NK cell delivery data processing method and system based on magnetic targeting

Abstract

The invention relates to the technical field of medical engineering, in particular to an NK cell delivery data processing method and system based on magnetic targeting. The method comprises the following steps: inquiring an immune receptor protein structure of natural killer cells; injecting a sodium chloride solution, adding an immune receptor protein structure, and reacting to generate an aptamer structure; the method comprises the following steps: combining streptavidin in a pre-prepared glycosyl magnetic microsphere to obtain a high-affinity magnetic bead; carrying out molecular docking on the high-affinity magnetic beads and the aptamer structure, recording a docking score, and generating an optimized aptamer; recording the magnetic bead mass ratio of the optimized aptamer; capturing the natural killer cells by adopting the optimized aptamer, and shooting to obtain a fluorescent microscopic image; and recording the dyeing condition of the fluorescent microscopic image, determining the survival state of the NK cells, marking the position information of the living cells, and generating an NK cell delivery efficiency table by using the mass ratio of the magnetic beads. Based on a medical engineering technology, the homing rate and the delivery efficiency of the NK cells at the liver injury part are improved.

Description

Technical Field

[0001] This invention relates to the field of medical engineering technology, and in particular to a data processing method and system for NK cell delivery based on magnetic targeting. Background Technology

[0002] The existing NK cell therapy and delivery technologies face several major challenges: low NK cell delivery efficiency, a lack of effective targeting mechanisms leading to uneven cell distribution in vivo and significant variations in therapeutic efficacy; cell enrichment methods often rely on physical filtration or non-specific adsorption, resulting in poor selectivity and low capture purity; aptamer design lacks systematic screening and affinity optimization, hindering high-specificity recognition of target cell surface markers and impacting targeting accuracy; and the uneven particle size distribution and poor surface functionalization stability of magnetic microspheres limit their enrichment capacity and biocompatibility in complex in vivo environments. Furthermore, current data processing methods cannot accurately quantify and analyze cell identification, image staining, and enrichment effects during delivery, lacking dynamic monitoring and feedback mechanisms, which hinders the evaluation of delivery system stability and safety. These issues severely restrict the clinical translation efficiency and reliability of NK cell targeted therapy in the intervention of major diseases such as liver injury. Summary of the Invention

[0003] Therefore, it is necessary for the present invention to provide a magnetically targeted NK cell delivery data processing method and system to solve at least one of the above-mentioned technical problems.

[0004] To achieve the above objectives, a data processing method for NK cell delivery based on magnetic targeting includes the following steps: Step S1: Locate the immune receptor protein structure of natural killer cells; inject sodium chloride solution and add the immune receptor protein structure to generate aptamer structure; Step S2: In the pre-prepared glycosyl magnetic microspheres, streptavidin is combined to obtain high-affinity magnetic beads; the high-affinity magnetic beads are molecularly docked with the aptamer structure, the docking score is recorded, and the optimized aptamer is generated; Step S3: Record the magnetic bead mass ratio of the optimized aptamer; capture natural killer cells using the optimized aptamer and obtain fluorescence microscopy images; Step S4: Record the staining of the fluorescence microscopy images, determine the survival status of NK cells, mark the location information of live cells, and generate an NK cell delivery efficiency table using the magnetic bead mass ratio.

[0005] The beneficial effects of this invention are as follows: (1) By constructing the immune receptor protein structure of natural killer cells, injecting sodium chloride solution and introducing the protein structure to react, the automatic screening and preliminary construction of aptamer structures can be achieved. This method simulates the in vivo recognition mechanism, improves the binding specificity of aptamers to receptors on the surface of target cells, and provides a high-affinity molecular basis for subsequent cell capture.

[0006] (2) Introducing streptavidin into pre-fabricated glycosyl magnetic microspheres can produce high-affinity magnetic beads. By performing molecular docking with aptamer structures, optimized aptamers with higher docking scores can be screened, thereby improving recognition efficiency and magnetic targeting capability. This step is highly controllable and reproducible, which helps maintain a consistent affinity level in large-scale preparation and ensures the stability of experimental results.

[0007] (3) Based on the recorded mass ratio of magnetic beads to aptamers, a standardized parameter system is established to improve cell binding efficiency and reduce non-specific adsorption during the capture of natural killer cells. Combined with fluorescence microscopy, staining images can be accurately acquired, enabling highly sensitive visual detection and screening of natural killer cells, further enhancing the reliability and practicality of the overall capture system.

[0008] (4) By analyzing the staining of live cells in fluorescence images, the survival status of NK cells can be accurately identified. Combined with the magnetic bead mass ratio, a delivery efficiency evaluation model is constructed to generate an NK cell delivery efficiency table. This table can provide quantitative support for subsequent magnetic targeted delivery experiments, improve the scientific nature of data processing and the accuracy of decision-making, and meet the research and clinical application needs of high-throughput cell screening and targeted delivery. Attached Figure Description

[0009] Other features, objects, and advantages of the invention will become more apparent from the following detailed description of non-limiting embodiments with reference to the accompanying drawings: Figure 1 This is a schematic diagram of the steps in the magnetic targeting-based NK cell delivery data processing method of the present invention; Figure 2 This is a schematic diagram of the modules of the magnetically targeted NK cell delivery data processing system of the present invention; Figure 3 This is a schematic diagram of the immune receptor protein structure and aptamer structure in this invention; Figure 4 This is a schematic diagram of the docking between the immune receptor protein structure and the aptamer in this invention; Figure 5 This is an image taken using a fluorescence microscope in this invention; The realization of the objective, functional features and advantages of the present invention will be further explained in conjunction with the embodiments and with reference to the accompanying drawings. Detailed Implementation

[0010] The technical method of this invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some, not all, of the embodiments of this invention. Based on the embodiments of this invention, all other embodiments obtained by those skilled in the art without inventive effort are within the scope of protection of this invention.

[0011] Furthermore, the accompanying drawings are merely illustrative of the invention and are not necessarily drawn to scale. The same reference numerals in the drawings denote the same or similar parts, and therefore repeated descriptions of them will be omitted. Some block diagrams shown in the drawings are functional entities and do not necessarily correspond to physically or logically independent entities. These functional entities can be implemented in software, in one or more hardware modules or integrated circuits, or in different network and / or processor methods and / or microcontroller methods.

[0012] It should be understood that although the terms "first," "second," etc., may be used herein to describe various units, these units should not be limited by these terms. These terms are used merely to distinguish one unit from another. For example, without departing from the scope of the exemplary embodiments, a first unit may be referred to as a second unit, and similarly, a second unit may be referred to as a first unit. The term "and / or" as used herein includes any and all combinations of one or more of the associated listed items.

[0013] To achieve the above objectives, please refer to Figures 1 to 5 This invention provides a data processing method for NK cell delivery based on magnetic targeting, the method comprising the following steps: Step S1: Locate the immune receptor protein structure of natural killer cells; inject sodium chloride solution and add the immune receptor protein structure to generate aptamer structure; In one embodiment, three-dimensional structural data (e.g., PDBID: 1HYR) of the NKG2D receptor protein expressed on human natural killer (NK) cells were obtained from NCBI and PDB databases, and the protein structure was imported into molecular modeling software (e.g., AutoDockVina). A 0.9% sodium chloride solution was prepared as the reaction environment, and the receptor structure was added to the solution. By simulating the molecular self-assembly process, the reaction was carried out for 24 hours under specific pH (7.2) and temperature (37°C) conditions, and finally, aptamer structural models with preliminary affinity were screened out.

[0014] In another embodiment, three different NK cell receptors (NKG2D, NKp30, and NKp44) were selected for modeling, and sodium chloride solutions with concentrations of 0.6%, 0.9%, and 1.2%, respectively, were injected. After treatment under standard reaction conditions (pH 7.4, temperature 36°C) for 12 hours, nine aptamer structures were initially generated, and their energy stability range was recorded as −6.5 kcal / mol to −9.3 kcal / mol. Subsequently, an affinity below −8.0 kcal / mol was used as the screening criterion for the next stage.

[0015] Step S2: In the pre-prepared glycosyl magnetic microspheres, streptavidin is combined to obtain high-affinity magnetic beads; the high-affinity magnetic beads are molecularly docked with the aptamer structure, the docking score is recorded, and the optimized aptamer is generated; In one embodiment, commercially available glycosyl magnetic microspheres (approximately 1 μm in diameter) were selected. Streptavidin molecules were introduced through surface functional groups (carboxyl groups) to form biotin-avidin reaction sites. The aptamer structure generated in the previous step was labeled with biotin end groups and then conjugated with magnetic beads to form high-affinity magnetic beads. Subsequently, a molecular docking algorithm (such as RosettaDock) was used to dock the magnetic bead-aptamer complex with an NK cell membrane model to obtain docking scores. The scores were then normalized, and the optimized aptamer with the highest score was selected as a candidate for subsequent experiments.

[0016] In another embodiment, assuming a total of 100 mg of glycosyl magnetic microspheres were prepared, 50 mg of which was inoculated with streptavidin and then bound to 5 candidate aptamer structures, with 10 μg of each structure used. A total of 25 docking experiments were performed, and the docking score for each group was recorded, ranging from 82.4 to 97.1. A screening threshold of 90 was set; 7 groups with docking scores greater than this threshold proceeded to the next round of experiments. This process significantly improved the NK cell recognition efficiency in subsequent experiments.

[0017] Step S3: Record the magnetic bead mass ratio of the optimized aptamer; capture natural killer cells using the optimized aptamer and obtain fluorescence microscopy images; In one embodiment, the mass ratio between the aptamer and the magnetic beads was recorded and optimized using a precision electronic balance (e.g., 1:5000), and this ratio was maintained for cell capture experiments. It was then co-incubated with NK cells (concentration 10). 6 (cells / mL, incubation time 30 min, 37℃), fluorescent staining was performed using FITC-labeled aptamers, and images were captured using an inverted fluorescence microscope (excitation wavelength 488 nm) to obtain images of the target cell binding state for subsequent analysis.

[0018] In another embodiment, assuming a total magnetic bead mass of 10 mg and an aptamer dosage of 2 μg, the mass ratio was 1:5000. Three groups of experiments were conducted, using mass ratios of 1:2000, 1:5000, and 1:8000 for cell incubation, respectively, and the fluorescence image brightness and cell binding density were observed. The results showed that the average cell surface fluorescence intensity (average gray value 180) was highest at a mass ratio of 1:5000, indicating the optimal binding efficiency; this ratio was selected as the standard operating parameter.

[0019] Of particular importance is the recording of the magnetic bead mass ratio of the optimized aptamer, including: Before preparing the experimental group, multiple mass ratio gradients of aptamers to glycosyl magnetic microspheres (such as 1:1, 2:1, 3:1, 4:1, 5:1) were set up to keep the mass of the magnetic beads constant and only the amount of aptamers added was adjusted. In one embodiment, before preparing the experimental groups, multiple mass ratio gradients of aptamers to glycosyl magnetic microspheres (e.g., 1:1, 2:1, 3:1, 4:1, 5:1) are set, where the mass of magnetic beads remains constant (e.g., 10 mg), and only the mass of aptamers added is adjusted, for example, adding 10 μg, 20 μg, 30 μg, 40 μg, and 50 μg of aptamers to construct different experimental groups. All materials are preheated and thoroughly mixed before the experiment to ensure consistency of experimental conditions under different mass ratio gradients. In another embodiment, five mass ratios are assumed to be set: 1:1, 2:1, 3:1, 4:1, and 5:1, with the mass of magnetic beads fixed at 10 mg, corresponding to aptamer addition masses of 10, 20, 30, 40, and 50 μg, respectively. This ensures that only the mass ratio differs among the five groups, thus enabling a single-factor evaluation of the effect of the ratio on binding efficiency.

[0020] After preparation, each group of samples was magnetically separated and washed in sequence, and fluorescently labeled probes were added. The average intensity of the fluorescence signal of each group was recorded as an indicator of binding efficiency. In one embodiment, after sample preparation, each experimental group underwent standardized magnetic separation and three buffer washes to remove unbound components. Then, an equal volume and concentration of fluorescently labeled probe solution (such as FITC-labeled oligonucleotides) were added, and the samples were incubated at 37°C for 30 minutes to ensure sufficient label binding. After incubation, a final wash was performed, and the average fluorescence signal intensity of each sample group was read using a fluorescence microplate reader as a quantitative indicator of aptamer-magnetic bead binding efficiency. In another embodiment, assuming the measured average fluorescence intensities (in au) for the five groups are 48.2, 72.6, 89.1, 83.5, and 59.8, respectively, showing that the binding signal initially increases and then decreases with increasing mass ratio, a preliminary inference suggests that a 3:1 ratio is optimal.

[0021] Enter the mass ratios corresponding to each group of experiments in the first column (e.g., 1, 2, 3, 4, 5). This column of data will serve as the horizontal axis (X-axis) of the bar chart, representing the mass ratio gradient of the aptamer to the magnetic beads. In the second column, enter the mean fluorescence intensity corresponding to each mass ratio. This column of data will serve as the vertical axis (Y-axis), representing the binding efficiency signal intensity. Click "Insert" → "Bar Chart", select the "Clustered Bar Chart" style, and the coordinate system and bar chart will be automatically generated to record the magnetic bead mass ratios of the optimized aptamer.

[0022] In one embodiment, the experimental data is input into an Excel worksheet. The first column contains the five corresponding mass ratio gradients (1, 2, 3, 4, 5), serving as the X-axis to represent the ratio of aptamers to magnetic beads. The second column contains the average fluorescence intensity measured for each group (e.g., 48.2, 72.6, 89.1, 83.5, 59.8), serving as the Y-axis to represent the binding efficiency intensity. Clicking "Insert" - "Chart" - "Clustered Column Chart" automatically generates a standard coordinate graph in Excel. The horizontal axis represents the mass ratio, and the vertical axis represents the fluorescence signal intensity, visually displaying the differences in binding efficiency between groups. In another embodiment, assuming the data has been input into an Excel spreadsheet as described above, by selecting two data columns and inserting a column chart, the graph shows that the signal intensity is highest and the binding efficiency is optimal when the mass ratio is 3:1. This graph can be used for rapid decision-making and result reporting for optimizing experimental ratios.

[0023] Step S4: Record the staining of the fluorescence microscopy images, determine the survival status of NK cells, mark the location information of live cells, and generate an NK cell delivery efficiency table using the magnetic bead mass ratio.

[0024] In one embodiment, the captured images are imported into image analysis software (such as ImageJ), and the staining status is determined by the fluorescence intensity of the green channel. A double staining method (PI / CFSE) is used to distinguish between dead and live cells, and the location of live cells is automatically marked based on fluorescence threshold settings (PI red > 120 for dead cells, CFSE green > 150 for live cells). The NK cell delivery efficiency is calculated by combining the magnetic bead mass ratio and the number of cells successfully bound per unit time, and a delivery efficiency table is finally output (e.g., number of cells successfully delivered per mg of magnetic beads = 350 cells / mg).

[0025] In another embodiment, it is assumed that 30 fluorescence images were acquired in the experiment, with a total number of viable cells of 6300. Three different magnetic bead mass ratios were used: 1:4000, 1:5000, and 1:6000, delivering 1800, 2300, and 2200 cells respectively. The calculated delivery efficiencies were 450 cells / mg, 575 cells / mg, and 550 cells / mg. The data were plotted as an NK cell delivery efficiency table to aid in selecting the optimal magnetic bead loading strategy.

[0026] Preferably, in step S1, adding sodium chloride solution and placing an immune receptor protein structure to generate an aptamer structure includes: Add sodium chloride solution to the simulated environment; In one embodiment, a 10nm × 10nm × 10nm three-dimensional cubic simulation chamber was constructed in the GROMACS 2021.3 environment. Solvation was performed using the TIP3P water molecule model, generating approximately 33,000 water molecules. The temperature was set at 310K and the pressure at 1 atm, and the system was controlled using Berendsen temperature coupling and Parrinello-Rahman pressure coupling. To simulate physiological conditions, sodium chloride was added to the system at a concentration of 0.15 mol / L, with approximately 90 pairs of Na⁺ and Cl⁻ ions added accordingly to maintain system electroneutrality and ensure that the overall ionic strength was consistent with human extracellular fluid. This step ensured subsequent interactions of biomolecules under near-realistic conditions. After the addition of ions, the system was subjected to energy minimization using the Steepest Descent method until the maximum force was less than 1000 kJ / mol / nm. Subsequently, a 100 ps NVT pre-equilibration and a 200 ps NPT equilibration were performed to stabilize the system.

[0027] The electrolyte strength of sodium chloride solution was calculated, immune receptor protein structures were added, charge rearrangement was simulated, changes in protein structure were recorded, and aptamer structures were generated.

[0028] In one embodiment, after system equilibration, the immune receptor protein structure NKG2D (PDBID: 4SOU) of human natural killer cells was loaded and placed in the center of the box, ensuring a minimum distance of 1 nm from the boundary. The protein was parameterized using the AMBER99SB-ILDN force field, and hydrogen bonds were constrained using the LINCS algorithm. After adding the protein, a 5000-step energy minimization was performed again, followed by a 2 ns molecular dynamics simulation. During the simulation, the protein conformation was recorded every 100 ps, ​​resulting in 20 frames. RMSD and dipole moment change analysis revealed significant changes in the charge distribution in the C-terminal region of the protein, indicating that the local electric fields of Na⁺ and Cl⁻ affected the charge rearrangement in this region. The free energy of the 20 extracted structures was calculated using the MM-PBSA method, and the conformation with the lowest free energy was selected as the aptamer structure. This conformation forms a new stable salt bridge near the Lys76-Arg84 domain, which may enhance its ability to recognize target ligands.

[0029] In another embodiment, assuming the simulation system box is 12nm × 12nm × 12nm, with a total volume of 1728nm³, adding 0.2mol / L NaCl requires approximately 210 pairs of Na⁺ / Cl⁻ ions. An SPC / E water model is used, and the simulation temperature is 298K. Electrolyte strength... According to the formula: The protein was loaded as the NCR1 structure (PDBID: 1P6T). After 5000 minimization steps, a 4ns simulation was run, with conformation extracted every 200ps, for a total of 20 frames. Analysis showed that the RMSD stabilized between 1.2–1.5nm, the protein charge center shifted by approximately 0.3nm, and the dipole moment increased by approximately 12%. The structure in frame 14 was ultimately selected as the aptamer structure. Its intramolecular electrostatic potential surface plot showed that the negatively charged region near the β7 fold expanded, which may improve the local affinity for binding to cationic ligands.

[0030] Preferably, in step S2, the conjugation of streptavidin in the pre-prepared glycosyl magnetic microspheres to obtain high-affinity magnetic beads includes: Mix the glycosyl magnetic microspheres using a vortex mixer, place them in a centrifuge tube, add buffer solution, and vortex to mix. Place the tube on a magnetic separator to enrich the magnetic beads, remove the supernatant, and repeat the washing process with buffer solution. In one embodiment, 1 mL of glycosyl-functionalized magnetic microspheres (approximately 500 nm in diameter, 10 mg / mL concentration) was placed into a 2 mL centrifuge tube and thoroughly mixed using a vortex mixer (2500 rpm) for 15 seconds to ensure uniform dispersion of the magnetic beads. Then, 1 mL of 1×PBS buffer was added, and the mixture was vortexed for another 10 seconds to maintain a neutral and isotonic environment on the surface of the magnetic beads. The mixture was placed in a magnetic separator and allowed to stand for approximately 60 seconds until the magnetic beads were completely adsorbed to the tube wall. The supernatant was then carefully discarded. The process of adding 1 mL of PBS buffer, vortexing, enriching, and discarding the supernatant was repeated three times to remove impurities and residual free sugars. After washing, the magnetic beads were in a pre-activation state and kept moist to prevent drying and affecting subsequent reactions.

[0031] In another embodiment, assuming simultaneous processing with 8 centrifuge tubes, 0.8 mL of glycosyl magnetic beads (total 6.4 mL) was added to each tube, with buffer added at a 1:1 ratio. The vortexing time for each centrifuge tube was fixed at 20 seconds, and the shaking frequency was set to 3000 rpm to ensure efficient homogenization. During magnetic rack separation, the magnetic beads were observed to be completely adsorbed within 45–60 seconds, with approximately 0.75 mL of liquid discarded each time. After three rounds of washing, the total volume removed was approximately 2.25 mL, and the residual magnetic bead volume remained between 0.5–0.6 mL, facilitating precise reaction volume control in subsequent activation steps.

[0032] Add carboxyl activator and N-hydroxysuccinimide, vortex to mix, and incubate at room temperature for 0.5-0.7 h to activate the carboxyl groups on the surface of the magnetic beads and bind streptavidin to obtain high-affinity magnetic beads.

[0033] In one embodiment, a freshly prepared carboxyl activation system was added to the cleaned glycosyl magnetic microspheres: 0.2 mL of MEDC (1-ethyl-3-(3-dimethylaminopropyl)carbodiimide) and 0.1 mL of MNHS (N-hydroxysuccinimide) buffer solution (pH=6.0, primarily MES buffer). The reaction system volume was maintained at 500 μL, and the mixture was vortexed for 10 seconds to ensure homogeneity. The mixture was then incubated at room temperature (20 rpm) for 40 minutes to complete carboxyl activation. 200 μL of 1 mg / mL streptavidin solution was then added, and the reaction was continued for 30 minutes to achieve covalent coupling. After the reaction was complete, the microspheres were washed three times with PBS to remove unbound streptavidin and reaction byproducts. The resulting magnetic beads were high-affinity glycosyl-streptavidin functional magnetic beads.

[0034] In another embodiment, assuming a reaction volume of 600 μL per tube, and EDC and NHS volumes of 0.12 mmol and 0.06 mmol, respectively, prepared in 0.05 MMES buffer at pH 5.8, the mixture was rapidly shaken for 15 seconds and incubated in a constant temperature water bath (25°C, horizontal rotation 25 rpm) for 45 minutes. Subsequently, 0.5 mL of streptavidin solution (1.5 mg / mL, prepared in Tris buffer) was added, extending the total reaction time to 70 minutes. During this process, the magnetic beads formed a stable covalently bound layer, with a yield of approximately 85% of the initial magnetic bead mass. Subsequent BCA protein quantification showed that the streptavidin binding amount on the magnetic bead surface was approximately 25 μg / mg of magnetic beads.

[0035] Preferably, the glycosyl magnetic microspheres are mixed using a vortex mixer, placed in a centrifuge tube, and buffer solution is added. The vortex mixing process includes: Before mixing, place the glycosyl magnetic microspheres in a clean centrifuge tube and vortex them continuously at a speed of 1500rpm-2000rpm for 10-15 seconds. In one embodiment, pretreated glycosyl magnetic microspheres (concentration 10 mg / mL) were selected, and 1.0 mL was placed into a sterilized 2 mL polypropylene centrifuge tube. The tube was then placed in a vortex mixer, and the speed was set to 1800 rpm for 12 seconds. This step helps to break up any potential agglomeration of the microspheres, resulting in a more uniform dispersion and ensuring consistent subsequent liquid mixing. During the operation, the microspheres were observed to be uniformly suspended without significant precipitation, indicating good premixing.

[0036] In another embodiment, assuming the sample consisted of six centrifuge tubes, with 0.8 mL of glycosyl magnetic microspheres added to each tube, the oscillation parameters were set to 2000 rpm and the duration to the maximum value of 15 seconds. Under these conditions, the estimated shear strength was approximately 1.5 × 10³ s⁻¹, suitable for breaking up particle aggregation. Simulation data showed that the premixing uniformity improved by approximately 28%, the subsequent surface area of ​​the microspheres interacting with the buffer solution significantly increased, and the initial droplet contact angle decreased to 35°, which is beneficial for rapid wetting.

[0037] During the addition of buffer solution, a staged injection method is adopted. In the first stage, 30%-40% of the total volume of buffer solution is added, and vortexing is performed again for 5-7 seconds to form a preliminary wetting layer on the surface of the microspheres. In one embodiment, the premixed glycosyl magnetic microsphere suspension was taken, and 1.0 mL of PBS buffer was added as planned, in two stages. In the first stage, 400 μL (40% of the total buffer volume) was added, followed immediately by a second round of vortexing at 1600 rpm for 6 seconds. After vortexing, the liquid surface in the tube was observed to be clear and free of particle deposition, indicating that the buffer had fully penetrated the microsphere surface, forming a stable wetting layer, which helps improve subsequent mixing efficiency and reaction consistency.

[0038] In another embodiment, assuming the total buffer volume in the experimental setup is 1.2 mL, the first-stage injection volume is selected as 30%, i.e., 360 μL. The vortex parameters are set to 1500 rpm for 7 seconds. After mixing, the D90 particle size change measured by a particle size analyzer is less than 5%, initially confirming that the microspheres maintain good dispersibility. Image tracking data shows that the initial binding rate of the microspheres to the buffer solution increases to more than three times that of the original static adsorption.

[0039] In the second stage, add the remaining 60%-70% of the buffer solution. Immediately after adding the buffer solution, perform a second vortexing for at least 15 seconds to form a stable suspension.

[0040] In one embodiment, after the first stage of injection and mixing, the remaining 600 μL of PBS buffer (60% of the total volume) is added. The injection process is completed within 5 seconds, followed immediately by a second vortex mixing at 2000 rpm for 18 seconds. After mixing, the system is observed to be a homogeneous milky white liquid without precipitation or aggregation, indicating that a stable suspension has been successfully formed. Under these conditions, the microspheres maintain high activity in subsequent reactions, making it suitable for functionalization or coupling steps.

[0041] In another embodiment, assuming the remaining buffer volume is 840 μL (original total volume 1.2 mL, 30% already added), the vortexing time after the second-stage injection is set to 20 seconds at a rotation speed of 1900 rpm. Simulation evaluation shows that the system reaches maximum mixing within 10 seconds after vortexing, the viscosity distribution tends to be uniform, and the resuspension rate reaches 96.7%. The stability of this suspension system can be maintained for more than 60 minutes at room temperature, providing sufficient time for subsequent reaction.

[0042] Preferably, the operation of enriching magnetic beads includes: During the enrichment process, the sample solution to be tested is injected into a centrifuge tube pre-filled with magnetic microspheres, and the injection rate is controlled at 1-2 mL / min. In one embodiment, a 2 mL polypropylene centrifuge tube was selected, and 1 mg of glycosyl-functionalized magnetic microspheres (suspended in PBS buffer, total volume 200 μL) were pre-added. Using a syringe pump (Harvard Apparatus PHDUltra), 0.5 mL of the serum sample to be tested was slowly injected into the centrifuge tube at a flow rate of 1.5 mL / min. During injection, a constant temperature of 25°C was maintained to ensure that the microspheres fully contacted the target molecules in the sample for efficient capture. After injection, the mixture was gently vortexed for 5 seconds to prevent microsphere sedimentation and promote a uniform reaction.

[0043] In another embodiment, it is assumed that four centrifuge tubes are used, each pre-loaded with 1.2 mg of magnetic microspheres, with a volume of 200 μL. The sample to be tested (0.8 mL per tube) is simultaneously injected at a rate of 2 mL / min using a multichannel syringe. Simulation data show that the relatively fast injection rate does not cause uncontrolled microsphere aggregation in high-viscosity samples (such as plasma), and the average mixing homogeneity remains above 87%. Within a 5-minute reaction window, the occupancy rate of the binding sites on the magnetic microsphere surface reaches 92%, laying the foundation for subsequent magnetic separation.

[0044] A magnetic device is installed on the outside of the centrifuge tube. By applying a stable magnetic field, magnetic microspheres are directionally adsorbed from the solution to one side of the tube wall, forming a visible magnetic bead aggregation zone.

[0045] In one embodiment, after sample injection, the centrifuge tube is placed on a magnetic rack equipped with a neodymium iron boron permanent magnet (size: 50×10×3mm, surface magnetic flux density approximately 3500G). The magnet is fixed to the outer right wall of the centrifuge tube, approximately 1.5cm from the bottom. Within one minute of applying the magnetic field, the magnetic microspheres rapidly and directionally aggregate on one side of the tube wall, forming a clearly visible black magnetic bead band, approximately 1mm wide and extending approximately 4mm vertically along the tube wall. The aggregation is confirmed to be stable, with no obvious free microspheres remaining in the liquid.

[0046] In another embodiment, assuming the use of an electrically controlled magnet, the magnetic field strength was adjusted to the range of 0–0.4T, and the experimental conditions were set as a continuous constant magnetic field of 0.3T. In the five sample groups, the average time for magnetic bead aggregation was 42 seconds, the bead thickness was approximately 1.2 mm, and the aggregation efficiency reached 96%. Image analysis showed that after aggregation, more than 90% of the magnetic beads were distributed within a range of 2.8 ± 0.3 mm from the center of the magnet. This directional adsorption process can significantly improve the efficiency and repeatability of subsequent elution steps.

[0047] Preferably, the combination of streptavidin to obtain high-affinity magnetic beads includes: After the magnetic particles in the centrifuge tube settle, remove the liquid on top of the centrifuge tube, wash repeatedly with buffer solution, add streptavidin, continue to vortex mix, and place in an environment of 35℃-37℃ to rotate and mix for reaction. In one embodiment, after the magnetic particles reacted with the sample, a 2 mL polypropylene centrifuge tube was placed on a magnetic rack. Within 5 minutes, the magnetic beads settled and aggregated on the tube wall. After removing the supernatant of 1.5 mL, 1 mL of PBS buffer (pH 7.4) was added, vortexed for 10 seconds, and washed three times. Then, 200 μL of streptavidin (concentration: 0.1 mg / mL) was added, and vortexed for another 15 seconds to ensure sufficient binding with the glycosyl groups on the microsphere surface. The centrifuge tube was then placed in a 37°C water bath shaker and rotated at 120 rpm for 30 minutes to ensure a uniform and complete reaction.

[0048] In another embodiment, four groups of samples were used, and streptavidin was added to each sample and reacted at 35°C, 36°C, 37°C, and 38°C for a uniform reaction time of 40 minutes. The mixing speeds were set at 100, 120, 140, and 160 rpm for comparison. The results showed that the magnetic bead binding saturation was highest at 37°C and 120 rpm, reaching 93%; while at 35°C and 160 rpm, the binding saturation decreased to approximately 78%. This hypothetical data illustrates that temperature and mixing rate have a coupled effect on streptavidin grafting efficiency, allowing for optimization of process parameters.

[0049] After the reaction is complete, place the centrifuge tube on a magnetic separator, remove the supernatant, add the blocking solution, and rotate to mix the reaction at 35℃-37℃. In one embodiment, after the streptavidin reaction is complete, the centrifuge tube is placed back on the magnetic separator and allowed to stand for 5 minutes. The magnetic beads re-aggregate to the tube wall. The supernatant (approximately 180 μL) is then slowly removed, and 1 mL of blocking buffer (PBS buffer containing 1% BSA) is added. After vortexing, the tube is immediately transferred to a 37°C constant-temperature shaker and vortexed at 120 rpm for 30 minutes to block any remaining unreacted sites on the surface of the magnetic beads and reduce non-specific binding.

[0050] In another embodiment, three groups of samples were treated with three different concentrations of blocking solution (0.5%, 1%, and 2% BSA) for comparative experiments at blocking times of 20, 30, and 40 minutes. The results showed that the background adsorption rate of the magnetic beads was lowest at 1% BSA concentration after 30 minutes of blocking, at only 6.2%; while the background adsorption rate increased to 18.7% under the 0.5% BSA / 20-minute condition. This set of hypothetical data illustrates that the concentration and time of the blocking solution need to be synergistically controlled to effectively improve the selectivity of the magnetic beads and the accuracy of subsequent detection.

[0051] After mixing and reaction, the high-affinity magnetic beads are preserved using a preservation solution.

[0052] In one embodiment, after the sealing step, the blocking solution was removed again by magnetic separation, and 1 mL of preservation solution (PBS buffer at pH 7.4) containing 0.02% sodium azide was added for long-term storage. After thorough vortexing, the mixture was transferred to a 2 mL low-adsorption EP tube and stored at 4°C in the dark. Tests showed that under these conditions, the magnetic beads retained more than 85% affinity activity after 30 days of storage, making them suitable for subsequent capture experiments.

[0053] In another embodiment, it was assumed that high-affinity magnetic beads were stored at 4°C, -20°C, and room temperature, and their binding capacity was measured at days 0, 7, 15, and 30. The results showed that the affinity of the magnetic beads decreased the slowest under 4°C storage conditions (retaining 86% after 30 days); while room temperature storage only resulted in a decrease in activity to 60% after 7 days; although -20°C freezing maintained activity, repeated freeze-thaw cycles led to an increase in aggregation rate to 28%. These hypothetical data indicate that storage temperature and the number of freeze-thaw cycles significantly affect the stability of the magnetic beads, and 4°C storage in the dark is recommended as the optimal method.

[0054] Preferably, step S2, which involves molecular docking of the high-affinity magnetic beads with the aptamer structure, includes: After centrifuging the high-affinity magnetic beads at 10000-12000g for 5-7 minutes, molecular docking was performed with the aptamer structure. Sterile water was added for fusion, and the mixture was placed in a freezer for 5-10 minutes. The docking score was recorded to generate an optimized aptamer with a stable tertiary structure.

[0055] In one embodiment, 1 mg of magnetic beads was placed in a sterile EP tube, 1 mL of pre-chilled sterile PBS buffer was added, and the mixture was gently pipetted to mix. The tube was then centrifuged at 11000 g for 6 minutes in a refrigerated centrifuge. After centrifugation, the supernatant was carefully aspirated, retaining the magnetic bead precipitate. Then, 50 μL of a 10 μM biotin-modified aptamer solution was added and mixed to allow for specific binding to the streptavidin on the surface of the magnetic beads. Sterile ultrapure water was added to bring the total volume to 200 μL, and the EP tube was immediately placed in a –20°C freezer for 8 minutes to enhance binding stability. After cooling, the sample was removed and allowed to warm slowly to room temperature for 5 minutes, then mixed again for subsequent cell capture experiments.

[0056] Preferably, the recorded docking score includes: Different amounts of high-affinity magnetic beads (0.2 nmol-1.0 nmol) were added to the prepared high-affinity magnetic beads for optimization. The beads were rotated and incubated at a preset temperature for 0.5 h-0.8 h, and the mass ratio of the magnetic beads was recorded. In one embodiment, pre-prepared high-affinity glycosyl magnetic beads were used as the base material and dispensed into multiple 1.5 mL sterile EP tubes. Five different amounts of biotinylated high-affinity aptamers (0.2 nmol, 0.4 nmol, 0.6 nmol, 0.8 nmol, and 1.0 nmol) were added to each tube. The samples were placed in a thermostatic mixer and incubated at 37°C with a rotation speed of 500 rpm for 0.5 to 0.8 hours. After incubation, the magnetic beads were separated using a magnetic rack, the supernatant was removed, and the sample was washed twice with PBS. The precipitate was collected. The total mass of the bound magnetic beads was measured, and the ratio between the added aptamer mass and the magnetic bead mass (i.e., the magnetic bead mass ratio) was recorded to screen the group with the highest binding efficiency, which served as the optimal combination for subsequent experiments.

[0057] In another embodiment, assuming the amount of biotinylated aptamer added to the magnetic beads is 0.2, 0.4, 0.6, 0.8, and 1.0 nmol, the corresponding post-binding masses of the magnetic beads are 1.2, 1.9, 2.5, 2.8, and 2.7 μg, respectively. The calculated magnetic bead mass ratios are 6.0, 4.75, 4.17, 3.5, and 2.7. Among these, the group with a magnetic bead mass ratio of 4.17 (i.e., 0.6 nmol corresponding to 2.5 μg) shows the best binding efficiency, and therefore this can be used as a recommended parameter for subsequent magnetic bead aptamer construction.

[0058] Add 0.6 nmol-0.8 nmol of biotinylated high-affinity magnetic beads to the prepared high-affinity magnetic beads, and incubate them by rotation for 0.25 h-2.5 h under preset temperature conditions for optimization, and record the optimization results; In one embodiment, based on the optimized magnetic bead mass ratio (e.g., 0.6 nmol: 2.5 μg), incubation time gradients (0.25 h, 0.5 h, 1 h, 1.5 h, 2 h, 2.5 h) were set to investigate the effect of incubation time on aptamer binding efficiency. Each group was incubated at 37 °C under the same rotating conditions. After incubation, samples were taken, and the binding efficiency was determined using a fluorescence labeling method. The relationship between binding intensity and fluorescence signal intensity at each time point was recorded to further confirm the optimal incubation time range.

[0059] In another embodiment, assuming 0.8 nmol of aptamer was used for incubation with high-affinity magnetic beads, the incubation time was set at 37°C for 0.25 h, 0.5 h, 1 h, 2 h, and 2.5 h, respectively, and the fluorescence intensities were measured to be 22.1, 38.7, 42.3, 42.8, and 41.9 (in relative fluorescence units, RFU). The results showed that the maximum binding efficiency was reached at 1 h, and the subsequent growth was slow or even slightly decreased, indicating that about 1 h is the optimal incubation time, which can ensure sufficient aptamer binding while saving time.

[0060] Among them, the magnetic bead mass ratio and the incubation optimization results are used as docking scores.

[0061] In one embodiment, the mass ratio of magnetic beads from different batches was calculated by measuring their mass. Under standardized incubation conditions, the magnetic beads were subjected to a binding reaction with target cells, and quantitative indicators of the incubation effect were obtained using flow cytometry or fluorescence microscopy. After normalizing the magnetic bead mass ratio and the incubation effect indicators, a weighted summation was used to calculate the docking score, with the weighting parameters set according to experimental requirements. This docking score was used to comprehensively evaluate the performance of the magnetic beads and the optimization effect of the incubation conditions, guiding the subsequent adjustment of the magnetic bead-aptamer binding scheme.

[0062] In another embodiment, assuming the mass ratios of five groups of magnetic beads are measured to be 85.2%, 78.6%, 91.4%, 69.8%, and 80.5%, respectively, the corresponding optimized incubation results (normalized by fluorescence intensity, with a maximum score of 100) are 88, 75, 92, 68, and 81, respectively. Two indicators are linearly normalized to values ​​between 0 and 1, with the highest magnetic bead mass ratio being 1 and the lowest being 0. The incubation results are processed similarly. Docking scores are calculated using a 50% weighting, resulting in scores of 0.83, 0.42, 1.00, 0.00, and 0.54, respectively. This quantitative scoring method provides an objective basis for screening optimal magnetic beads and incubation conditions, thereby improving cell capture efficiency and binding stability.

[0063] Preferably, step S3, which involves using an optimized aptamer to capture natural killer cells and obtaining fluorescence microscopic images, includes: Thaw the frozen natural killer cells at 35℃-37℃, centrifuge and discard the supernatant, wash with buffer, add the pre-prepared cell live / dead staining combination solution, in which the ratio of fluorescein ethyl ester: propidium iodide: buffer is 1:1:1000, and incubate at 36℃-37℃ in the dark for 25min-30min. In one embodiment, cryopreserved natural killer (NK) cells were rapidly thawed in a 35°C-37°C water bath, then immediately transferred to centrifuge tubes and centrifuged at 400g for 5 minutes, discarding the supernatant. The cells were washed once with pre-chilled PBS buffer, centrifuged again, and the supernatant was discarded. A pre-prepared cell live / dead cell staining solution was added, with a volume ratio of fluorescein ethyl ester (Calcein AM), propidium iodide (PI), and buffer solution of 1:1:1000. The cells were incubated at 36°C-37°C in the dark for 25 to 30 minutes, gently mixing during this time to allow the dye to fully penetrate the cell membrane and bind to internal components, thus distinguishing between live and dead cells.

[0064] In another embodiment, it is assumed that each vial of cryopreserved NK cells contains approximately 1 × 10⁻⁶ cells. 6 Cells were revived using 1 mL of cryopreservation solution, incubated at 35.5°C for 2 minutes, and immediately centrifuged at 450 g for 5 minutes. The cells were then washed once with PBS. For staining, 2 μL each of fluorescein ethyl ester and propidium iodide were added to 2 mL of buffer and mixed thoroughly. The mixture was then used for staining at 36.2°C for 28 minutes. After staining, the cell state was quickly observed using an inverted microscope in preparation for subsequent binding experiments.

[0065] The prepared optimized aptamer was mixed with the stained natural killer cell suspension and incubated with shaking at 35℃-37℃ for 40-45 minutes. The supernatant was removed magnetically, and the mixture was resuspended in buffer solution. The resulting product was photographed under a fluorescence microscope.

[0066] In one embodiment, the prepared optimized aptamer (e.g., biotinylated M-CD16-apt aptamer coupled to the surface of glycosyl magnetic microspheres via streptavidin) was mixed with stained NK cell suspension at a 1:1 volume ratio and incubated in a constant temperature shaker at 35°C-37°C for 40 to 45 minutes. After incubation, the magnetic beads were adsorbed using a magnetic rack and the supernatant was removed, followed by resuspension with PBS buffer. The resulting magnetic bead-cell complex was dropped onto a glass slide and four sets of images were captured sequentially under a fluorescence microscope: (a) selecting the FITC channel (optical channel for fluorescence signals emitted by fluorescein dye) to observe Calcein-labeled live cell images; (b) selecting the PI channel to record PI-stained dead cell signals; (c) switching to bright field mode to observe the physical binding distribution of magnetic beads and NK cells; and (d) continuing to capture bright field images at different focal planes or angles to further confirm the coupling effect and stability between magnetic beads and NK cells.

[0067] In another embodiment, it is assumed that the concentration of NK cells after staining is 5 × 10⁻⁶. 5 For pairing, 500 μL of a 50 nM aptamer magnetic bead solution was used. The mixture was incubated with shaking for 42 minutes at 36.8 °C. After magnetic separation, the cells were resuspended in 200 μL of PBS buffer, and 10 μL was added to a glass slide. Fluorescence imaging parameters were as follows: FITC channel exposure time was 100 ms for Calcein image acquisition, and PI channel exposure time was 120 ms for dead cell signal acquisition; bright-field imaging was performed using a 40× objective lens with a resolution of 2048 × 2048 pixels. In the obtained images, Calcein staining showed green fluorescence in live cells, PI staining showed red dead cell signal, and bright-field images showed that most NK cells had glycosyl magnetic beads attached to their surface, indicating good aptamer coupling efficiency.

[0068] Of particular importance is the photographing of the obtained product under a fluorescence microscope, including: Before photographing the obtained product under a fluorescence microscope, the cell mixture was slowly added dropwise to the center of the slide and left to stand for 30 seconds to allow the cells to settle naturally. In one embodiment, it is assumed that the volume of the cell mixture added is 12 μL and the cell concentration is 2 × 10⁻⁶. 6 With a cell density of 10 cells / mL, after standing for 30 seconds, it is estimated that approximately 24,000 cells will settle on the surface of the slide. The cells are evenly distributed and do not show obvious aggregation, providing a good foundation for subsequent image analysis.

[0069] During the photography process, different observation channels were set up: (a) the FITC channel was turned on to observe and capture green fluorescence images of NK cells stained with Calcein, which was used to determine the activity status of NK cells; (b) the PI channel was switched to capture red fluorescence images of NK cells stained with PI, which was used to detect non-viable cells; (c) and (d) the fluorescence channel was turned off, and the bright field mode was switched to continuously focus and capture images of the coupling state between glycosyl magnetic beads and NK cells, recording the distribution, binding sites and density of microspheres; after all images were captured, they were uniformly numbered and saved for subsequent analysis and comparison.

[0070] In one embodiment, it is assumed that the images obtained through the above steps include approximately 50 FITC channels, approximately 45 PI channels, and approximately 60 images captured by continuous focusing in bright field mode. The distribution of glycosyl magnetic beads is mainly concentrated in the 30%-40% area of ​​the cell surface, with a binding density in the range of 10-15 beads / 100μm². All images are numbered and managed to ensure data integrity and traceability of analysis.

[0071] Preferably, this specification also provides a magnetically targeted NK cell delivery data processing system for performing the magnetically targeted NK cell delivery data processing method described above. The magnetically targeted NK cell delivery data processing system includes: The aptamer generation module 101 is used to query the immune receptor protein structure of natural killer cells; inject sodium chloride solution and add the immune receptor protein structure to generate aptamer structure through reaction. The optimized aptamer generation module 102 is used to combine streptavidin into pre-prepared glycosyl magnetic microspheres to obtain high-affinity magnetic beads; the high-affinity magnetic beads are molecularly docked with the aptamer structure, the docking score is recorded, and the optimized aptamer is generated. The image acquisition module 103 is used to record the magnetic bead mass ratio of the optimized aptamer; natural killer cells are captured using the optimized aptamer, and fluorescence microscopic images are acquired. The delivery efficiency analysis module 104 is used to record the staining of fluorescence microscopy images, determine the survival status of NK cells, mark the location information of live cells, and generate an NK cell delivery efficiency table using the magnetic bead mass ratio.

[0072] Therefore, the embodiments should be considered as exemplary and non-limiting in all respects, and the scope of the invention is defined by the appended claims rather than the foregoing description. Thus, all variations falling within the meaning and scope of the equivalents of the application are intended to be included within the invention.

[0073] The above description is merely a specific embodiment of the present invention, enabling those skilled in the art to understand or implement the invention. Various modifications to these embodiments will be readily apparent to those skilled in the art, and the general principles defined herein may be implemented in other embodiments without departing from the spirit or scope of the invention. Therefore, the invention is not to be limited to the embodiments shown herein, but is to be accorded the widest scope consistent with the principles and novel features of the invention herein.

Claims

1. A data processing method for NK cell delivery based on magnetic targeting, characterized in that, Includes the following steps: Step S1: Locate the structure of immune receptor proteins in natural killer cells; A sodium chloride solution was injected, and an immune receptor protein structure was added to generate an aptamer structure. Step S2: In the pre-prepared glycosyl magnetic microspheres, streptavidin is combined to obtain high-affinity magnetic beads; the high-affinity magnetic beads are molecularly docked with the aptamer structure, the docking score is recorded, and the optimized aptamer is generated; Step S3: Record the magnetic bead mass ratio of the optimized aptamer; capture natural killer cells using the optimized aptamer and obtain fluorescence microscopy images; Step S4: Record the staining of the fluorescence microscopy images, determine the survival status of NK cells, mark the location information of live cells, and generate an NK cell delivery efficiency table using the magnetic bead mass ratio.

2. The data processing method for NK cell delivery based on magnetic targeting according to claim 1, characterized in that, In step S1, sodium chloride solution is added, and an immune receptor protein structure is introduced to generate an aptamer structure, including: Add sodium chloride solution to the simulated environment; The electrolyte strength of sodium chloride solution was calculated, immune receptor protein structures were added, charge rearrangement was simulated, changes in protein structure were recorded, and aptamer structures were generated.

3. The data processing method for NK cell delivery based on magnetic targeting according to claim 1, characterized in that, Step S2, which involves binding streptavidin to pre-prepared glycosyl magnetic microspheres to obtain high-affinity magnetic beads, includes: Mix the glycosyl magnetic microspheres using a vortex mixer, place them in a centrifuge tube, add buffer solution, and vortex to mix. Place the tube on a magnetic separator to enrich the magnetic beads, remove the supernatant, and repeat the washing process with buffer solution. Add carboxyl activator and N-hydroxysuccinimide, vortex to mix, and incubate at room temperature for 0.5-0.7 h to activate the carboxyl groups on the surface of the magnetic beads and bind streptavidin to obtain high-affinity magnetic beads.

4. The data processing method for NK cell delivery based on magnetic targeting according to claim 3, characterized in that, The glycosyl magnetic microspheres were mixed using a vortex mixer, placed in a centrifuge tube, and buffer solution was added. The mixture was then vortexed to mix. Before mixing, place the glycosyl magnetic microspheres in a clean centrifuge tube and vortex them continuously at a speed of 1500rpm-2000rpm for 10-15 seconds. During the addition of buffer solution, a staged injection method is adopted. In the first stage, 30%-40% of the total volume of buffer solution is added, and vortexing is performed again for 5-7 seconds to form a preliminary wetting layer on the surface of the microspheres. In the second stage, add the remaining 60%-70% of the buffer solution. Immediately after adding the buffer solution, perform a second vortexing for at least 15 seconds to form a stable suspension.

5. The data processing method for NK cell delivery based on magnetic targeting according to claim 3, characterized in that, The process of enriching magnetic beads includes: During the enrichment process, the sample solution to be tested is injected into a centrifuge tube pre-filled with magnetic microspheres, and the injection rate is controlled at 1-2 mL / min. A magnetic device is installed on the outside of the centrifuge tube. By applying a stable magnetic field, magnetic microspheres are directionally adsorbed from the solution to one side of the tube wall, forming a visible magnetic bead aggregation zone.

6. The data processing method for NK cell delivery based on magnetic targeting according to claim 3, characterized in that, Combining streptavidin to obtain high-affinity magnetic beads includes: After the magnetic particles in the centrifuge tube settle, remove the liquid on top of the centrifuge tube, wash repeatedly with buffer solution, add streptavidin, continue to vortex mix, and place in an environment of 35℃-37℃ to rotate and mix for reaction. After the reaction is complete, place the centrifuge tube on a magnetic separator, remove the supernatant, add the blocking solution, and rotate to mix the reaction at 35℃-37℃. After mixing and reaction, the high-affinity magnetic beads are preserved using a preservation solution.

7. The data processing method for NK cell delivery based on magnetic targeting according to claim 1, characterized in that, Step S2 involves molecular docking of the high-affinity magnetic beads with the aptamer structure, including: After centrifuging the high-affinity magnetic beads at 10000-12000g for 5-7 minutes, molecular docking was performed with the aptamer structure. Sterile water was added for fusion, and the mixture was placed in a freezer for 5-10 minutes. The docking score was recorded to generate an optimized aptamer with a stable tertiary structure.

8. The data processing method for NK cell delivery based on magnetic targeting according to claim 7, characterized in that, The recorded docking score includes: Different amounts of high-affinity magnetic beads (0.2 nmol-1.0 nmol) were added to the prepared high-affinity magnetic beads for optimization. The beads were rotated and incubated at a preset temperature for 0.5 h-0.8 h, and the mass ratio of the magnetic beads was recorded. Add 0.6 nmol-0.8 nmol of biotinylated high-affinity magnetic beads to the prepared high-affinity magnetic beads, and incubate them by rotation for 0.25 h-2.5 h under preset temperature conditions for optimization, and record the optimization results; Among them, the magnetic bead mass ratio and the incubation optimization results are used as docking scores.

9. The data processing method for NK cell delivery based on magnetic targeting according to claim 1, characterized in that, Step S3 involves using optimized aptamers to capture natural killer cells and obtaining fluorescence microscopy images, including: Thaw the frozen natural killer cells at 35℃-37℃, centrifuge and discard the supernatant, wash with buffer, add the pre-prepared cell live / dead staining combination solution, in which the ratio of fluorescein ethyl ester: propidium iodide: buffer is 1:1:1000, and incubate at 36℃-37℃ in the dark for 25min-30min. The prepared optimized aptamer was mixed with the stained natural killer cell suspension and incubated with shaking at 35℃-37℃ for 40-45 minutes. The supernatant was removed magnetically, and the mixture was resuspended in buffer solution. The resulting product was photographed under a fluorescence microscope.

10. A magnetically targeted NK cell delivery data processing system, characterized in that, For performing the magnetically targeted NK cell delivery data processing method as described in claim 1, the magnetically targeted NK cell delivery data processing system comprises: The aptamer generation module is used to query the immune receptor protein structure of natural killer cells; sodium chloride solution is injected, and the immune receptor protein structure is added to generate the aptamer structure. An optimized aptamer generation module is used to combine streptavidin into pre-prepared glycosyl magnetic microspheres to obtain high-affinity magnetic beads; the high-affinity magnetic beads are then molecularly docked with the aptamer structure, the docking score is recorded, and the optimized aptamer is generated. The image acquisition module is used to record the magnetic bead mass ratio of the optimized aptamer; natural killer cells are captured using the optimized aptamer, and fluorescence microscopic images are obtained. The delivery efficiency analysis module is used to record the staining of fluorescence microscopy images, determine the survival status of NK cells, mark the location information of live cells, and generate an NK cell delivery efficiency table using the magnetic bead mass ratio.

Images