Embedding method and application of punctured tissue

Abstract

The present invention discloses a method and application for embedding punctured tissue. The embedding method comprises the following steps: covering the surface of the punctured tissue with OCT embedding glue, turning the punctured tissue until it is completely wrapped by the OCT embedding glue; then clipping the punctured tissue wrapped with OCT embedding glue and placing it on a platform without OCT embedding glue; then spreading the punctured tissue on a gasket paper, pressing to ensure that the punctured tissue and the gasket paper are completely in contact, and covering the surface of the punctured tissue with OCT embedding glue; placing the gasket paper with the punctured tissue in a clean embedding tank, lightly pressing the gasket paper around so that the gasket paper fits the bottom of the embedding tank; squeezing out the OCT embedding glue to fill the embedding tank, placing it in a pre-cooled freezing embedding instrument, covering it with a freezing tray, pressing an endothermic hammer, and timing; when the timing ends, taking out the freezing tray, and embedding is completed. The method is simple to operate, and the punctured tissue can be clearly seen after embedding, which is convenient for cutting the entire tissue and is conducive to frozen sectioning.

Description

Technical Field

[0001] The present invention belongs to the field of inspection and detection, and particularly relates to an embedding method for punctured tissue and its application. Background Art

[0002] Biopsy is an important method for diagnosing disease. Open incisions, which involve larger wounds, yield more biopsy tissue, facilitating testing. However, these procedures require a longer recovery period and carry a higher risk of infection. Puncture biopsy, with its smaller wounds and smaller biopsies, increases the difficulty of testing, but facilitates patient recovery and offers a higher safety factor.

[0003] Intraoperative frozen pathology diagnosis is an emergency consultation with a pathologist during surgery to diagnose disease issues related to the surgical plan. The issuance of an intraoperative frozen pathology diagnosis report requires sampling, frozen embedding, frozen sectioning, frozen HE staining, and microscopic observation. Among them, the thickness of conventional frozen tissue sampling is 3-4mm. When frozen embedding, it only needs to be embedded according to the required cutting surface. Slightly unevenness can be achieved by deep incision to completely cut the tissue or the lesion. However, the diameter of the punctured tissue is small, and uneven embedding will result in incomplete cutting, which may lead to missed diagnosis. Therefore, conventional frozen embedding methods are not suitable for punctured tissue.

[0004] Currently, there are only two methods for cryo-embedding puncture tissue, which are referred to as puncture tissue embedding operation 1 and puncture tissue embedding operation 2 for ease of distinction. In puncture tissue embedding operation 1, the specimen tip is frozen with cryo-embedding gel. After the cryo-embedding gel completely solidifies, the tip is placed in a cryo-microtome and trimmed to create a flat surface. This surface provides a flat support for the puncture strip, marking the relative position of the specimen tip. The specimen tip is then removed, and the puncture tissue strips are laid onto this flat surface. Cryo-embedding gel is then added to the puncture strip, allowed to solidify, and the specimen tip is returned to its marked position. Coarse and fine trimming are then performed, and the section is cut. Puncture tissue embedding operation 2 is to freeze the sample head with cryo-embedding gel. When the cryo-embedding gel is in a translucent semi-solidified state, use a freezing hammer to press on the cryo-embedding gel. The cryo-embedding gel is compressed and frozen to form a plane. Then, the punctured tissue strips are laid on the plane. Then, a layer of cryo-embedding gel is evenly applied on the puncture strips until the specimen is completely covered. The specimen is then gently flattened with a freezing hammer and sliced. Both embedding methods have obvious defects: the first defect is that during the embedding process, when the punctured tissue is placed on the low-temperature plane formed by the frozen section embedding agent on the sample head, the contact area between the fresh punctured tissue and the low-temperature plane will instantly solidify and cannot be moved, which may cause the punctured tissue to be embedded unevenly, such as an "arch bridge" shape, with both ends fixed and the middle arched. When slicing, the arched part will be trimmed off and the section cannot be made; the second defect is that the plane provided by the solidified embedding glue on the sample head is used to support the punctured tissue, and the diameter of the punctured tissue is not uniform, which causes the bottom surface of the punctured tissue to be flat but the cross-section to be uneven. If the punctured tissue is to be cut completely, a lot of tissue will be lost; the third defect is that after the punctured tissue is flattened by these two embedding methods, frozen embedding glue must be added to the punctured tissue as soon as possible, otherwise the punctured tissue will not be able to fuse with the subsequently added embedding glue after solidification, and the punctured tissue will be separated from the subsequently added embedding glue and cannot be made into a section.

[0005] Therefore, it is necessary to develop an embedding technology that is more conducive to puncture tissue preparation. Summary of the Invention

[0006] The primary purpose of the present invention is to overcome the shortcomings and deficiencies of the prior art and to provide a method for embedding punctured tissue.

[0007] Another object of the present invention is to apply the embedding method of the punctured tissue to frozen sections.

[0008] The purpose of the present invention is achieved through the following technical solution: A method for embedding punctured tissue, comprising the following steps:

[0009] (1) Precool the freezing embedding apparatus to below -20°C and clean the embedding tank;

[0010] (2) Place the punctured tissue on the platform, cover the surface of the punctured tissue with OCT embedding gel, and turn the punctured tissue over until it is completely wrapped with the OCT embedding gel; then remove the punctured tissue wrapped with OCT embedding gel and place it on a platform without OCT embedding gel to reduce the amount of OCT embedding gel wrapped around the punctured tissue;

[0011] (3) Lay the punctured tissue obtained in step (2) flat on the gasket paper, press to ensure that the punctured tissue and the gasket paper are completely in contact, and cover the surface of the punctured tissue with OCT embedding glue;

[0012] (4) Place the puncture tissue-filled spacer obtained in step (3) into the embedding tank, gently press the spacer around the periphery to make the spacer fit the bottom of the embedding tank; squeeze out the OCT embedding glue to fill the embedding tank, and place it in a pre-cooled freezing embedding instrument; cover the freezing tray, press the endothermic hammer, and start timing;

[0013] (5) When the timer ends, remove the frozen tray and the embedding is completed.

[0014] The pre-cooling temperature in step (1) is preferably -30°C to -40°C; more preferably -35°C.

[0015] The punctured tissue described in step (2) includes the punctured tissue obtained by coarse needle puncture and the punctured tissue obtained by fine needle puncture.

[0016] The platform described in step (2) is a positioning shovel, a photographing board, or other clean flat surface.

[0017] The spacer paper described in step (3) is a thin sheet of paper that is light-transparent and soft, so as to facilitate observation and not damage the blade during slicing; preferably, the spacer paper has a diameter of 3 cm.

[0018] The pressing in step (3) is preferably performed using tweezers.

[0019] The amount of OCT embedding glue used in step (3) is preferably just enough to cover the punctured tissue.

[0020] The timing duration in step (4) is preferably 25 to 35 seconds, more preferably 30 seconds.

[0021] Application of the above-mentioned embedding method for punctured tissue in preparing frozen tissue sections.

[0022] The present invention has the following advantages and effects compared to the prior art:

[0023] (1) The present invention uses a freezing embedding apparatus, which can reduce the formation of tissue ice crystals and improve the quality of slices.

[0024] (2) The present invention wraps the punctured tissue with cryoembedding gel in advance, thereby avoiding the punctured tissue and the cryoembedding gel from separating and being unable to prepare slices.

[0025] (3) The present invention uses a small sample paper as a support, and the position of the punctured tissue can be placed at will.

[0026] (4) The present invention uses small sample gasket paper for leveling, which can make the punctured tissue section flat and avoid missed diagnosis due to incomplete cutting.

[0027] (5) The embedded tissue of the present invention has a smooth cross section, which reduces the waste of punctured tissue and retains more tissue for the final diagnosis after freezing in paraffin. BRIEF DESCRIPTION OF THE DRAWINGS

[0028] Figure 1 This is a photograph of the punctured tissue obtained by coarse needle puncture covered with OCT embedding gel.

[0029] Figure 2 This is a photograph showing the removal of excess OCT embedding gel from around the punctured tissue obtained by coarse needle puncture.

[0030] Figure 3 This is a photograph of the puncture tissue obtained by coarse needle puncture placed on a small sample spacer paper.

[0031] Figure 4 This is a photo of a small sample of gasket paper.

[0032] Figure 5 This is a photograph of a small sample of tissue obtained by coarse needle puncture being placed on a paper pad into the embedding tank of a cryo-embedding apparatus.

[0033] Figure 6 This is a photo of the embedding tank being covered with a freezing tray and pressed with a heat absorbing hammer.

[0034] Figure 7 This is a photograph of the punctured tissue obtained by coarse needle puncture after embedding.

[0035] Figure 8 This is a cross-sectional view of the punctured tissue obtained by coarse needle puncture.

[0036] Figure 9 This is a photograph of a slide obtained after staining of a tissue section obtained by coarse needle aspiration.

[0037] Figure 10 This is a photograph of the punctured tissue obtained by fine needle aspiration.

[0038] Figure 11 This is a photograph of the punctured tissue obtained by fine needle aspiration covered with OCT embedding gel.

[0039] Figure 12This is a photograph of the removal of excess OCT embedding gel around the punctured tissue obtained by fine needle aspiration.

[0040] Figure 13 This is a photograph of a paper pad containing a small sample of tissue obtained by fine needle aspiration.

[0041] Figure 14 This is a photograph of OCT embedding gel filling the embedding tank containing the punctured tissue and small sample spacer paper.

[0042] Figure 15 This is a photograph of the punctured tissue obtained by fine needle aspiration after embedding.

[0043] Figure 16 This is a cross-sectional view of the punctured tissue obtained by fine needle aspiration.

[0044] Figure 17 This is a photograph of a slide obtained after staining of a fine needle aspiration tissue section. DETAILED DESCRIPTION

[0045] The present invention will be described in further detail below with reference to the embodiments and drawings, but the embodiments of the present invention are not limited thereto.

[0046] Example 1

[0047] A method for embedding puncture tissue obtained by coarse needle puncture, comprising the following steps:

[0048] (1) Precool the freezing embedding apparatus (Milestone, Italy) to -35°C and clean the embedding tank;

[0049] (2) Place the puncture strip on the positioning spatula, squeeze a small amount of OCT embedding gel on the surface of the punctured tissue, and use tweezers to flip the puncture strip until it is completely covered with OCT embedding gel (such as Figure 1 Then, remove each puncture strip and reduce the amount of OCT embedding glue wrapped in the puncture strip (as shown in Figure 2 The purpose of this step is to wrap the punctured tissue with embedding glue to facilitate smooth fusion with the subsequent embedding glue; less embedded glue is wrapped to facilitate the punctured tissue to fit as closely as possible with the small sample spacer paper;

[0050] (3) Take out the puncture strip and lay it flat on the small sample spacer paper. Use tweezers to press it properly to ensure that the puncture tissue and the small sample spacer paper are completely in contact (such as Figure 3 As shown), squeeze out a little OCT embedding gel to cover the surface of the puncture strip; small sample spacer paper as shown Figure 4 Shown is a light blue thin sheet of paper with a diameter of 30 mm;

[0051] (4) Clamp the small sample gasket paper with the puncture tissue obtained in step (3) and place it into the embedding tank of the freezing embedding instrument. Gently press the small sample gasket paper around to make the small sample gasket paper fit the bottom of the embedding tank. Then quickly squeeze out the OCT embedding glue to fill the embedding tank (such as Figure 5 Cover with the freezing tray and press the heat absorbing hammer (as shown); Figure 6 As shown), time 30s;

[0052] (5) When the timer is up, take out the freezing tray and the embedding is complete. The embedded puncture tissue is shown in Figure 7. The shape of the puncture tissue can be seen through the small sample gasket paper. Put it into the freezing microtome, adjust the direction of the chuck, and trim it carefully to cut the whole tissue. Figure 8 The frozen sections obtained are as follows: Figure 9 shown.

[0053] Example 2

[0054] A method for embedding puncture tissue obtained by fine needle puncture, comprising the following steps:

[0055] (1) Pre-cool the freezing embedding apparatus to -35°C and clean the embedding tank;

[0056] (2) Place the puncture strip on the photographic plate ( Figure 10 ), squeeze out a small amount of OCT embedding glue on the surface of the punctured tissue ( Figure 11 ), use tweezers to turn the puncture strips until they are completely wrapped by OCT embedding gel, and then take out each puncture strip to reduce the amount of OCT embedding gel wrapped in the puncture strips ( Figure 12 );

[0057] (3) Clip out the puncture strip and lay it flat on the small sample pad paper ( Figure 13 ), squeeze out a little OCT embedding gel to cover the surface of the puncture strip;

[0058] (4) Clamp the small sample gasket paper and place it in the embedding tank of the freezing embedding instrument. First, gently press the punctured tissue to ensure that the small sample gasket paper fits the punctured tissue. Then gently press the small sample gasket paper around to make the small sample gasket paper fit the bottom of the embedding tank. Then quickly squeeze out the OCT embedding glue to fill the embedding tank. Figure 14 ), cover with a freezing tray, press on the endothermic hammer, and time for 30 seconds;

[0059] (5) When the timer is up, take out the frozen tray and embedding is completed; the punctured tissue after embedding is as follows: Figure 15 As shown, the shape of the punctured tissue can be seen through the small sample spacer paper; put it into the freezing microtome, adjust the direction of the chuck, and finely trim to cut the entire tissue, as shown in the figure. Figure 16 The frozen sections obtained were as follows: Figure 17 shown.

[0060] It can be seen that the present invention has the following advantages over the two existing embedding methods for punctured tissue: the first advantage is that the punctured tissue is pre-wrapped with a frozen section embedding agent instead of being placed directly (such as punctured tissue embedding operation 1 and punctured tissue embedding operation 2), thereby avoiding the separation of the punctured tissue and the subsequently added frozen section embedding agent, and can be sliced ​​smoothly; the second advantage is that the punctured tissue can be placed at any position before being placed in the embedding tank of the frozen embedding instrument, thereby preventing the punctured tissue from crossing after embedding, which may result in the punctured tissue being unable to be completely cut, while the existing punctured tissue embedding method, due to the pre-wrapping of the frozen section embedding agent The sample head is wrapped, so when the punctured tissue is placed on the low-temperature plane formed by the frozen section embedding agent on the sample head, the contact area will instantly solidify when the fresh punctured tissue comes into contact with the low-temperature plane, making it impossible to move and adjust the position of the punctured tissue. After placement, the frozen section embedding agent must be added quickly. The embedding effect depends on the operator's proficiency. The third advantage is that the punctured tissue can be clearly seen after embedding, which is convenient for cutting the entire tissue. The fourth advantage, which is also the biggest advantage, is that the punctured tissue can be embedded flat, which is conducive to cutting the entire punctured tissue and reducing tissue waste, leaving more tissue for the final diagnosis in paraffin after freezing.

[0061] The above embodiments are preferred implementation modes of the present invention, but the implementation modes of the present invention are not limited to the above embodiments. Any other changes, modifications, substitutions, combinations, and simplifications that do not deviate from the spirit and principles of the present invention should be considered as equivalent replacement methods and are included in the scope of protection of the present invention.

Claims

1. A method for embedding punctured tissue, characterized in that The following steps are involved: (1) Precool the freezing embedding apparatus to below -20°C and clean the embedding tank; (2) Place the punctured tissue on the platform, cover the surface of the punctured tissue with a small amount of OCT embedding gel, and turn the punctured tissue over until it is completely wrapped in the OCT embedding gel; then remove the punctured tissue wrapped in OCT embedding gel and place it on a platform without OCT embedding gel to reduce the amount of OCT embedding gel wrapped in the punctured tissue; (3) Spread the punctured tissue obtained in step (2) flat on the gasket paper, press it with tweezers to ensure that the punctured tissue and the gasket paper are completely in contact, and then cover the surface of the punctured tissue with a small amount of OCT embedding glue; the amount of OCT embedding glue used should just cover the punctured tissue; (4) Place the spacer paper with the punctured tissue obtained in step (3) in the embedding tank, and gently press the spacer paper around to make it fit the bottom of the embedding tank; Squeeze out OCT embedding gel to fill the embedding tank, place it in a pre-cooled freezing embedding instrument; cover with freezing tray, press the endothermic hammer, and start timing; (5) When the timer ends, remove the frozen tray and embedding is completed.

2. The embedding method of punctured tissue according to claim 1, characterized in that: The pre-cooling temperature in step (1) is -30°C to -40°C; The punctured tissue described in step (2) includes the punctured tissue obtained by coarse needle puncture and the punctured tissue obtained by fine needle puncture.

3. The embedding method of punctured tissue according to claim 2, characterized in that: The pre-cooling temperature in step (1) is -35°C.

4. The embedding method of punctured tissue according to claim 1, characterized in that: The platform described in step (2) is a positioning shovel or a photographing board.

5. The embedding method of punctured tissue according to claim 1, characterized in that: The gasket paper described in step (3) is a thin sheet of paper that is light-transparent and soft.

6. The embedding method of punctured tissue according to claim 1, characterized in that: The timing described in step (4) is 25 to 35 seconds.

7. The embedding method for punctured tissue according to claim 6, characterized in that: The timing described in step (4) is 30 seconds.

8. Use of the embedding method for punctured tissue according to any one of claims 1 to 7 in preparing frozen tissue sections.

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