A blood purification adsorption catheter and its preparation method

Abstract

This invention belongs to the field of biomedical adsorption materials technology, and discloses a blood purification adsorption catheter and its preparation method. The invention uses polydimethylsiloxane polymer as the main body, with two elliptical intersecting blades as a mixer structural unit. After curing with PDMS polymer, a catheter body with a specific mixer structure embedded in the inner wall is prepared. Subsequently, the inner wall of the catheter is modified by reversible addition-fracture transfer radical polymerization to form a polymer brush layer. By coupling nanoantibodies to the polymer brush, a catheter material with antigen adsorption function is prepared. The static mixer structure in the catheter material prepared by this invention can significantly enhance the diffusion and convection of target toxin molecules in the fluid, effectively improving the efficiency of specific adsorption of toxin molecules by the antibody-functionalized catheter inner wall, and is expected to be applied in clinical blood purification treatment for the removal of medium and large molecular toxins, including inflammatory factors.

Description

Technical Field

[0001] This invention belongs to the field of biomedical adsorption materials technology, specifically relating to a blood purification adsorption conduit with a static mixer structure for removing inflammatory factor-type blood toxins and its preparation method. Background Technology

[0002] Inflammatory factors are highly active, soluble protein molecules with multiple functions, including mediating and regulating immune, inflammatory, and hematopoietic processes. They can be classified into interleukins (ILs), interferons (IFNs), tumor necrosis factor superfamily (TNF), colony-stimulating factors (CSFs), chemokines, and growth factors (GFs). Studies have shown that inflammatory factors play an important role in the development of various diseases such as psoriasis, arthritis, and severe infectious diseases like sepsis. In patients with these diseases, the levels of inflammatory factors are elevated tens or even hundreds of times in the blood, and the removal of these inflammatory factors leads to better clinical prognosis.

[0003] Currently, clinical blood purification therapy for removing large molecular toxins such as inflammatory factors mainly includes hemodialysis and perfusion techniques. These methods primarily utilize principles such as convection, diffusion, and hydrophobic adsorption to remove toxin molecules. However, these methods are non-selective in removing blood toxins; that is, blood components with similar molecular properties can also be broadly removed. Therefore, the clinical removal effect of existing methods on inflammatory factors is not ideal, and multiple methods, such as convection, diffusion, and perfusion, are usually combined to maximize the removal effect. Even so, existing non-specific removal methods still have low efficiency in removing inflammatory factor toxins. Studies have shown that after a single treatment, the removal rate of inflammatory factors is only about 5%-30%. Due to the continuous production of inflammatory factors, the level of inflammatory factors in the body of critically ill patients may even increase after treatment. Furthermore, the combined use of multiple treatment techniques increases the amount of blood in the patient's extracorporeal circulation, which may increase the risk of bleeding and hemodynamic instability. Therefore, developing a novel catheter material that combines blood transfusion and toxin removal functions in blood purification therapy could effectively reduce the treatment risks for patients.

[0004] To enable materials to selectively recognize and bind to blood toxins, thereby improving the removal efficiency of toxin molecules, this invention uses nanobodies as functional ligands to functionalize the inner wall of catheter materials. Nanobodies are the smallest known antibody fragments capable of specifically recognizing antigen molecules, possessing advantages such as small molecular weight, high affinity, good stability, and easy preparation at low cost. Currently, blood perfusion adsorbents using anti-β2M nanobodies as ligands and agarose gel as a medium have been reported, demonstrating significant specific removal effects on β2M [C. Huang, J. Ren, F. Ji, S. Muyldermans, L. Jia, Nanobody-Based high-performance immunosorbent for selective beta2-microglobulin purification from blood, Acta Biomaterialia 107(2020)232-241.]. Due to the inherent physicochemical properties of medium and large protein molecules, their diffusion in blood is slow. When blood flows through a catheter, it may not be able to diffuse to the inner wall of the catheter modified with nanobodies in a short time, thus affecting binding and removal. Static mixers effectively mix and agitate fluids flowing through pipes, improving mass transfer of target molecules and increasing contact between molecules and the functionalized pipe wall, thereby further enhancing removal efficiency. By combining nanobodies with functionalized modifications and static mixers to prepare catheter materials, highly efficient and specific removal of large molecular toxins such as inflammatory factors from blood can be achieved. Summary of the Invention

[0005] This invention addresses the shortcomings of existing blood purification treatments, such as insufficient and inefficient removal of large molecular toxins among inflammatory factors. It provides a specific inflammatory factor adsorption catheter using nanobodies as functional ligands. By immobilizing and modifying nanobodies on the inner wall of the catheter and incorporating a special static mixer structure, effective mixing of the blood flowing through the catheter is achieved, enhancing mass transfer of the target toxin molecules. This increases the effective contact and specific binding between the nanobodies modified on the inner wall of the catheter and the target toxin molecules, thereby strengthening the toxin removal effect. In vitro perfusion experiments show that the catheter of this invention can efficiently and accurately remove inflammatory factors from the blood, and can combine blood transfusion and toxin removal functions in blood purification therapy.

[0006] A blood purification adsorption catheter and its preparation method include the following steps:

[0007] (1) Pour Sylgard 184 premixed adhesive into the sandwich formed by the peelable shell and the cylindrical mold; after curing, remove the shell, immerse the cured polymer material and the mold in an organic solvent, and swell at room temperature so that the volume of the cured material expands to facilitate the removal of the mold. After removing the mold, dry the tubular material to remove the residual organic solvent and restore the volume before swelling to obtain a conduit with a mixer unit.

[0008] Sylgard184 premixed adhesive comprises two components: dimethylsiloxane monomer and crosslinking agent. The two are mixed at a weight ratio of 10:1 and cured to form polydimethylsiloxane (PDMS). The curing temperature is room temperature to 100°C and the curing time is 1 to 5 hours. In the preferred case, curing at 60°C for 2 hours is selected.

[0009] The conduit with mixer units comprises multiple mixer units evenly arranged within it. Each mixer unit consists of two intersecting, spaced blades. The blades are shaped like halves of an ellipse divided along its major axis, which is defined as the blade length. The two intersecting blades are of opposite lengths and form an intersecting angle. The formula for calculating the blade length and the intersecting angle is as follows: Blade length = Inner diameter / Sin(90° - 0.5 × intersecting angle). Preferably, the intersecting angle between the two blades within a single mixer unit is 90°, and the width of each mixer unit is 7.4 mm. Therefore, the minimum center-to-center spacing between adjacent mixer units is 7.4 mm, and preferably 18.5 mm.

[0010] The conduit with mixer units is formed by using a pre-designed mold made of cylindrical hard photolithography resin material manufactured by 3D printing technology. The inner surface of the mold contains grooves that correspond to the blade structure. After casting with PDMS, the mold is removed, and corresponding protruding mixer units are generated on the inner wall of the PDMS conduit.

[0011] The organic solvent is any one of toluene, kerosene, dichloromethane, and chloroform, with toluene being preferred. The swelling time is 0.5-2 hours, preferably 1 hour.

[0012] The drying process can be either ambient temperature and pressure drying or vacuum-controlled ambient temperature drying, with vacuum-controlled ambient temperature drying being preferred.

[0013] (2) The outer wall of the conduit with the mixer unit is wrapped and hydrophilicated in a plasma cleaner. Then, the inner wall of the conduit is aminated using a silane coupling agent. The modified amino group is used to fix and modify the RAFT chain transfer agent onto the inner surface of the conduit.

[0014] The hydrophilization treatment uses high-purity oxygen as the gas, and the time is based on the water contact angle of the material surface, which is less than 40°. Furthermore, the plasma hydrophilization treatment time is 1-5 minutes, with a power of 100-250 watts, preferably 2 minutes and 150 watts.

[0015] The silane coupling agent is 3-aminopropyltrimethoxysilane, and anhydrous ethanol of the silane coupling agent with a volume fraction of 5-20% (preferably 10%) is used as the amination agent, and the amination time is 5 hours.

[0016] The RAFT chain transfer agent is 4-cyano-4-(thiobenzoylthio)valeric acid. The coupling reaction of the RAFT chain transfer agent is carried out in MES buffer at pH=6 using the carbodiimide method (NHS-EDC). The concentration of the RAFT chain transfer agent in the reaction system is 5-30 mg / mL, and the coupling reaction time is 12-24 hours, preferably 20 mg / mL, with a coupling reaction time of 12 hours.

[0017] (3) The catheter obtained in step (2) is prepared by RAFT polymerization under anaerobic conditions to form a catheter with polymer brush modification on the inner wall;

[0018] The monomer used in the RAFT polymerization reaction is hydroxyethyl methacrylate (HEMA), and the reaction time is 12-48 hours, preferably 24 hours.

[0019] (4) The catheter obtained in step (3) is immersed in an acetone solution containing a chemical activating agent for activation. Then the activated material is taken out and immersed in a phosphate buffer (pH 7.4) containing nanobodies for ligand coupling. After washing, the final product is obtained.

[0020] The chemical activating agent is N,N'-carbonyldiimidazole (CDI), with a CDI concentration of 0.05-0.2 mol / L; the activation time is at least 2 hours. All cleaning processes involve at least three thorough cleanings.

[0021] The nanobody is an antibody protein capable of specifically recognizing inflammatory factors, with an affinity constant of 10. -9 For concentrations above M, the nanobody concentration ranges from 0.1 to 4.5 mg / mL during conjugation, and the conjugation time is 5-24 hours. Preferably, the concentration is 0.5 mg / mL and the conjugation time is 12 hours.

[0022] The catheter material prepared using the above method can enhance blood disturbance and mixing mass transfer through its static mixer structure when blood flows through it. This increases the contact between target molecules in the blood and the functional nanobodies coupled to the inner wall, thereby improving the binding and adsorption of target molecules on the inner wall surface. This catheter material can be used for the specific removal of harmful protein toxins such as inflammatory factors in blood purification therapy.

[0023] Another aspect of the present invention is to protect the adsorption catheter material prepared by the preparation method described above, and the application of the catheter material in removing macromolecular toxoids from inflammatory factors in blood purification.

[0024] The beneficial effects of this invention are as follows: This invention uses a specially designed mold to manufacture a static mixer PDMS conduit material with cross-shaped blades on the inner wall, which enhances the mixing and mass transfer of fluids flowing through the conduit. By immobilizing nanobodies on the inner wall of the conduit, it can be applied to blood purification therapy to specifically remove corresponding inflammatory cytokine toxins from the blood, which is helpful in the treatment of severe infections or diseases related to cytokine storms. Attached Figure Description

[0025] Figure 1 This is a schematic diagram of the catheter structure of the present invention.

[0026] Figure 2 The graph shows the relationship between the blade crossing angle, the mixing coefficient calculated by fluid simulation, and the inlet and outlet pressure difference of the duct material.

[0027] Figure 3 The graph shows the relationship between the mixing coefficients obtained from fluid simulation calculations and different mixer unit spacings.

[0028] Figure 4 a is a flowchart of surface RAFT polymerization to form pHEMA molecular brush modification and nanoantibody conjugation; Figure 4 b is an atomic force microscope image of blank PDMS material; Figure 4 c is an atomic force microscope image of the PDMS material after RAFT polymer brush layer modification.

[0029] Figure 5 The graph shows the relationship between the adsorption capacity of IL-17A on nanobody conjugates of different concentrations and planar PDMS materials.

[0030] Figure 6 a is a schematic diagram of the process for manufacturing conduit materials using molds; Figure 6 Image b shows the conduit material with four different mixer unit spacings manufactured.

[0031] Figure 7 a is a schematic diagram of a simulated extracorporeal blood perfusion therapy experiment; Figure 7Figure b shows a comparison of the adsorption performance of four different mixer unit spacer materials.

[0032] Figure 8 This image shows the adsorption effect of the catheter material on inflammatory factors at different flow rates.

[0033] In the figure: 1. The conduit wall is 1 mm thick; 2. The semi-elliptical mixer blades are 1 mm thick; 3. The vertical distance between the two intersecting blades is fixed at 0.5 mm; 4. The intersecting angle of the two intersecting blades in space; 5. The inner diameter of the conduit is fixed at 6 mm. Detailed Implementation

[0034] The following non-limiting embodiments are intended to enable those skilled in the art to more fully understand the present invention, but do not limit the invention in any way. Unless otherwise specified, the experimental methods used in the embodiments are conventional methods, and the materials and reagents used are commercially available unless otherwise specified.

[0035] To facilitate understanding of this invention, the terminology used is defined as follows. In the event of any inconsistency between the terms and phrases mentioned and their commonly known meanings, the meaning as expressed in this invention shall prevail.

[0036] As described in this invention, the term "mixer unit" refers to a structural unit consisting of two guide vanes intersecting at a certain angle, embedded in the inner wall of a pipe. In this invention, "interval between adjacent mixer units" refers to the distance between the center points of two adjacent mixer units.

[0037] The theoretical range of the intersection angle between the two guide vanes in the aforementioned mixer unit is 0 to 180°. By using fluid simulation technology, the mixing effect of the mixer can be optimized by changing the intersection angle of the two vanes in the mixer unit and the spacing between adjacent mixer units, thereby improving the adsorption effect of the conduit material on the target molecules.

[0038] Example 1

[0039] like Figure 1 As shown, the conduit material of the present invention consists of a conduit body 1 and a mixer unit 2 embedded in the inner wall of the conduit. The outer diameter of the conduit is 8 mm, and the inner diameter is 6 mm. The mixer unit 2 consists of two spatially intersecting semi-elliptical blades, and the spatial distance 3 between the two intersecting blades is set to 0.5 mm. The thickness of the blades is set to 1 mm. When the aforementioned four parameters are fixed, the change in the spatial intersection angle 4 of the two blades in the mixer unit can cause a change in the length 2 of the major axis of the semi-elliptical blades. The calculation formula between the length 2 of the blade major axis and the spatial intersection angle 4 of the two blades is as follows: length 2 of the blade major axis = inner diameter of the conduit / Sin(90° - 0.5 × intersection angle 4).

[0040] like Figure 1As shown, in this embodiment, a series of conduit models were constructed using Solidworks modeling software. Each model contains two mixer units, which are rotated 90° along the conduit's central axis. Only the blade crossing angle (4°) within the mixer units is changed across different models, while all other parameters remain the same, resulting in a series of conduit material models with mixer units possessing different parameters. Ansys Fluent 2021R2 software was used to perform fluid simulation calculations on the models, and the mixing coefficient for the fluid along the entire conduit length was calculated using the following formula:

[0041]

[0042] In the formula, v and w represent the values ​​when the fluid flows through Figure 1 The duct shown in the front view represents the velocity components of the fluid in the x and y directions at a certain point within the duct, where k (1~n) represents the set of all points within the duct. In addition, the pressure drop at the inlet and outlet of the pipe can be directly calculated by simulation software. After fluid simulation calculations, the mixing efficiency of the duct and the pressure drop at the inlet and outlet of the pipe for models with different mixer blade cross angles are as follows: Figure 2 As shown. In blood purification therapy, for safety reasons, it is necessary to avoid excessive pressure drop across the catheter to prevent irreversible damage to the patient. Based on simulation calculations of mixing efficiency, a blade crossover angle of 90° maintains a high level of mixing efficiency while minimizing pressure drop. Therefore, 90° was chosen as the crossover angle between the two blades in the mixer unit.

[0043] Example 2

[0044] In this embodiment, the cross angle of the two blades in the mixer unit obtained in Embodiment 1, which is 90°, is used as a fixed parameter to optimize the spacing between adjacent mixer units. The specific optimization method is as follows:

[0045] like Figure 1 As shown, the conduit material consists of a conduit body 1 and a mixer unit 2 embedded in the inner wall of the conduit. The outer diameter of the conduit is 8 mm, and the inner diameter is 6 mm. Each mixer unit consists of two spatially intersecting elliptical blades, with a spatial distance 3 of 0.5 mm between the two intersecting blades. The blade thickness is 1 mm. Solidworks modeling software was used for modeling. Each model contains two mixer units, with the two blades within each mixer unit intersecting at a 90° angle. Adjacent mixer units are rotated 90° along the central axis of the conduit, changing only the spacing between adjacent mixer units to obtain a series of models. Ansys Fluent 2021 R2 software was used to perform fluid simulation calculations on the models, and the mixing coefficient of the fluid within the conduit length was calculated using the following formula:

[0046]

[0047] In the formula, v and w represent the values ​​when the fluid flows through Figure 1 The duct shown in the front view represents the velocity components of the fluid in the x and y directions at a certain point inside the duct, where k (1~n) represents the set of all points within the duct. Fluid simulation results for duct models with different mixer element spacing are shown below. Figure 3 As shown in the figure. Simulation results indicate that when the mixer unit spacing is 18.5 mm or greater, the mixing efficiency reaches its peak and remains at a relatively stable level. Since the adsorption effect of toxins is related not only to the mixing efficiency but also to the inner surface area of ​​the conduit, a larger inner surface area allows for greater nano-antibody conjugation, thus further improving toxin removal. Therefore, compared to other mixer unit spacings greater than 18.5 mm, selecting 18.5 mm as the mixer unit spacing maintains a relatively good mixing effect while ensuring the maximum number of mixers per unit length and the largest inner surface area.

[0048] Example 3

[0049] This example uses a planar PDMS material prepared with Sylgard 184 premixed adhesive for surface RAFT polymer brush coating modification. This includes, for example... Figure 4 Steps shown in a:

[0050] (1) Preparation of planar PDMS. After uniformly mixing the two components of Sylgard 184 premixed adhesive A and B at a mass ratio of 10:1, pour the total weight of the mixed adhesive (7g) into a clean round-bottomed plate and cure at 60℃ for 2 hours. Remove the material and cut the planar PDMS into 1cm×1cm pieces using a utility knife for later use.

[0051] (2) The planar PDMS obtained in step (1) was subjected to plasma hydrophilization treatment, with high-purity oxygen treatment for 1 minute and power of 150 watts. The treated PDMS material was then quickly immersed in an ethanol solution of 10% by volume of 3-aminopropyltrimethoxysilane and reacted with magnetic stirring at 150 rpm for 5 hours at room temperature.

[0052] (3) Take the material prepared in step (2) above and soak it in MES buffer at pH 6 containing 20 mg / mL 4-cyano-4-(thiobenzoylthio)valerate, 1 mol / L N-hydroxysuccinimide (NHS) and 2 mol / L 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC), and incubate at 35°C in the dark with shaking for 12 hours.

[0053] (4) After thoroughly washing the material from step (3) with anhydrous ethanol, immerse it in anhydrous ethanol containing 1 mol / L HEMA monomer and 10 mg / mL 4,4'-azobis(4-cyanopentanoic acid). Deoxygenate with high-purity nitrogen for 30 minutes, then place the above reaction system in an oil bath and heat at 70°C, stirring at 150 rpm for 12-48 hours. After the reaction, characterize the surface of the material using atomic force microscopy (AFM).

[0054] AFM characterization results are as follows Figure 4 As shown, compared to unmodified PDMS ( Figure 4 (b) The surface roughness of the material increased significantly after the RAFT polymerization reaction, indicating that the pHEMA polymer brush layer was successfully modified onto the PDMS surface. Table 1 shows the measured surface roughness values ​​of PDMS after RAFT polymerization at different times. After 24 hours of polymerization, the average roughness reached 155 nm ( Figure 4 c). Extending the polymerization time to 48 hours did not significantly increase the average roughness. Therefore, 24 hours was subsequently used as the RAFT polymerization reaction time.

[0055] Example 4

[0056] Based on the optimal polymerization time determined in Example 3 (24 hours), this example uses planar PDMS material and performs RAFT polymerization for 24 hours to modify the surface polymer brush layer. In this example, human interleukin-17A (IL-17A) is used as a model inflammatory factor toxin for removal, and correspondingly, anti-human IL-17A nanobodies are used as ligands to prepare the catheter material. The anti-human interleukin-17A (IL-17A) nanobodies are conjugated and optimized on the polymer brush-modified PDMS catheter material, mainly including the following steps:

[0057] (1) Anti-human IL-17A nanobody was manufactured according to the method described in the patent application number 2023106819650.

[0058] (2) Prepare planar PDMS and perform RAFT surface modification: The method is the same as in Example 3.

[0059] (3) The RAFT-modified 1cm×1cm planar PDMS material was immersed in acetone with a CDI concentration of 0.1mol / L and incubated at 35°C with shaking for 8 hours. Then, the material was quickly rinsed with deionized water and immersed in phosphate buffer containing anti-human IL-17A nanobodies, with the nanobodies concentrations set at 0.05, 0.2, 0.5, 1.5, and 4.5 mg / mL, respectively. After incubation at room temperature with shaking for 12 hours, the nanobodies-conjugated material was rinsed three times with phosphate buffer and stored at 4°C.

[0060] (4) Dilute antigen IL-17A to 25 μg / mL with phosphate buffer. Cut the planar PDMS material conjugated with nanobody in step (3) into small pieces and immerse them in the 25 μg / mL IL-17A solution, shaking and incubating for 1 hour. Then wash the material three times with phosphate buffer containing Tween 20, and then incubate the material with 100 μL glycine hydrochloride buffer (pH 1.5) with shaking for 5 minutes. Determine the IL-17A content in glycine hydrochloride buffer using the Coomassie brilliant blue method.

[0061] The adsorption capacity of IL-17A per square centimeter of nanobody-functionalized material can be calculated based on the IL-17A content eluted from glycine hydrochloric acid solution. The relationship between different nanobody conjugation concentrations and the adsorption capacity of the material for IL-17A is shown below. Figure 5 As shown, it can be observed that when the conjugation concentration of the nanobody is 0.5 mg / mL, the resulting material exhibits the highest adsorption capacity for IL-17A, reaching 7 μg / mL. 2 about.

[0062] Example 5

[0063] Based on the simulation results of Example 2, this example uses molds of different specifications to prepare four different mixer unit spacing conduit materials, which are used to further describe and clarify the preparation and function of the mixer conduit materials of the present invention. (Including...) Figure 6 The following steps are shown in section a:

[0064] (1) Select four molds with a diameter of 6 mm and an adjacent mixer unit spacing of 7.4, 11.1, 14.8 and 18.5 mm. Each mold also has 3 mixer unit structures. Insert the molds into the detachable tubular shell with an inner diameter of 8 mm. Pour Sylgard 184 premixed adhesive that has been mixed evenly and de-bubbled into the sandwich formed by the shell and the mold. Cure at 60°C for 2 hours.

[0065] (2) After peeling off the outer shell, place the remaining mold and the cured gel material into toluene and let it swell at room temperature for 1 hour; then remove the mold.

[0066] (3) The swollen catheter material was dried in a room temperature vacuum oven to remove the residual toluene reagent. The volume of the catheter material was restored, and four different specifications of catheter materials were obtained.

[0067] Different specifications of catheters prepared, such as Figure 6 As shown in b, the spacing between adjacent mixing units differs significantly among the four types of conduits, and the mixer unit composed of cross blades corresponds well with the groove in the mold (white). The results indicate that conduit materials with embedded mixer unit structures can be successfully prepared using the above method.

[0068] Example 6

[0069] In this embodiment, four types of catheters with different mixer unit spacings prepared in Example 5 were used for RAFT polymer brush modification and anti-human IL-17A nanobody functionalization. The effectiveness of these nanobodies in adsorbing IL-17A from blood was further elucidated and verified. The steps are as follows:

[0070] (1) Preparation of conduit materials for four different mixer unit spacings:

[0071] The method is the same as in Example 5.

[0072] (2) The catheter material obtained in step (1) above is modified with a RAFT polymer brush and conjugated with an anti-human IL-17A nanobody:

[0073] The method is the same as in Examples 3 and 4, except that the concentration of the nanobody conjugation is 0.5 mg / mL.

[0074] (3) Use the catheter material obtained in step (2) above to conduct an irrigation experiment as follows: Figure 7 a) 50 mL of human blood supplemented with human IL-17A was placed in a beaker and slowly stirred with a magnetic stirrer. The catheter was then connected to the peristaltic pump tubing. The perfusion rate was 50 mL / min, and the perfusion time was 1 hour. The adsorption performance of four catheters with different mixer unit spacings for IL-17A was compared by comparing the human IL-17A content in the blood before and after perfusion.

[0075] The adsorption results of the catheter are as follows Figure 7 As shown in b, compared with the empty conduit material without a mixer, under the same conditions, the adsorption effect of the four conduits with different adjacent mixer unit spacings is significantly enhanced for IL-17A. The adsorption effect is best when the adjacent mixer unit spacing is equal to 18.5 mm. This result indicates that under this specification, the static mixer has the best mixing efficiency for fluid flowing through the conduit, which is basically consistent with the results calculated in Example 2.

[0076] Example 7

[0077] This embodiment simulates the blood perfusion rate in a clinical treatment scenario to verify the adsorption effect of the blood purification adsorption catheter material with a static mixer structure of the present invention on blood IL-17A. The experimental steps are as follows:

[0078] (1) Preparation of catheter material: In this example, catheter material with an interval of 18.5 mm between adjacent mixer units was selected for the experiment.

[0079] The method is the same as in Example 1.

[0080] (2) The catheter material obtained in step (1) above is subjected to RAFT modification and nanobody conjugation:

[0081] The method is the same as in Examples 3 and 4, except that the concentration of nanoantibody conjugate is 0.5 mg / mL.

[0082] (3) Perform perfusion adsorption experiments using the catheter material obtained in step (2) above. Place 50 mL of human blood containing human IL-17A into a beaker and stir slowly with a magnetic stirrer. Connect the catheter to the peristaltic pump tubing as follows. Figure 7 As shown in Figure a, the peristaltic pump flow rates were set to 50, 100, and 150 mL / min, corresponding to cycle times of 1 hour, 30 minutes, and 20 minutes, respectively, to ensure a consistent number of cycles. After adsorption, the concentration of IL-17A in the blood before and after adsorption was measured.

[0083] The adsorption results of blood IL-17A at different flow rates are as follows: Figure 8 As shown, the specific removal rate was over 85% when the flow rate was 50 ml / mL; however, the specific removal rate of IL-17A by the catheter decreased with increasing flow rate. Considering that the blood perfusion time in clinical practice is usually much longer than the perfusion time in this embodiment, there is still considerable room for improvement in the removal effect of the catheter material on IL-17A at the above three flow rates in actual clinical applications.

[0084] Table 1 shows the surface roughness of the polymer brush layer corresponding to different RAFT polymerization times.

[0085] Aggregation time (hours) 12 24 48 Roughness (nanometers) 129.6±7.6 155.3±6.0 158.0±10.8 .

Claims

1. A method for preparing a blood purification adsorption catheter, characterized in that, Includes the following steps: (1) Pour Sylgard 184 premix into the sandwich formed by the peelable shell and the cylindrical mold; After curing, the outer shell is removed, and the cured polymer material and the mold are immersed in an organic solvent and swelled at room temperature. This allows the cured material to expand in volume, making it easier to remove the mold. The tubular material after removing the mold is dried to remove the residual organic solvent and restore its original volume before swelling, resulting in a conduit with a mixer unit. (2) Wrap the outer wall of the conduit with the mixer unit, hydrophilize it in a plasma cleaner, and then use a silane coupling agent to amide the inner wall of the conduit; use the modified amino group to fix and modify the RAFT chain transfer agent on the inner surface of the conduit. (3) The catheter obtained in step (2) is prepared into a catheter with a polymer brush modified on the inner wall by RAFT polymerization under anaerobic conditions; the monomer used in the RAFT polymerization reaction is hydroxyethyl methacrylate. (4) The catheter obtained in step (3) is immersed in an acetone solution containing a chemical activating agent for activation. Then the activated material is taken out and immersed in a phosphate buffer containing nanobodies at pH 7.4 for ligand coupling. After washing, the final product is obtained. The aforementioned conduit with mixer units consists of multiple mixer units evenly arranged within the conduit. Each mixer unit comprises two intersecting, spaced blades. The blades are shaped as half-structures divided along the major axis of an ellipse, with the major axis defined as the blade length. The two intersecting blades are of opposite lengths and form an intersecting angle. The formula for calculating the blade length and the intersecting angle is as follows: Blade length = Inner diameter / Sin(90° - 0.5 × intersecting angle). The conduit with the mixer unit is formed by means of a pre-designed mold made of cylindrical hard photolithography resin material manufactured by 3D printing technology. The inner surface of the mold contains grooves opposite to the blade structure. After being cast with PDMS, the mold is removed, and corresponding protruding mixer units are generated on the inner wall of the PDMS conduit.

2. The method for preparing a blood purification adsorption catheter according to claim 1, characterized in that, The Sylgard184 premixed adhesive comprises two components: dimethylsiloxane monomer and crosslinking agent. The two are mixed in a weight ratio of 10:1 and cured to form polydimethylsiloxane. The curing temperature is room temperature to 100°C and the curing time is 1 to 5 hours.

3. The method for preparing a blood purification adsorption catheter according to claim 1, characterized in that, The organic solvent is any one of toluene, kerosene, dichloromethane, and chloroform, and the swelling time is 0.5-2 hours.

4. The method for preparing a blood purification adsorption catheter according to claim 1, characterized in that, The gas used in the hydrophilization treatment is high-purity oxygen, and the time is based on the water contact angle of the material surface as the standard for hydrophilicity, which is less than 40°. The silane coupling agent is 3-aminopropyltrimethoxysilane, and anhydrous ethanol of the silane coupling agent with a volume fraction of 5-20% is used as the amination agent, and the amination time is 5 hours. The RAFT chain transfer agent is 4-cyano-4-(thiobenzoylthio)valerate. The coupling reaction of the RAFT chain transfer agent is carried out in MES buffer at pH=6 using the carbodiimide method. The concentration of the RAFT chain transfer agent in the reaction system is 5~30 mg / mL, and the coupling reaction time is 12~24 hours.

5. The method for preparing a blood purification adsorption catheter according to claim 1, characterized in that, The RAFT polymerization reaction takes 12-48 hours.

6. The method for preparing a blood purification adsorption catheter according to claim 1, characterized in that, The chemical activating agent is N,N'-carbonyldiimidazole, with a CDI concentration of 0.05-0.2 mol / L; the activation time is at least 2 hours, and the cleaning is performed thoroughly at least 3 times.

7. The method for preparing a blood purification adsorption catheter according to claim 1, characterized in that, The nanobody described is a nanobody that can specifically recognize inflammatory factors, with an affinity constant of 10. -9 For M and above, the concentration range of nanobody during conjugation is 0.1~4.5 mg / mL, and the conjugation time is 5-24 hours.

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