Light-cured resin with adjustable refractive index, preparation method of light-cured resin and application of light-cured resin in tissue embedding

Abstract

The invention provides light-cured resin with an adjustable refractive index, a preparation method of the light-cured resin and application of the light-cured resin in tissue embedding. The light-cured resin is prepared from the following raw material components: 0.5%-5% of a photoinitiator, 60%-90% of a polymerizable monomer, 1%-20% of a high refractive index monomer and 4%-20% of a diluent. The light-cured resin disclosed by the invention does not contain toxic and harmful volatile organic solvents; the product has small viscosity, and can quickly and uniformly permeate into tissues as a mounting medium; a semi-interpenetrating network structure is formed after curing, so that the volume shrinkage can be effectively reduced, and the sealing piece quality is improved; the curing time can be shortened to 1 min, and the method is expected to be applied to automatic mounting tablets; the cured mounting medium is high in transparency and adjustable in refractive index.

Description

Technical Field

[0001] The invention belongs to the technical field of tissue embedding, and in particular relates to a light-curing resin with adjustable refractive index, a preparation method thereof and application in tissue embedding. Background Art

[0002] In the disciplines of pathology and forensic medicine, when studying, observing and judging the morphological changes of cell tissues, it is generally necessary to fix and slice the tissues. During the slicing process, the tissue structure needs not to be destroyed. If there is an implant, the original tissue structure morphology between the tissue and the implant needs to be maintained. Slicing includes paraffin sectioning, frozen sectioning, ultra-thin sectioning and other techniques. Among them, in hard tissue sectioning, it is mainly for bone tissue, bone tissue with implants, other tissue specimens embedded with hard implants, or bone tissue that cannot be decalcified after being labeled with bone-loving fluorescein during the animal experiment stage, through dehydration, infiltration, and embedding. Fixing tissue has always been a challenge in hard tissue sectioning. Hard tissue embedding and sectioning technology generally requires the following special processing steps: fixation, infiltration dehydration, embedding, and precise sectioning to obtain thin slices. In terms of the quality and quantity of regenerated bone tissue, hard tissue embedding technology can maintain the integrity of calcification deposition and the integrity of the original bone morphology.

[0003] Photocurable resins are a special class of polymer materials that can rapidly transform from liquid to solid under ultraviolet light or other light sources. This transformation is achieved through a photoinduced polymerization reaction, in which the monomer or prepolymer molecules in the resin are chemically linked to form a polymer with a three-dimensional network structure. Photocurable resins are widely used in 3D printing, printing photosensitive plates, microchip circuit patterns, UV coatings, UV inks, UV adhesives and other fields. The advantages of photocurable resins for hard tissue embedding mainly include:

[0004] (1) Rapid curing: Photocurable resin can cure quickly under ultraviolet light, which greatly shortens the time of tissue embedding. Good tissue structure preservation: Photocurable resin can well maintain the original structure and morphology of the tissue, which is very important for subsequent histological research and analysis.

[0005] (2) Strong adaptability: Photocurable resins are suitable for a variety of hard tissue samples, including bone tissue, teeth, and implanted non-metallic biomaterials.

[0006] (3) Easy to operate: Compared with some traditional embedding materials, the operation of light-curing resin is simpler, faster and easier to master.

[0007] (4) High resolution: Tissue sections embedded in photocurable resin can achieve higher resolution, which is conducive to the observation of fine structures.

[0008] (5) Reduce sample damage: Since the curing process of photocurable resin does not require high temperature, it reduces damage to heat-sensitive samples.

[0009] (6) Suitable for automated processing: The use of photocurable resin can be combined with automated slicing and grinding equipment to improve the efficiency and repeatability of the experiment.

[0010] (7) Good chemical stability: Photocurable resin has good chemical stability and is not easily affected by environmental factors, which is conducive to long-term preservation of samples.

[0011] (8) Transparency: Photocurable resins have good transparency, which helps to observe the natural structure of the sample without staining.

[0012] (9) Multiple uses: Photocurable resins can be used not only for traditional histological research, but also for hard tissue samples that require special treatment, such as the study of teeth and bone tissue. High refractive index mounting media can be directly applied to fluorescently labeled cells or tissue samples on microscope slides to improve the clarity and resolution of imaging. They are used to seal stained tissue sections to obtain clearer microscopic results and help preserve tissue specimens for a long time to prevent oxidation and fading.

[0013] In short, high refractive index mounting agents are important in improving imaging quality, protecting fluorescent labeled samples, and in specific industrial applications. Different application scenarios have different requirements for the refractive index, and the refractive index of the resin needs to be easily and conveniently adjustable.

[0014] The embedding agents in the prior art usually contain harmful volatile solvents, have high viscosity, uneven penetration, long curing time, usually up to 30 minutes, severe volume shrinkage, affecting the quality of the sealing, require manual operation, are inefficient, and have a narrow adjustable range of refractive index.

[0015] Therefore, there is an urgent need to provide a light-curable resin with adjustable refractive index for tissue embedding. Summary of the invention

[0016] Based on the defects of the prior art, the first purpose of the present invention is to provide a photocurable resin with adjustable refractive index; the second purpose of the present invention is to provide a method for preparing the photocurable resin with adjustable refractive index; the third purpose of the present invention is to provide the application of the photocurable resin with adjustable refractive index in tissue embedding.

[0017] The purpose of the present invention is achieved through the following technical solutions:

[0018] In one aspect, the present invention provides a photocurable resin with an adjustable refractive index. The raw material components for preparing the photocurable resin include, based on 100% by mass:

[0019]

[0020] In the above-mentioned photocurable resin, preferably, the photoinitiator is selected from 2,4,6-trimethylbenzoyl diphenylphosphine oxide (TPO) and / or 2,4,6-trimethylbenzoyl ethylphosphonate (TPO-L); but not limited thereto.

[0021] In the above-mentioned photocurable resin, preferably, the polymerizable monomer is selected from methacrylate monomers.

[0022] In the above-mentioned photocurable resin, preferably, the methacrylate monomer is selected from one or more combinations of hydroxyethyl methacrylate, polyethylene glycol methacrylate, isobornyl methacrylate, methyl methacrylate, ethyl methacrylate, butyl methacrylate, isobutyl methacrylate, 2-ethylhexyl methacrylate, glycidyl methacrylate, phenoxyethyl methacrylate, dimethylaminoethyl methacrylate and trifluoroethyl methacrylate; but it is not limited to this.

[0023] In the above-mentioned photocurable resin, preferably, the molecular weight of the polyethylene glycol methacrylate is 200-2000.

[0024] In the above-mentioned photocurable resin, preferably, the high refractive index monomer is selected from one or more combinations of 2-methyl-2-acrylic acid cyclothioethane methyl ester, 2-(methylthio)ethyl methacrylate, 2-(methylseleno)ethyl methacrylate, 4-methylthiobenzyl methacrylate and 4-methylselenobenzyl methacrylate; but it is not limited to this.

[0025] In the above-mentioned photocurable resin, preferably, the diluent is selected from one or more of polyethylene glycol, polypropylene glycol and polyethyleneimine; but not limited thereto.

[0026] In the above-mentioned photocurable resin, preferably, the molecular weight of the diluent is 300-5000.

[0027] On the other hand, the present invention also provides a method for preparing the above-mentioned photocurable resin, the method for preparing the photocurable resin comprising:

[0028] Add polymerizable monomer, high refractive index monomer, diluent and photoinitiator into a stirring kettle in sequence, and stir evenly to obtain a photocurable resin.

[0029] In another aspect, the present invention also provides use of the above-mentioned photocurable resin in tissue embedding.

[0030] The above description is only an overview of the technical solution of the present invention. In order to more clearly understand the technical means of the present invention and implement it according to the contents of the specification, the following is a detailed description of the preferred embodiments of the present invention in conjunction with the accompanying drawings. BRIEF DESCRIPTION OF THE DRAWINGS

[0031] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings required for use in the embodiments or the description of the prior art will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying creative work.

[0032] Figure 1 Comparison of the sealing effects of the two groups of sealing agents, Example 5 of the present invention and the control group, by conventional HE staining (low power microscope), A is Example 5 of the present invention, and B is the control group.

[0033] Figure 2 Comparison of the sealing effects of the two groups of sealing agents, Example 5 of the present invention and the control group, by conventional HE staining (high power microscope), A is Example 5 of the present invention, and B is the control group.

[0034] Beneficial effects of the present invention:

[0035] The photocurable resin of the present invention:

[0036] (1) It does not contain toxic or harmful volatile organic solvents, which can prevent harmful solvents from causing harm to people and corrosion to machines;

[0037] (2) The product has low viscosity and can penetrate into tissues quickly and evenly as a sealing agent;

[0038] (3) Compared with the existing non-photocurable embedding agent, the photocuring agent of the present invention forms a semi-interpenetrating network structure after curing, which can effectively reduce volume shrinkage and improve the quality of the sealing film;

[0039] (4) Accelerate the sealing time. Compared with the existing non-photocurable embedding agents, the curing time of the photocuring agent of the present invention can be shortened to 1 minute, and it is expected to be applied to automated sealing agents;

[0040] (5) The refractive index can be precisely adjusted according to the content of sulfur-containing and selenium-containing monomers, which effectively improves the refractive index of the resin. The cured sealing material has high transparency and an adjustable refractive index.

[0041] The above description is only an overview of the technical solution of the present invention. In order to more clearly understand the technical means of the present invention and implement it according to the contents of the specification, the preferred embodiments of the present invention are described in detail below. DETAILED DESCRIPTION

[0042] The technical solutions in the embodiments of the present invention will be described clearly and completely below. Obviously, the described embodiments are only a part of the embodiments of the present invention, rather than all the embodiments. Based on the embodiments in the present invention, all other embodiments obtained by ordinary technicians in the field without creative work are within the scope of protection of the present invention. The process, conditions, reagents, experimental methods, etc. for implementing the present invention, except for the contents specifically mentioned below, are all common knowledge and common common sense in the art, and the present invention does not particularly limit the contents.

[0043] Embodiment 1:

[0044] This embodiment provides a photocurable resin with adjustable refractive index. The raw material components for preparing the photocurable resin include, by mass percentage:

[0045] Polymerizable monomer (80%):

[0046] Hydroxyethyl Methacrylate 30%

[0047]

[0048] The preparation method of the light-curable resin with adjustable refractive index is as follows:

[0049] According to the above-mentioned mass ratio, hydroxyethyl methacrylate, polyethylene glycol methacrylate (molecular weight 400), isobornyl methacrylate, 2-(methylthio)ethyl methacrylate, polyethyleneimine (molecular weight 600) and 2,4,6 (trimethylbenzoyl) diphenylphosphine oxide (TPO) are added into a stirring tank in sequence, and stirred evenly to obtain an embedding resin, i.e., a photocurable resin with adjustable refractive index.

[0050] This embodiment also provides the application of the light-curable resin with adjustable refractive index in tissue embedding.

[0051] Embodiment 2:

[0052] This embodiment provides a photocurable resin with adjustable refractive index. The raw material components for preparing the photocurable resin include, by mass percentage:

[0053]

[0054] The preparation method of the light-curable resin with adjustable refractive index is as follows:

[0055] According to the above-mentioned mass ratio, hydroxyethyl methacrylate, polyethylene glycol methacrylate (molecular weight 400), isobornyl methacrylate, 2-(methylthio)ethyl methacrylate, polyethyleneimine (molecular weight 1000) and 2,4,6 (trimethylbenzoyl) diphenylphosphine oxide (TPO) are added into a stirring tank in sequence, and stirred evenly to obtain an embedding resin, i.e., a photocurable resin with adjustable refractive index.

[0056] This embodiment also provides the application of the light-curable resin with adjustable refractive index in tissue embedding.

[0057] Embodiment 3:

[0058] This embodiment provides a photocurable resin with adjustable refractive index. The raw material components for preparing the photocurable resin include, by mass percentage:

[0059] Polymerizable monomer (80%):

[0060]

[0061] The preparation method of the light-curable resin with adjustable refractive index is as follows:

[0062] According to the above-mentioned mass ratio, hydroxyethyl methacrylate, polyethylene glycol methacrylate (molecular weight 400), isobornyl methacrylate, 2-(methylthio)ethyl methacrylate, polyethyleneimine (molecular weight 2000) and 2,4,6 (trimethylbenzoyl) diphenylphosphine oxide (TPO) are added into a stirring tank in sequence, and stirred evenly to obtain an embedding resin, i.e., a photocurable resin with adjustable refractive index.

[0063] This embodiment also provides the application of the light-curable resin with adjustable refractive index in tissue embedding.

[0064] Embodiment 4:

[0065] This embodiment provides a photocurable resin with adjustable refractive index. The raw material components for preparing the photocurable resin include, by mass percentage:

[0066] Polymerizable monomer (80%):

[0067] Hydroxyethyl Methacrylate 40%

[0068] Isobornyl methacrylate 40%

[0069] High refractive index monomer: 2-(methylthio)ethyl methacrylate 10%

[0070] Diluent: polyethylene glycol (molecular weight 400) 9.5%

[0071] Photoinitiator: 2,4,6 (trimethylbenzoyl) diphenylphosphine oxide 0.5%

[0072] The preparation method of the light-curable resin with adjustable refractive index is as follows:

[0073] According to the above mass ratio, hydroxyethyl methacrylate, isobornyl methacrylate, 2-(methylthio)ethyl methacrylate, polyethylene glycol (molecular weight 400) and 2,4,6 (trimethylbenzoyl) diphenylphosphine oxide (TPO) are added to a stirring tank in sequence, and stirred evenly to obtain an embedding resin, i.e., a photocurable resin with adjustable refractive index.

[0074] This embodiment also provides the application of the light-curable resin with adjustable refractive index in tissue embedding.

[0075] Embodiment 5:

[0076] This embodiment provides a photocurable resin with adjustable refractive index. The raw material components for preparing the photocurable resin include, by mass percentage:

[0077] Polymerizable monomer (80%):

[0078] Hydroxyethyl Methacrylate 40%

[0079] Isobornyl methacrylate 40%

[0080] High refractive index monomer: 2-(methylseleno)ethyl methacrylate 15%

[0081] Diluent: polyethylene glycol (molecular weight 2000) 4.5%

[0082] Photoinitiator: 2,4,6 (trimethylbenzoyl) diphenylphosphine oxide 0.5%

[0083] The preparation method of the light-curable resin with adjustable refractive index is as follows:

[0084] According to the above-mentioned mass ratio, hydroxyethyl methacrylate, isobornyl methacrylate, 2-(methylseleno)ethyl methacrylate, polyethylene glycol (molecular weight 2000) and 2,4,6 (trimethylbenzoyl) diphenylphosphine oxide (TPO) are added into a stirring tank in sequence, and stirred evenly to obtain an embedding resin, i.e., a photocurable resin with adjustable refractive index.

[0085] This embodiment also provides the application of the light-curable resin with adjustable refractive index in tissue embedding.

[0086] The viscosity before photocuring, photocuring time and refractive index of the photocurable resins prepared in the above Examples 1 to 5 were tested using an M-2000V ellipsometer. The test results are shown in Table 1 below.

[0087] Table 1:

[0088]

[0089] It can be seen from the experimental data in Table 1 that the viscosity of the light curing agent can be adjusted by adding a diluent, which is conducive to adapting to the operation of various automatic cover slide machines. The refractive index of the embedding agent can be adjusted by adding a high refractive index monomer. By adjusting the ratio, the refractive index of the embedding agent can be accurately adjusted between 1.51 and 1.68. Since it does not contain low-boiling point toxic and harmful organic solvents, it can protect the health of experimental operators and does not damage the automatic cover slide machine.

[0090] Embedding experiment of tissue sections:

[0091] 400 pathological sections were randomly selected; and a sealing test was performed using commercial conventional neutral gum (20% xylene) and the environmentally friendly light curing agent prepared in Example 5.

[0092] The randomly taken pathological sections were divided into two groups, A and B, and then placed in a 60°C oven for 30 minutes, followed by conventional HE staining. After staining, they were placed in a fume hood to dry until half dry, and finally manually sealed by the same pathology technician, where the A group sections were sealed with traditional neutral gum, and the B group sections were sealed with the environmentally friendly light curing agent of Example 5. The same pathology technician sealed the A and B group sections, using a dropper to draw a proper amount of reagent and drop it on the slide, and then cover it with a cover glass at an appropriate bevel angle. The A group sections were placed in a fume hood at room temperature, and the B group sections were placed under ultraviolet light for irradiation and sealing. The experimental results are shown in Table 1 below. Figure 1 and Figure 2 shown.

[0093] Table 1: Comparison of sealing effects of two groups of sealing agents

[0094] feature Group A(N=200) B(N=200) <![CDATA[X 2 Value]]> P-value none 148 131 bubble Small amount 37 31 56.214 <0.05 Large 15 38 Glue overflow have 32 69 27.196 <0.05 none 168 131 Gum distribution Uniform 182 144 26.238 <0.05 Uneven 18 56 transparency Clarity 185 152 24.373 <0.05 Unclear 15 48 After drying Yellowing 11 15 9.313 <0.05 none 189 185

[0095] The results showed that the A group was sealed by natural drying, and the B group was sealed within 1 minute under the irradiation of 60W ultraviolet light with bright gum. Through parallel observation and comparison between the two groups, it was found that there was no significant difference in the various indicators of the pathological sections after sealing with the sealing agent in the A and B groups. The overall section sealing agent was evenly distributed, with less glue overflow and good transparency. A few turned yellow after drying, but it did not affect the overall reading effect. The tissue cells showed a clear structure under the microscope, and the cells were clearly red and blue in conventional HE staining ( Figure 1 , Figure 2 The comparison of the two groups of slices is shown in Table 1. After drying, both groups of slices can be stored for a long time for pathological reading and archiving.

[0096] The present invention adopts an environmentally friendly light-curing agent to perform ultraviolet curing after sealing, and proves that the use of light-curing environmentally friendly sealing agent to replace traditional neutral gum sealing agent is a good choice without affecting the quality of pathological film reading, and is a pollution-free and safe reagent worthy of promotion.

[0097] The present invention uses specific embodiments to illustrate the principles and implementation methods of the present invention. The description of the above embodiments is only used to help understand the method of the present invention and its core idea. At the same time, for those skilled in the art, according to the idea of ​​the present invention, there will be changes in the specific implementation methods and application scope. In summary, the content of this specification should not be understood as a limitation on the present invention.

Claims

1. A photocurable resin with adjustable refractive index, characterized in that: The raw material components for preparing the photocurable resin include, based on 100% by mass:

2. The photocurable resin according to claim 1, characterized in that: The photoinitiator is selected from 2,4,6 (trimethylbenzoyl) diphenylphosphine oxide and / or ethyl 2,4,6-trimethylbenzoylphosphonate.

3. The photocurable resin according to claim 1, characterized in that: The polymerizable monomer is selected from methacrylate monomers.

4. The photocurable resin according to claim 3, characterized in that: The methacrylate monomer is selected from one or more combinations of hydroxyethyl methacrylate, polyethylene glycol methacrylate, isobornyl methacrylate, methyl methacrylate, ethyl methacrylate, butyl methacrylate, isobutyl methacrylate, 2-ethylhexyl methacrylate, glycidyl methacrylate, phenoxyethyl methacrylate, dimethylaminoethyl methacrylate and trifluoroethyl methacrylate.

5. The photocurable resin according to claim 4, characterized in that: The molecular weight of the polyethylene glycol methacrylate is 200-2000.

6. The photocurable resin according to claim 1, characterized in that: The high refractive index monomer is selected from one or more combinations of 2-methyl-2-acrylic acid cyclothioethane methyl ester, 2-(methylthio)ethyl methacrylate, 2-(methylseleno)ethyl methacrylate, 4-methylthiobenzyl methacrylate and 4-methylselenobenzyl methacrylate.

7. The photocurable resin according to claim 1, characterized in that: The diluent is selected from one or more of polyethylene glycol, polypropylene glycol and polyethyleneimine.

8. The photocurable resin according to claim 1 or 7, characterized in that: The molecular weight of the diluent is 300-5000.

9. The method for preparing a photocurable resin according to any one of claims 1 to 8, characterized in that: The photocurable resin preparation method comprises: Add polymerizable monomer, high refractive index monomer, diluent and photoinitiator into a stirring kettle in sequence, and stir evenly to obtain photocurable resin.

10. Use of the photocurable resin according to any one of claims 1 to 8 or the photocurable resin prepared by the preparation method according to claim 9 in tissue embedding.

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