative treatment plans.

[0005] To achieve the above objectives, the present invention provides the following technical solution.

[0006] This invention provides a method for preparing a SERS-enhanced substrate, characterized by the following specific steps:

[0007] S1. Preparation of gold nanoparticles: Heat deionized water under stirring, and after boiling, add 1% chloroauric acid solution; while continuing to boil, add 4% trisodium citrate solution, react for half an hour, and cool to room temperature under stirring; gold nanoparticles are obtained, centrifuged and washed three times at 7000 rpm, then resuspended in deionized water and stored at 4℃ for later use.

[0008] S2. Core-shell structure preparation: 70% zirconium propoxide, N,N An intermediate was obtained by reacting a mixture of dimethylformamide and glacial acetic acid at 130°C for 2 hours. Subsequently, the intermediate was mixed with gold nanoparticles prepared in S1 and 1,4-benzenedicarboxylic acid at room temperature. After thorough mixing, zinc nitrate was added, and after 18 hours, a zirconium-gold composite structure was obtained. A PVDF membrane with a size of 1 cm × 1 cm was treated with oxygen isoparticles for 0.5 hours, washed, and then incubated with a 2% polyacetylimide aqueous solution for 1 hour. After thorough washing with deionized water, it was placed in a zirconium-gold composite structure solution at 100°C and reacted for 24 hours to form a monolayer film on the surface, thus obtaining the SERS substrate.

[0009] S3, Utilization N,N The SERS substrate prepared in S2 was further modified with '-di(2-aminoethyl)-1,3-propanediamine: a 20% glycerol-ethanol solution was prepared to dissolve... N,N '-Di(2-aminoethyl)-1,3-propanediamine molecule, after placing the SERS substrate prepared in S2 into it for 1-2 hours, rinsed with ethanol and deionized water successively, dried with nitrogen and stored in a refrigerator at 4°C, to obtain the ERS-enhanced substrate.

[0010] The present invention also provides an application of the SERS-enhanced substrate obtained by the preparation method described above in the preparation of products for detecting IDH mutations in gliomas.

[0011] Furthermore, the product detects IDH mutations in glioma homogenate samples.

[0012] Further, the prepared sample homogenate was dropped onto the SERS-enhanced substrate and mixed. After incubation for 30-60 seconds, SERS detection was performed using a laser with a wavelength of 785 nm and an integration time of 10 seconds. Before spectral acquisition, a 520 cm⁻¹ silicon chip was used. -1 The spectrometer was calibrated using Raman spectral bands; the bands at 512 and 1526 cm⁻¹ were determined in the IDH mutation samples. -1 A strong characteristic peak appeared at the point; by combining the ratio analysis model, the judgment model can be obtained to determine whether there is a mutation in the sample to be tested.

[0013] Furthermore, the judgment model: ,in, I 512 The characteristic peak is at 512 cm⁻¹. -1 Strength at that location, I 1003 The characteristic peak is at 1003 cm⁻¹. -1 Strength at that location, I 1526 The characteristic peak is at 1526 cm⁻¹. -1 The intensity at the location; set the threshold to 1. When R is greater than the threshold, the sample is predicted as mutant, and when R is less than the threshold, the sample is predicted as wild.

[0014] This invention also provides a method for detecting IDH mutations in gliomas based on Raman spectroscopy, characterized by comprising the following steps:

[0015] Step 1, Preparation of SERS-enhanced substrate:

[0016] S1. Preparation of gold nanoparticles: Heat deionized water under stirring, and after boiling, add 1% chloroauric acid solution; while continuing to boil, add 4% trisodium citrate solution, react for half an hour, and cool to room temperature under stirring; gold nanoparticles are obtained, centrifuged and washed three times at 7000 rpm, then resuspended in deionized water and stored at 4℃ for later use.

[0017] S2. Core-shell structure preparation: 70% zirconium propoxide, N,N An intermediate was obtained by reacting a mixture of dimethylformamide and glacial acetic acid at 130°C for 2 hours. Subsequently, the intermediate was mixed with gold nanoparticles prepared in S1 and 1,4-benzenedicarboxylic acid at room temperature. After thorough mixing, zinc nitrate was added, and after 18 hours, a zirconium-gold composite structure was obtained. A PVDF membrane with a size of 1 cm × 1 cm was treated with oxygen isoparticles for 0.5 hours, washed, and then incubated with a 2% polyacetylimide aqueous solution for 1 hour. After thorough washing with deionized water, it was placed in a zirconium-gold composite structure solution at 100°C and reacted for 24 hours to form a monolayer film on the surface, thus obtaining the SERS substrate.

[0018] S3, Utilization N,NThe SERS substrate prepared in S2 was further modified with '-di(2-aminoethyl)-1,3-propanediamine: a 20% glycerol-ethanol solution was prepared to dissolve... N,N '-Di(2-aminoethyl)-1,3-propanediamine molecule, after placing the SERS substrate prepared in S2 into it for 1-2 hours, rinsed with ethanol and deionized water successively, dried with nitrogen and stored in a refrigerator at 4°C to obtain the ERS-enhanced substrate;

[0019] Step 2, Glioma sample preparation: Add 0.86% ice-cold saline to the obtained glioma sample according to the ratio, and homogenize the sample using a homogenizer until the glioma sample is a white and uniform solution.

[0020] Step 3: Detection using a Raman spectrometer: The homogenate obtained in Step 2 is dropped onto the SERS-enhanced substrate obtained in Step 1, mixed, and incubated for 30-60 seconds. SERS detection is then performed using a laser with a wavelength of 785 nm and an integration time of 10 seconds. Before spectral acquisition, a 520 cm⁻¹ laser on a silicon chip is used. -1 The spectrometer was calibrated using Raman spectral bands; the bands at 512 and 1526 cm⁻¹ were determined in the IDH mutation samples. -1 A strong characteristic peak appeared in the sample, while it was barely visible in the wild-type sample.

[0021] Furthermore, the ratio of the gel sample to ice-cold saline in step 2 is 1g:9mL.

[0022] Furthermore, the step also includes: obtaining a judgment model using a ratio analysis model, wherein the judgment model... ,in, I 512 The characteristic peak is at 512 cm⁻¹. -1 Strength at that location, I 1003 The characteristic peak is at 1003 cm⁻¹. -1 Strength at that location, I 1526 The characteristic peak is at 1526 cm⁻¹. -1 The intensity at the location; set the threshold to 1. When R is greater than the threshold, the sample is predicted as mutant, and when R is less than the threshold, the sample is predicted as wild.

[0023] The present invention also provides an electronic device, including a memory and a processor, wherein the memory stores a computer program executable on the processor, characterized in that the processor, when executing the program, implements the steps in any of the above-described methods for detecting IDH mutations in gliomas based on Raman spectroscopy.

[0024] The present invention also provides a computer-readable storage medium having a computer program stored thereon, characterized in that, when the computer program is executed by a processor, it implements the steps in any of the above-described methods for detecting IDH mutations in gliomas based on Raman spectroscopy.

[0025] The beneficial effects of the present invention compared with the prior art.

[0026] This invention proposes a method to predict whether gliomas in human brain tissue have IDH mutations, by utilizing preferred Raman substrate materials, characteristic spectra, and classification methods.

[0027] The technical solution provided in this application fully utilizes the advantages of Raman spectroscopy liquid phase detection, high sensitivity, and fingerprint spectrum detection, which helps to quickly identify intraoperative molecular pathology and provides effective guidance for the selection of surgical and subsequent treatment plans.

[0028] This technology can provide results within 2 minutes, greatly improving the efficiency of clinical diagnosis and enabling intraoperative molecular pathological diagnosis.

[0029] This invention prepares a core-shell structure based on gold nanoparticles and UiO-66, utilizing... N,N After modification with '-di(2-aminoethyl)-1,3-propanediamine, the presence or absence of IDH mutations in gliomas was determined using ratio analysis, thus providing a reference for the selection of clinical treatment options. This method has the advantages of being rapid, accurate, and highly sensitive. Attached Figure Description

[0030] Figure 1 Scanning electron microscope image of the SERS substrate.

[0031] Figure 2 Signal comparison of IDH mutant samples before and after SERS substrate modification.

[0032] Figure 3 SERS profiles and differentially expressed sites of glioma samples with IDH mutations and IDH wild-type.

[0033] Figure 4 SERS maps and differentially expressed sites of IDH-mutant and IDH-wild-type glioma samples and glioma cells.

[0034] Figure 5 Tissue sample prediction results (1 represents IDH mutant, 0 represents IDH wild type).

[0035] Figure 6 Cell sample prediction results (1 represents IDH mutant, 0 represents IDH wild type). Detailed Implementation

[0036] The following embodiments will help to understand the present invention, but these embodiments are only for illustrative purposes and the present invention is not limited thereto. The operating methods in the embodiments are all conventional operating methods in this technical field.

[0037] Example 1: Preparation of SERS-enhanced substrate.

[0038] Preparation of gold nanoparticles. 200 mL of deionized water was heated with stirring until boiling. 8 mL of 1% (w / v) chloroauric acid solution was added. While continuing to boil, 4 mL of 4% (w / v) trisodium citrate solution was added. The reaction was allowed to proceed for half an hour, then cooled to room temperature with stirring to obtain 15 nm gold nanoparticles. These nanoparticles were centrifuged at 7000 rpm for 10 min, washed three times, resuspended in deionized water, and stored at 4 °C for later use.

[0039] Core-shell structure preparation. 100 μL of 70% zirconium n-propoxide (dissolved in n-propanol) and 10 mL of... N,N A zirconium-gold composite structure was obtained by reacting a mixture of dimethylformamide and 4 mL of glacial acetic acid at 130 °C for 2 h. Subsequently, 3 mL of the intermediate was mixed with 1.5 mL of gold nanoparticles and 0.1 g of 1,4-benzenedicarboxylic acid at room temperature and stirred thoroughly. After thorough mixing, 0.6 g of zinc nitrate was added, and the reaction proceeded for 18 h to obtain a zirconium-gold composite structure. A 1 cm × 1 cm PVDF membrane was treated with oxygen isoparticles for 0.5 h, washed, and then incubated with a 2% polyacetylimide aqueous solution for 1 h. After thorough washing with deionized water, the membrane was placed in a zirconium-gold composite structure solution at 100 °C for 24 h, forming a monolayer film on the surface, thus obtaining the SERS substrate. To improve the detection performance of the SERS substrate for IDH mutations (e.g., ... Figure 2 ),use N,N The SERS substrate was further modified with '-di(2-aminoethyl)-1,3-propanediamine. A 20% glycerol-ethanol solution was prepared to dissolve the modifying molecule, and a 10 mM modification solution was prepared. The substrate was immersed in this solution for 1–2 hours, followed by rinsing with ethanol and deionized water, drying under nitrogen, and storing at 4°C. The scanning electron microscopy results of the SERS substrate are shown below. Figure 1 As shown.

[0040] Example 2: Preparation of glioma samples.

[0041] The glioma sample and glioma cell sample (cell line from clinical patients, the name of the cell line provided by the hospital is GSC4) obtained from clinical surgery were weighed at about 10mg and added to 0.86% ice-cold physiological saline at a ratio of 1g:9mL. The sample was homogenized using a homogenizer with the parameters set as follows: 80% power, 10s of sonication, and 10s interval. The sample was then sonicated until it became a white homogeneous solution.

[0042] The obtained homogenate was dropped onto the SERS-enhanced substrate and stirred with a 1.0 mL pipette tip to distribute it evenly. The liquid layer thickness was about 50 μm. After incubation for 30 to 60 seconds, Raman detection was performed.

[0043] The weight of the glioma sample is sufficient to obtain a adequate SERS signal. The pipette tip is used to thoroughly mix the sample to ensure the signal is as uniform as possible, and the liquid layer thickness is reduced to minimize Raman signal loss caused by sample accumulation. Incubation for 30-60 seconds is the experimentally validated reaction time between the SERS substrate and the homogenized sample; excessive incubation time can lead to significant sample evaporation and alter the original signal. Compared to PBS and ultrapure water, ice-cold physiological saline (8.6g sodium chloride dissolved in 1L ultrapure water and pre-cooled to 4°C) ensures greater sample stability without affecting the SERS signal.

[0044] Example 3: Detection of multidrug-resistant tumor cells.

[0045] In Example 2, the incubated cells were analyzed using Raman spectroscopy. The instrument used was a RenishawinVia confocal Raman spectrometer with a laser wavelength of 785 nm, a maximum excitation power of 14 mW, and an integration time of 10 s. Before spectral acquisition, a 520 cm⁻¹ silicon chip was used. -1 The spectrometer was calibrated using the Raman spectral bands. At least three random points were selected on each sample for scanning to obtain the average Raman signal (e.g., ...). Figure 3 and 4 (As shown).

[0046] The selected wavelengths were those with low noise levels during detection, while the power was chosen to ensure SERS results were obtained without damaging cells. Experiments verified that SERS results were obtained at 512 and 1526 cm⁻¹ in IDH-mutant samples. -1 A strong characteristic peak appeared at this location, while it was barely visible in the wild-type sample. The stable 1003 cm⁻¹... -1 The intensity of the characteristic peak at 1526 cm⁻¹ was used as an internal reference to establish the values ​​at 1003 cm⁻¹. -1 Peak intensity ratio model can efficiently identify IDH mutations.

[0047] Specifically as follows:

[0048] Raman signals were acquired from the samples prepared in Example 2. The spectra were preprocessed using baseline estimation and denoising based on sparsity, and statistical prediction was performed using a ratio analysis model. The prediction formula is as follows:

[0049] .

[0050] in, I 512 The characteristic peak is at 512 cm⁻¹. -1 Strength at that location,I 1003 The characteristic peak is at 1003 cm⁻¹. -1 Strength at that location, I 1526 The characteristic peak is at 1526 cm⁻¹. -1 The intensity at a given point. With a threshold of 1, when R is greater than the threshold, the sample is predicted as mutant; when R is less than the threshold, the sample is predicted as wild-type.

[0051] The predicted results were compared with the results of tumor mutation determination obtained from DNA sequencing. The results showed that the predicted values ​​corresponded 100% with the actual values.

[0052] Example 4.

[0053] The specific embodiments of the present invention will be described in detail below with reference to the accompanying drawings. It should be understood that the specific embodiments described herein are for illustration and explanation only and are not intended to limit the present invention.

[0054] 1. Prepare the Raman substrate as described above.

[0055] 2. Glioma tissue samples were homogenized by ultrasound and centrifuged to obtain the supernatant.

[0056] 3. Raman signal acquisition was performed on the samples. The spectra were preprocessed using sparsity-based baseline estimation and denoising, and statistical prediction was performed using a ratio analysis model. The prediction formula is as follows:

[0057] .

[0058] Among them, I512 is the characteristic peak at 512 cm⁻¹. -1 The intensity at 1003 cm⁻¹ is a characteristic peak. -1 The intensity at 1526 cm⁻¹ is characteristic of I1526. -1 The intensity at a given point. With a threshold of 1, when R is greater than the threshold, the sample is predicted as mutant; when R is less than the threshold, the sample is predicted as wild-type.

[0059] 4. Based on the obtained prediction results, a comparison was made with the results of DNA sequencing to determine whether tumor mutations were present. The results showed that the predicted values ​​and actual values ​​corresponded 100% completely (e.g., ...). Figure 5 ).

[0060] Example 5.

[0061] 1. Prepare the Raman substrate as described above.

[0062] 2. Glioma cell samples were homogenized by ultrasound and the supernatant was obtained by centrifugation.

[0063] 3. Raman signal acquisition was performed on the samples. The spectra were preprocessed using sparsity-based baseline estimation and denoising, and statistical prediction was performed using a ratio analysis model. The prediction formula is as follows:

[0064] .

[0065] Among them, I512 is the characteristic peak at 512 cm⁻¹. -1 The intensity at 1003 cm⁻¹ is a characteristic peak. -1 The intensity at 1526 cm⁻¹ is characteristic of I1526. -1 The intensity at a given point. With a threshold of 1, when R is greater than the threshold, the sample is predicted as mutant; when R is less than the threshold, the sample is predicted as wild-type.

[0066] Based on the obtained prediction results, a comparison was made with the results of DNA sequencing to determine whether tumor mutations were present. The results showed that the predicted values ​​and actual values ​​corresponded 100% completely (e.g., ...). Figure 6 ).

Claims

1. The application of a SERS-enhanced substrate in the preparation of products for detecting IDH mutations in gliomas, characterized in that, The specific steps for preparing SERS-enhanced substrates are as follows: S1. Preparation of gold nanoparticles: Heat deionized water under stirring, and after boiling, add 1% chloroauric acid solution; while continuing to boil, add 4% trisodium citrate solution, react for half an hour, and cool to room temperature under stirring; gold nanoparticles are obtained, centrifuged and washed three times at 7000 rpm, then resuspended in deionized water and stored at 4℃ for later use. S2. Core-shell structure preparation: 70% zirconium propoxide, N,N An intermediate was obtained by reacting a mixture of dimethylformamide and glacial acetic acid at 130°C for 2 hours. Subsequently, the intermediate was mixed with gold nanoparticles prepared in S1 and 1,4-benzenedicarboxylic acid at room temperature. After thorough mixing, zinc nitrate was added, and after 18 hours, a zirconium-gold composite structure was obtained. A PVDF membrane with a size of 1 cm × 1 cm was treated with oxygen plasma for 0.5 hours, washed, and then incubated with a 2% polyacetylimide aqueous solution for 1 hour. After thorough washing with deionized water, it was placed in a zirconium-gold composite structure solution at 100°C and reacted for 24 hours to form a monolayer film on the surface, thus obtaining the SERS substrate. S3, Utilization N,N The SERS substrate prepared in S2 was further modified with '-di(2-aminoethyl)-1,3-propanediamine: a 20% glycerol-ethanol solution was prepared to dissolve... N,N '-Di(2-aminoethyl)-1,3-propanediamine molecule, after placing the SERS substrate prepared in S2 into it for 1-2 hours, rinsed with ethanol and deionized water successively, dried with nitrogen and stored in a refrigerator at 4°C, to obtain the SERS-enhanced substrate.

2. The application according to claim 1, characterized in that, The product is used to detect IDH mutations in glioma homogenate samples.

3. The application according to claim 2, characterized in that, Preparation of glioma homogenate samples: The obtained glioma samples were mixed with 0.86% ice-cold saline according to the specified ratio, and homogenized using a homogenizer until the glioma samples were a white and homogeneous solution, thus obtaining the homogenate.

4. An electronic device comprising a memory and a processor, the memory storing a computer program executable on the processor, characterized in that, When the processor executes the program, it performs the steps in the following method: Step 1, Preparation of SERS-enhanced substrate: S1. Preparation of gold nanoparticles: Heat deionized water under stirring, and after boiling, add 1% chloroauric acid solution; while continuing to boil, add 4% trisodium citrate solution, react for half an hour, and cool to room temperature under stirring; gold nanoparticles are obtained, centrifuged and washed three times at 7000 rpm, then resuspended in deionized water and stored at 4℃ for later use. S2. Core-shell structure preparation: 70% zirconium propoxide, N,N An intermediate was obtained by reacting a mixture of dimethylformamide and glacial acetic acid at 130°C for 2 hours. Subsequently, the intermediate was mixed with gold nanoparticles prepared in S1 and 1,4-benzenedicarboxylic acid at room temperature. After thorough mixing, zinc nitrate was added, and after 18 hours, a zirconium-gold composite structure was obtained. A PVDF membrane with a size of 1 cm × 1 cm was treated with oxygen plasma for 0.5 hours, washed, and then incubated with a 2% polyacetylimide aqueous solution for 1 hour. After thorough washing with deionized water, it was placed in a zirconium-gold composite structure solution at 100°C and reacted for 24 hours to form a monolayer film on the surface, thus obtaining the SERS substrate. S3, Utilization N,N The SERS substrate prepared in S2 was further modified with '-di(2-aminoethyl)-1,3-propanediamine: a 20% glycerol-ethanol solution was prepared to dissolve... N,N '-Di(2-aminoethyl)-1,3-propanediamine molecule, after placing the SERS substrate prepared in S2 into it for 1-2 hours, it was rinsed with ethanol and deionized water, dried with nitrogen and stored in a 4°C refrigerator to obtain the SERS-enhanced substrate; Step 2, Glioma sample preparation: Add 0.86% ice-cold saline to the obtained glioma sample according to the ratio, and homogenize the sample using a homogenizer until the glioma sample is a white and uniform solution to obtain a homogenate. Step 3: Detection using a Raman spectrometer: The homogenate obtained in Step 2 is dropped onto the SERS-enhanced substrate obtained in Step 1, mixed, and incubated for 30-60 seconds. SERS detection is then performed using a laser with a wavelength of 785 nm and an integration time of 10 seconds. Before spectral acquisition, a 520 cm⁻¹ laser on a silicon chip is used. -1 The spectrometer was calibrated using Raman spectral bands; the bands at 512 and 1526 cm⁻¹ were determined in the IDH mutation samples. -1 A strong characteristic peak appeared in the sample, while it was barely visible in the wild-type sample.

5. The electronic device according to claim 4, characterized in that, In step 2, the ratio of glioma sample to ice-cold saline is 1g:9mL.

6. The electronic device according to claim 4, characterized in that, The method further includes the following steps: obtaining a judgment model using a ratio analysis model, wherein the judgment model is: ,in, I 512 The characteristic peak is at 512 cm⁻¹. -1 Strength at that location, I 1003 The characteristic peak is at 1003 cm⁻¹. -1 Strength at that location, I 1526 The characteristic peak is at 1526 cm⁻¹. -1 The intensity at the location; set the threshold to 1. When R is greater than the threshold, the sample is predicted as mutant, and when R is less than the threshold, the sample is predicted as wild.

7. A computer-readable storage medium having a computer program stored thereon, characterized in that, When the computer program is executed by the processor, it performs the steps in the following method: Step 1, Preparation of SERS-enhanced substrate: S1. Preparation of gold nanoparticles: Heat deionized water under stirring, and after boiling, add 1% chloroauric acid solution; while continuing to boil, add 4% trisodium citrate solution, react for half an hour, and cool to room temperature under stirring; gold nanoparticles are obtained, centrifuged and washed three times at 7000 rpm, then resuspended in deionized water and stored at 4℃ for later use. S2. Core-shell structure preparation: 70% zirconium propoxide, N,N An intermediate was obtained by reacting a mixture of dimethylformamide and glacial acetic acid at 130°C for 2 hours. Subsequently, the intermediate was mixed with gold nanoparticles prepared in S1 and 1,4-benzenedicarboxylic acid at room temperature. After thorough mixing, zinc nitrate was added, and after 18 hours, a zirconium-gold composite structure was obtained. A PVDF membrane with a size of 1 cm × 1 cm was treated with oxygen plasma for 0.5 hours, washed, and then incubated with a 2% polyacetylimide aqueous solution for 1 hour. After thorough washing with deionized water, it was placed in a zirconium-gold composite structure solution at 100°C and reacted for 24 hours to form a monolayer film on the surface, thus obtaining the SERS substrate. S3, Utilization N,N The SERS substrate prepared in S2 was further modified with '-di(2-aminoethyl)-1,3-propanediamine: a 20% glycerol-ethanol solution was prepared to dissolve... N,N '-Di(2-aminoethyl)-1,3-propanediamine molecule, after placing the SERS substrate prepared in S2 into it for 1-2 hours, it was rinsed with ethanol and deionized water, dried with nitrogen and stored in a 4°C refrigerator to obtain the SERS-enhanced substrate; Step 2, Glioma sample preparation: Add 0.86% ice-cold saline to the obtained glioma sample according to the ratio, and homogenize the sample using a homogenizer until the glioma sample is a white and uniform solution to obtain a homogenate. Step 3: Detection using a Raman spectrometer: The homogenate obtained in Step 2 is dropped onto the SERS-enhanced substrate obtained in Step 1, mixed, and incubated for 30-60 seconds. SERS detection is then performed using a laser with a wavelength of 785 nm and an integration time of 10 seconds. Before spectral acquisition, a 520 cm⁻¹ laser on a silicon chip is used. -1 The spectrometer was calibrated using Raman spectral bands; the bands at 512 and 1526 cm⁻¹ were determined in the IDH mutation samples. -1 A strong characteristic peak appeared in the sample, while it was barely visible in the wild-type sample.

8. The computer-readable storage medium according to claim 7, characterized in that, In step 2, the ratio of glioma sample to ice-cold saline is 1g:9mL.

9. The computer-readable storage medium according to claim 8, characterized in that, The method further includes the following steps: obtaining a judgment model using a ratio analysis model, wherein the judgment model is: ,in, I 512 The characteristic peak is at 512 cm⁻¹. -1 Strength at that location, I 1003 The characteristic peak is at 1003 cm⁻¹. -1 Strength at that location, I 1526 The characteristic peak is at 1526 cm⁻¹. -1 The intensity at the location; set the threshold to 1. When R is greater than the threshold, the sample is predicted as mutant, and when R is less than the threshold, the sample is predicted as wild.

Images