CN117782751A - Machine dyeing method for immunohistochemical multi-tissue different indexes - Google Patents
Machine dyeing method for immunohistochemical multi-tissue different indexes Download PDFInfo
- Publication number
- CN117782751A CN117782751A CN202311836878.4A CN202311836878A CN117782751A CN 117782751 A CN117782751 A CN 117782751A CN 202311836878 A CN202311836878 A CN 202311836878A CN 117782751 A CN117782751 A CN 117782751A
- Authority
- CN
- China
- Prior art keywords
- tissue
- glass slide
- slices
- cover film
- staining
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
- 238000000034 method Methods 0.000 title claims abstract description 35
- 238000004043 dyeing Methods 0.000 title claims abstract description 20
- 230000002055 immunohistochemical effect Effects 0.000 title claims abstract description 14
- 239000011521 glass Substances 0.000 claims abstract description 65
- 239000007788 liquid Substances 0.000 claims abstract description 27
- 230000000903 blocking effect Effects 0.000 claims abstract description 22
- 239000013039 cover film Substances 0.000 claims abstract description 19
- 239000007787 solid Substances 0.000 claims abstract description 19
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 18
- 238000011534 incubation Methods 0.000 claims abstract description 8
- 238000010186 staining Methods 0.000 claims description 14
- WZUVPPKBWHMQCE-UHFFFAOYSA-N Haematoxylin Chemical compound C12=CC(O)=C(O)C=C2CC2(O)C1C1=CC=C(O)C(O)=C1OC2 WZUVPPKBWHMQCE-UHFFFAOYSA-N 0.000 claims description 6
- 238000002791 soaking Methods 0.000 claims description 6
- 239000012747 synergistic agent Substances 0.000 claims description 3
- 238000005406 washing Methods 0.000 claims description 3
- 230000008439 repair process Effects 0.000 claims description 2
- 239000003153 chemical reaction reagent Substances 0.000 description 8
- 230000008569 process Effects 0.000 description 6
- 238000003364 immunohistochemistry Methods 0.000 description 5
- 230000002209 hydrophobic effect Effects 0.000 description 3
- 238000011532 immunohistochemical staining Methods 0.000 description 3
- 230000007170 pathology Effects 0.000 description 3
- MHAJPDPJQMAIIY-UHFFFAOYSA-N Hydrogen peroxide Chemical compound OO MHAJPDPJQMAIIY-UHFFFAOYSA-N 0.000 description 2
- 230000007547 defect Effects 0.000 description 2
- 230000000694 effects Effects 0.000 description 2
- 238000005516 engineering process Methods 0.000 description 2
- 230000007246 mechanism Effects 0.000 description 2
- 108010008886 CAM 5.2 antigen Proteins 0.000 description 1
- 239000004215 Carbon black (E152) Substances 0.000 description 1
- 206010028980 Neoplasm Diseases 0.000 description 1
- 108700020962 Peroxidase Proteins 0.000 description 1
- 102000003992 Peroxidases Human genes 0.000 description 1
- 238000009825 accumulation Methods 0.000 description 1
- 239000000427 antigen Substances 0.000 description 1
- 102000036639 antigens Human genes 0.000 description 1
- 108091007433 antigens Proteins 0.000 description 1
- 239000012472 biological sample Substances 0.000 description 1
- 210000005013 brain tissue Anatomy 0.000 description 1
- 201000011510 cancer Diseases 0.000 description 1
- 238000003759 clinical diagnosis Methods 0.000 description 1
- 239000002131 composite material Substances 0.000 description 1
- 238000012864 cross contamination Methods 0.000 description 1
- 238000001514 detection method Methods 0.000 description 1
- 238000012137 double-staining Methods 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 230000005484 gravity Effects 0.000 description 1
- 230000000887 hydrating effect Effects 0.000 description 1
- 230000036571 hydration Effects 0.000 description 1
- 238000006703 hydration reaction Methods 0.000 description 1
- 229930195733 hydrocarbon Natural products 0.000 description 1
- 150000002430 hydrocarbons Chemical class 0.000 description 1
- 238000010166 immunofluorescence Methods 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 239000003960 organic solvent Substances 0.000 description 1
- 230000001575 pathological effect Effects 0.000 description 1
- 238000007789 sealing Methods 0.000 description 1
- 238000007447 staining method Methods 0.000 description 1
Landscapes
- Sampling And Sample Adjustment (AREA)
- Investigating Or Analysing Biological Materials (AREA)
Abstract
The invention discloses an immunohistochemical multi-tissue different index mechanical dyeing method, which comprises the following steps: step 1, a piece is fished, and a glass slide drags a plurality of tissue slices; step 2, baking slices; step 3, dewaxing, blocking and repairing a plurality of tissue slices of a glass slide; step 4, carrying out circle setting and water blocking on each tissue slice on the glass slide; step 5, adding corresponding primary antibody indexes to each tissue slice according to the corresponding indexes of dyeing of different tissue slices, and performing primary antibody incubation; step 6, combining the glass slide with a solid liquid cover film, and placing the glass slide and the solid liquid cover film into a specimen incubation bin; and 7, performing water removal and blocking rings on a plurality of tissues of a glass slide. The method can realize the advantages that a plurality of (two or more) tissue slices are fished on one glass slide, and different tissue slices are respectively dyed with different indexes.
Description
Technical Field
The invention relates to the field of pathology detection, in particular to a machine-dyeing method for immunohistochemical multi-tissue different indexes.
Background
Immunohistochemistry (immunohistochemistry) is widely used in clinical and diagnostic applications, such as clinical diagnosis of cancer. In immunohistochemistry, tissue sections (tissues) are generally required to be dyed (machine-dyed) on a full-automatic immunohistochemical dyeing machine, and the following modes generally exist in the prior dyeing: 1. single-staining, namely fishing out a tissue slice from a glass slide and taking the tissue slice as an index; 2. double-staining, namely fishing out a tissue slice by a glass slide, and making two indexes according to the sequence; 3. immunofluorescence multicolor immunohistochemistry, a tissue section is fished by a glass slide, and a plurality of indexes (such as 2-7 indexes) are continuously carried out; 4. the photo is positively dyed, a glass slide drags out a plurality of dot positive tissues, and the same index is dyed. The dyeing method cannot be realized: a plurality of (two or more) tissue slices are fished on a glass slide, and different tissue slices are respectively dyed with different indexes.
The technical scheme adopted in the prior art for realizing different indexes of dyeing different tissue slices is that different tissue slices are fished on different glass slides, and then indexes are respectively dyed, for example CN103975230A published by 2014, 8, 6, and the invention application of a method, equipment and a system for dyeing biological samples. According to the technical scheme, although the effect that different tissue slices are respectively dyed with different indexes can be achieved, the different tissue slices are distributed on different glass slides, so that the following defects exist: 1. one tissue and one glass slide are respectively and independently added with general reagents, and the general reagents have large dosage and high cost; 2. tissue slices of the same case are distributed on different glass slides, the reading is complicated, and because the number of the slices is more, a plurality of slices of the same case are dyed on a plurality of platforms, the slice outlet time is different, so that the doctor has low efficiency of reading the slices; 3. after pathological staining of the tissue sections, the tissue sections are permanently stored, and after accumulation, one tissue is one slide glass, and a great amount of space is required for storing the existing tissue sections in a pathology department.
Disclosure of Invention
The invention provides an immunohistochemical multi-tissue different index mechanical dyeing method, which overcomes the defects in the background technology.
The technical scheme adopted for solving the technical problems is as follows: an immunohistochemical multi-tissue different index mechanical staining method, comprising:
step 1, a piece is fished, and a glass slide drags a plurality of tissue slices;
step 2, baking slices;
step 3, dewaxing, blocking and repairing a plurality of tissue slices of a glass slide;
step 4, carrying out circle setting and water blocking on each tissue slice on the glass slide;
step 5, adding corresponding primary antibody indexes to each tissue slice according to the corresponding indexes of dyeing of different tissue slices, and performing primary antibody incubation;
step 6, combining the glass slide with a solid liquid cover film, and placing the glass slide and the solid liquid cover film into a specimen incubation bin;
and 7, performing water removal and blocking rings on a plurality of tissues of a glass slide.
In one embodiment: in step 5, each tissue is circled on the slide with a water blocking pen.
In one embodiment: in the step 3, the roasted tissue slices are combined with a solid liquid cover film to dewax, block and repair a plurality of tissue slices of a glass slide; in step 4, the slide glass and the solid liquid cover film are detached, and each tissue slice is looped and blocked on the slide glass.
In one embodiment: and 7, performing water-blocking ring removal on a plurality of tissues of a glass slide, and washing cleanly.
In one embodiment: further comprises:
and 8, continuing the subsequent immunohistochemical procedure, wherein the subsequent immunohistochemical procedure comprises the steps of adding secondary antibody, synergistic agent, DAB color development liquid, hematoxylin counterstain, hydration and the like.
In one embodiment: the glass slide is combined with the solid liquid cover film, a capillary gap space is formed between the glass slide and the solid liquid cover film, the tissue slice is positioned in the capillary gap space, and the capillary gap space is obliquely arranged relative to the vertical direction.
In one embodiment: the repairing of the step 3 adopts soaking type water bath thermal repairing, and a plurality of tissue slices on the glass slide are completely soaked in repairing liquid for repairing by a soaking method.
Compared with the background technology, the technical proposal has the following advantages:
can realize the advantages that a plurality of (two or more) tissue slices are fished out on a glass slide, and different tissue slices are respectively dyed with different indexes, wherein: the processes and experimental conditions of the steps 2, 3 and 7 are the same, and other reagents can be saved greatly by sharing the process, so that the dyeing cost is greatly reduced; in the step 4, the circle setting and water blocking are carried out to avoid the influence of cross contamination of corresponding indexes (individual indexes) of different tissue slices; the tissue slices of the same case are more concentrated, the slice outlet efficiency of the tissue slices of the same case is higher, the slice reading is more convenient, and the slice reading efficiency of a doctor of a pathology department is improved; a plurality of tissues are placed on one glass slide, so that the storage space of tissue slices can be effectively saved. And each tissue is circled and blocked on the glass slide by using a non-oily water blocking pen, so that the cost is low and the blocking effect is good. In addition, the process in the step 8 and experimental conditions are the same, and other reagents, especially high-cost reagents such as secondary antibody, synergy and DAB, can be greatly saved by the common process, so that the dyeing cost is greatly reduced.
Drawings
FIG. 1 is a photograph of a plurality of tissue sections of a slide prior to staining the corresponding indices, respectively.
FIG. 2 is a photograph of a plurality of tissue sections of a slide after staining the corresponding indicators, respectively.
Detailed Description
A method of mechanochromatic staining of multiple tissues for immunohistochemistry, comprising:
step 1, taking out slices, namely taking out a plurality of tissue slices by using a glass slide, wherein each tissue slice can be a tissue slice of the same case or a tissue slice of different cases; the plurality of tissue slices, such as two tissue slices, and the specific number of scoops (the number of tissue slices) is arranged according to the size of the tissue slices; wherein the step 1 is generally preceded by conventional 'material drawing and embedding, and slicing by a slicer after embedding into wax blocks';
step 2, baking a plurality of tissue slices of a glass slide, wherein the tissue slices can be placed on a special baking machine for baking, or baked on a full-automatic immunohistochemical dyeing machine of HG1 series of various types, which is produced by Beijing Haigelai biotechnology Co., ltd; wherein: the baking sheet aims to adhere the fished sheet to a glass slide.
Step 3, the baked tissue slices are combined with a solid liquid cover film, and a plurality of tissue slices of a glass slide are subjected to dewaxing, blocking and repairing by using an HG1 series full-automatic immunohistochemical staining machine of various types manufactured by Beijing Haigelai biotechnology Co., ltd; the technical process and experimental conditions of dewaxing, blocking and repairing of each tissue slice are the same, and the common working procedure is performed, so that the equipment cost can be reduced, the dosage can be reduced and the efficiency can be improved compared with the same mechanism for performing single tissue slice according to single glass slide; wherein: dewaxing is to dewax the tissue section with dewaxing liquid to eliminate wax so that subsequent antigen and antibody can combine well; the blocking was aimed at adding hydrogen peroxide to the tissue sections to reduce non-specific background staining by endogenous peroxidases.
Step 4, after repairing, taking out the glass slide, separating the glass slide from the solid liquid cover film, and carrying out circle setting water blocking on each tissue slice on the glass slide, wherein the concrete steps are as follows: each tissue is circled and blocked on the glass slide by a hydrophobic water blocking pen, and the hydrophobic water blocking pen can be quickly dissolved in an organic solvent after being completely drawn, and the hydrophobic water blocking pen is made of hydrocarbon-containing composite materials;
step 5, according to the corresponding indexes of dyeing different tissue slices, respectively dripping corresponding primary antibodies to each tissue slice, wherein the method has the advantages that an appropriate amount of primary antibodies can be respectively dripped according to the tissue size (compared with the method of uniformly adding primary antibodies when dyeing according to the same mechanism of a single tissue slice of a single glass slide, the primary antibody adding amount of a small and medium tissue slice can be saved, the cost of the primary antibodies is reduced), and primary antibody incubation at corresponding temperature and corresponding time is performed; referring to FIGS. 1 and 2, two sections of brain tissue were stained with CAM5.2 and CK18, respectively, in the same case, FIG. 1 before staining and FIG. 2 after staining;
step 6, the glass slide is recombined with the solid liquid cover film and put into a specimen incubation bin of an HG1 series model full-automatic immunohistochemical staining machine manufactured by Beijing Haigelai biotechnology Co., ltd for subsequent on-machine experiment;
step 7, performing water blocking ring removal on a plurality of tissue slices of a glass slide after loading, and washing cleanly; wherein, the technical mode and the technological condition of each tissue slice water-blocking ring are the same, and the process is shared;
and 8, continuously running an immunohistochemical follow-up procedure on an HG1 series model full-automatic immunohistochemical staining machine produced by Beijing Haigelai biotechnology limited company, wherein the subsequent procedure comprises the steps of adding secondary antibody, synergistic agent, DAB color development liquid, hematoxylin counterstaining, hydrating and the like, and taking out a glass slide for sealing after finishing the procedure.
The glass slide is combined with the solid liquid cover film, a capillary gap space is formed between the glass slide and the solid liquid cover film, the tissue slices are located in the capillary gap space, the capillary gap space is obliquely arranged relative to the vertical direction, firstly, tissue protection is facilitated, secondly, the reagent naturally flows down through gravity and forms liquid surface tension through the capillary gap, so that the reagent is paved on the tissue slices on the glass slide, and the condition that one glass slide is fully covered with different tissue slices on one glass slide after being added with a general reagent can be met.
The repairing of the step 3 adopts soaking type water bath thermal repairing, and a plurality of tissues on the glass slide can be completely soaked in repairing liquid for complete and full repairing by a soaking method, so that a plurality of tissue slices can be uniformly and fully repaired at the same time, and the effect of high-quality dyeing can be equally realized.
The foregoing description is only illustrative of the preferred embodiments of the present invention, and therefore should not be taken as limiting the scope of the invention, for all changes and modifications that come within the meaning and range of equivalency of the claims and specification are therefore intended to be embraced therein.
Claims (7)
1. The machine dyeing method for immunohistochemical multi-tissue different indexes is characterized by comprising the following steps of: comprising the following steps:
step 1, a piece is fished, and a glass slide drags a plurality of tissue slices;
step 2, baking slices;
step 3, dewaxing, blocking and repairing a plurality of tissue slices of a glass slide;
step 4, carrying out circle setting and water blocking on each tissue slice on the glass slide;
step 5, adding corresponding primary antibody indexes to each tissue slice according to the corresponding indexes of dyeing of different tissue slices, and performing primary antibody incubation;
step 6, combining the glass slide with a solid liquid cover film, and placing the glass slide and the solid liquid cover film into a specimen incubation bin;
and 7, performing water removal and blocking rings on a plurality of tissues of a glass slide.
2. The method for mechanochromatic staining of multiple tissue different indices according to claim 1, wherein: in step 5, each tissue is circled on the slide with a water blocking pen.
3. The method for mechanochromatic staining of multiple tissue different indices according to claim 1, wherein: in the step 3, the roasted tissue slices are combined with a solid liquid cover film to dewax, block and repair a plurality of tissue slices of a glass slide; in step 4, the slide glass and the solid liquid cover film are detached, and each tissue slice is looped and blocked on the slide glass.
4. The method for mechanochromatic staining of multiple tissue different indices according to claim 1, wherein: further comprises:
and 8, continuing the subsequent immunohistochemical procedure, wherein the subsequent immunohistochemical procedure comprises the addition of secondary antibody, synergistic agent, DAB color development liquid and hematoxylin.
5. The method for mechanochromatic staining of multiple tissue different indices according to claim 1, wherein: and 7, performing water-blocking ring removal on a plurality of tissues of a glass slide, and washing cleanly.
6. The method for mechanochromatic staining of multiple tissue different indices according to claim 1, wherein: the glass slide is combined with the solid liquid cover film, a capillary gap space is formed between the glass slide and the solid liquid cover film, the tissue slice is positioned in the capillary gap space, and the capillary gap space is obliquely arranged relative to the vertical direction.
7. The method for mechanochromatic staining of multiple tissue different indices according to claim 1, wherein: the repairing of the step 3 adopts soaking type water bath thermal repairing, and a plurality of tissue slices on the glass slide are completely soaked in repairing liquid for repairing by a soaking method.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202311836878.4A CN117782751A (en) | 2023-12-28 | 2023-12-28 | Machine dyeing method for immunohistochemical multi-tissue different indexes |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202311836878.4A CN117782751A (en) | 2023-12-28 | 2023-12-28 | Machine dyeing method for immunohistochemical multi-tissue different indexes |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN117782751A true CN117782751A (en) | 2024-03-29 |
Family
ID=90398015
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202311836878.4A Pending CN117782751A (en) | 2023-12-28 | 2023-12-28 | Machine dyeing method for immunohistochemical multi-tissue different indexes |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN117782751A (en) |
-
2023
- 2023-12-28 CN CN202311836878.4A patent/CN117782751A/en active Pending
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN1195585C (en) | Apparatus and methods for efficient processing of biological samples on slides | |
| CN103930761B (en) | Unmixing binary system for pre-processing the biological specimen of embedding | |
| Hsieh et al. | Papillary-cystic pattern is characteristic in mammary analogue secretory carcinomas but is rarely observed in acinic cell carcinomas of the salivary gland | |
| CN108918518B (en) | Method for observing same cell morphology by common optical, fluorescence and scanning electron microscope | |
| CN105784991A (en) | Device and method for repeated immunostaining of same tissue section | |
| Zhang et al. | Application of immunohistochemistry technique in hydrobiological studies | |
| Schiavon et al. | Evaluation of reliability of FISH versus brightfield dual-probe in situ hybridization (BDISH) for frontline assessment of HER2 status in breast cancer samples in a community setting: influence of poor tissue preservation | |
| Lim et al. | Fluorescence in situ hybridization on tissue sections | |
| Rustad | An interference microscopical and cytochemical analysis of local mass changes in the mitotic apparatus during mitosis | |
| US20050026297A1 (en) | Kits and methods for preparing gell samples optmimized for dual staining | |
| CN117782751A (en) | Machine dyeing method for immunohistochemical multi-tissue different indexes | |
| Exbrayat | Classical methods of visualization | |
| US20170284909A1 (en) | Method for preparing liquid-state dripping or coating pathological quality control product and uses thereof | |
| Sathawane et al. | Nuances of the Papanicolaou stain | |
| CN110050070A (en) | Full-automatic method for extracting nucleic acid for tissue sample | |
| Komatsu et al. | Application of liquid-based preparation to fine needle aspiration cytology in breast cancer | |
| CN106092707A (en) | A kind of apparatus and method for same tissue slice being carried out repeatedly immunostaining | |
| CN114076707A (en) | Paraffin embedded tissue sample processing method and kit for tumor prognosis evaluation | |
| CN106048047A (en) | Method for identifying marine nematodes by denatured gradient gel electrophoresis | |
| CN112798322A (en) | 3D cell microsphere paraffin section method | |
| Kalantari-Hesari et al. | Modified methods to simplification histochemical, immunohistochemical, and hematoxylin-eosin staining | |
| CN111307556A (en) | Method for performing immunofluorescence staining after HE section fades | |
| CN107941587A (en) | A kind of method that improved Golgi Cox dyeing prepares brain tissue paraffin section | |
| Gabriel et al. | Evaluation of a New Mordant Based Haematoxylin Dye (Haematoxylin X) for Use in Clinical Pathology | |
| WO2023195221A1 (en) | Flat plate for microscopic specimen preparation use, and method for preparing microscopic specimen |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination |