Tissue fluorescence motif with controllable depth structure as well as preparation method and application of tissue fluorescence motif

Abstract

The invention discloses a tissue fluorescence motif with a controllable depth structure and a preparation method and application thereof, and belongs to the technical field of medical imaging and optical detection. The tissue fluorescence motif is prepared from a multi-component optical simulation material and a mold, and the material is prepared from the following raw materials in percentage: 0.01 to 0.05 percent of melanin solution, 0.01 to 0.1 percent of polystyrene microspheres, 0.03 to 0.08 percent of fluorescent agent, 1.0 to 1.5 percent of coagulator and the balance of liquid matrix. The tissue fluorescence motif has the beneficial effects that the tissue fluorescence motif can simultaneously simulate the absorption, scattering and endogenous fluorescence characteristics of tissues, so that the optical behavior of the tissue fluorescence motif is closer to that of real biological tissues. The design of the die body with controllable structured depth small holes enables exogenous fluorescence to be distributed at different depths according to requirements, so that depth gradient fluorescence signals in real tissues are simulated. And the preparation process is simple, high in repeatability, easy to standardize and capable of realizing batch production.

Description

Technical Field

[0001] This invention belongs to the field of medical imaging and optical detection technology, and relates to tissue optical phantoms for performance evaluation, calibration, standardization and verification of fluorescence imaging systems. Specifically, it relates to a tissue fluorescence phantom with controllable depth structure and its preparation method and application. Background Technology

[0002] Fluorescence imaging technology has been widely used in clinical infection detection, tumor imaging, photodynamic therapy agent distribution detection, and biochemical metabolism research. To accurately evaluate the depth penetration capability, quantitative fluorescence accuracy, and image processing algorithm performance of fluorescence imaging devices, calibration and validation using tissue optical phantoms are typically required. Existing technologies have the following drawbacks: 1. Most phantoms only possess absorption and scattering characteristics, lacking tunable fluorescence features. Traditional phantoms such as agar phantoms, PVA-C phantoms, and Intralipid phantoms are only used for simulating optical parameters and cannot reproduce the intrinsic fluorescence of solid tissues (such as NADH). 2. Most fluorescent phantoms have a uniform overall structure and cannot simulate fluorescence signals at different tissue depths. Due to the lack of depth structure, they are difficult to use for depth resolution performance testing of devices. 3. There is a lack of phantom preparation methods with precise and repeatable geometric structures. 4. Phantom structures that can support porous depths and simultaneously hold multiple fluorescent reagents have not been established, resulting in functional limitations and failing to meet the overall performance validation requirements of imaging devices at different wavelengths. Therefore, there is an urgent need for a tissue fluorescence phantom that can simultaneously simulate absorption, scattering, and endogenous fluorescence and has a controllable depth pore structure to solve the problem of the lack of standardized testing for the depth imaging performance of equipment.

[0003] Chinese patent application CN116124752A discloses a tissue biomimetic phantom based on multispectral modulation and its generation method. Although it can modulate the spectral characteristics of the target fluorescent molecules, the phantom lacks endogenous background fluorescent substances and necessary absorbers and scatterers, and cannot realistically simulate the complex optical environment of biological tissues. At the same time, the phantom is a uniform monolithic structure and lacks a channel system with controllable depth, so it cannot simulate fluorescence distribution at different depths on the same phantom, nor does it support dynamic replacement of different fluorescent reagents during the test, making it difficult to meet the actual needs of depth resolution and comprehensive performance evaluation of fluorescence imaging equipment. Summary of the Invention

[0004] The technical problem to be solved by this invention is how to solve the problems of limited simulation function and poor accuracy of simulation results in existing tissue fluorescence phantoms.

[0005] This invention aims to solve the following technical problems existing in current tissue fluorescent phantoms: 1. Difficulty in simultaneously simulating the absorption, scattering, and endogenous fluorescence characteristics of real tissues: Existing phantoms often only simulate one of the scattering or absorption characteristics of tissues, or only add exogenous fluorescence while lacking endogenous fluorescence components, resulting in significant differences between the simulation results and the optical behavior of real tissues, affecting the accuracy of imaging equipment evaluation.

[0006] 2. Inability to simulate fluorescence signal distribution at different depths in a structured and controllable manner: Existing phantoms are usually uniform monolithic structures, which cannot form controllable and repeatable small-scale spatial structures within the phantom to simulate fluorescence signals generated at different depths in tissues. Therefore, it is impossible to systematically evaluate the deep detection capability and depth resolution performance of the device.

[0007] 3. The phantom preparation process is complex and not conducive to standardization and mass production: Existing methods require manual layering and repeated casting, which are cumbersome and greatly affected by human factors, resulting in poor consistency between phantoms and making it difficult to meet the needs of laboratory standardization, equipment manufacturers' mass production and clinical validation.

[0008] 4. The phantom has a single function and cannot support the multi-index performance evaluation of imaging equipment: Traditional phantoms are mostly used for simple luminescence simulation. They cannot simultaneously realize absorption, scattering, endogenous fluorescence modulation and exogenous fluorescence depth simulation in the same phantom. They cannot meet the comprehensive evaluation of the equipment in terms of excitation efficiency, emission and collection efficiency, depth decay curve, imaging algorithm robustness and other aspects.

[0009] The present invention solves the above-mentioned technical problems through the following technical means: The first aspect of the present invention provides a multi-component optical simulation material, which is composed of the following raw materials in percentage: 0.01~0.05% (w / v) melanin solution, 0.01~0.1% (w / v) polystyrene microspheres, 0.03~0.08% (w / v) fluorescent agent, 1.0~1.5% (w / v) coagulant, and the remainder being a liquid matrix.

[0010] Melanin solution: As an absorbent, a concentration range of 0.01–0.05% (w / v) can effectively simulate the typical absorption coefficient of biological tissue (μa≈0.1–1 cm⁻¹). -1 ).

[0011] Polystyrene microspheres (particle size 0.5–2 μm, 0.01–0.1%): used as a scattering agent to simulate the reduced scattering coefficient of tissues (μs'≈5–20 cm⁻¹). -1 ).

[0012] Fluorescent agents: make the autofluorescence intensity of the solidified phantom close to the background fluorescence of skin or mucous membrane tissue.

[0013] Preferably, the fluorescent agent is reduced coenzyme I (NADH) or flavin adenine dinucleotide (FAD).

[0014] Preferably, the coagulant is agar or gelatin.

[0015] Preferably, the liquid matrix is ​​purified water or PBS solution.

[0016] PBS solution: Phosphate buffer solution.

[0017] A second aspect of the present invention provides a method for preparing a tissue fluorescent phantom using the above-mentioned material, comprising the following steps: (1) Mix the melanin solution, polystyrene microspheres, coagulant and liquid matrix, heat, and then add fluorescent agent and mix well; (2) Pour the mixture obtained in (1) into mold A, cover mold B, let stand until completely solidified, and then demold to obtain the product.

[0018] Preferably, the heating specifically refers to heating to boiling.

[0019] Preferably, mold A is matched with mold B, and mold B has multiple columns of different heights.

[0020] A third aspect of the present invention provides a tissue fluorescent phantom prepared by the above method.

[0021] A fourth aspect of the present invention proposes the application of the above-mentioned tissue fluorescence phantom in the evaluation of imaging devices.

[0022] Preferably, the evaluation items for imaging equipment include, but are not limited to, the lateral resolution, depth imaging performance and penetration capability, fluorescence quantitative accuracy, multispectral imaging algorithm verification, and signal-to-noise ratio (SNR) evaluation of the imaging system.

[0023] The fifth aspect of the present invention provides a mold assembly for preparing tissue fluorescent phantoms, comprising mold A (lower mold) and mold B (upper mold), wherein mold A is matched with mold B, and mold B is provided with multiple columns of different heights (the height of the columns is determined according to whether the observed object is located in the epidermis, dermis, subcutaneous tissue or deep tissue); the columns are conical columns with a draft angle of 1° to 3°.

[0024] Preferably, the material of mold A is selected from heat-resistant glass and stainless steel; the shape of mold A is any one of cylinder, cuboid, cube, and cone; mold A is used to hold liquid mold material.

[0025] Preferably, the mold B is used to insert into the mold body to form small holes of different depths.

[0026] Preferably, the column is a tapered column with a bottom diameter of 5-10 mm and a draft angle of 1°-3° (this draft angle can ensure the integrity of the hole when it is pulled out, and also ensure that it can be easily pulled out); more preferably, the bottom diameter is 7 mm and the draft angle is 1.5°.

[0027] The beneficial effects of this invention are as follows: 1. This invention provides a tissue fluorescence phantom capable of simultaneously simulating the absorption, scattering, and endogenous fluorescence characteristics of tissue, making its optical behavior more closely resemble that of real biological tissue. It provides a phantom design with controllable structured depth pores, allowing exogenous fluorescence to be distributed at different depths as needed, thereby simulating the depth gradient fluorescence signal in real tissue. It provides a tissue fluorescence phantom preparation method that is simple, highly reproducible, easily standardized, and suitable for mass production. It provides a functional tissue fluorescence phantom capable of supporting multi-dimensional performance evaluation of imaging equipment (such as depth resolution, sensitivity, signal-to-noise ratio, and imaging algorithm performance).

[0028] 2. The phantom of the present invention has the following advantages: (1) Comprehensive simulation of optical properties: The absorption, scattering and fluorescence properties of the phantom can be precisely controlled by melanin, microspheres, NADH / FAD, etc., which are close to the optical properties of real tissue.

[0029] (2) Depth controllable, structured fluorescence source: Multiple depth pores are formed in one step by inserting columns of varying heights. It is repeatable and standardized. The target fluorescent agent can be added according to the fluorescence properties of the tissue to be studied. (3) Simple preparation and mass production: Only one casting is required, and the repeatability is good. (4) Multifunctional imaging evaluation platform: It can be used for: penetration depth evaluation, imaging sensitivity testing, optical system depth of focus and resolution evaluation, fluorescence quantitative linearity testing, and calibration of different fluorescence wavelength channels.

[0030] 3. A multi-component optical phantom system capable of simultaneously simulating tissue absorption, scattering, and endogenous fluorescence characteristics. This invention constructs a tunable optical material system composed of a liquid matrix, an absorber, a scattering agent, a base fluorescent agent, and a coagulant, which can simultaneously and accurately simulate the absorption coefficient, scattering coefficient, and background fluorescence characteristics of real tissues in a single phantom.

[0031] 4. Existing technologies cannot achieve controllable simulation of three types of optical properties in the same phantom. This invention utilizes an upper mold with columns of different heights to achieve a "one-time molding structured fluorescent phantom with controllable hole depth." The invention uniquely employs a mold B with columns of different heights, used in conjunction with mold A, to directly form multiple hole structures of different depths in a single casting process.

[0032] 5. Overcoming the limitations of existing phantoms that require multiple castings or cannot form repeatable depth structures. By adding fluorescent reagents through small holes, "controllable depth fluorescence signal simulation" is achieved, supporting multi-index evaluation of imaging equipment. The small hole structure is used to accommodate different fluorescent reagents, thereby constructing fluorescence signals of multiple depths and intensities, which can simulate interstitial fluorescence attenuation and penetration behavior, and can be used for performance testing of fluorescence imaging equipment such as sensitivity, penetration depth, and quantitative capability. 6. Existing phantoms cannot provide a structured, quantified, and controllable depth fluorescence simulation system.

[0033] Of course, implementing any product or method of the present invention does not necessarily require achieving all of the advantages described above at the same time. Attached Figure Description

[0034] Figure 1 This is a schematic diagram of the structure of the mold assembly 1 in Embodiment 1 of the present invention; Figure 2 This is a schematic diagram of the structure of the mold assembly 2 in Embodiment 2 of the present invention; Figure 3 This is a flowchart illustrating the preparation and application of the phantom in Embodiment 1 of the present invention; Figure 4 This is an image showing the imaging effect of the phantom matched with different fluorescent agents in Embodiment 1 of the present invention. Detailed Implementation

[0035] To make the objectives, technical solutions, and advantages of the embodiments of the present invention clearer, the technical solutions of the present invention will be clearly and completely described below in conjunction with the embodiments of the present invention. Obviously, the described embodiments are only some embodiments of the present invention, not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention. Unless otherwise defined, the technical terms used below have the same meaning as understood by those skilled in the art.

[0036] Unless otherwise specified, the test materials and reagents used in the following examples are commercially available or prepared by known methods.

[0037] Unless otherwise specified, all techniques or conditions described in the embodiments can be performed in accordance with the techniques or conditions described in the literature in this field or in the product manual. Unless otherwise specified, the quantitative experiments in the following embodiments are all repeated three times or more, and the results are averaged.

[0038] Example 1: (Preparation and application flowchart as shown) Figure 3 (As shown) A multi-component optical simulation material is composed of the following percentages of raw materials: 0.03% (w / v) melanin solution, 0.05% (w / v) polystyrene microspheres, 0.04% (w / v) fluorescent agent, 1.2% (w / v) coagulant, and the remainder being a liquid matrix; the melanin solution is polydopamine (PDA), the fluorescent agent is reduced coenzyme I (NADH), the coagulant is agar, and the liquid matrix is ​​purified water.

[0039] (This material system can simultaneously achieve realistic simulation of three types of optical properties: absorption, scattering, and autofluorescence in a single phantom.) A mold assembly 1 for preparing tissue fluorescent phantoms (such as...) Figure 1 As shown, the test equipment includes mold A (lower mold) and mold B (upper mold), which are matched. Mold A is cylindrical and made of stainless steel. Mold B contains 12 columns of different heights (the number of columns can be selected according to the testing purpose of the test equipment). The columns are tapered with a bottom diameter and a draft angle of 1.5°. The top of mold B also has four overflow chambers to automatically collect excess liquid after extrusion, ensuring consistent mold thickness. Mold A and mold B are precisely aligned via slots.

[0040] (Using mold assembly 1, we can obtain fluorescent phantoms of different depths in a planar structure, and obtain small pores of multiple depths in a single molding process.) The method for preparing a tissue fluorescent phantom using the multi-component optical simulation material and mold combination 1 of this embodiment includes the following steps: Step 1: Mixing and melting materials (1) Add the following to the blue-capped bottle: Purified water; Melanin solution (according to target concentration); Polystyrene microsphere suspension (at target concentration); Agar powder; (2) Shake gently to disperse the solid initially; (3) Loosen the bottle cap and heat it in the microwave until it boils; (4) Remove and shake well; (5) Repeat the heating-shaking process above 2-3 times until the agar is completely dissolved. Step 2: Add substrate fluorescent agent Add the pre-prepared NADH solution to the bottle and shake gently to obtain a stable and detectable tissue background fluorescence signal.

[0041] Step 3: Casting and Molding (1) Quickly pour the hot liquid into mold A and use a bulb syringe / dropper to remove the air bubbles floating on the surface; (2) Immediately cover mold B so that the entire column is immersed in the liquid; (3) Excess liquid automatically flows into the leakage chamber; (4) Let stand at room temperature until completely solidified (cooling at 4-10℃ is the fastest and most effective).

[0042] Step 4: Demolding Vertically pulling out mold B yields a tissue fluorescent phantom containing multiple small holes of different depths.

[0043] Specific applications: (1) Addition of fluorescent reagent Different fluorescent dyes (such as porphyrin, ICG, FIGC, Cy5, etc.) were added to small holes at different depths to create simulations: the imaging effects of different fluorescent agents are shown in the following figures. Figure 4 As shown.

[0044] Superficial fluorescence Mid-layer fluorescence Deep fluorescence Multilayer mixed fluorescence (2) Application in imaging equipment evaluation Phantoms can be used for the following evaluation items: Lateral resolution of fluorescence imaging systems Depth imaging performance and penetration capability Fluorescence quantification accuracy Multispectral imaging algorithm verification Imaging system signal-to-noise ratio (SNR) evaluation Because the depth structure is fixed, it can be used for standardized comparison between devices or batches.

[0045] Comparative Example 1: This comparative example aims to highlight the advantages of the present invention in terms of the completeness of optical property simulation and the controllability of internal structure by comparing it with existing uniformly structured optical phantoms, so as to prove that the present invention solves the technical problems of lack of standardization in depth resolution testing and insufficient simulation realism in the prior art.

[0046]

[0047] Comparative experiment Experimental hypothesis: To evaluate the depth penetration capability of fluorescence imaging equipment (i.e., whether fluorescence signals can be effectively detected at different depths).

[0048]

[0049] Example 2: A multi-component optical simulation material is composed of the following percentages of raw materials: 0.01% (w / v) melanin solution, 0.1% (w / v) polystyrene microspheres, 0.03% (w / v) fluorescent agent, 1.5% (w / v) coagulant, and the remainder being a liquid matrix; the melanin solution is polydopamine (PDA), the fluorescent agent is flavin adenine dinucleotide (FAD), the coagulant is gelatin, and the liquid matrix is ​​PBS solution.

[0050] A mold assembly 2 for preparing tissue fluorescent phantoms (such as...) Figure 2 As shown, the test equipment includes mold A (lower mold) and mold B (upper mold), which are matched. Mold A is cylindrical and made of heat-resistant glass. Mold B contains eight columns of different heights (the number of columns can be selected according to the testing purpose of the test equipment). The columns are tapered with a bottom diameter equal to a draft angle of 1°. Mold B has a slope, and the top of mold B also has four overflow chambers for automatically collecting excess liquid to ensure consistent mold thickness. Mold A and mold B are precisely aligned via slots.

[0051] (Using mold combination 2, fluorescent phantoms of different depths of tissue on the slope can be obtained, and small holes of multiple depths can be obtained in a one-time molding process.) Example 3: A multi-component optical simulation material is composed of the following percentages of raw materials: 0.05% (w / v) melanin solution, 0.01% (w / v) polystyrene microspheres, 0.08% (w / v) fluorescent agent, 1.0% (w / v) coagulant, and the remainder being a liquid matrix. The melanin solution is polydopamine (PDA), the fluorescent agent is flavin adenine dinucleotide (FAD), the coagulant is gelatin, and the liquid matrix is ​​PBS solution.

[0052] Summarize Existing phantoms are structurally homogeneous but lack complete optical property simulation (lacking intrinsic fluorescence). Therefore, they cannot provide a structured, depth-controlled testing environment for fluorescence imaging devices, and in particular, they cannot meet the standardized evaluation requirements for the deep detection capabilities and depth resolution performance of such devices.

[0053] This invention overcomes the shortcomings of existing technologies by introducing a controllable depth structure prepared by "one-time molding of column array" and combining it with a complete optical simulation material system of absorption + scattering + intrinsic fluorescence. It achieves high-fidelity and high-repeatability simulation of the depth gradient fluorescence signal of biological tissues, and provides a standardized and functional tool for the performance evaluation of fluorescence imaging equipment.

[0054] The above embodiments are only used to illustrate the technical solutions of the present invention, and are not intended to limit it. Although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that modifications can still be made to the technical solutions described in the foregoing embodiments, or equivalent substitutions can be made to some of the technical features. Such modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the spirit and scope of the technical solutions of the embodiments of the present invention.

Claims

1. A multi-component optical simulation material, characterized in that, The material is composed of the following percentages of raw materials: 0.01~0.05% melanin solution, 0.01~0.1% polystyrene microspheres, 0.03~0.08% fluorescent agent, 1.0~1.5% coagulant, and the remainder is liquid matrix.

2. The multi-component optical simulation material according to claim 1, characterized in that, The fluorescent agent is reduced coenzyme I or flavin adenine dinucleotide; the coagulant is agar or gelatin; and the liquid matrix is ​​purified water or PBS solution.

3. The method for preparing tissue fluorescent phantoms using the multi-component optical simulation material according to claim 1, characterized in that, Includes the following steps: (1) Mix the melanin solution, polystyrene microspheres, coagulant and liquid matrix, heat, and then add fluorescent agent and mix well; (2) Pour the mixture obtained in (1) into mold A, cover mold B, let stand until completely solidified, and then demold to obtain the product.

4. The method according to claim 3, characterized in that, The heating process specifically involves heating to boiling.

5. The method according to claim 3, characterized in that, The mold A is matched with the mold B, and the mold B has multiple columns of different heights.

6. The tissue fluorescent phantom prepared by the method according to any one of claims 3-5.

7. The application of the tissue fluorescence phantom as described in claim 6 in the evaluation of imaging equipment.

8. The application according to claim 7, characterized in that, The evaluation items for imaging equipment include the lateral resolution, depth imaging performance and penetration capability, fluorescence quantitative accuracy, multispectral imaging algorithm verification, and signal-to-noise ratio evaluation of the imaging system.

9. A mold assembly for preparing tissue fluorescent phantoms, characterized in that, It includes mold A and mold B, which are matched. Mold B has multiple columns of different heights. The columns are tapered columns with a draft angle of 1° to 3°.

10. The mold assembly according to claim 9, characterized in that, The material of mold A is selected from heat-resistant glass and stainless steel; the shape of mold A is any one of cylinder, cuboid, cube, and cone; mold A is used to hold liquid mold material; mold B is used to insert the mold to form small holes of different depths.

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