Method for preparing a micro-sample pathological digital slide

Abstract

The application relates to a preparation method of a micro sample pathological digital slide, which realizes rapid positioning and focusing by collecting a digital image of a micro sample through a contrast focusing mode, and solves the positioning and focusing problems of the micro sample under a slide scanner. The application provides a more effective tool for pathologists and medical researchers to study the pathological characteristics of micro samples. The micro samples can be more accurately analyzed and researched, which helps to deeply understand the occurrence mechanism, development process and treatment response of diseases, provides strong support for medical research, and promotes the continuous development of the pathology discipline. The micro samples can be better processed, helping doctors to more accurately judge the nature and stage of tumors and provide a basis for formulating a treatment plan. The prepared high-quality pathological digital slide image is easier to transmit and share, and facilitates remote consultation and academic exchange among pathologists.

Description

Technical Field

[0001] This disclosure relates to the field of biological experimental technology, and in particular to a method for preparing a digital slide for pathological microsamples. Background Technology

[0002] Digital pathology slides (WSI) are a technological product that transforms traditional pathology slides into digital images through digital scanning and processing. Digital pathology slides have wide applications in current medical, research, and teaching fields. In terms of storage and management, traditional pathology slides occupy a large amount of physical space and are prone to damage, fading, or loss during long-term storage. Digital slides, on the other hand, are stored digitally on computer hard drives, servers, or cloud storage. This not only greatly saves physical storage space but also easily enables the orderly management of massive amounts of pathology images. Through a database system, the required pathology slide images can be quickly retrieved, and the stability of digital storage ensures that image quality is unaffected by time and is readily available for retrieval.

[0003] Digital slides significantly enhance diagnostic accuracy. Computer software enables sophisticated image analysis and processing, such as quantitative analysis of cell morphology, size, nucleocytoplasmic ratio, tissue structure, and staining characteristics. This quantitative data provides pathologists with a more objective and accurate basis for diagnosis. Furthermore, digital slides facilitate comparative observation of multiple sections, clearly revealing the manifestation of lesions at different levels. They also allow for three-dimensional reconstruction, showcasing pathological structures from a three-dimensional perspective, giving doctors a more comprehensive understanding of the lesions and reducing misdiagnosis and missed diagnosis.

[0004] Information sharing and exchange are another significant advantage of digital slides. Through the internet, digital slides can be rapidly transmitted between pathologists in different hospitals and regions. During remote consultations, experts can visually examine pathological sections without physically touching the slides, conducting diagnoses and discussions as if they were on-site. In academic exchange activities, researchers can easily share special or typical cases, promoting the dissemination and exchange of medical knowledge.

[0005] Digital slides have laid the foundation for the application of artificial intelligence in pathological diagnosis. The vast amount of digital slide data provides valuable material for training AI algorithms. By learning patterns of different pathological features, these algorithms can provide guidance to doctors during the diagnostic process, effectively improving diagnostic efficiency and quality, especially for labor-intensive and repetitive tasks.

[0006] In medical education, digital slides have demonstrated excellent teaching effectiveness. Teachers can use them to show students clear details of pathological slides, and through the software's annotation functions, clearly point out key pathological features to guide student learning. Furthermore, digital slides make examinations and assessments more convenient; teachers can quickly prepare assessment content and accurately evaluate students' mastery of pathological knowledge, thereby improving teaching quality.

[0007] The main challenge in studying ultra-small pathological samples, such as organoids, clinical biopsies, and micro-organs, is their small size, typically ranging from a few hundred micrometers to a few millimeters. While slide scanners can produce digital slide images with micrometer-level resolution, they cannot accurately identify tissue samples smaller than 0.5 mm in diameter before scanning begins. These small pathological samples present a significant challenge to the preparation of digital slides. If the sample is too small (less than 0.5 mm in diameter), it is difficult to locate and provide sufficient focus (≥5), resulting in blurring, defocusing, scanning blind spots, and severe background interference, ultimately leading to the failure of digital slide preparation (see attached image). Figure 1 and attached Figure 2 The image shown is a digital slide image of a 0.5 mm liver sample. However, liver samples from the same animal source, with a diameter of 5 mm, yield clear HE-stained digital slide images with no background interference (see attached image). Figure 3 and attached Figure 4 (Digital slide image of a 5mm liver sample shown).

[0008] Therefore, small samples are difficult to locate and focus under a slide scanner. It is necessary to improve the preparation method of small sample pathological digital slides to solve the problem of difficulty in locating and focusing small samples under a slide scanner. By improving the slide preparation method, the samples can be more easily identified and aligned on the slide. Summary of the Invention

[0009] To address the aforementioned issues, this application proposes a method for preparing digital slides for pathological micro-samples. This method involves adding a contrast enhancer to the micro-sample or slicing it together with a large sample to locate and display the micro-sample.

[0010] This application proposes a method for preparing digital slides for small-sample pathology, comprising the following steps:

[0011] S1. Extract and preprocess small samples from animal model tissues;

[0012] S2. Prepare sample positioning markers and slice them together with the micro sample to form a pathological digital slide of the micro sample;

[0013] S3. Perform digital scanning on the pathological digital slide, locate and focus the micro sample based on the sample positioning marker, and acquire WSI slice images of the micro sample;

[0014] S4. Perform image processing on the WSI slice image and locate the micro sample based on the sample positioning marker.

[0015] In another aspect, this application proposes a method for preparing digital slides for small-sample pathology, comprising the following steps:

[0016] S1. Extract and preprocess small samples from animal model tissues;

[0017] S2. Slice the micro sample to form a pathological digital slide of the micro sample, and add sample positioning markers to the pathological digital slide to provide positioning and focusing location markers for the micro sample on the pathological digital slide;

[0018] S3. Perform digital scanning on the pathological digital slide, locate and focus the micro sample based on the sample positioning marker, and acquire WSI slice images of the micro sample;

[0019] S4. Perform image processing on the WSI slice image and locate the micro sample based on the sample positioning marker.

[0020] Technical effects of the present invention:

[0021] This application uses a contrast-focusing method to acquire digital images of tiny samples, enabling rapid positioning and focusing. This solves the positioning and focusing problems of tiny samples under a slide scanner, reduces scanning blind spots, and improves image quality, providing strong support for the pathological analysis of tiny samples.

[0022] This invention has the following technical advantages:

[0023] 1. Technological improvements and breakthroughs:

[0024] Improving positioning and focusing accuracy: Due to their extremely small size, tiny samples are difficult to accurately locate and focus on under a slide scanner during routine pathological slide preparation and observation. This invention improves the preparation process by using larger samples for auxiliary positioning and adding special markings, allowing for clear identification of tiny samples on the slide. This enables rapid and accurate location and focusing of the sample during scanning, significantly improving the convenience and accuracy of sample observation and reducing the risk of misdiagnosis or missed diagnosis due to positioning difficulties.

[0025] Reducing scanning blind spots: Tiny samples are prone to uneven distribution on the slide, causing some areas to be missed during scanning, forming scanning blind spots. New preparation methods can make the sample more evenly distributed on the slide, or special slide preparation techniques can ensure that the scanner can fully acquire the sample information, avoiding the impact of missing information on diagnostic and research results.

[0026] Enhancing image quality: Small samples have limited image information and may not have high contrast with the background. By adding appropriate contrast enhancers during the preparation process, such as adding brightly stained organs like the pancreas, kidneys, and liver, or by optimizing sample processing steps, increasing the staining concentration, and extending the staining time, the staining contrast of the sample can be made more obvious, highlighting the pathological features of the sample, making the distinction between the sample and the background more obvious, improving the clarity and recognizability of the image, and enabling pathologists to observe and analyze the pathological features of the sample more accurately.

[0027] 2. Scientific research and medical applications:

[0028] Advancing the development of pathology research: This invention provides pathologists and medical researchers with a more effective tool to study the pathological characteristics of tiny samples. In medical research, often only extremely small samples can be obtained, such as fine-needle aspiration samples or minute tissue sections. This invention allows for more accurate analysis and research of these tiny samples, contributing to a deeper understanding of disease mechanisms, progression, and responses to treatment. It provides strong support for medical research and drives the continuous development of pathology.

[0029] Assisting in disease diagnosis: In clinical diagnosis, for some diseases where it is difficult to obtain large quantities of samples, the microsample pathology digital slide preparation method can improve the accuracy and reliability of diagnosis. For example, in the early diagnosis of tumors, sometimes only a small number of cell samples can be obtained through fine-needle aspiration. This method can better handle these tiny samples, helping doctors to more accurately determine the nature and stage of the tumor, and providing a basis for developing a treatment plan.

[0030] Facilitating remote consultations and data sharing: The high-quality digital slide images of pathology produced are easier to transmit and share, facilitating remote consultations and academic exchanges among pathologists. Doctors in different regions can share images of small-sample digital slides of pathology via the network, collaboratively discussing cases and improving diagnostic accuracy and efficiency. Simultaneously, it also facilitates the establishment of pathology databases, providing fundamental data for large-scale clinical research and disease surveillance.

[0031] Other features and aspects of this disclosure will become clear from the following detailed description of exemplary embodiments with reference to the accompanying drawings. Attached Figure Description

[0032] The accompanying drawings, which are included in and form part of this specification, illustrate exemplary embodiments, features, and aspects of this disclosure together with the specification and serve to explain the principles of this disclosure.

[0033] Figure 1 A schematic diagram of a digital slide image of a liver sample measuring 0.5 mm is shown.

[0034] Figure 2 Shown as Figure 1 Enlarged schematic diagram;

[0035] Figure 3 A schematic diagram of a digital slide image of a 5mm liver sample is shown.

[0036] Figure 4 Shown as Figure 3 Enlarged schematic diagram;

[0037] Figure 5 The image shown is a schematic diagram of a large (lower) and a small (upper) liver sample in a low magnification WSI image of the present invention.

[0038] Figure 6 Shown as Figure 5 A magnified schematic diagram of WSI for small to medium-sized samples;

[0039] Figure 7 Shown as Figure 5 A magnified schematic diagram of WSI for medium to large sample sizes;

[0040] Figure 8 The diagram shows a low-magnification pathological tissue illustration of the optic nerve on a digital slide of an ICR mouse.

[0041] Figure 9 The diagram shows a high-magnification pathological tissue illustration of the optic nerve on a digital slide of an ICR mouse. Detailed Implementation

[0042] Various exemplary embodiments, features, and aspects of this disclosure will now be described in detail with reference to the accompanying drawings. The same reference numerals in the drawings denote elements that have the same or similar functions. Although various aspects of the embodiments are shown in the drawings, they are not necessarily drawn to scale unless specifically indicated otherwise.

[0043] The term “exemplary” as used herein means “serving as an example, embodiment, or illustration.” Any embodiment illustrated herein as “exemplary” is not necessarily to be construed as superior to or better than other embodiments.

[0044] Furthermore, to better illustrate this disclosure, numerous specific details are set forth in the following detailed description. Those skilled in the art will understand that this disclosure can be practiced without certain specific details. In some instances, means, components, and circuits well known to those skilled in the art have not been described in detail in order to highlight the main points of this disclosure.

[0045] The experimental reagents and equipment involved in this invention can be purchased commercially or prepared independently; this invention does not impose any limitations. For some experimental steps, such as slicing, digital slide scanning imaging, and image processing, they can be performed in conjunction with corresponding imaging equipment and image processing system software.

[0046] In this embodiment, a tiny sample refers to an animal cell tissue of 0.5 mm in size.

[0047] Large sample refers to a 5mm contrast sample tissue, which can be understood in conjunction with the background description.

[0048] Example 1

[0049] This method involves setting up a comparison sample for the micro sample, slicing it together with the micro sample to form a slide, and then digitally scanning and imaging it. The scanning device then uses the comparison sample to locate and focus the micro sample for imaging.

[0050] This application proposes a method for preparing digital slides for small-sample pathology, comprising the following steps:

[0051] S1. Extract and preprocess small samples from animal model tissues;

[0052] S2. Prepare sample positioning markers and slice them together with the micro sample to form a pathological digital slide of the micro sample;

[0053] S3. Perform digital scanning on the pathological digital slide, locate and focus the micro sample based on the sample positioning marker, and acquire WSI slice images of the micro sample;

[0054] S4. Perform image processing on the WSI slice image and locate the micro sample based on the sample positioning marker.

[0055] This embodiment uses the preparation of liver tissue slides from ICR mice as an example.

[0056] Digital scanning was performed using a Leica Aperio AT2 slide scanner at 20X speed.

[0057] The specific preparation steps are as follows:

[0058] 1. ICR mice were euthanized by cervical dislocation, and the left lobe of the liver was quickly removed from the abdominal cavity and placed in 10% buffered neutral formalin solution for fixation overnight.

[0059] 2. Liver tissue was harvested and cut into liver tissue blocks with diameters of 0.5 mm (small sample) and 5 mm (large sample).

[0060] 3. Gradient dehydration: The tissue was sequentially immersed in a gradient of ethanol concentrations of 60%, 70%, 80%, 90%, 95% ethanol I, 95% ethanol II, anhydrous ethanol I, and anhydrous ethanol II for 60 min each.

[0061] 4. Clearing: After gradient dehydration, the tissue was immersed in clearing agent I and clearing agent II for 20 minutes each.

[0062] 5. Paraffin Immersion: After the tissue has been transparent, it is immersed in paraffin I, paraffin II and paraffin III in sequence for 40 minutes each.

[0063] 6. Paraffin embedding: Small samples and large samples are embedded side by side in the same paraffin block, and the sections are routinely 3μm thick.

[0064] 7. HE staining: Bake the slide at 65℃ for 10 min, clear with xylene I and xylene II for 8 min each, treat with anhydrous ethanol I and anhydrous ethanol II for 5 min each, treat with gradient alcohols (90% ethanol, 80% ethanol I, and 70% ethanol) for 3 min each, rinse with tap water for 3 min, stain with hematoxylin for 7 min, rinse with tap water for 3 min, differentiate with 0.5% hydrochloric acid ethanol for 8 s, rinse with tap water for 3 min, blue with 0.5% dilute ammonia for 1 min, rinse with tap water for 2 min, treat with 80% ethanol II and 95% ethanol for 3 min each, stain with eosin for 1 min, treat with 95% ethanol II and 95% ethanol III for 80 s each, treat with anhydrous ethanol III and anhydrous ethanol IV for 5 min each, and then treat with xylene III for 10 min. Mount the slide and air dry for later use.

[0065] Reagent preparation method:

[0066] Preparation of hematoxylin staining solution: First, dissolve potassium aluminum sulfate in 200ml of ultrapure water by heating. Then, dissolve hematoxylin in 10ml of anhydrous ethanol and pour it into the dissolved potassium aluminum sulfate solution and boil for 1 minute. After cooling slightly, slowly add 1g of red mercuric oxide and continue heating and stirring until the staining solution turns deep purple-red. Cool with cold water, filter with filter paper, and add 5ml of acetic acid.

[0067] 0.5% Hydrochloric Acid Ethanol: Take 300ml of anhydrous ethanol, add it to 100ml of water, mix well, and you will get 75% ethanol. Take 2ml of concentrated hydrochloric acid, slowly add 398ml of 75% ethanol, mix well, and you will get 75% ethanol.

[0068] 0.5% dilute ammonia: Add 5 drops of concentrated ammonia to 450ml of water, mix well, and it's ready.

[0069] 8. After drying, use a Leica Aperio AT2 slide scanner to perform a 20X digital slide scan.

[0070] The scanning steps are as follows:

[0071] Instrument self-test: The slide scanner is powered on and performs a self-test. The instrument initializes until the LED screen displays "ScannerReady".

[0072] Preparing the slides: Place the slides on the slide tray with the coverslip facing up and the label side facing outwards;

[0073] Parameter settings: Set the scan magnification to 20X in the software;

[0074] Start Scan: Click Start, select Pre-take Snapshot, adjust the scan area frame to cover all tissues, and drag the green quadrilateral dot of the control area to a clear area without tissues. At the same time, increase the focus on the tissue, with ≥4 focus points on large samples and ≥1 focus point on small samples.

[0075] Image processing: Upload the captured images to the computer and process them using scanning software;

[0076] Save images: Save the processed images for later analysis or printing.

[0077] 9. Results: Both large and small liver samples were successfully scanned using multiple slides.

[0078] like Figure 5 As shown, in the low-magnification WSI image, large liver samples (lower part of the image) and small liver samples (upper part of the image) are visible, with clear boundaries and bright colors.

[0079] Depend on Figure 6 and Figure 7 The high-magnification image of WSI shown in the image reveals that in both large and small liver samples, the central vein is filled with red blood cells, the hepatocytes are fully formed, the nuclei are round, the nucleoli are prominent and exhibit obvious eosinophilia, located in the center of the hepatocytes, and the cytoplasm is eosinophilic. Digital slides for small-sample liver pathology are prepared with excellent results.

[0080] Therefore, this application addresses the difficulty in locating and focusing tiny samples under a slide scanner by improving slide preparation methods, making samples easier to identify and align on the slide. Appropriate contrast-enhancing materials are selected based on the characteristics of the tiny samples. For cellular tiny samples (such as organoid cells or cells in aspiration samples), for example, when processing clinical aspiration samples (tiny samples), if the puncture site is the liver, a small portion of normal animal liver tissue can be selected (large sample). During slide preparation, the large sample and the tiny sample are placed side-by-side, using the large sample and the target tiny sample together to create a clear identification of the tiny sample on the slide. This makes the distribution of samples on the slide more conducive to scanning, reducing scanning blind spots caused by small samples. Simultaneously, it addresses the problems of limited image information and background interference caused by tiny samples by adding appropriate contrast-enhancing materials or optimizing sample processing steps during preparation, improving image clarity and recognition, and ensuring accurate presentation of pathological features. Large samples can be easily identified under the scanner using conventional staining or labeling methods, allowing the position of the tiny sample to be quickly determined through its relative relationship with the large sample, thus achieving rapid positioning and accurate focusing.

[0081] The improved slide preparation method described above can effectively solve the problems of positioning and focusing of small samples under a slide scanner, reduce scanning blind spots, and improve image quality, providing strong support for the pathological analysis of small samples.

[0082] It should be noted that although animal liver has been used as an example in the above description, those skilled in the art will understand that this disclosure is not limited thereto. In fact, users can flexibly set experimental subjects according to actual application scenarios, as long as the technical functions of this application can be achieved in accordance with the above techniques.

[0083] This embodiment utilizes a large sample to assist in the localization of a small sample. For example, in the processing of small samples from liver biopsies, normal animal liver tissue is used as the large sample. Besides using a large sample from the same tissue, other samples with structures or properties similar to the small sample can be selected as comparative markers. Different auxiliary sample types can be chosen: Alternatively, other known samples with similar characteristics or structures to the small sample can be selected as auxiliary markers. For example, if the small sample is a trace sample of cells from a specific disease, a cell sample with similar lesions but at a more typical and easily identifiable stage can be selected as a reference for auxiliary localization. This may be more beneficial for comparing and localizing the small sample. For example:

[0084] Preparation of digital slides for the optic nerve of ICR mice

[0085] ICR mice were euthanized by cervical dislocation. The skull was quickly opened to remove the optic nerve, and the left lobe of the liver was removed from the abdominal cavity. The liver was then fixed overnight in 10% buffered neutral formalin solution. The next day, the optic nerve was embedded as a whole, and a piece of liver was placed in the embedding cassette. The tissue was then dehydrated sequentially with 70%, 80%, 90%, 95%, and 100% ethanol solutions, cleared in xylene solution, and then embedded in paraffin to form a paraffin block. 3μm sections were prepared and stained with hematoxylin and eosin. The sections were stained according to the present invention. Digital scanning and analysis of the sections revealed that the optic nerve of ICR mice is a bundle of nerve fibers connecting the eye and brain, mainly composed of axons from retinal ganglion cells. HE staining clearly revealed its structure. Figure 8 As shown, the optic nerve is tiny (approximately 230 μm in diameter), and due to its cellular composition, it stains very lightly with HE (hematoxylin and eosin) staining, making it slightly difficult to distinguish with the naked eye. In contrast, the liver stains brightly, providing a clear contrast. Figure 9 As shown, it is mainly composed of optic nerve fibers, optic nerve myelin sheath, glial cells and blood vessels.

[0086] Therefore, this invention utilizes a large sample to assist in the localization of a small sample and clarifies the relative positions of the large and small samples (e.g., placing them at a specific distance) to achieve assisted localization. This method effectively solves the problem of difficulty in localizing and focusing small samples under a slide scanner, ensuring that the small samples have clear markings on the slide, thereby achieving rapid localization and accurate focusing. This method is innovative and practical, and may be a key part of the scope of this invention because it provides a simple and effective strategy for solving the problem of small sample localization, unlike traditional methods that rely solely on the characteristics of the sample itself or complex instruments.

[0087] Example 2

[0088] Based on the implementation principle of Example 1, this application, in another aspect, proposes a method for preparing a digital slide for pathological microscopy, comprising the following steps:

[0089] S1. Extract and preprocess small samples from animal model tissues;

[0090] S2. Slice the micro sample to form a pathological digital slide of the micro sample, and add sample positioning markers to the pathological digital slide to provide positioning and focusing location markers for the micro sample on the pathological digital slide;

[0091] S3. Perform digital scanning on the pathological digital slide, locate and focus the micro sample based on the sample positioning marker, and acquire WSI slice images of the micro sample;

[0092] S4. Perform image processing on the WSI slice image and locate the micro sample based on the sample positioning marker.

[0093] Based on Example 1, the present invention can also employ other positioning marking methods. For example:

[0094] Using physical markers for localization: In addition to relying on additional samples for localization, physical markers can be added to the slide to help locate tiny samples. For example, tiny metal or fluorescent markers can be placed at specific locations on the slide. The approximate location of the tiny sample can be determined by the instrument's identification of these markers, followed by precise scanning and focusing. This method avoids interference that might be introduced by additional samples, and the markers can be precisely designed and placed as needed.

[0095] Choosing other contrast-enhancing substances: The invention mentions adding contrast-enhancing substances to improve image clarity and recognition. Besides existing chemical stains and other substances, novel nanomaterials can be explored as contrast enhancers. For example, some nanoparticles possess unique optical properties, capable of generating strong scattering or fluorescence signals under specific wavelengths of light. Combining these with small samples can significantly enhance the contrast between the sample and the background. Alternatively, specific antibodies combined with fluorescent labels can be used to enhance the display of specific components. Corresponding antibodies can be selected for labeling different pathological features to improve the recognition of specific pathological structures in small samples.

[0096] The specific physical markers, contrast agents, or contrast enhancers can be selected by the experimenters.

[0097] Therefore, this invention adds appropriate contrast-enhancing substances (such as selecting suitable staining agents for different types of small samples) and optimizes sample processing steps (such as precise control of washing and dehydration processes) during the preparation process to improve image clarity and recognizability, ensuring accurate presentation of pathological features. This approach overcomes the diagnostic difficulties caused by limited image information and background interference in small samples, representing a significant technological improvement in the field of pathological slide preparation and providing technical support for the preparation of high-quality digital pathological slides.

[0098] Example 3

[0099] By combining Examples 1 and 2, the two preparation modes mentioned above can be integrated during the glass slide preparation process to obtain digital images with higher image clarity and contrast of small samples.

[0100] Therefore, the present invention has the following technical advantages:

[0101] 1. Improve the accuracy of positioning and focusing:

[0102] Clearly define the location of the sample: Due to their extremely small size, tiny samples are difficult to locate accurately during conventional slide preparation and scanning. The invented method can use large samples for auxiliary positioning, slide marking, or special slide materials to make the tiny samples clearly marked on the slide, so that the scanner can quickly and accurately locate and focus on them, reducing the risk of misdiagnosis or missed diagnosis due to positioning difficulties.

[0103] Reduced operational errors: Enables pathologists or researchers to more easily obtain clear images of small samples when using digital slides for analysis, improving the efficiency and accuracy of diagnosis and research, and reducing operational errors caused by the difficulty in locating and focusing samples.

[0104] 2. Reduce scanning blind spots:

[0105] Uniform sample distribution: Tiny samples are prone to uneven distribution on the slide, causing some areas to be missed during scanning and forming scanning blind spots. The improved preparation method allows for a more uniform distribution of the sample on the slide, ensuring that the scanner can fully acquire the sample information and avoid affecting diagnostic and research results due to missing information.

[0106] Multi-angle scanning coverage: For a large number of small samples or samples with complex distribution, multi-angle scanning can more comprehensively cover the samples, reduce information omissions caused by small and unevenly distributed samples, and improve the information integrity of digital slides.

[0107] 3. Improve image quality:

[0108] Enhanced contrast: Small samples have limited image information and may not have high contrast with the background. Adding contrast-enhancing substances or optimizing sample processing steps can highlight the pathological features of the sample, make the distinction between the sample and the background more obvious, and improve the clarity and recognizability of the image.

[0109] 4. Facilitates data management and sharing:

[0110] Standardized preparation process: The invented method can standardize the preparation process of digital slides for small-sample pathology, ensuring consistency in the digital slides prepared by different laboratories or medical institutions, and facilitating data management and comparison. This is of great significance for conducting multi-center studies, establishing pathology databases, and enabling remote consultations.

[0111] Promoting information sharing: High-quality digital slide images are easier to transmit and share, facilitating remote consultations and academic exchanges among pathologists, which helps promote the development of pathology and improve the efficiency of medical resource utilization.

[0112] The various embodiments of this disclosure have been described above. These descriptions are exemplary and not exhaustive, nor are they limited to the disclosed embodiments. Many modifications and variations will be apparent to those skilled in the art without departing from the scope and spirit of the described embodiments. The terminology used herein is chosen to best explain the principles, practical application, or technical improvements to the embodiments in the market, or to enable others skilled in the art to understand the embodiments disclosed herein.

Claims

1. A method for preparing a digital slide for pathological examination of a small sample, characterized in that, Includes the following steps: S1. Extract and preprocess small samples from animal model tissues; S2. Prepare sample positioning markers and slice them together with the micro sample to form a pathological digital slide of the micro sample; S3. Perform digital scanning on the pathological digital slide, locate and focus the micro sample based on the sample positioning marker, and acquire WSI slice images of the micro sample; S4. Perform image processing on the WSI slice image, and locate the micro sample based on the sample positioning marker; The sample positioning marker is a 5mm large sample from the same tissue as the micro sample; or The sample location marker is another cell sample that is different from the microsample but has the same or similar sample cell morphology characteristics.

2. The method for preparing a digital slide for micro-sample pathology according to claim 1, characterized in that, Both the micro-sample and the large sample were embedded in paraffin.

3. The method for preparing a digital slide for micro-sample pathology according to claim 1, characterized in that, The sample localization marker is a contrast enhancer used to perform component imaging on the tiny sample.

4. The method for preparing a digital slide for micro-sample pathology according to claim 3, characterized in that, The contrast enhancer is one or more of the following contrast agents: Chemical staining agents, fluorescent contrast agents, or nano-enhancing agents.

Images