Specimen for displaying longitudinal partition film structure and preparation method and application thereof

Abstract

The invention provides a specimen for displaying a medial membrane structure and a preparation method and application thereof, and belongs to the technical field of medical tissue specimen preparation. The method comprises the following steps: fixing a fresh tissue block in a mediastinal region in a stationary liquid, washing with water, and dehydrating to obtain a dehydrated tissue block; carrying out photopolymerization curing on the dehydrated tissue block to obtain an embedded specimen; adhering the embedded specimen to a glass slide A by adopting an adhesive, and then positioning and polishing the embedded specimen; adhering a glass slide B to the polished surface of the embedded specimen by using an adhesive to obtain an original specimen; and cutting, grinding, polishing and dyeing the original specimen to obtain the specimen showing the longitudinal partition film structure. According to the longitudinal septum membrane hardening specimen prepared by the invention, anatomical characteristics constructed by membrane structure fibers are relatively comprehensively displayed in a large-scale region with compact tissue gaps, and the bottleneck problem which puzzles the research of a membrane structure in a longitudinal septum region for a long time can be solved.

Description

Technical Field

[0001] The invention belongs to the technical field of medical tissue specimen preparation, and in particular relates to a specimen showing a mediastinal membrane structure, a preparation method and an application thereof. Background Art

[0002] Existing anatomical specimens and methods have limitations in displaying the membranous structure of the mediastinum. The most common formalin-fixed gross anatomical specimens cannot observe the complete mediastinal thin fine structure; while the microscopic histological sectioning and staining methods can display the membranous structure of the mediastinum in a small range through continuous sectioning of hard tissue, but cannot display large-scale anatomical features; although recent plasticized specimens and tomographic plasticized specimens can display a large range of fibrous anatomical structures, due to the thin structure, deformation and extrusion during production, the membranous structure of the mediastinum and the adjacent organs are closely attached, and the tissue gaps are too tight, making them difficult to identify in plasticized specimens. At present, there are no specimens and methods that can more comprehensively display the anatomical characteristics of the membranous structure at the macroscopic level in terms of fine structures, especially in areas with dense gaps. Summary of the invention

[0003] In view of this, the object of the present invention is to provide a specimen showing the structure of the mediastinum membrane and a preparation method and application thereof.

[0004] In order to achieve the above-mentioned object of the invention, the present invention provides the following technical solutions:

[0005] The present invention provides a method for preparing a specimen showing a mediastinal membrane structure, comprising the following steps:

[0006] 1) Fixing the fresh tissue block in the mediastinum region in a fixative, washing with water, and dehydrating to obtain a dehydrated tissue block;

[0007] 2) curing the dehydrated tissue block by photopolymerization to obtain an embedded specimen;

[0008] 3) Adhesive is used to adhere the embedded specimen to the slide A, and then the embedded specimen is positioned and polished;

[0009] 4) Adhere the glass slide B to the polished surface of the embedded specimen with an adhesive to obtain the original specimen; cut, grind, polish, and stain the original specimen to obtain a specimen showing the structure of the mediastinum membrane.

[0010] Preferably, the dehydration in step 1) is divided into two steps, the first step is to use alcohol solutions with volume fractions of 45-55%, 65-72%, 73-78%, 80-88%, 88-98%, 100%, 100%, 100% for dehydration 8 times, the second step is to use anhydrous ethanol and photopolymerization resin infiltration embedding solution T7200VLC for dehydration 3 times, the volume ratio of anhydrous ethanol and photopolymerization resin infiltration embedding solution T7200VLC is 7:3, 1:1, 3:7 respectively; after the dehydration, the photopolymerization resin infiltration embedding solution T7200VLC is used for infiltration 3 times in sequence; the time of each infiltration is 6-8d;

[0011] The volume of the dehydration solution each time is 8 to 12 times the volume of the fresh tissue block in the mediastinum region, and the dehydration time each time is 4 to 8 hours.

[0012] Preferably, the fixing solution in step 1) is a paraformaldehyde solution, and the concentration of the paraformaldehyde solution is 30 to 50 g / L;

[0013] The fixing time is 42 to 54 hours, and the washing time is 18 to 30 hours.

[0014] Preferably, the adhesive in step 3) is Technovit 4000 adhesive;

[0015] The positioning method includes: positioning on the embedded specimen by laser;

[0016] The polishing method comprises: exposing the surface of the embedded specimen by cutting, and polishing the surface of the embedded specimen.

[0017] Preferably, the adhesive in step 4) is Technovit 7210VLC adhesive;

[0018] The cutting method comprises: cutting the original specimen into slices with a thickness of 80 to 120 μm from the glass slide B side of the original specimen.

[0019] Preferably, the method of grinding the disc in step 4) comprises: setting the thickness of the grinding disc for different specimens, and then grinding the disc on various levels of P320, P800, P1200, and P2500 silicon carbide grinding disc paper.

[0020] Preferably, the polishing method in step 4) comprises: polishing the slices with P4000 silicon carbide grinding paper.

[0021] Preferably, the dyeing method comprises: dyeing the polished slices with a dye solution, washing with water, and sealing with neutral gum.

[0022] The present invention also provides a specimen showing the mediastinum membrane structure prepared by the preparation method, wherein the thickness of the specimen showing the mediastinum membrane structure is 5 to 10 μm.

[0023] The present invention also provides a specimen showing the mediastinal membrane structure prepared by the preparation method or the application of the specimen showing the mediastinal membrane structure in pathological research and diagnosis.

[0024] Compared with the prior art, the present invention has the following beneficial effects:

[0025] The membranous sclerosis specimen of the mediastinum prepared by the preparation method of the present invention realizes the anatomical characteristics of the fiber structure of the membrane structure that is more comprehensively displayed in a large area with dense tissue gaps, and can solve the bottleneck problem of the long-standing research on the membrane structure of the mediastinum region, including the starting and ending or attachment sites, distribution, running direction, and relationship between the surrounding adjacent structures and organs of the membrane structure. The present invention has a wide range of application values ​​in biomedical teaching, life science popularization, biomedical research, clinical and sports rehabilitation training, etc., and has extremely high potential social and economic benefits. BRIEF DESCRIPTION OF THE DRAWINGS

[0026] Figure 1 It is a macro-micro histological change diagram of the horizontal section of the upper opening of the thorax;

[0027] Figure 2 It is the distribution map of the membrane structure fibers around the esophagus at the upper thoracic opening;

[0028] Figure 3 It is the distribution map of the membrane structure around the esophagus at the height of the bronchial hilum;

[0029] Figure 4 It is the distribution map of the perioesophageal membrane structure at the inferior pulmonary vein height. DETAILED DESCRIPTION

[0030] The present invention provides a method for preparing a specimen showing a mediastinal membrane structure, comprising the following steps:

[0031] 1) Fixing the fresh tissue block in the mediastinum region in a fixative, washing with water, and dehydrating to obtain a dehydrated tissue block;

[0032] 2) curing the dehydrated tissue block by photopolymerization to obtain an embedded specimen;

[0033] 3) Adhesive is used to adhere the embedded specimen to the slide A, and then the embedded specimen is positioned and polished;

[0034] 4) Adhere the glass slide B to the polished surface of the embedded specimen with an adhesive to obtain the original specimen; cut, grind, polish, and stain the original specimen to obtain a specimen showing the structure of the mediastinum membrane.

[0035] In the present invention, the fresh tissue block of the mediastinum region is fixed in a fixative, washed with water, and then dehydrated to obtain a dehydrated tissue block. The fixative is preferably a paraformaldehyde solution, and the concentration of the paraformaldehyde solution is preferably 30 to 50 g / L, more preferably 35 to 45 g / L; the fixation time is preferably 42 to 54 hours, more preferably 45 to 51 hours; the washing time is preferably 18 to 30 hours, more preferably 21 to 27 hours; the dehydration is preferably carried out by using EXAKT 510 dehydration instrument; the dehydration is divided into two steps, the first step is preferably to use the alcohol solution with a volume fraction of 45-55%, 65-72%, 73-78%, 80-88%, 88-98%, 100%, 100%, 100% for dehydration 8 times, and further preferably to use the alcohol solution with a volume fraction of 48-52%, 68-71%, 74-76%, 83-86%, 92-97%, 100%, 100%, 100% for dehydration 8 times; the second step is preferably to use anhydrous ethanol and photopolymerization resin infiltration embedding solution T7200VLC The mixed dehydration was performed for 3 times, and the volume ratios of the anhydrous ethanol and the photopolymerizable resin infiltration embedding solution T7200VLC were 7:3, 1:1, and 3:7, respectively; the volume of the dehydration solution each time was preferably 8 to 12 times the volume of the fresh tissue block in the mediastinum region, and more preferably 9 to 11 times the volume of the fresh tissue block in the mediastinum region; the time for each dehydration was preferably 4 to 8 hours, and more preferably 5 to 7 hours; after the dehydration, the photopolymerizable resin infiltration embedding solution T7200VLC was used for 3 infiltrations in sequence, and the time for each infiltration was preferably 6 to 8 days, and more preferably 7 days.

[0036] In the present invention, the dehydrated tissue block is subjected to photopolymerization curing to obtain an embedded specimen. The photopolymerization curing is preferably performed in a photocuring embedding machine, and the photocuring embedding machine is preferably an EXAKT 520 photocuring embedding machine.

[0037] In the present invention, an adhesive is used to adhere the embedded specimen to the glass slide A, and then the embedded specimen is positioned and polished. The adhesive is preferably Technovit4000 adhesive; the positioning method includes: positioning on the embedded specimen by laser; vacuum adsorbing the glass slide A on the slicer fixture before positioning, and the slicer is preferably an EXAKT300CP slicer; the polishing method includes: exposing the surface of the embedded specimen by cutting, and polishing the surface of the embedded specimen; the cutting is preferably done using a diamond saw blade; before polishing, the glass slide A is adsorbed on a grinder, and the grinder is preferably an EXAKT 400CS grinder.

[0038] In the present invention, a glass slide B is adhered to the polished surface of the embedded specimen by an adhesive to obtain an original specimen; the original specimen is cut, ground, polished, and stained to obtain a specimen showing the structure of the mediastinal membrane. The adhesive is preferably Technovit 7210VLC adhesive; the adhesion is preferably performed on an EXAKT 401 slide bonding device; the cutting method includes: cutting the original specimen into slices with a thickness of 80 to 120 μm from the glass slide B side of the original specimen, and the thickness of the slice is further preferably 90 to 110 μm; before the slicing, the glass slide A side of the original specimen is adsorbed on the slicer fixture, and the slicer is preferably an EXAKT 300CP slicer; the cutting is positioned by an automatic stepping positioning device. The grinding method includes: setting the thickness of the grinding disc for different specimens, and then grinding the disc on various levels of P320, P800, P1200, and P2500 silicon carbide grinding disc paper; before grinding, adsorbing the slide B of the original specimen on a fully automatic digital display grinding machine with a rolling grinding disc device and an electronic measurement control system, and the fully automatic digital display grinding machine is preferably an EXAKT 400CS fully automatic digital display grinding machine; the polishing method includes: polishing the slices with P4000 silicon carbide grinding disc paper. The dyeing method includes: dyeing the polished slices with a dye solution, washing with water, and sealing with neutral gum. The staining adopts Regaud hematoxylin staining solution or Weigert iron hematoxylin staining solution to stain the nucleus, and the time of the nuclear staining is preferably 5-10 minutes, and more preferably 6-9 minutes; after the nuclear staining, 1% hydrochloric acid alcohol is used for differentiation, and the differentiation time is preferably 5-15 seconds, and more preferably 8-12 seconds; after differentiation, washing with water, after washing with water, 0.1-1% lithium carbonate is used to increase the degree of anti-blueing, and the concentration of the lithium carbonate is preferably 0.3-0.8%, after washing with water, Masson fuchsin acid fuchsin solution is used for 5-10 minutes, and the fuchsin time is preferably 6-9 minutes; then washing with glacial acetic acid aqueous solution, differentiation with phosphomolybdic acid aqueous solution for 3-5 minutes, and the differentiation time is preferably 3.5-4.5 minutes, and then washing with glacial acetic acid aqueous solution; then dyeing with aniline blue aqueous solution or 1% light green aqueous solution for 1-2 minutes, and the dyeing time is preferably 1.2-1.8 minutes, washing with glacial acetic acid aqueous solution, dehydration with 95% ethanol and anhydrous ethanol in turn, xylene transparent, and neutral gum sealing after transparency. The entire process of cutting, grinding and polishing is controlled by an AW100 (micrometer) measurement control system, which can make the thickness of each part of the slice uniform (difference does not exceed ±0.01 μm) and the surface of the slice smooth. After focusing under an optical microscope, the relationship between various tissues and cell results can be clearly displayed.

[0039] The present invention also provides a specimen showing the mediastinum membrane structure prepared by the preparation method, wherein the thickness of the specimen showing the mediastinum membrane structure is 5 to 10 μm.

[0040] The present invention also provides a specimen showing the mediastinal membrane structure prepared by the preparation method or the application of the specimen showing the mediastinal membrane structure in pathological research and diagnosis.

[0041] The technical solutions provided by the present invention are described in detail below in conjunction with the embodiments, but they should not be construed as limiting the protection scope of the present invention.

[0042] Example 1

[0043] 1. Dissection: After the human body is preserved and fixed with 10% formalin solution, the membranous structure and its related structures in the target area are dissected and sampled to obtain the mediastinum area specimen.

[0044] 2. Tissue fixation: Fresh tissue blocks of mediastinal specimens were fixed in 40 g / L paraformaldehyde fixative for 48 hours and then rinsed in running water for 24 hours.

[0045] 3. Dehydration: Dehydrate with alcohol solutions from low to high concentrations (50%, 70%, 75%, 85%, 95%, 100%, 100%, 100%) in an EXAKT 510 dehydrator, and then dehydrate with anhydrous ethanol and photopolymerization resin infiltration embedding solution T7200VLC, the volume ratio of anhydrous ethanol and photopolymerization resin infiltration embedding solution T7200VLC is 7:3, 1:1, 3:7 respectively; then infiltrate with photopolymerization resin infiltration embedding solution T7200VLC three times in sequence, each infiltration time is 7 days. The volume of each dehydration solution is 10 times the volume of the fresh tissue block of the mediastinal specimen, and the dehydration time is 6 hours.

[0046] 4. After embedding and dehydration, the tissue block is placed in a special mold and photopolymerized and cured in an EXAKT 520 light-curing embedding machine.

[0047] 5. After the tissue block is plasticized and polymerized, the special mold is removed and the embedded specimen is bonded to slide A with Technovit4000 adhesive. Slide A is vacuum-adsorbed on the EXAKT300CP slicer fixture, positioned on the embedded specimen by laser, and cut with a diamond saw blade to expose the specimen surface. Slide A is adsorbed on the EXAKT 400CS grinder to polish the specimen surface. Slide B is bonded to the polished surface of the specimen on the EXAKT 401 slide bonding device with Technovit 7210VLC adhesive. Slide A, embedded specimen and slide B are combined into a sandwich-like structure. The slide A side of the sandwich structure is adsorbed on the EXAKT 300CP slicer fixture, and a slice with a thickness of 100μm is cut from the slide B side by the automatic stepping positioning device. Adsorb the slide B with the slice attached to the EXAKT 400CS fully automatic digital display grinding machine with a rolling grinding disc device and an electronic measurement control system, and pre-set the thickness of the grinding disc for different specimens. Grind the slices to the predetermined thickness on various levels of P320, P800, P1200, and P2500 silicon carbide grinding disc paper, and finally polish the slices with P4000 silicon carbide grinding disc paper until there are no scratches. The entire process is controlled by the AW100 (micrometer) measurement control system. According to the experimental observation requirements, the final thickness of the slice can be 5 to 10 μm.

[0048] 6. Staining: Use Regaud hematoxylin stain or Weigert iron hematoxylin stain to stain the nucleus for 8 minutes; differentiate with 1% hydrochloric acid alcohol for 10 seconds, and wash thoroughly with water; add 0.5% lithium carbonate for 5 minutes to increase the degree of anti-blueing, and wash with water; use Masson Ponceau acid fuchsin solution for 8 minutes; wash with 2% glacial acetic acid aqueous solution for 1 minute; differentiate with 1% phosphomolybdic acid aqueous solution for 4 minutes, and wash with 2% glacial acetic acid aqueous solution for 1 minute; without washing, directly stain with aniline blue aqueous solution or 1% light green aqueous solution for 1.5 minutes; wash with 0.2% glacial acetic acid aqueous solution for 1 minute; dehydrate with 95% ethanol for 3 seconds, dehydrate with anhydrous ethanol 3 times for 5 seconds each time, make it clear with xylene 3 times for 1 minute each time, and seal with neutral gum.

[0049] Example 2

[0050] 1. Dissection: After the human body is preserved and fixed with 10% formalin solution, the membranous structure and its related structures in the target area are dissected and sampled to obtain the mediastinum area specimen.

[0051] 2. Tissue fixation: Fresh tissue blocks of mediastinal specimens were fixed in 30 g / L paraformaldehyde fixative for 42 hours and then rinsed in running water for 18 hours.

[0052] 3. Dehydration: Dehydrate with alcohol solutions from low to high concentrations (45%, 65%, 73%, 80%, 88%, 100%, 100%, 100%) in an EXAKT 510 dehydrator, and then dehydrate with anhydrous ethanol and photopolymerization resin infiltration embedding solution T7200VLC, the volume ratio of anhydrous ethanol and photopolymerization resin infiltration embedding solution T7200VLC is 7:3, 1:1, 3:7 respectively; then infiltrate with photopolymerization resin infiltration embedding solution T7200VLC three times in sequence, each infiltration time is 6 days. The volume of each dehydration solution is 8 times the volume of the fresh tissue block of the mediastinal specimen, and the dehydration time is 4 hours.

[0053] 4. After embedding and dehydration, the tissue block is placed in a special mold and photopolymerized and cured in an EXAKT 520 light-curing embedding machine.

[0054] 5. After the tissue block is plasticized and polymerized, the special mold is removed and the embedded specimen is bonded to slide A with Technovit4000 adhesive. Slide A is vacuum-adsorbed on the EXAKT300CP slicer fixture, positioned on the embedded specimen by laser, and cut with a diamond saw blade to expose the specimen surface. Slide A is adsorbed on the EXAKT 400CS grinder to polish the specimen surface. Slide B is bonded to the polished surface of the specimen on the EXAKT 401 slide bonding device with Technovit 7210VLC adhesive. Slide A, embedded specimen and slide B are combined into a sandwich-like structure. The slide A side of the sandwich structure is adsorbed on the EXAKT 300CP slicer fixture, and a slice with a thickness of 80μm is cut from the slide B side by the automatic stepping positioning device. Adsorb the slide B with the slice attached to the EXAKT 400CS fully automatic digital display grinding machine with a rolling grinding disc device and an electronic measurement control system, and pre-set the thickness of the grinding disc for different specimens. Grind the slices to the predetermined thickness on various levels of P320, P800, P1200, and P2500 silicon carbide grinding disc paper, and finally polish the slices with P4000 silicon carbide grinding disc paper until there are no scratches. The entire process is controlled by the AW100 (micrometer) measurement control system. According to the experimental observation requirements, the final thickness of the slice can be 5 to 10 μm.

[0055] 6. Staining: Use Regaud hematoxylin stain or Weigert iron hematoxylin stain to stain the nucleus for 5 minutes; differentiate with 1% hydrochloric acid alcohol for 5 seconds, and wash thoroughly with water; add 0.1% lithium carbonate for 5 minutes to increase the degree of anti-blueing, and wash with water; use Masson Ponceau acid fuchsin solution for 5 minutes; wash with 2% glacial acetic acid aqueous solution for 1 minute; differentiate with 1% phosphomolybdic acid aqueous solution for 3 minutes, and wash with 2% glacial acetic acid aqueous solution for 1 minute; without washing, directly stain with aniline blue aqueous solution or 1% light green aqueous solution for 1 minute; wash with 0.2% glacial acetic acid aqueous solution for 1 minute; dehydrate with 95% ethanol for 2 seconds, dehydrate with anhydrous ethanol 3 times for 5 seconds each time, make it clear with xylene 3 times for 1 minute each time, and seal with neutral gum.

[0056] Example 3

[0057] 1. Dissection: After the human body is preserved and fixed with 10% formalin solution, the membranous structure and its related structures in the target area are dissected and sampled to obtain the mediastinum area specimen.

[0058] 2. Tissue fixation: Fresh tissue blocks of mediastinal specimens were fixed in 50 g / L paraformaldehyde fixative for 54 hours and then rinsed in running water for 30 hours.

[0059] 3. Dehydration: Dehydrate with alcohol solutions from low to high concentrations (55%, 72%, 78%, 88%, 98%, 100%, 100%, 100%) in an EXAKT 510 dehydrator, and then dehydrate with anhydrous ethanol and photopolymerization resin infiltration embedding solution T7200VLC, the volume ratio of anhydrous ethanol and photopolymerization resin infiltration embedding solution T7200VLC is 7:3, 1:1, 3:7 respectively; then infiltrate with photopolymerization resin infiltration embedding solution T7200VLC three times in sequence, each infiltration time is 8 days. The volume of each dehydration solution is 12 times the volume of the fresh tissue block of the mediastinum specimen, and each dehydration time is 8 hours.

[0060] 4. After embedding and dehydration, the tissue block is placed in a special mold and photopolymerized and cured in an EXAKT 520 light-curing embedding machine.

[0061] 5. After the tissue block is plasticized and polymerized, the special mold is removed and the embedded specimen is bonded to slide A with Technovit4000 adhesive. Slide A is vacuum-adsorbed on the EXAKT300CP slicer fixture, positioned on the embedded specimen by laser, and cut with a diamond saw blade to expose the specimen surface. Slide A is adsorbed on the EXAKT 400CS grinder to polish the specimen surface. Slide B is bonded to the polished surface of the specimen on the EXAKT 401 slide bonding device with Technovit 7210VLC adhesive. Slide A, embedded specimen and slide B are combined into a sandwich-like structure. The slide A side of the sandwich structure is adsorbed on the EXAKT 300CP slicer fixture, and a slice with a thickness of 120μm is cut from the slide B side by the automatic stepping positioning device. Adsorb the slide B with the slice attached to the EXAKT 400CS fully automatic digital display grinding machine with a rolling grinding disc device and an electronic measurement control system, and pre-set the thickness of the grinding disc for different specimens. Grind the slices to the predetermined thickness on various levels of P320, P800, P1200, and P2500 silicon carbide grinding disc paper, and finally polish the slices with P4000 silicon carbide grinding disc paper until there are no scratches. The entire process is controlled by the AW100 (micrometer) measurement control system. According to the experimental observation requirements, the final thickness of the slice can be 5 to 10 μm.

[0062] 6. Staining: Use Regaud hematoxylin stain or Weigert iron hematoxylin stain to stain the nucleus for 10 minutes; differentiate with 1% hydrochloric acid alcohol for 15 seconds, and wash thoroughly with water; add 1% lithium carbonate for 5 minutes to increase the degree of anti-blueing, and wash with water; use Masson Ponceau acid fuchsin solution for 10 minutes; wash with 2% glacial acetic acid aqueous solution for 1 minute; differentiate with 1% phosphomolybdic acid aqueous solution for 5 minutes, and wash with 2% glacial acetic acid aqueous solution for 1 minute; without washing, directly stain with aniline blue aqueous solution or 1% light green aqueous solution for 2 minutes; wash with 0.2% glacial acetic acid aqueous solution for 1 minute; dehydrate with 95% ethanol for 3 seconds, dehydrate with anhydrous ethanol 3 times for 5 seconds each time, make it clear with xylene 3 times for 1 minute each time, and seal with neutral gum.

[0063] Experimental Example 1

[0064] The stained specimen prepared in Example 1 was sliced ​​and the relationship between the tissues and the cell structure were observed under an optical microscope.

[0065] Experimental results: Figure 1 to Figure 4 shown. Figure 1 For the overall picture, Figures 2 to 4 It is a local picture.

[0066] Depend on Figure 1It can be seen that the visceral fascia (▲), pretracheal fascia (tracheal sheath ↓), vascular sheath (*), prevertebral fascia (△) and parietal pleura (>) can be seen on both sides of the upper thoracic opening. The recurrent laryngeal nerve (◆), thoracic duct (TD), vagus nerve (LVN); esophagus (Eso), trachea (Traschea), right common carotid artery (RCCA), right subclavian artery (RSCA), left common carotid artery (LCCA), and left subclavian artery (LSCA) can be seen on both sides.

[0067] Depend on Figure 2 It can be seen that the two sides of the esophagus are divided into deep and superficial layers. The deep layer covers the trachea and the outside of the recurrent laryngeal nerve to the left and continues to the front of the trachea, and connects with the opposite side to form the pretracheal fascia; a vascular sheath is formed on the outside of the trachea to wrap around the left common carotid artery, and this vascular sheath continues upward with the carotid sheath in the neck; the superficial layer of fascia wraps the left subclavian artery to the left front and outside. On the right side of the esophagus, the superficial part of the visceral fascia is connected to the right mediastinal pleura. At the height of the upper opening of the thorax, the thoracic duct is located deep between the left subclavian artery and the left common carotid artery, sandwiched between the two layers of fascia of the visceral fascia on the left side of the esophagus.

[0068] Depend on Figure 3 It can be seen that the visceral fascia located in front and behind the esophagus is thin, wrapping around the esophagus to form an esophageal sheath, which is connected to the mediastinal pleura on both sides. The vascular sheath wraps around the descending aorta and thoracic duct, and covers the azygos vein to the right. Visceral fascia, pretracheal fascia, vascular sheath, prevertebral fascia and mediastinal pleura can be seen. Thoracic duct, lymph nodes, azygos vein, arterio-esophageal ligament, paraesophageal space.

[0069] Depend on Figure 4 It can be seen that the anterior wall of the esophagus is adjacent to the pericardial cavity and the left atrium, and is wrapped by the visceral fascia from the front and back. The right side of the esophagus is not covered by fascia, and its anterior and posterior fascial layers are connected to the right mediastinal pleura respectively. On the left side of the esophagus, the fascia gradually approaches and fuses into a single layer, spanning the front of the descending aorta and connecting to the left mediastinal pleura. The vascular sheath surrounds the descending aorta and thoracic duct, and extends to the right to cover the azygos vein. Visceral fascia, vascular sheath, prevertebral fascia and mediastinal pleura can be seen. Thoracic duct, azygos vein, parietal pericardium, pericardial cavity, visceral pericardium, atrial muscle, arterio-esophageal ligament, paraesophageal space.

[0070] It can be seen from the above embodiments and experimental examples that the specimens prepared by the preparation method of the present invention can more comprehensively display the anatomical characteristics of the thin film structure fiber structure in a large area with dense tissue gaps, including the starting and ending points or attachment sites, distribution, running direction, and the relationship between the surrounding adjacent structural organs of the thin film structure.

[0071] The above is only a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principle of the present invention. These improvements and modifications should also be regarded as the scope of protection of the present invention.

Claims

1. A method for preparing a specimen showing the structure of the mediastinum membrane, characterized in that: The following steps are involved: 1) Fixing the fresh tissue block in the mediastinum region in a fixative, washing with water, and dehydrating to obtain a dehydrated tissue block; 2) performing photopolymerization curing on the dehydrated tissue block to obtain an embedded specimen; 3) Adhesive is used to adhere the embedded specimen to the slide A, and then the embedded specimen is positioned and polished; 4) Adhere the glass slide B to the polished surface of the embedded specimen with an adhesive to obtain the original specimen; cut, grind, polish, and stain the original specimen to obtain a specimen showing the structure of the mediastinum membrane.

2. The preparation method according to claim 1, characterized in that: Step 1) The dehydration is divided into two steps. The first step is to use alcohol solutions with volume fractions of 45-55%, 65-72%, 73-78%, 80-88%, 88-98%, 100%, 100%, and 100% for dehydration 8 times in sequence. The second step is to use anhydrous ethanol and photopolymerization resin infiltration embedding solution T7200VLC for dehydration 3 times. The volume ratio of anhydrous ethanol to photopolymerization resin infiltration embedding solution T7200VLC is 7:3, 1:1, and 3:7 in sequence. After the dehydration, the photopolymerization resin infiltration embedding solution T7200VLC is used for infiltration 3 times in sequence. The time of each infiltration is 6 to 8 days. The volume of the dehydration solution each time is 8 to 12 times the volume of the fresh tissue block in the mediastinum region, and the dehydration time each time is 4 to 8 hours.

3. The preparation method according to claim 1, characterized in that: Step 1) The fixing solution is a paraformaldehyde solution, and the concentration of the paraformaldehyde solution is 30-50 g / L; The fixing time is 42 to 54 hours, and the washing time is 18 to 30 hours.

4. The preparation method according to claim 1, characterized in that: Step 3) The adhesive is Technovit 4000 adhesive; The positioning method includes: positioning on the embedded specimen by laser; The polishing method comprises: exposing the surface of the embedded specimen by cutting, and polishing the surface of the embedded specimen.

5. The preparation method according to claim 1, characterized in that: Step 4) The adhesive is Technovit 7210 VLC adhesive; The cutting method comprises: cutting the original specimen into slices with a thickness of 80 to 120 μm from the glass slide B side of the original specimen.

6. The preparation method according to claim 1, characterized in that: Step 4) The method for grinding the disc comprises: setting the thickness of the disc according to different specimens, and then grinding the disc on various levels of P320, P800, P1200, and P2500 silicon carbide grinding papers.

7. The preparation method according to claim 1, characterized in that: Step 4) The polishing method comprises: polishing the slices with P4000 silicon carbide grinding paper.

8. The preparation method according to claim 1, characterized in that: Step 4) The staining method comprises: staining the polished slices with a dye solution, washing with water, and sealing with neutral gum.

9. The specimen exhibiting the structure of the mediastinum membrane prepared by the preparation method according to any one of claims 1 to 8, characterized in that: The thickness of the specimen showing the mediastinum membrane structure is 5 to 10 μm.

10. Use of the specimen exhibiting the mediastinal membrane structure obtained by the preparation method according to any one of claims 1 to 8 or the specimen exhibiting the mediastinal membrane structure according to claim 9 in pathological research and diagnosis.

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