A puncture sample fixing staining solution and its preparation method and application method

Abstract

The present invention discloses a biopsy specimen fixing and staining solution, and its preparation and application methods. The solution comprises the following components, measured by weight: 100-200 parts of acetone, 200-400 parts of glacial acetic acid, 200-400 parts of eosin Y, 50-200 parts of eosin stain, 50-200 parts of formalin, 1-10 parts of dimethyl sulfoxide, and 50-100 parts of zinc chloride. The biopsy specimen fixing and staining solution of the present invention has good staining effect and stable coloring.

Description

Technical Field

[0001] The invention relates to the field of biotechnology, in particular to a puncture sample fixing and staining solution, belonging to the technical field of specimen processing. Background Art

[0002] Pathology is a discipline based on the observation of cell morphology, providing an important basis for clinical diagnosis. With the development of medical technology, tissue puncture technology is becoming increasingly widely used to facilitate disease diagnosis and reduce patient suffering. However, this is accompanied by difficulties in fixation and staining of punctured tissue. Compared with conventional samples, puncture samples are smaller in size and are thin strips. If the punctured tissue is not stained or stained insufficiently during the pathology sampling process, after the fixation, dehydration, clearing, and wax immersion procedures, the tissue will appear white. During the subsequent embedding and sectioning process, the tissue and paraffin wax will be indistinguishable, resulting in tissue omission or loss during the sectioning process, making the work difficult to proceed. Therefore, the staining and labeling steps of the punctured tissue are particularly important.

[0003] The existing technology usually involves adding eosin dye to the tissue fixative, which can color the puncture sample to a certain extent and play a role in discovery and identification before making a tissue wax block. However, after the puncture sample stained with eosin is fixed, dehydrated, transparent, wax-impregnated, embedded, etc. to form a wax block, the eosin color on some puncture samples fades or becomes lighter. During the process of making pathological sections of the tissue in the wax block, it cannot play a role in discovery and identification of the puncture sample.

[0004] In summary, the present invention provides a rapid fixation staining solution for puncture samples, which has better coloring and clinical practicality compared with the existing technology. The application of the present invention is of great significance to improving the development of pathology and clinical pathology diagnosis and treatment in my country. Summary of the Invention

[0005] To overcome the shortcomings of the prior art, the present invention provides a rapid fixation and staining solution for puncture samples, as well as a preparation method and application method thereof, which are characterized by low cost, easy configuration, convenient use, and good coloring. The present invention discloses a rapid fixation and staining solution for puncture samples, which comprises the following components, in parts by weight:

[0006] 13. 100-200 parts of acetone,

[0007] 14. 200-400 parts of glacial acetic acid,

[0008] 15. Eosin Y 200-400 parts,

[0009] 16. 50-200 parts of eosin staining solution,

[0010] 17. 50-200 portions of formalin

[0011] 1-10 parts of dimethyl sulfoxide,

[0012] 50-100 parts of zinc chloride.

[0013] In some embodiments, a rapid fixation and staining solution for puncture samples comprises the following components, in parts by weight:

[0014] 100 parts of acetone,

[0015] 200 parts of glacial acetic acid,

[0016] 300 parts of Eosin Y,

[0017] 50 parts of eosin staining solution,

[0018] 100 parts of formalin,

[0019] 10 parts of dimethyl sulfoxide,

[0020] 100 parts of zinc chloride.

[0021] In some embodiments, the pH value of a rapid fixation and staining solution for puncture samples is less than 4.5.

[0022] The present invention also discloses a method for preparing a puncture sample fixing staining solution, comprising the following steps:

[0023] Step 1: Mix 200-400 parts of glacial acetic acid, 200-400 parts of eosin Y, 50-200 parts of eosin stain, and 50-100 parts of zinc chloride to obtain solution A;

[0024] Step 2: Thoroughly mix 100-200 parts of acetone, 50-200 parts of formalin, and 1-10 parts of dimethyl sulfoxide to obtain solution B;

[0025] Step 3: Ultrasonic mixing of solution A in step 1 and solution B in step 2 is performed to obtain a stationary solution.

[0026] Optionally, the ultrasonic power of the ultrasonic mixing in step 1 is 100 W, the ultrasonic time is 10 to 20 minutes, and the mixing temperature is room temperature.

[0027] The present invention also discloses a method for staining and fixing a puncture sample, comprising the following steps:

[0028] Step 1: Use tweezers to lay the punctured tissue parallel to the filter paper;

[0029] Step 2: Add the above-mentioned fixative staining solution dropwise to the punctured tissue and fix and stain for 5 to 30 seconds;

[0030] Step 3: Place the fixed and stained puncture tissue into the embedding frame;

[0031] Step 4: Dehydration, transparency, wax dipping and sectioning. BRIEF DESCRIPTION OF THE DRAWINGS

[0032] In order to more clearly illustrate the technical solutions of the embodiments of the present invention, the following briefly introduces the drawings required for use in the embodiments. It should be understood that the following drawings only illustrate certain embodiments of the present invention and therefore should not be regarded as limiting the scope. For ordinary technicians in this field, other relevant drawings can be obtained based on these drawings without paying any creative work.

[0033] Figure 1 HE staining of the paraffin sections prepared by fixation and staining in Example 4 shows uniform staining, no squeezing or cracks between cells, and a good preparation effect.

[0034] Figure 2 HE staining of the paraffin sections prepared by fixation and staining in Comparative Example 1 shows that the staining is uniform, there is no squeezing or cracks between cells, and the preparation effect is good. DETAILED DESCRIPTION

[0035] To make the purpose, technical solutions and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention are described clearly and completely below. Where specific conditions are not specified in the embodiments, conventional conditions or conditions recommended by the manufacturer are used. Where the manufacturer of the reagents or instruments is not specified, all are conventional products that can be purchased commercially.

[0036] The features and performance of the present invention are further described in detail below with reference to the embodiments.

[0037] Example 1 Preparation of puncture sample fixation staining solution:

[0038] 200 parts of glacial acetic acid, 300 parts of eosin Y, 100 parts of eosin dye solution and 50 parts of zinc chloride were ultrasonically mixed to obtain solution A, wherein the ultrasonic power was 100 W, the ultrasonic time was 10 to 20 minutes, and the mixing temperature was room temperature;

[0039] Mix 200 parts of acetone, 100 parts of formalin and 1 part of dimethyl sulfoxide thoroughly to obtain solution B;

[0040] Liquid A and liquid B were ultrasonically mixed to obtain a stationary solution, with an ultrasonic power of 100 W, an ultrasonic time of 10 to 20 min, a mixing temperature of room temperature, and a pH of 4.0.

[0041] Example 2

[0042] 400 parts of glacial acetic acid, 200 parts of eosin Y, 200 parts of eosin dye solution and 80 parts of zinc chloride were ultrasonically mixed to obtain solution A, wherein the ultrasonic power was 100 W, the ultrasonic time was 10 to 25 minutes, and the mixing temperature was room temperature;

[0043] Mix 100 parts of acetone, 100 parts of formalin and 10 parts of dimethyl sulfoxide thoroughly to obtain solution B;

[0044] Liquid A and liquid B were ultrasonically mixed to obtain a stationary solution, with an ultrasonic power of 100 W, an ultrasonic time of 10 to 20 min, a mixing temperature of room temperature, and a pH of 3.5.

[0045] Example 3

[0046] 150 parts of glacial acetic acid, 400 parts of eosin Y, 50 parts of eosin dye solution and 100 parts of zinc chloride were ultrasonically mixed to obtain solution A, wherein the ultrasonic power was 100 W, the ultrasonic time was 10 to 20 minutes, and the mixing temperature was room temperature;

[0047] Mix 150 parts of acetone, 100 parts of formalin and 8 parts of dimethyl sulfoxide thoroughly to obtain solution B;

[0048] Liquid A and liquid B were ultrasonically mixed to obtain a stationary solution, with an ultrasonic power of 100 W, an ultrasonic time of 10 to 20 min, a mixing temperature of room temperature, and a pH of 4.5.

[0049] Example 4

[0050] 200 parts of glacial acetic acid, 300 parts of eosin Y, 50 parts of eosin dye solution and 100 parts of zinc chloride were ultrasonically mixed to obtain solution A, wherein the ultrasonic power was 100 W, the ultrasonic time was 10 to 20 minutes, and the mixing temperature was room temperature;

[0051] Mix 100 parts of acetone, 100 parts of formalin and 10 parts of dimethyl sulfoxide thoroughly to obtain solution B;

[0052] Liquid A and liquid B were ultrasonically mixed to obtain a stationary solution, with an ultrasonic power of 100 W, an ultrasonic time of 10 to 20 min, a mixing temperature of room temperature, and a pH of 4.5.

[0053] Comparative Example 1 Alcohol-soluble eosin stain

[0054] Eosin reagent (No. BSBA-4022) was purchased from Beijin Zhongshan Jinqiao Company.

[0055] Comparative Example 2

[0056] 200 parts of glacial acetic acid, 300 parts of eosin Y, and 50 parts of eosin dye solution were ultrasonically mixed to obtain solution A, wherein the ultrasonic power was 100 W, the ultrasonic time was 10-20 min, and the mixing temperature was room temperature;

[0057] Mix 100 parts of acetone, 100 parts of formalin and 10 parts of dimethyl sulfoxide thoroughly to obtain solution B;

[0058] Liquid A and liquid B were ultrasonically mixed to obtain a stationary solution, with an ultrasonic power of 100 W, an ultrasonic time of 10 to 20 min, a mixing temperature of room temperature, and a pH of 4.0.

[0059] Comparative Example 3

[0060] 200 parts of glacial acetic acid, 300 parts of eosin Y, 50 parts of eosin dye solution and 100 parts of zinc chloride were ultrasonically mixed to obtain solution A, wherein the ultrasonic power was 100 W, the ultrasonic time was 10 to 20 minutes, and the mixing temperature was room temperature;

[0061] Mix 100 parts of acetone and 100 parts of formalin thoroughly to obtain solution B;

[0062] Liquid A and liquid B were ultrasonically mixed to obtain a stationary solution, with an ultrasonic power of 100 W, an ultrasonic time of 10 to 20 min, a mixing temperature of room temperature, and a pH of 4.5.

[0063] Comparative Example 4

[0064] 200 parts of glacial acetic acid, 300 parts of eosin Y, and 50 parts of eosin dye solution were ultrasonically mixed to obtain solution A, wherein the ultrasonic power was 100 W, the ultrasonic time was 10-20 min, and the mixing temperature was room temperature;

[0065] Mix 100 parts of acetone and 100 parts of formalin thoroughly to obtain solution B;

[0066] Liquid A and liquid B were ultrasonically mixed to obtain a stationary solution, with an ultrasonic power of 100 W, an ultrasonic time of 10 to 20 min, a mixing temperature of room temperature, and a pH of 4.5.

[0067] Experimental Example 1

[0068] The tonsil puncture sample tissue was stained using the fixing staining solution prepared in Examples 1 to 4 and Comparative Examples 1 to 4 above. The specific operation was as follows:

[0069] Step 1: Use tweezers to lay the punctured tissue parallel to the filter paper;

[0070] Step 2: Add the fixative staining solution to the punctured tissue and fix and stain for 5 to 30 seconds;

[0071] Step 3: Place the fixed and stained puncture tissue into the embedding frame;

[0072] Step 4: Dehydration, transparency, wax dipping and sectioning.

[0073] Here are the results:

[0074] Staining quality Example 1 The staining color is deep, and the embedded tissue is easier to distinguish from paraffin Example 2 The staining color is deep, and the embedded tissue is easier to distinguish from paraffin Example 3 The staining color is deep, and the embedded tissue is easier to distinguish from paraffin Example 4 The staining color is deep, and the embedded tissue contrasts sharply with the paraffin Comparative Example 1 The staining is light, and the embedded tissue needs to be carefully distinguished from the paraffin Comparative Example 2 The staining is light, and it is difficult to distinguish between embedded tissue and paraffin Comparative Example 3 The staining is light, and it is difficult to distinguish between embedded tissue and paraffin Comparative Example 4 Light staining, difficult to distinguish between embedded tissue and paraffin

[0075] It can be seen from the test results in the above table that the puncture sample fixation staining solution prepared by the present invention has a good staining effect and can improve the efficiency of clinical sample preparation.

[0076] Experimental Example 2

[0077] Paraffin sections prepared in Example 4 and Comparative Example 1 were subjected to conventional HE staining, and the tissue sections prepared by the two staining and fixation methods were observed under a microscope. The results showed that the tissue structures of the sections prepared by the two methods were clear and the color contrast was sharp. Figure 1 This is the HE staining picture of the slice prepared in Example 4. Figure 2 This is the HE staining picture of the slice prepared in Comparative Example 1.

[0078] The foregoing description is merely a preferred embodiment of the present invention and is not intended to limit the present invention. Those skilled in the art will readily appreciate that various modifications and variations of the present invention are possible. Any modifications, equivalent substitutions, or improvements made within the spirit and principles of the present invention are intended to be within the scope of protection of the present invention.

Claims

1. A puncture sample fixing and staining solution, characterized in that: In parts by weight, it comprises the following ingredients: 100 parts of acetone, 200 parts of glacial acetic acid, 300 parts of Eosin Y, 50 parts of eosin staining solution, 100 parts of formalin, 10 parts of dimethyl sulfoxide, 100 parts of zinc chloride, The pH value of the fixing and staining solution is less than 4.

5.

2. A method for preparing a fixed dye solution as claimed in claim 1, characterized in that: The following steps are involved: Step 1: Mix 200 parts of glacial acetic acid, 300 parts of eosin Y, 50 parts of eosin stain, and 100 parts of zinc chloride to obtain solution A; Step 2: Thoroughly mix 100 parts of acetone, 100 parts of formalin, and 10 parts of dimethyl sulfoxide to obtain solution B; Step 3: Ultrasonic mixing of solution A in step 1 and solution B in step 2 is performed to obtain a stationary solution.

3. The method for preparing a fixed dye solution according to claim 2, wherein In step 1, the ultrasonic power of the ultrasonic mixing is 200 W, the ultrasonic time is 10 to 20 minutes, and the mixing is carried out at room temperature.

4. A method for staining and fixing a puncture sample, using the fixing and staining solution according to claim 1, characterized in that: The following steps are involved: Step 1: Use tweezers to lay the punctured tissue parallel to the filter paper; Step 2: Add the fixative staining solution to the punctured tissue and fix and stain for 5-30 seconds; Step 3: Place the fixed and stained puncture tissue into the embedding frame; Step 4: Dehydration, transparency, wax dipping and sectioning.

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