CN112326961A - Analysis method and storage device for proportion of PD-L1 positive tumor cells in non-small cell lung cancer - Google Patents
Analysis method and storage device for proportion of PD-L1 positive tumor cells in non-small cell lung cancer Download PDFInfo
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Abstract
The invention relates to the field of computers, in particular to an analysis method and storage equipment for the proportion of PD-L1 positive tumor cells in non-small cell lung cancer. The method for analyzing the proportion of PD-L1 positive tumor cells in the non-small cell lung cancer comprises the following steps: obtaining a section to be dyed; performing immunohistochemical staining operation of tumor cell markers; performing immunohistochemical staining operation of a specific marker; acquiring a first region of interest, acquiring characteristic points, and calculating the total number of tumor cells according to the number of the characteristic points; according to the second region of interest, obtaining characteristic points, and calculating the number of tumor cells stained by the specific marker according to the characteristic points; and calculating the fraction of the positive proportion of the tumor cells according to the total number of the tumor cells and the number of the tumor cells stained by the specific marker. The whole process does not need a pathologist to carry out manual interpretation, so that the obtained result is more accurate and objective, and the efficiency is higher.
Description
Technical Field
The invention relates to the field of computers, in particular to an analysis method and storage equipment for the proportion of PD-L1 positive tumor cells in non-small cell lung cancer.
Background
The pathological doctor has long training period, low income and poor working environment, and more importantly, needs the combination of a large amount of knowledge, not only needs clinical knowledge, but also needs the training of pathological specialties. The classification of pathology has higher resolution under a microscope, the classification of various diseases has tens of thousands, the difference between classification and well-known classification needs to be remembered, the differentiation and diagnosis of many similar diseases need to be distinguished at the cellular level, a large amount of technical data and a large amount of special techniques need to be remembered, and more importantly, a pathologist needs to rely on the intelligence of a pathologist to convert a two-dimensional image into a disease diagnosis and convert the two-dimensional image into the interpretation of the occurrence and development processes of the diseases, and the process is not easy to implement, so that the pathologist is difficult to culture.
With the rapid development of precise medicine, the use of targeted drugs depends on the diagnosis of pathologists who gradually move from behind to the front end of the medical procedure. However, the requirements of precise medicine for pathological diagnosis are higher than ever, and qualitative, semi-quantitative and quantitative requirements in the past pose higher challenges for pathologists.
The immunotherapy of tumor is the most popular research topic in the current precise medical field, and the immunotherapy of tumor has entered a completely new era in recent 5 years, namely the current popular tumor immunotherapy technology, namely monoclonal antibody immune checkpoint inhibition. Among them, the immune checkpoint inhibition technique of the programmed death receptor (PD-1) and its ligand (PD-L1) is a novel and popular research direction for current tumor immunotherapy. PD-1/PD-L1 inhibitors have now been approved in the FDA and china for a number of indications including malignant melanoma, metastatic squamous cell carcinoma of the head and neck, non-small cell lung cancer, renal cell carcinoma, classical hodgkin lymphoma, urothelial cancer, colorectal cancer, liver cancer, and gastric cancer, among others.
The subjects (tumor cells/immune cells) and criteria (Cut off values) were read differently for the different PD-L1 detection systems. This makes PD-L1 likely to face the following problems during use: 1) tissue samples are limited, and different drugs need to be detected by using different detection kits; 2) the detection platform is various, and different automatic instruments are required to be equipped in departments, and different dyeing procedures are mastered; 3) the cost is high; 4) the pathological diagnosticians need to carry out various PD-L1 interpretation training, and the interpretation difficulty and the daily workload are increased.
In the past, a pathologist usually needs to judge an H & E stained section, normal tissues, necrotic tissues and the like are eliminated, the tumor cell position is found out finally, the tumor cell is counted (the number of cells is large and only estimation is carried out), then PD-L1 positive cells are found out at the tumor cell position in another PD-L1 stained section, counting is carried out, and the result of TPS (the ratio of the number of PD-L1 positive tumor cells to the total number of tumor cells) is obtained finally, wherein the counting result is usually influenced by the qualification and subjective factors of the pathologist, and the result consistency is poor.
Disclosure of Invention
Therefore, an analysis method for the proportion of PD-L1 positive tumor cells in the non-small cell lung cancer needs to be provided, so that the problems that the existing calculation method needs a pathologist to calculate artificially, and is low in efficiency, poor in accuracy and high in threshold are solved. The specific technical scheme is as follows:
a method for analyzing the proportion of PD-L1 positive tumor cells in non-small cell lung cancer comprises the following steps:
obtaining a section to be dyed;
performing immunohistochemical staining operation of tumor cell markers on part of the section to be stained;
carrying out specific marker immunohistochemical staining operation on part of the section to be stained;
acquiring a first region of interest of a stained section after immunohistochemical staining of a tumor cell marker, acquiring feature points in the first region of interest, and calculating the total number of tumor cells according to the number of the feature points in the first region of interest;
acquiring a second region of interest of the stained section of the specific marker after immunohistochemical staining according to the first region of interest position, acquiring characteristic points in the second region of interest, and calculating the number of tumor cells stained by the specific marker according to the characteristic points in the second region of interest;
and calculating the fraction of the positive proportion of the tumor cells according to the total number of the tumor cells and the number of the tumor cells stained by the specific marker.
Further, the step of performing immunohistochemical staining operation of tumor cell markers on a part of the section to be stained further comprises the following steps:
spin-drying the section to be dyed, dripping cocktail mixed primary antibody diluted in a proper proportion, incubating for 1 hour at room temperature (25 ℃), flushing PBS for 3 multiplied by 3 minutes, dripping secondary antibody, incubating for 15 to 30 minutes at room temperature, flushing PBS for 3 multiplied by 3 minutes, throwing away PBS, developing for 3 to 10 minutes by using DAB developing solution which is freshly prepared, counterstaining hematoxylin for 25 seconds, enabling PBS to return blue for 30 seconds, dehydrating according to an alcohol gradient of 85% (3 minutes) -95% (3 minutes) -100% (3 minutes), finally enabling xylene to be transparent for 3 minutes, sealing with neutral gum, positioning the antibody on a tumor cell membrane, staining in a brown yellow color, and staining the cell nucleus in a blue color after counterstaining with hematoxylin.
Further, the step of obtaining the first region of interest of the stained section after the tumor cell marker immunohistochemical staining further comprises the steps of:
extracting a first region of interest by taking preset colors as features of interest, wherein the preset colors include but are not limited to: brown-yellow;
the step of acquiring the feature points in the first region of interest further comprises the following steps:
and taking a blue point in the first region of interest as a characteristic point.
Further, the step of performing specific marker immunohistochemical staining operation on a part of the section to be stained further comprises the steps of:
spin-drying the section, adding diluted primary antibody (PD-L1) in proper proportion, incubating for 1 hour at room temperature (25 ℃), washing for 3X 3 minutes by PBS, adding secondary antibody, incubating for 15-30 minutes at room temperature, washing for 3X 3 minutes by PBS, throwing away PBS, developing for 3-10 minutes by DAB developing solution which is prepared freshly, counterstaining for 25 seconds by hematoxylin, rewetting for 30 seconds by PBS, dehydrating according to alcohol gradient of 85% (3 minutes) -95% (3 minutes) -100% (3 minutes), finally, clearing for 3 minutes by xylene, sealing with neutral gum, positioning PD-L1 antibody on cell membranes of tumor cells and immune cells, staining in brown yellow, and staining cell nuclei in blue after counterstaining by hematoxylin.
Further, the "acquiring the feature points in the second region of interest" further includes:
and taking the brown color point and the blue color point in the second region of interest as characteristic points.
Further, the "acquiring a section to be stained" further includes the steps of:
the method comprises the following steps of trimming a plurality of cancer tissue wax blocks for a preset time, continuously slicing the cancer tissue wax blocks to a thickness of 3 mu m, floating the continuous slices in cold water, naturally unfolding the slices, transferring the separated slices into warm water at 45 ℃ for spreading for 30 seconds, pasting the slices by polylysine-treated glass slides according to the slicing sequence, placing the prepared tissue chips into a 65 ℃ oven for baking for 2 hours, taking out the tissue chips, cooling the tissue chips at room temperature, and placing the tissue chips into a-4 ℃ refrigerator for storage.
Further, the "calculating the fraction of positive proportion of tumor cells according to the total number of tumor cells and the number of tumor cells stained by the specific marker" further comprises the steps of:
tumor cell positive proportion score ═(number of tumor cells stained by any intensity PD-L1 immunohistochemical marker)/total number of tumor cells) × 100%.
Further, the "acquiring a second region of interest of the stained section after immunohistochemical staining of the specific marker according to the first region of interest position" further includes the steps of:
the first region of interest is located at the same position on the slice as the second region of interest.
Further, after the step of performing immunohistochemical staining operation of tumor cell markers on the part of the section to be stained, the method further comprises the following operation: scanning the section subjected to immunohistochemical staining of the tumor cell marker to obtain a first picture;
after the specific marker immunohistochemical staining operation is carried out on part of the section to be stained, the method further comprises the following steps: the immunohistochemically stained section of the specific marker was scanned to obtain a second picture.
In order to solve the technical problem, the storage device is further provided, and the specific technical scheme is as follows:
a storage device having stored therein a set of instructions for performing:
acquiring a first picture obtained by scanning a section subjected to tumor cell marker immunohistochemical staining, acquiring a first region of interest of the stained section subjected to tumor cell marker immunohistochemical staining on the first picture, acquiring feature points in the first region of interest, and calculating the total number of tumor cells according to the number of the feature points in the first region of interest;
acquiring a second picture obtained by scanning the immunohistochemically stained section of the specific marker, acquiring a second region of interest of the immunohistochemically stained section of the specific marker on the second picture according to the position of the first region of interest, acquiring characteristic points in the second region of interest, and calculating the number of tumor cells stained by the specific marker according to the characteristic points in the second region of interest;
and calculating the fraction of the positive proportion of the tumor cells according to the total number of the tumor cells and the number of the tumor cells stained by the specific marker.
The invention has the beneficial effects that: by obtaining a section to be stained; performing immunohistochemical staining operation of tumor cell markers on part of the section to be stained; carrying out specific marker immunohistochemical staining operation on part of the section to be stained; acquiring a first region of interest of a stained section after immunohistochemical staining of a tumor cell marker, acquiring feature points in the first region of interest, and calculating the total number of tumor cells according to the number of the feature points in the first region of interest; acquiring a second region of interest of the stained section of the specific marker after immunohistochemical staining according to the first region of interest position, acquiring characteristic points in the second region of interest, and calculating the number of tumor cells stained by the specific marker according to the characteristic points in the second region of interest; calculating the fraction of positive proportion of tumor cells according to the total number of the tumor cells and the number of the tumor cells stained by the specific immunohistochemical marker. In the whole process, the total number of the tumor cells and the number of the tumor cells stained by the specific marker are calculated by a program without manual interpretation by a pathologist, so that the obtained result is more accurate and objective, and the efficiency is higher.
In addition, the existing pathological AI image analysis methods need to label a large number of pathological features of a large number of tissue samples by depending on pathologists, and adopt machine learning to obtain the features, but due to the heterogeneity of tumor tissues, the learning and analysis of the pathological features of the tissues by the machine are difficult and time-consuming. The tumor cell marker is subjected to immunohistochemical staining, and the tumor cell region can be labeled by using a single color characteristic, so that the processes of machine learning pathological characteristics and image analysis are greatly simplified.
Drawings
FIG. 1 is a flowchart of a method for analyzing the proportion of PD-L1-positive tumor cells in non-small cell lung cancer according to an embodiment;
fig. 2 is a schematic block diagram of a storage device according to an embodiment.
Description of reference numerals:
200. a storage device.
Detailed Description
To explain technical contents, structural features, and objects and effects of the technical solutions in detail, the following detailed description is given with reference to the accompanying drawings in conjunction with the embodiments.
Referring to fig. 1, in this embodiment, a specific embodiment of the method for analyzing the proportion of PD-L1 positive tumor cells in non-small cell lung cancer is as follows:
first, the following explanation will be made for terms that will appear in the present embodiment:
TPS: tumor cell positive Proportion Score, Tumor cell positive Proportion Score.
In this embodiment, the specific marker is PD-L1, but in other embodiments, any other specific marker may be used, and the tumor cell marker is cocktail mixed primary antibody, which is not limited. The following is a detailed description:
step S101: and obtaining a section to be stained. The method specifically comprises the following steps: the method comprises the following steps of trimming a plurality of cancer tissue wax blocks for a preset time, continuously slicing the cancer tissue wax blocks to a thickness of 3 mu m, floating the continuous slices in cold water, naturally unfolding the slices, transferring the separated slices into warm water at 45 ℃ for spreading for 30 seconds, pasting the slices by polylysine-treated glass slides according to the slicing sequence, placing the prepared tissue chips into a 65 ℃ oven for baking for 2 hours, taking out the tissue chips, cooling the tissue chips at room temperature, and placing the tissue chips into a-4 ℃ refrigerator for storage. In the present embodiment, 10 cases of lung adenocarcinoma tissues were taken. In other embodiments, the number of the to-be-stained sections and the type of the to-be-stained sections can be selected according to a specific practical application scenario, and are not limited.
After the section to be stained is prepared, the immunohistochemical staining operation is performed on the section, and in the present embodiment, 2 to 3 consecutive sections are preferably taken to perform immunohistochemical staining of tumor cell markers and immunohistochemical staining of specific markers. Hereinafter, step S102 and step S103 are explained in detail, and it should be noted that step S102 and step S103 do not have a sequential step relationship, and may be performed simultaneously or may be performed before any step.
Step S102: and (3) performing immunohistochemical staining operation of tumor cell markers on part of the section to be stained. The method specifically comprises the following steps: spin-drying the section to be dyed, dripping cocktail mixed primary antibody diluted in a proper proportion, incubating for 1 hour at room temperature (25 ℃), flushing PBS for 3 multiplied by 3 minutes, dripping secondary antibody, incubating for 15 to 30 minutes at room temperature, flushing PBS for 3 multiplied by 3 minutes, throwing away PBS, developing for 3 to 10 minutes by using DAB developing solution which is freshly prepared, counterstaining hematoxylin for 25 seconds, enabling PBS to return blue for 30 seconds, dehydrating according to an alcohol gradient of 85% (3 minutes) -95% (3 minutes) -100% (3 minutes), finally enabling xylene to be transparent for 3 minutes, sealing with neutral gum, positioning the antibody on a tumor cell membrane, staining in a brown yellow color, and staining the cell nucleus in a blue color after counterstaining with hematoxylin.
Step S103: and (3) carrying out specific marker immunohistochemical staining operation on part of the section to be stained. The method specifically comprises the following steps: spin-drying the section, adding diluted primary antibody (PD-L1) in proper proportion, incubating for 1 hour at room temperature (25 ℃), washing for 3X 3 minutes by PBS, adding secondary antibody, incubating for 15-30 minutes at room temperature, washing for 3X 3 minutes by PBS, throwing away PBS, developing for 3-10 minutes by DAB developing solution which is prepared freshly, counterstaining for 25 seconds by hematoxylin, rewetting for 30 seconds by PBS, dehydrating according to alcohol gradient of 85% (3 minutes) -95% (3 minutes) -100% (3 minutes), finally, clearing for 3 minutes by xylene, sealing with neutral gum, positioning PD-L1 antibody on cell membranes of tumor cells and immune cells, staining in brown yellow, and staining cell nuclei in blue after counterstaining by hematoxylin.
Step S104: acquiring a first region of interest of a stained section after tumor cell marker immunohistochemical staining, acquiring feature points in the first region of interest, and calculating the total number of tumor cells according to the number of the feature points in the first region of interest. In this embodiment, the first region of interest is extracted with a preset color as the feature of interest, where the preset color includes, but is not limited to: the color of the first region of interest can be extracted by another process, and as the future process improves, no limitation is made on the color, wherein the immunohistochemistry steps of different colors are substantially consistent, and the difference is only the color of the first region of interest: "develop color for 3-10 minutes with DAB color development liquid prepared freshly"; red- "develop color for 15-20 minutes with freshly prepared AP-Red developing solution"; red- "develop color for 10-30 minutes with AEC color solution prepared freshly"; blue black- "develop 10-30 minutes with BCIP/NBT developing solution prepared freshly". The core technology of the present application is to mark out the tumor cell region, i.e. the first region of interest, by cocktail staining. And taking a blue point in the first region of interest as a characteristic point. The method specifically comprises the following steps: the immunohistochemically stained section of the tumor cell marker was scanned to obtain a first image. On the first picture, brown yellow is taken as an interesting feature, a first interesting region, namely a tumor tissue region, is identified and extracted, and morphological feature points of the region are extracted. And identifying single cells by taking blue as a characteristic point in the first region of interest, and calculating the total number of tumor cells according to the number of blue points.
Step S105: and acquiring a second region of interest of the stained section of the specific marker after immunohistochemical staining according to the first region of interest position, acquiring characteristic points in the second region of interest, and calculating the number of tumor cells stained by the specific marker according to the characteristic points in the second region of interest. In the present embodiment, the brown-yellow point and the blue point in the second region of interest are taken as feature points. The method specifically comprises the following steps: the immunohistochemically stained section of the specific marker was scanned to obtain a second picture. On the second picture, the morphological characteristics of the first region of interest in the step S104 are mapped into the PD-L1 immunohistochemically stained section at the same scale and angle, a second region of interest of the PD-L1 immunohistochemically stained picture is obtained (i.e. the position of the first region of interest on the section is the same as the position of the second region of interest on the section), the tumor cells stained by PD-L1 are identified in the region with the brown and blue as the characteristics of interest, and the number is counted.
Step S106: and calculating the fraction of the positive proportion of the tumor cells according to the total number of the tumor cells and the number of the tumor cells stained by the specific marker. The method specifically comprises the following steps:
tumor cell positive proportion score ═(number of tumor cells stained by any intensity PD-L1 immunohistochemical marker)/total number of tumor cells) × 100%.
Because of the large number of tumor cells, the pathologist generally obtains the number of tumor cells by estimation, as can be seen from the comparison results in the following table, the pathologist generally estimates the number of tumor cells to be lower than the AI counting result, and the accurate counting is basically consistent with the AI counting result. In the aspect of judging the number of PD-L1 positive tumor cells, under the condition that the number of PD-L1 positive tumor cells is less, the judgment result of a pathologist is basically consistent with the result of AI counting, but when the number of PD-L1 positive tumor cells is more, the judgment result of the pathologist is lower than the result of AI counting.
The interpretation result of the lung adenocarcinoma PD-L1 is finally embodied in the form of TPS, and has grade 3 scoring standard, including 1) no PD-L1 expression, TPS < 1%; 2) expression PD-L1, TPS between 1% and 49%; 3) PD-L1 is highly expressed, and TPS is more than or equal to 50 percent. The counting method of the invention is basically consistent with TPS results obtained by counting one by a pathologist, and the TPS results estimated by the pathologist are obviously higher than the TPS results obtained by counting one by the counting method of the invention and the pathologist in No. 4, 5 and 9 tissues.
By obtaining a section to be stained; performing immunohistochemical staining operation of tumor cell markers on part of the section to be stained; carrying out specific marker immunohistochemical staining operation on part of the section to be stained; acquiring a first region of interest of a stained section after immunohistochemical staining of a tumor cell marker, acquiring feature points in the first region of interest, and calculating the total number of tumor cells according to the number of the feature points in the first region of interest; acquiring a second region of interest of the stained section of the specific marker after immunohistochemical staining according to the first region of interest position, acquiring characteristic points in the second region of interest, and calculating the number of tumor cells stained by the specific marker according to the characteristic points in the second region of interest; calculating the fraction of positive proportion of tumor cells according to the total number of the tumor cells and the number of the tumor cells stained by the specific immunohistochemical marker. In the whole process, the total number of the tumor cells and the number of the tumor cells stained by the specific marker are calculated by a program without manual interpretation by a pathologist, so that the obtained result is more accurate and objective, and the efficiency is higher.
Referring to fig. 2, in this embodiment, a specific implementation of a storage device 200 is as follows, where the storage device 200 includes but is not limited to: personal computers, servers, general purpose computers, special purpose computers, network devices, embedded devices, programmable devices, and the like. The method specifically comprises the following steps:
a storage device 200 having stored therein a set of instructions for performing:
acquiring a first picture obtained by scanning a section subjected to tumor cell marker immunohistochemical staining, acquiring a first region of interest of the stained section subjected to tumor cell marker immunohistochemical staining on the first picture, acquiring feature points in the first region of interest, and calculating the total number of tumor cells according to the number of the feature points in the first region of interest;
acquiring a second picture obtained by scanning the immunohistochemically stained section of the specific marker, acquiring a second region of interest of the immunohistochemically stained section of the specific marker on the second picture according to the position of the first region of interest, acquiring characteristic points in the second region of interest, and calculating the number of tumor cells stained by the specific marker according to the characteristic points in the second region of interest;
and calculating the fraction of the positive proportion of the tumor cells according to the total number of the tumor cells and the number of the tumor cells stained by the specific marker.
In the whole process, the total number of tumor cells and the number of the tumor cells stained by the specific marker are calculated by the instruction set in the storage device 200, and a pathologist does not need to manually interpret, so that the obtained result is more accurate and objective, the efficiency is higher, and the cost is lower.
It should be noted that, although the above embodiments have been described herein, the invention is not limited thereto. Therefore, based on the innovative concepts of the present invention, the technical solutions of the present invention can be directly or indirectly applied to other related technical fields by making changes and modifications to the embodiments described herein, or by using equivalent structures or equivalent processes performed in the content of the present specification and the attached drawings, which are included in the scope of the present invention.
Claims (10)
1. A method for analyzing the proportion of PD-L1 positive tumor cells in non-small cell lung cancer, which is characterized by comprising the following steps:
obtaining a section to be dyed;
performing immunohistochemical staining operation of tumor cell markers on part of the section to be stained;
carrying out specific marker immunohistochemical staining operation on part of the section to be stained;
acquiring a first region of interest of a stained section after immunohistochemical staining of a tumor cell marker, acquiring feature points in the first region of interest, and calculating the total number of tumor cells according to the number of the feature points in the first region of interest;
acquiring a second region of interest of the stained section of the specific marker after immunohistochemical staining according to the first region of interest position, acquiring characteristic points in the second region of interest, and calculating the number of tumor cells stained by the specific marker according to the characteristic points in the second region of interest;
and calculating the fraction of the positive proportion of the tumor cells according to the total number of the tumor cells and the number of the tumor cells stained by the specific marker.
2. The method for analyzing proportion of PD-L1 positive tumor cells in non-small cell lung cancer according to claim 1, wherein said "performing tumor cell marker immunohistochemical staining procedure on part of the section to be stained" further comprises the steps of:
spin-drying the section to be dyed, dripping cocktail mixed primary antibody diluted in a proper proportion, incubating for 1 hour at room temperature (25 ℃), flushing PBS for 3 multiplied by 3 minutes, dripping secondary antibody, incubating for 15 to 30 minutes at room temperature, flushing PBS for 3 multiplied by 3 minutes, throwing away PBS, developing for 3 to 10 minutes by using DAB developing solution which is freshly prepared, counterstaining hematoxylin for 25 seconds, enabling PBS to return blue for 30 seconds, dehydrating according to an alcohol gradient of 85% (3 minutes) -95% (3 minutes) -100% (3 minutes), finally enabling xylene to be transparent for 3 minutes, sealing with neutral gum, positioning the antibody on a tumor cell membrane, staining in a brown yellow color, and staining the cell nucleus in a blue color after counterstaining with hematoxylin.
3. The method for analyzing proportion of PD-L1 positive tumor cells in non-small cell lung cancer according to claim 2, wherein the step of obtaining the first region of interest of the stained section after immunohistochemical staining of the tumor cell markers further comprises the steps of:
extracting a first region of interest by taking preset colors as features of interest, wherein the preset colors include but are not limited to: brown-yellow;
the step of acquiring the feature points in the first region of interest further comprises the following steps:
and taking a blue point in the first region of interest as a characteristic point.
4. The method for analyzing proportion of PD-L1 positive tumor cells in non-small cell lung cancer according to claim 1, wherein said "performing specific marker immunohistochemical staining procedure on part of the section to be stained" further comprises the steps of:
spin-drying the section, adding diluted primary antibody (PD-L1) in proper proportion, incubating for 1 hour at room temperature (25 ℃), washing for 3X 3 minutes by PBS, adding secondary antibody, incubating for 15-30 minutes at room temperature, washing for 3X 3 minutes by PBS, throwing away PBS, developing for 3-10 minutes by DAB developing solution which is prepared freshly, counterstaining for 25 seconds by hematoxylin, rewetting for 30 seconds by PBS, dehydrating according to alcohol gradient of 85% (3 minutes) -95% (3 minutes) -100% (3 minutes), finally, clearing for 3 minutes by xylene, sealing with neutral gum, positioning PD-L1 antibody on cell membranes of tumor cells and immune cells, staining in brown yellow, and staining cell nuclei in blue after counterstaining by hematoxylin.
5. The method for analyzing proportion of PD-L1 positive tumor cells in non-small cell lung cancer according to claim 4, wherein the "obtaining feature points in the second region of interest" further comprises the steps of:
and taking the brown color point and the blue color point in the second region of interest as characteristic points.
6. The method for analyzing proportion of PD-L1 positive tumor cells in non-small cell lung cancer according to claim 1, wherein the "obtaining the section to be stained" further comprises the steps of:
the method comprises the following steps of trimming a plurality of cancer tissue wax blocks for a preset time, continuously slicing the cancer tissue wax blocks to a thickness of 3 mu m, floating the continuous slices in cold water, naturally unfolding the slices, transferring the separated slices into warm water at 45 ℃ for spreading for 30 seconds, pasting the slices by polylysine-treated glass slides according to the slicing sequence, placing the prepared tissue chips into a 65 ℃ oven for baking for 2 hours, taking out the tissue chips, cooling the tissue chips at room temperature, and placing the tissue chips into a-4 ℃ refrigerator for storage.
7. The method of claim 1, wherein the step of obtaining a fraction of positive tumor ratios of tumor cells based on the total number of tumor cells and the number of tumor cells stained with the specific marker comprises the steps of:
tumor cell positive proportion score ═(number of tumor cells stained by any intensity PD-L1 immunohistochemical marker)/total number of tumor cells) × 100%.
8. The method for analyzing proportion of PD-L1 positive tumor cells in non-small cell lung cancer according to claim 1, wherein the "obtaining the second region of interest of the stained section after specific marker immunohistochemical staining according to the first region of interest position" further comprises the steps of:
the first region of interest is located at the same position on the slice as the second region of interest.
9. The method for analyzing proportion of PD-L1 positive tumor cells in non-small cell lung cancer according to claim 1, wherein the step of "immunohistochemical staining of tumor cell marker on part of the section to be stained" further comprises the following steps: scanning the section subjected to immunohistochemical staining of the tumor cell marker to obtain a first picture;
after the specific marker immunohistochemical staining operation is carried out on part of the section to be stained, the method further comprises the following steps: the immunohistochemically stained section of the specific marker was scanned to obtain a second picture.
10. A storage device having a set of instructions stored therein, the set of instructions being operable to perform:
acquiring a first picture obtained by scanning a section subjected to tumor cell marker immunohistochemical staining, acquiring a first region of interest of the stained section subjected to tumor cell marker immunohistochemical staining on the first picture, acquiring feature points in the first region of interest, and calculating the total number of tumor cells according to the number of the feature points in the first region of interest;
obtaining a second picture obtained by scanning the immunohistochemically stained section of the specific marker, obtaining a second region of interest of the immunohistochemically stained section of the immunohistochemical marker on the second picture according to the position of the first region of interest, obtaining characteristic points in the second region of interest, and calculating the number of tumor cells stained by the immunohistochemical marker according to the characteristic points in the second region of interest;
and calculating the fraction of the positive proportion of the tumor cells according to the total number of the tumor cells and the number of the tumor cells stained by the specific marker.
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