Silica gel microvessel phantom model and manufacturing method and application thereof

Abstract

This invention belongs to the fields of tissue engineering and biotechnology, specifically relating to a silicone microvascular phantom model, its fabrication method, and its applications. The fabrication method includes the following steps: etching convex textures onto a metal template, and dripping hot melt adhesive at the endpoints of the convex textures; placing a smooth panel into a metal container, laying the metal template flat on the smooth panel with the convex texture facing upwards; dissolving silicone and pouring it into the container, removing air bubbles, and after the silicone solidifies, separating the silicone from the metal template to obtain a first silicone block; pouring the silicone into the metal container, removing air bubbles, and solidifying to obtain a second silicone block; placing the first silicone block on the surface of the second silicone block and solidifying to obtain a silicone composite; inserting a needle connected to a conduit into the cavity of the vascular texture of the silicone composite to reinforce the interface, thus obtaining the final product. The microvascular phantom model prepared by this method has a diameter of less than 100 micrometers and can be preserved for a long time and reused repeatedly.

Description

Technical Field

[0001] This invention belongs to the fields of tissue engineering and biotechnology, specifically relating to a silicone microvascular phantom model, its fabrication method, and its application. Background Technology

[0002] Traditional ultrasound, constrained by the diffraction limit, has a resolution of approximately half the wavelength, typically between 100 and 300 micrometers, unable to resolve smaller structures. However, ultrasound super-resolution imaging based on ultrasound localization microscopy, by locating, tracking, and accumulating ultrasound contrast agents, can achieve a resolution of tens of micrometers, far below the diffraction limit of traditional ultrasound. Since 2015, when Tanter's group combined ultrafast ultrasound imaging with microbubble-based super-resolution ultrasound imaging to obtain ultra-high resolution brain blood flow imaging at tens of micrometers, ultrasound super-resolution technology has rapidly developed, with numerous new algorithms emerging, including microbubble localization algorithms, microbubble tracking algorithms, and motion correction algorithms for super-resolution results. However, because existing microvascular phantoms cannot simultaneously achieve diameters below one hundred micrometers and specified vessel shapes, all of these algorithms are affected to varying degrees by the lack of experimental data for the phantoms.

[0003] Existing microvascular phantoms suitable for ultrasound super-resolution imaging can be mainly divided into two categories. One category primarily uses fixed template imprinting, which allows for shape control but cannot achieve microvascular phantoms with dimensions of tens of micrometers. The other category primarily uses stretched template imprinting, allowing for vessel diameters of tens of micrometers, but the vessel shape cannot be precisely controlled. Furthermore, most microvascular phantoms are limited by the materials used in their fabrication, preventing long-term preservation and repeated use. In addition, existing technologies also include the following methods for preparing microvascular phantoms:

[0004] (1) Using metal wire as a mold + gelatin preparation:

[0005] First, a non-branched metal wire is fixed to a container frame. Then, a mixture of gelatin and water is heated in another container until completely dissolved. Heating continues until all air bubbles are removed from the gelatin. This mixture is then poured into the container with the fixed metal wire, ensuring the wire is completely enclosed within the gelatin. After the gelatin solidifies, the wire is removed from the frame, leaving a cavity where the wire once was. This cavity can be used as a microvascular phantom. However, this method can produce microvascular phantoms up to 88 micrometers in size, but because the shape mold is a metal wire, it cannot create complex structures or specific shapes. Gelatin has a short shelf life, becoming unusable after approximately 2-3 days. It also has poor light transmittance; without the addition of additional scattering particles, the gelatin block is a pale yellow to yellowish-brown opaque solid, making it difficult to integrate with optical imaging.

[0006] (2) Preparation using 3D printed molds + polyvinyl alcohol:

[0007] Microvascular models were designed using computer-aided design software, and 3D printing technology was used to obtain realistic cardiovascular molds. The 3D-printed mold was fixed in the center of a container, and an aqueous solution of polyvinyl alcohol (PVA) and graphite powder was injected into the container. After curing, the PVA was divided in two along the vascular mold, and the middle vascular mold was removed. After removing the mold, the two halves of PVA with vascular patterns were reattached to obtain a microvascular phantom. However, this method is limited by the precision of 3D printing and cannot achieve vascular patterns with a diameter less than 200 micrometers.

[0008] (3) Lathe-machined convex aluminum block as mold + gelatin preparation:

[0009] Two grooves are cut into an aluminum block using a CNC machine tool. Raised, bifurcated vascular patterns are cut into one groove, while the other remains flat. Liquid gelatin, heated and dissolved, is poured into the grooves. After solidification, the textured gelatin is removed and bonded to a smooth, untextured piece of gelatin. It is then placed in a refrigerator to cure. Once cured, the microvascular phantom is obtained. However, this method is limited by the machining precision of the lathe, making it impossible to achieve vascular patterns smaller than 100 micrometers in diameter, and all parts of the vascular patterns are straight lines. This gelatin phantom has a shelf life of 5 days; after this time, mold may grow, compromising the phantom's integrity. Gelatin has poor light transmittance; without the addition of additional scattering particles, the gelatin block is a pale yellow to yellowish-brown opaque solid, making it difficult to integrate with optical imaging.

[0010] (4) 3D printed filament mold + manual filament drawing and bonding:

[0011] First, 3D printers were used to create acrylonitrile-butadiene-styrene copolymer microrods, or ABS rods, with a diameter of 600 micrometers. These microrods, being thermoplastic, were artificially stretched and thinned after printing to obtain a series of ABS rods with a minimum diameter of 50 micrometers. The ABS rods were then sorted by size and joined together, using acetone as an adhesive for connecting two rods. After the vascular mold was joined, it was placed in a container, and a hydrogel tissue simulation material solution was poured in and cooled until solidified. The phantom block containing the mold was then placed in an ultrasonic cleaner, where acetone was poured in and the mixture was cooled to dissolve the ABS rods. Once dissolved, a phantom model with microvessels was obtained. However, due to the inherent uncertainties of artificial stretching, the shape of each part of the microvessels and the spacing between adjacent vessels could not be precisely controlled. Summary of the Invention

[0012] To address the aforementioned technical problems, this invention provides a method for fabricating a silicone microvascular phantom model. The microvascular phantom model prepared by this method has a diameter of less than 100 micrometers and can be preserved for a long time and reused repeatedly.

[0013] The specific technical solution provided by this invention is as follows:

[0014] This invention provides a method for fabricating a silicone microvascular phantom model, comprising the following steps:

[0015] The desired convex texture is etched on the metal template, and hot melt adhesive is dripped at the endpoints of the convex texture.

[0016] Place the light panel into the metal container, lay the metal template flat on the light panel, and keep the side with the etched convex texture facing upwards;

[0017] After the silicone is melted, it is poured into the metal container, air bubbles are removed, and after the silicone is cured, the silicone is separated from the metal template to obtain the first silicone block containing vascular patterns.

[0018] Pour the silicone into a metal container, remove air bubbles, and allow it to solidify to obtain a second silicone block.

[0019] A first silicone block containing vascular patterns is placed on the surface of a second silicone block, and the convex textured surface on the first silicone block is made to adhere to the surface of the second silicone block. After curing, a silicone composite is obtained.

[0020] Insert a needle connected to a tube into the cavity of the vascular pattern of the silicone composite, and reinforce the interface with hot melt adhesive to obtain a silicone microvascular phantom model.

[0021] In a preferred embodiment of the present invention, the width of the convex texture is 40μm~200μm and the height is 60μm~140μm.

[0022] In a preferred embodiment of the present invention, the diameter of the hot melt adhesive droplet is 3 mm.

[0023] In a preferred embodiment of the present invention, the light panel is a glossy glass panel.

[0024] In a preferred embodiment of the present invention, the metal template is a stainless steel plate and the metal container is a stainless steel container.

[0025] In a preferred embodiment of the present invention, the air bubble removal is performed by vacuuming the equipment in a vacuum drying oven.

[0026] More preferably, the visible air bubbles are punctured.

[0027] In a preferred embodiment of the present invention, the curing is performed until the silicone surface exhibits stable stringing exceeding 1 cm.

[0028] In a second aspect, this invention provides a silicone microvascular phantom model prepared according to the above method. Its diameter ranges from a minimum of 47.2 micrometers to a maximum of several hundred micrometers. These figures are based on measurements obtained through a limited number of experiments, but are not a specific limitation on the diameter of the prepared microvascular phantom.

[0029] In a third aspect, the present invention provides the application of the aforementioned silicone microvascular phantom model in ultrasound contrast imaging.

[0030] In a preferred embodiment of the present invention, the silicone microvascular phantom model is used to observe ultrasound contrast agent microbubbles.

[0031] In a fourth aspect, the present invention provides the application of the aforementioned silicone microvascular phantom model in simultaneous ultrasound and optical imaging.

[0032] Compared with the prior art, the technical advantages of the present invention are as follows:

[0033] 1. The method of this invention only requires commissioning the fabrication of templates with embossed textures of different shapes to produce microvascular phantoms of specified shapes. Controllable parameters include vessel diameter, curvature, slope, connectivity, number of bifurcations, bifurcation angle, and vessel spacing. The diameter of the prepared microvascular phantom can be maintained below 100 micrometers, with a minimum of below 50 micrometers, comparable to the size of microvessels in the real human body. The imaging effects of various ultrasound imaging methods, including high-resolution ultrasound imaging, on microvascular networks with diameters of tens of micrometers can be verified on this phantom. Furthermore, by controlling variables, a specific parameter, such as vessel diameter, vessel spacing, bifurcation angle, or curvature, can be gradually modified to quantitatively analyze the impact of that parameter on the ultrasound imaging method.

[0034] 2. This invention uses metal templates and containers during manufacturing, avoiding the problems that some materials can cause liquid silicone to fail to solidify, and the fact that most 3D printing materials are photosensitive resins, which can also lead to liquid silicone not solidifying. Furthermore, during ultrasonic imaging, it is necessary to ensure that the convex textures are on the same plane. If materials such as plastic are used, due to insufficient mechanical strength, plastic deformation will occur after repeated use, and the convex textures will not be on the same plane in subsequent uses. Metal templates and containers can avoid this problem.

[0035] 3. The microvascular phantom prepared by this invention enables the observation of ultrasound contrast agent microbubbles using microfluidic methods. Because the diameter of this microvascular phantom is only tens of micrometers, the injection of extremely low concentrations of ultrasound contrast agent into the microvessel allows for controlled flow of the liquid within the tube, enabling the separation and observation of the movement of individual microbubbles under an optical microscope. Applying an external sound field during the microbubble movement allows for the observation of changes in the microbubbles within a specified sound field, thereby analyzing the properties of the contrast agent and the effect of the sound field on the contrast agent.

[0036] 4. The microvascular phantoms prepared by this invention can be stored for a long time and reused repeatedly. Once prepared, the phantom model has no risk of dehydration or spoilage and can be stored at room temperature for an extended period.

[0037] 5. The microvascular phantom prepared by this invention can achieve simultaneous ultrasound and optical imaging. By connecting an externally triggered high-speed camera and an optical microscope, optical images can be acquired on the same imaging plane simultaneously with ultrasound imaging. Attached Figure Description

[0038] Figure 1 This is a schematic diagram of different convex textures etched on a stainless steel plate.

[0039] Figure 2 This is an enlarged schematic diagram of the "Y"-shaped convex texture etched on a stainless steel plate.

[0040] Figure 3 This is a schematic diagram of hot melt adhesive being dripped at the end of a convex texture.

[0041] Figure 4 This is a diagram showing a stainless steel flat-bottomed container holding a smooth glass plate.

[0042] Figure 5 This is a schematic diagram of a template laid flat on two glass plates.

[0043] Figure 6 This is a schematic diagram before silica gel degassing.

[0044] Figure 7 This is a schematic diagram after silica gel degassing.

[0045] Figure 8 This is a schematic diagram of fully cured silicone.

[0046] Figure 9 This is a schematic diagram of silicone containing vascular patterns after separation from the container.

[0047] Figure 10 This is a schematic diagram of the wavy, convex textured silicone after it has been separated from the container.

[0048] Figure 11 This is a schematic diagram of the silicone with a "Y"-shaped convex texture after it has been separated from the container.

[0049] Figure 12 This is a schematic diagram showing stringing on the surface of silicone.

[0050] Figure 13 This is a schematic diagram showing the bonding of a silicone block with vascular patterns to cured silicone.

[0051] Figure 14 This is a schematic diagram showing the connection between a needle with a silicone tube and a silicone phantom.

[0052] Figures 15-16It is a 3D drawing used for processing metal templates with different convex textures.

[0053] Figure 17 This is an application diagram of ultrasound and optical image acquisition using fabricated microvascular phantoms; A~C are the optical imaging results of microvascular phantoms of different shapes, and D~F are the super-resolution imaging results of the corresponding microvascular phantoms. Detailed Implementation

[0054] To better explain and facilitate understanding of the present invention, the present invention will be described in detail below with reference to the accompanying drawings and specific embodiments.

[0055] The link to the silicone supplier involved in this invention embodiment is: https: / / item.taobao.com / item.htm?id=671502688551. This particular silicone model comes in two types, A and B. Both A and B are liquid when placed alone, but gradually solidify after being mixed. The mixing ratio of A to B is 1:1, allowing for an error of approximately 1%.

[0056] The metal template with embossed texture involved in the embodiments of this invention is a 3D drawing pre-made by the invention team using SolidWorks software, such as... Figures 15-16 As shown, the material is then processed by a contractor using a chemical etching method. This method can be used to obtain metal templates with different convex textures, thereby creating microvascular phantoms of specific shapes.

[0057] Example 1

[0058] A method for fabricating a silicone microvascular phantom model includes the following steps:

[0059] (1) Use chemical etching process to carve the required raised texture on the stainless steel template, such as Figures 1-2 The shape of the convex texture can be customized, with a width of approximately 40 micrometers and a height of approximately 60 micrometers. The stainless steel template is obtained through custom manufacturing.

[0060] (2) Use hot melt glue to dot a hemisphere with a diameter of about 3mm at the end of the convex texture of the template, such as... Figure 3 .

[0061] (3) Place two 52*76mm*1mm smooth glass plates at the bottom of a 14cm*8.4cm*2.2cm stainless steel flat-bottomed container, such as... Figure 4 .

[0062] (4) Place four stainless steel templates into a stainless steel container, lay them flat on two glass plates, with two templates on each glass plate, the raised texture side facing up, as shown. Figure 5 .

[0063] (5) Take 55 grams of each of silicone A and B. The amount can be adjusted according to the size of the container. Just make sure that the mass ratio of silicone A and B is 1:1 and the ratio deviation is within 1%. Add it to a brand new disposable plastic bottle and stir it evenly with a glass rod.

[0064] (6) Pour all the mixed silicone into a stainless steel container containing a stainless steel template. At this point, a large number of air bubbles are visible to the naked eye.

[0065] (7) Place the stainless steel container in a vacuum drying oven and evacuate it. When the liquid silica gel expands to the point of overflowing the container, turn off the vacuum pump and open the balancing valve. Wait for the internal and external air pressures to equalize before removing the container. Use a stainless steel needle to puncture any visible air bubbles.

[0066] (8) Repeat step (7) multiple times until the vacuum level reaches -0.95 and no visible bubbles are present in the liquid silica gel. Compare the results before and after degassing. Figure 6 Before degassing, Figure 7 After degassing.

[0067] (9) Remove the stainless steel container from the vacuum drying oven, put it in a bag, and place it flat on a calibrated optical level surface to cure naturally. Curing should be complete in about 18 hours. Figure 8 .

[0068] Uncured silicone is highly adhesive and will attract dust from the air. Dust can interfere with subsequent optical and ultrasonic imaging effects. Bags can prevent dust from falling onto the silicone surface.

[0069] Curing on an optical platform ensures that the upper and lower surfaces of the cured silicone are parallel. If it is not cured on an optical platform, the upper surface will be horizontal and the lower surface will be parallel to the surface on which it was placed, while the surface on which it was placed may not be horizontal. This results in the upper and lower surfaces not being parallel, making it difficult to focus during subsequent use.

[0070] (10) After complete curing, use a die and blade to cut along the edge of the stainless steel template to separate the silicone from the stainless steel template. Cut off the silicone containing the vascular pattern and remove it for later use. Figures 9-11 .

[0071] (11) After the silicone containing the blood vessel pattern has completely cured, in the same way as step (3), place two 52*76mm*1mm smooth glass plates into the bottom of another 14cm*8.4cm*2.2cm stainless steel flat-bottomed container.

[0072] (12) In the same step (5), take 55 grams of each of the two types of silicone, A and B, and add them into a disposable plastic bottle. Stir well with a glass rod.

[0073] (13) In the same step (6), pour all the mixed silica gel into a metal container. At this time, a large number of bubbles can be seen with the naked eye.

[0074] (14) Following step (7), place the stainless steel container in a vacuum drying oven and evacuate it. When the liquid silica gel expands to the point of overflowing the container, turn off the vacuum pump and open the balancing valve. Wait for the internal and external pressures to equalize before removing the container. Use a stainless steel needle to puncture any visible air bubbles.

[0075] (15) Repeat step (14) multiple times until no visible bubbles are present in the liquid silica gel when the vacuum degree reaches -0.95.

[0076] (16) Remove the stainless steel container and place it flat on a calibrated optical level surface to cure naturally. Curing should continue for approximately 6-8 hours, or until a needle punctures the silicone surface and produces a stable stringing pattern exceeding 1 cm. Figure 12 .

[0077] (17) Place the silicone block with the vascular pattern obtained in step (10) with the convex texture facing down on the surface of the silicone block obtained in step (16), in the same position as in step (4). After the silicone block is placed, wait one to two minutes until the silicone is visibly adhered. Then, use a plastic rod to gently tap the areas near the cavities at both ends of the convex texture that are not tightly adhered until they are tightly adhered. Figure 13 .

[0078] (18) Place the silicone bond obtained in step (17) together with the stainless steel container on a calibrated optical level surface for natural curing. It will be fully cured after about 10 hours. The upper surface of this silicone bond is called surface A, and the lower surface is called surface B.

[0079] (19) After complete curing, use a die and blade to remove the silicone phantom. Use double-sided tape to adhere it to the glass plate, with A side facing down and in contact with the double-sided tape, and B side facing up.

[0080] (20) Insert the needle connected to the silicone tube into the cavity of the silicone phantom from the side and back, and reinforce the joint with hot melt glue. In use, simply connect the external syringe to the silicone tube to inject liquid into the silicone phantom, such as... Figure 14 .

[0081] Example 2

[0082] A method for fabricating a silicone microvascular phantom model differs from Example 1 only in that the width of the convex texture is 200 μm and the height is 140 μm.

[0083] Example 3

[0084] A method for fabricating a silicone microvascular phantom model differs from Example 1 only in that the width of the convex texture is 150 μm and the height is 90 μm.

[0085] The silicone microvascular phantom models prepared in Examples 1 to 3 could still be used normally one and a half months after completion, proving their long-term stability at room temperature.

[0086] Ultrasound and optical image acquisition was performed using a fabricated silicone microvascular phantom model, such as... Figure 17 As shown. Figure 17 Images A through C show the optical imaging results of microvessel phantoms of different shapes. Based on the optical images and measurements, the minimum diameter of the phantom vessels in this batch of samples is 47.20 μm, and these phantoms achieve various vessel shapes and structures, including straight lines, curves, and bifurcations. Images D through F show the super-resolution imaging results of the corresponding microvessel phantoms. A solution containing contrast agent microbubbles flows within the vessel phantom model. Super-resolution images are obtained by locating, tracking, filtering, and accumulating the contrast agent microbubble data collected over a period of time. The colors in the super-resolution images reflect the density of contrast agent localization points in the microbubbles; brighter colors indicate a higher concentration of contrast agent localization at that point. The super-resolution images show that the shape and structure of the accumulated contrast agent localization points are basically consistent with the optical images, and most microbubble localization points exist within the actual microvessel texture, demonstrating that the microvessel phantoms provided by this invention can achieve ultrasound super-resolution imaging of vessels of specified shapes. Due to differences in imaging principles, ultrasound super-resolution images are susceptible to variations in image quality caused by localization, tracking, and reconstruction algorithms. Optical methods, on the other hand, can obtain extremely accurate information on the location and shape of blood vessels. Therefore, optical images can serve as the gold standard for evaluating the effectiveness of ultrasound imaging algorithms. Subsequent comparative analysis of the differences between super-resolution and optical images will allow for the evaluation of super-resolution imaging performance.

[0087] It should be understood that the above description of specific embodiments of the present invention is only for illustrating the technical approach and features of the present invention, and is intended to enable those skilled in the art to understand the content of the present invention and implement it accordingly. However, the present invention is not limited to the specific embodiments described above. All changes or modifications made within the scope of the claims of the present invention should be covered within the protection scope of the present invention.

Claims

1. A method for fabricating a silicone microvascular phantom model, characterized in that, Includes the following steps: The desired convex texture is etched on the metal template, and hot melt adhesive is dripped at the endpoints of the convex texture. Place the light panel into the metal container, lay the metal template flat on the light panel, and keep the side with the etched convex texture facing upwards; Pour the silicone into the metal container, remove air bubbles, and after the silicone has solidified, separate the silicone from the metal template to obtain the first silicone block containing vascular patterns. Pour the silicone into a metal container, remove air bubbles, and allow it to solidify to obtain a second silicone block. A first silicone block containing vascular patterns is placed on the surface of a second silicone block, and the convex textured surface on the first silicone block is made to adhere to the surface of the second silicone block. After curing, a silicone composite is obtained. Insert a needle connected to a tube into the cavity of the vascular pattern of the silicone composite, and reinforce the interface with hot melt adhesive to obtain a silicone microvascular phantom model.

2. The method for fabricating a silicone microvascular phantom model according to claim 1, characterized in that, The width of the convex texture is 40μm~200μm and the height is 60μm~140μm.

3. The method for fabricating a silicone microvascular phantom model according to claim 1, characterized in that, The diameter of the hot melt adhesive droplets is 3mm.

4. The method for fabricating a silicone microvascular phantom model according to claim 1, characterized in that, The light panel is a glossy glass panel.

5. The method for fabricating a silicone microvascular phantom model according to claim 1, characterized in that, The metal template is made of stainless steel, and the metal container is made of stainless steel.

6. The method for fabricating a silicone microvascular phantom model according to claim 1, characterized in that, The air bubble removal process involves placing the item in a vacuum drying oven and then applying a vacuum.

7. The method for fabricating a silicone microvascular phantom model according to claim 1, characterized in that, The curing process continues until the silicone surface exhibits stable stringing exceeding 1 cm.

8. A silicone microvascular phantom model prepared by the method according to any one of claims 1 to 7.

9. The application of the silicone microvascular phantom model of claim 8 in ultrasound contrast imaging.

10. The application of the silicone microvascular phantom model of claim 8 in simultaneous ultrasound and optical imaging.

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