Method for detecting sperm storage gland distribution information based on panoramic scanning

Abstract

The invention relates to a method for detecting sperm storage gland distribution information based on panoramic scanning, and belongs to the technical field of biological detection. According to the method, panoramic scanning is carried out on sperm storage gland tissue slices to generate digital panoramic slices, then the digital slices are segmented, recognized and counted, the number, density and area proportion of sperm storage glands are accurately calculated, and distribution information of the sperm storage glands is obtained. Compared with a traditional method which depends on an optical microscope to analyze a limited visual field, select a local image visual field for statistics and extrapolate and estimate overall information, the method has the advantages that full-visual-field coverage of sperm storage gland tissue can be realized, statistical errors are remarkably reduced, the accuracy and repeatability of results are improved, and the method is suitable for popularization and application. And a more efficient and accurate analysis means is provided for poultry reproductive biology research.

Description

Technical Field

[0001] The present invention belongs to the technical field of biological detection, and particularly relates to a method for detecting sperm storage tubule (SST) distribution information based on panoramic scanning. Background Art

[0002] In modern poultry breeding practices, the high-intensity selection for body weight and growth rate has led to a significant decline in the reproductive performance of poultry. Therefore, the evaluation of poultry reproductive performance has gradually become the focus of scientific research and production practices. As an important structure in the female poultry reproductive system, the sperm storage tubule (SST) directly affects the efficiency of sperm storage and release, thereby influencing the fertilization rate and the number of offspring. The quantity and distribution characteristics of SST not only determine the sperm storage capacity but also affect the motility and survival time of stored sperm, which is of great significance for improving the reproductive performance and breeding efficiency of female poultry.

[0003] The accurate evaluation of SST distribution information is crucial for understanding the reproductive physiological mechanism of poultry, optimizing breeding strategies, and improving poultry production efficiency. However, there are certain technical limitations in the current methods for SST statistics. Traditional statistical methods usually place the sections under a microscope and observe them. By manually selecting local fields of view and recording the number of SSTs. Subsequently, based on a small number of randomly selected images, the average number of SSTs is calculated and the overall SST density is estimated. This method has low technical repeatability and is limited by the randomness of field-of-view selection, which may lead to insufficient representativeness of the statistical results, large errors, and affect the accuracy of SST distribution information detection. Therefore, there is an urgent need for a method that can efficiently and accurately detect SST distribution information. Summary of the Invention

[0004] In view of this, the purpose of the present invention is to provide a method for detecting SST distribution information based on panoramic scanning, which can more comprehensively, efficiently, and accurately measure the distribution information of SSTs, making the obtained SST distribution information closer to the actual situation.

[0005] The present invention provides a method for detecting SST distribution information based on panoramic scanning, including the following steps:

[0006] Performing panoramic scanning on a sperm storage tubule tissue section to obtain a digital panoramic section;

[0007] After magnifying the digital panoramic section, dividing it into N images according to the mucosal structure for output, where N is an integer greater than 1;

[0008] Measuring and counting the number of sperm storage tubules, the area of sperm storage tubules, and the mucosal area in each image;

[0009] The distribution information of the seminal receptacles is calculated, and the distribution information of the seminal receptacles includes the total number of seminal receptacles, the average density of seminal receptacles, and the proportion of the average area of seminal receptacles:

[0010] The total number of seminal receptacles = the sum of the numbers of seminal receptacles in N images;

[0011] The average density of seminal receptacles = the total number of seminal receptacles / the sum of the mucosal areas of N images;

[0012] The proportion of the average area of seminal receptacles = the sum of the areas of seminal receptacles in N images / the sum of the mucosal areas of N images.

[0013] Preferably, the seminal receptacle tissue includes the seminal receptacle tissue of avian animals; the avian animals include chickens, ducks, geese, pigeons or turkeys.

[0014] Preferably, the seminal receptacle tissue includes the site of the uterovaginal junction or the gonadal tissue where sperm are stored.

[0015] Preferably, the section of the seminal receptacle tissue is a paraffin section.

[0016] Preferably, the method for preparing the paraffin section includes the following steps:

[0017] The seminal receptacle tissue is successively fixed with a first fixative, trimmed, and then fixed with a second fixative to obtain a fixed sample;

[0018] The fixed sample is successively dehydrated, embedded, sectioned, and spread to obtain a paraffin section of the seminal receptacle tissue.

[0019] Preferably, the thickness of the cross-section of the trimmed seminal receptacle block is 1-4 mm.

[0020] Preferably, the thickness of the section is 1-4 μm.

[0021] Preferably, the staining method for the section of the seminal receptacle tissue includes H&E staining, Masson staining, or safranin-fast green staining.

[0022] Preferably, the magnification of the digital panoramic section is 1×-10×.

[0023] The present invention also provides the application of the above method in the research of avian reproductive biology and reproductive physiology, the evaluation of avian reproductive ability, and / or the optimization of livestock and poultry breeding.

[0024] The beneficial effects of the present invention:

[0025] The method for detecting the distribution information of seminal vesicles based on panoramic scanning provided by the present invention, compared with the traditional method that relies on optical microscope analysis of a limited field of view, selects a partial image field for statistics and extrapolates to estimate the overall distribution information, can achieve full-field coverage of seminal vesicle tissue, significantly reduce statistical errors, and improve the accuracy and repeatability of the results. At the same time, through the digital imaging system, real-time viewing and dynamic observation of high-definition panoramic sections can be directly realized in the terminal software, effectively ensuring the image resolution (up to the micron level) and accurate restoration of morphological features, significantly improving the recognition of the microscopic tissue structure of seminal vesicles, and avoiding the technical defects of image edge distortion and uneven resolution in the traditional method that relies on manual microscope focusing, local field shooting, and manual image stitching. In addition, in terms of operation efficiency, through the automatic panoramic scanning and intelligent segmentation technology of the digital system, the processing time for a single section is only 3-4 minutes, which is 5-8 times more efficient than the traditional manual operation that requires focusing and shooting field by field, taking 20-30 minutes / slice, avoiding manual operation errors and greatly shortening the detection cycle, and providing a more efficient and accurate analysis method for avian reproductive biology research. BRIEF DESCRIPTION OF THE DRAWINGS

[0026] Figure 1 It is a technical roadmap for detecting the distribution information of seminal vesicles based on panoramic scanning in Example 1.

[0027] Figure 2 It is a comparison diagram of digital panoramic section output, analysis process and results in Example 1 and Comparative Example 1.

[0028] Figure 3 It is a comparison diagram of digital panoramic section output, analysis process and results in Example 2 and Comparative Example 2. DETAILED DESCRIPTION OF THE EMBODIMENTS

[0029] The present invention provides a method for detecting the distribution information of seminal vesicles based on panoramic scanning, including the following steps:

[0030] Perform panoramic scanning on the seminal vesicle tissue section to obtain a digital panoramic section;

[0031] After magnifying the digital panoramic section, divide it into N images according to the mucosal structure and output them, where N is an integer greater than 1;

[0032] Measure and count the number of seminal vesicles, the area of seminal vesicles, and the mucosal area in each image;

[0033] Calculate the distribution information of seminal vesicles, where the distribution information of seminal vesicles includes the total number of seminal vesicles, the average density of seminal vesicles, and the proportion of the average area of seminal vesicles:

[0034] The total number of seminal vesicles = the sum of the number of seminal vesicles in N images;

[0035] The average density of the sperm storage glands = the total number of sperm storage glands / the sum of the mucosal areas of N images;

[0036] The proportion of the average area of the sperm storage glands = the sum of the areas of the sperm storage glands in N images / the sum of the mucosal areas of N images.

[0037] In the present invention, the sperm storage gland tissue preferably includes the sperm storage gland tissue of avian animals, and the avian animals preferably include chickens, ducks, geese, pigeons or turkeys. In one embodiment, the avian animal can be a chicken or a duck, and the age of the chicken or duck is preferably the sexually mature period, such as a 18-week-old laying hen or a 70-week-old laying duck. The sperm storage gland tissue preferably includes the site of the uterovaginal junction or the gonadal tissue where sperm are stored.

[0038] In the present invention, the section of the sperm storage gland tissue is preferably a paraffin section. The preparation method of the paraffin section preferably includes the following steps: fixing the sperm storage gland tissue successively with a first fixing solution, trimming the block and fixing with a second fixing solution to obtain a fixed sample; dehydrating, embedding, sectioning and spreading the fixed sample successively to obtain a paraffin section of the sperm storage gland tissue.

[0039] In the present invention, the seminal vesicle tissue is preferably fresh seminal vesicle tissue with a complete morphology and no extrusion deformation. The seminal vesicle tissue is fixed with a first fixing solution, and the first fixing solution is preferably a 4 wt.% paraformaldehyde fixing solution. The volume ratio of the seminal vesicle to the paraformaldehyde fixing solution is preferably ≤1:10. In one embodiment, 1:5, 1:8 or 1:10 can be selected. Using the paraformaldehyde fixing solution in this ratio can effectively fix the seminal vesicle tissue. The fixing time of the first fixing solution is preferably ≥48 h. In one embodiment, 48 h, 60 h or 72 h can be selected. After fixation with the first fixing solution, trimming is carried out. The trimming is preferably carried out along the direction perpendicular to the seminal vesicle to facilitate obtaining a cross-section of the seminal vesicle. The thickness of the cross-section of the trimmed seminal vesicle block is preferably 1-4 mm. In one embodiment, 1 mm, 2 mm, 3 mm or 4 mm can be selected. After trimming, fixation is carried out with a second fixing solution. The second fixing solution is preferably a formalin solution, and the volume percentage concentration of the formalin solution is preferably 4%; the fixing time of the second fixing solution is preferably >1 day. In one embodiment, 2 days, 3 days, 4 days or 5 days can be selected; the present invention has no special limitation on the amount of the second fixing solution, and the amount of the conventional fixing solution in the art can be used. After fixation with the second fixing solution, a fixed sample is obtained, and the fixed sample is dehydrated. The dehydration step preferably includes successively soaking the fixed sample in 70% vol ethanol for 3 h, 85% vol ethanol for 3 h, 95% vol ethanol for 3 h, 95% vol ethanol for 3 h, 100% vol ethanol for 3 h, 100% vol ethanol for 3 h and 100% vol ethanol for 2 h to remove the moisture in the tissue sample. After dehydration, embedding is carried out. The present invention has no special limitation on the embedding step, and the conventional paraffin embedding step in the art can be used. After embedding, sectioning is carried out. The thickness of the section is preferably 1-4 μm. In one embodiment, 1 μm, 2 μm, 3 μm or 4 μm can be selected. After sectioning, spreading is carried out. After spreading, a baking process is preferably further included. The present invention has no special limitation on the spreading and baking processes, and the conventional spreading and baking steps in the art can be used for operation.

[0040] In the present invention, after baking the paraffin section of the seminal vesicle tissue, staining is preferably carried out. The staining method preferably includes H&E staining, Masson staining or safranin-fast green staining, and more preferably H&E staining. The present invention has no special limitation on the specific steps of the H&E staining, Masson staining or safranin-fast green staining, and the conventional staining steps in the art can be used for operation. In one embodiment, the H&E staining preferably includes the following steps: dewaxing and rehydrating the paraffin section, hematoxylin staining, eosin counterstaining, dehydrating and mounting. The present invention has no special limitation on the specific operation methods of the dewaxing and rehydrating, hematoxylin staining, eosin counterstaining, dehydrating and mounting, and the conventional operation methods in the art can be used.

[0041] After obtaining the stained section of the seminal receptacle tissue, perform panoramic scanning on the section of the seminal receptacle tissue. The panoramic scanning is preferably performed using a panoramic scanner. The present invention has no special limitation on the source and model of the panoramic scanner, and a conventional panoramic scanner in the art can be used; the present invention has no special limitation on the process of the panoramic scanning and the magnification of the objective lens, and automatic focusing can be performed according to the section.

[0042] After obtaining the digital panoramic section by panoramic scanning, magnify the digital panoramic section. The magnification factor is preferably 1× to 10×, more preferably 4× to 8×. In one embodiment, 4×, 5×, 6×, 7× or 8× can be selected. After magnification, divide it into N images along the mucosal structure gap for output. N is an integer greater than 1. The value of N in the present invention is preferably adjusted according to the magnification factor and the clarity of the seminal receptacle in the picture, so as to be able to clearly identify the seminal receptacle in the image. Then measure and count the number of seminal receptacles, the area of the seminal receptacle and the mucosal area within the field of view in each image. The number of seminal receptacles is preferably counted using image statistics software for each output image, and the counting of the number of seminal receptacles is preferably manual counting.

[0043] The present invention also provides the application of the above method in the research of avian reproductive biology and reproductive physiology, the evaluation of avian reproductive ability and / or the optimization of livestock and poultry breeding.

[0044] The method of the present invention can obtain the distribution information of all seminal receptacles in the whole section of the seminal receptacle tissue by panoramic scanning of the section of the seminal receptacle tissue, then dividing, counting and summing. The error is smaller, the result is more accurate, and it is closer to the actual value, which has important value for the research of avian reproductive biology and reproductive physiology, the evaluation of avian reproductive ability and / or the optimization of livestock and poultry breeding. The present invention has no special limitation on the specific process of the above application, as long as the method for detecting the distribution information of the seminal receptacle based on panoramic scanning in the present invention is used in the above application, it belongs to the protection scope of the present invention.

[0045] The technical solutions provided by the present invention will be described in detail below with reference to the embodiments, but they cannot be understood as limiting the protection scope of the present invention.

[0046] In the following embodiments, unless otherwise specified, they are all conventional methods.

[0047] In the following embodiments, the materials, reagents, etc. used, unless otherwise specified, can be obtained from commercial channels.

[0048] Example 1

[0049] Determination of the distribution information of the seminal receptacle of 18-week-old laying hens

[0050] The technical route of the measurement process is as follows Figure 1 shown, and the steps are as follows:

[0051] 1) Preparation of tissue sections of the sperm storage gland of 18-week-old laying hens:

[0052] After decapitating and sacrificing 18-week-old laying hens, carefully remove the sperm storage gland tissue using ophthalmic surgical scissors and forceps. Use forceps to pick up the outer lumen and rinse it in physiological saline to remove blood stains. Immediately immerse it in a 4 wt.% paraformaldehyde fixative for the first fixation for 72 h. The volume ratio of the sperm storage gland sample to the 4 wt.% paraformaldehyde fixative is 1:10.

[0053] After the first fixation is completed, take out the sperm storage gland tissue sample and trim the block perpendicular to the direction of the sperm storage gland. The thickness of the sperm storage gland block is approximately 3 mm. Place the trimmed block in an embedding cassette with the cross-section of the sperm storage gland tube facing down. Tighten the lid of the embedding cassette and place it in a large beaker containing 4% formalin by volume for 2 days. Take out the embedding cassette from the large beaker and start dehydration. The dehydration procedure is as follows: Immerse the sperm storage gland block in 70% vol ethanol for 3 h, 85% vol ethanol for 3 h, 95% vol ethanol for 3 h, 95% vol ethanol for 3 h, 100% vol ethanol for 3 h, 100% vol ethanol for 3 h, and 100% vol ethanol for 2 h in sequence to remove the water in the tissue sample.

[0054] After dehydration, infiltrate the wax in wax I, wax II, and wax III (wax I, wax II, and wax III are all paraffins melted at 65 °C) for 1 h in sequence. Then pour the paraffin melted at 60 °C into the mold and embed the sample. Note that the cutting surface should be perpendicular to the direction of the sperm storage gland. Place the sample close to the mold. After the paraffin solidifies at room temperature, place it in a -20 °C refrigerator and let it stand for 5 min to cool, and then the mold can be peeled off. Cut the tissue wax block on a microtome with a section thickness of 3 μm. Put the above tissue thin wax slices into a 39 °C constant temperature spreading machine. When the floating wax slices are flattened, vertically insert the numbered glass slides into the water. After the tissue thin wax slices adhere to the glass slides, make good sample marks.

[0055] Use a baking machine to bake the prepared tissue sections at 60 °C for 2 h, and then place the baked sections into a slide basket for hematoxylin-eosin staining. The staining steps are as follows: (1) Dewaxing and rehydration: Immerse the sections in an environmentally friendly dewaxing solution in two times, each time for 20 min. After immersion, successively immerse them in 100% vol ethanol for 5 min, 100% vol ethanol for 5 min, 75% vol ethanol for 5 min, and wash with tap water; (2) Hematoxylin staining: Immerse the sections in hematoxylin stain for 3 - 5 min, wash with tap water, differentiate with a differentiating solution, wash with tap water, blue with a bluing solution, and rinse with running water; (3) Eosin counterstaining: Immerse the sections in 95% vol ethanol for dehydration for 1 min, and then immerse them in eosin stain for 15 s; (4) Dehydration and mounting: Immerse the sections successively in 100% vol ethanol for 2 min, 100% vol ethanol for 2 min, 100% vol ethanol for 2 min, n-butanol I for 2 min, n-butanol II for 2 min, xylene I for 2 min, and xylene II for 2 min for clearing. Finally, take out the glass slide, wipe off the excess liquid, drop a drop of neutral balsam on the tissue section, cover it with a coverslip, and press it firmly to prevent the appearance of air bubbles. Air-dry it in a fume hood, and the section is then made. In the well-made seminal gland section, the seminal gland is transversely sectioned standardly and clearly visible, facilitating the subsequent statistical analysis of relevant histological characteristics after panoramic scanning.

[0056] 2) Panoramic scanning and data statistics:

[0057] Use a Pannoramic MIDI fully automatic digital pathology scanner to complete the panoramic scanning of the sections. To improve the image quality of the section scanning, the section needs to be checked and cleaned before scanning.

[0058] Specifically: Carefully wipe the section with an ultra-fine fiber lens cloth, which can effectively remove dust, water stains or fingerprints on the upper and lower surfaces. For stains that are not easily removed, wet them with water or an alcohol solution and then wipe and clean. After the section is completely dry, load it into the scanner to prevent residual liquid from entering the scanner.

[0059] After the section is cleaned, start preparing for scanning. Turn on the scanner, click the button to pop out the slide table, carefully place the section flat on the slide table to avoid a large defocused area during the scanning process. After the scanner previews the image, the size of the scanning area can be seen. Click the button to start scanning. The instrument will automatically focus, or manual focusing can also be performed according to needs. The change in the field of view indicates the scanning progress. After scanning is completed, click to view the panoramic image and check. After checking without errors, save the digital panoramic section. Click the pop-up button, take out the slide table of the scanner, remove the seminal gland section, and turn off the device. The scanning is then completed.

[0060] Open the saved digital panoramic sections in the computer Case-Viewer software, and scroll the mouse wheel to magnify the panoramic sections to 5× magnification for observing the morphological structure of the seminal receptacle tissue.

[0061] In the Case-Viewer software, use the annotation brush to divide the digital panoramic sections into 42 images with appropriate magnification that can be recognized by the statistical software along the mucosal structure gaps and output (such as Figure 2 ), measure and count the number of seminal receptacles, the area of the seminal receptacles, and the mucosal area in each image; the number of seminal receptacles is manually counted, and the results are shown in Table 1:

[0062] Table 1 Measurement results of the distribution information of seminal receptacles in 18-week-old laying hens in Example 1

[0063]

[0064] Note: The quantity (pcs) is the number of seminal receptacles.

[0065] The following calculation method is used to calculate the distribution information of the seminal receptacles:

[0066] Total number of seminal receptacles = sum of the number of seminal receptacles in N images;

[0067] Average density of seminal receptacles = total number of seminal receptacles / sum of the mucosal areas of N images;

[0068] Proportion of average area of seminal receptacles = sum of the areas of seminal receptacles in N images / sum of the mucosal areas of N images, and the calculation result of the proportion of average area of seminal receptacles is expressed as a percentage.

[0069] According to Figure 2 and Table 1, it can be obtained that: A total of 42 images are output by the method in Example 1. It can be calculated that the total number of seminal receptacles in 18-week-old laying hens is 292, the total area of the seminal receptacles is 0.835 mm 2 , and the total mucosal area is 23.550 mm 2 . Thus, it is finally obtained that the average density is 12.399 pcs / mm 2 , and the average area proportion is 3.55%.

[0070] Comparative Example 1

[0071] Use the traditional method to measure the distribution information of seminal receptacles in 18-week-old laying hens. The steps are as follows:

[0072] Estimate the distribution information of seminal vesicles in the seminal vesicle tissue section in step 1) of Example 1 using the traditional estimation method. The steps are as follows: Instead of the traditional process of observing the section under an optical microscope and selecting 10 local images for statistics, randomly select 10 local images from the digital panoramic section (magnification factor of 5×) obtained in step 2) of Example 1; the 10 selected local images are as Figure 2 shown.

[0073] Import the 10 local images into the Case-Viewer image processing software, and calculate the average seminal vesicle density and the average proportion of seminal vesicle area in the region. Among them, the seminal vesicle density of a single image = the number of seminal vesicles in a single image / the mucosal area of a single image, the average seminal vesicle density = the sum of the seminal vesicle densities of 10 images / 10, the proportion of seminal vesicle area in a single image = the area of seminal vesicles in a single image / the mucosal area of a single image * 100%, the average proportion of seminal vesicle area = the sum of the proportions of seminal vesicle areas of 10 images / 10. Among them, the number of seminal vesicles and the mucosal area of the image can be obtained by relying on the assistance of the Case-Viewer software for statistics. Multiply the calculated average seminal vesicle density by the total mucosal area in the digital panoramic section ( Figure 2 the total area of the segmented region in), to obtain the total number of seminal vesicles. The results are shown in Table 2.

[0074] Table 2 Measurement results of the distribution information of seminal vesicles in 18-week-old laying hens in Comparative Example 1

[0075]

[0076] Note: The quantity (pieces) is the number of seminal vesicles.

[0077] According to Figure 2 and Table 2, it can be obtained that: By using the method in Comparative Example 1 for estimation, the average density of the seminal vesicle tissue of 18-week-old laying hens is 19.312 pieces / mm 2 , the average proportion of area is 4.34%, the total mucosal area is 23.550 mm 2 , and the total number of seminal vesicles is 455.

[0078] Measure the total number of seminal vesicles, the average density of the seminal vesicle tissue, and the average proportion of area in the seminal vesicle tissue sample of the same 18-week-old laying hen. Compare the results obtained by the two methods in Example 1 and Comparative Example 1 as follows:

[0079] The total number of seminal vesicles is 292 and 455 respectively, with a difference of 163 between the two. The error rate of the total number of seminal vesicles measured by the two methods is 55.82%.

[0080] The average density of the seminal vesicle tissue is 12.399 pieces / mm 2 and 19.312 pieces / mm2 , with a difference of 6.913 per mm 2 , and the error rate of the average density of the seminal vesicles measured by the two methods is 55.75%.

[0081] The average area ratios of the seminal vesicle tissues are 3.55% and 4.34% respectively, with a difference of 0.79%. The error rate of the average area ratio of the seminal vesicles measured by the two methods is 22.25%.

[0082] Example 2

[0083] Determination of the distribution information of the seminal vesicles of laying ducks (70 weeks old), the steps are as follows:

[0084] 1) Preparation of tissue sections of the seminal vesicles of laying ducks at 70 weeks old:

[0085] The method for preparing tissue sections of the seminal vesicles of 18-week-old laying hens in step 1) of Example 1 was used to prepare tissue sections of the seminal vesicles of laying ducks at 70 weeks old, and tissue sections of the seminal vesicles of laying ducks at 70 weeks old were obtained.

[0086] 2) Panoramic scanning and data statistics:

[0087] The method in step 2) of Example 1 was used for panoramic scanning and data statistics, except that the obtained digital panoramic section was magnified 5× and then divided into 6 images (such as Figure 3 ), and the obtained distribution information of the seminal vesicles is shown in Table 3.

[0088] Table 3 Results of the determination of the distribution information of the seminal vesicles of laying ducks at 70 weeks old in Example 2

[0089]

[0090] Note: The quantity (pieces) is the number of seminal vesicles.

[0091] The calculation method in step 2) of Example 1 was used to calculate the distribution information of the seminal vesicles. According to Figure 3 and Table 3, it can be obtained that: by using the method in Example 2, the total number of seminal vesicles of laying ducks at 70 weeks old can be calculated to be 2249, and the average density is 16.743 per mm 2 , and the average area ratio is 4.88%.

[0092] Comparative Example 2

[0093] The traditional method was used to determine the distribution information of the seminal vesicles of laying ducks at 70 weeks old, and the steps are as follows:

[0094] Estimate the distribution information of seminal vesicles in the seminal vesicle tissue section in step 1) of Example 2 using the traditional estimation method. The steps are as follows: Instead of the traditional process of observing the section under an optical microscope and selecting 5 local images for statistics, randomly select 5 local images from the digital panoramic section (magnification 5×) obtained in step 2) of Example 2; the 5 selected local images are as Figure 3 shown.

[0095] Import the 5 local images into the Case-Viewer image processing software, and calculate the average density of seminal vesicles and the average proportion of the area of seminal vesicles in the area. Among them, the density of seminal vesicles in a single picture = the number of seminal vesicles in a single picture / the mucosal area of a single picture, the average density of seminal vesicles = the sum of the densities of seminal vesicles in 5 pictures / 5, the proportion of the area of seminal vesicles in a single picture = the area of seminal vesicles in a single picture / the mucosal area of a single picture * 100%, the average proportion of the area of seminal vesicles = the sum of the proportions of the area of seminal vesicles in 5 pictures / 5. Among them, the number of seminal vesicles and the mucosal area of the picture can be obtained by relying on the assistance of the Case-Viewer software for statistics. Multiply the calculated average density of seminal vesicles by the total mucosal area in the digital panoramic section ( Figure 3 the total area of the segmented area in), to obtain the total number of seminal vesicles. The results are shown in Table 4.

[0096] Table 4 Measurement results of the distribution information of seminal vesicles in 70-week-old laying hens in Comparative Example 2

[0097]

[0098] Note: The quantity (pcs) is the number of seminal vesicles.

[0099] According to Figure 3 and Table 4, it can be obtained that: By using the method in Comparative Example 2 for estimation, the average density of the seminal vesicle tissue of 70-week-old laying hens is 11.744 pieces / mm 2 , the average proportion of the area is 3.15%, the total mucosal area is 134.327 mm 2 , and the total number of seminal vesicles is 1578.

[0100] Measure the total number of seminal vesicles, the average density of seminal vesicle tissue, and the average proportion of the area in the seminal vesicle tissue sample of the same 70-week-old laying hen. Compare the results obtained by the two methods in Example 2 and Comparative Example 2 as follows:

[0101] The total number of seminal vesicles is 2249 and 1578 respectively, with a difference of 671 between the two, and the error rate of the total number of seminal vesicles measured by the two methods is 29.84%.

[0102] The average density of seminal vesicle tissue is 16.743 pieces / mm 2 and 11.744 pieces / mm 2, with a difference of 4.999 per / mm 2 , and the error rate of the average density of the seminal vesicles measured by the two methods was 29.86%.

[0103] The average area ratios of the seminal vesicle tissues were 4.88% and 3.15% respectively, with a difference of 1.73%, and the error rate of the average area ratio of the seminal vesicles measured by the two methods was 35.45%.

[0104] The above are only the preferred embodiments of the present invention. It should be noted that for those of ordinary skill in the art, without departing from the principle of the present invention, several improvements and modifications can be made, and these improvements and modifications should also be regarded as the protection scope of the present invention.

Claims

1. A method for detecting the distribution information of seminal vesicles based on panoramic scanning, characterized in that It includes the following steps: Perform panoramic scanning on the seminal gland tissue section to obtain a digital panoramic section; After magnifying the digital panoramic section, divide it into N images according to the mucosal structure and output them, where N is an integer greater than 1; Measure and count the number of seminal glands, the area of seminal glands, and the mucosal area in each image; Calculate the seminal gland distribution information, which includes the total number of seminal glands, the average density of seminal glands, and the proportion of the average area of seminal glands; The total number of seminal glands = the sum of the number of seminal glands in N images; The average density of seminal glands = the total number of seminal glands / the sum of the mucosal areas of N images; The proportion of the average area of seminal glands = the sum of the areas of seminal glands in N images / the sum of the mucosal areas of N images.

2. The method according to claim 1, wherein The seminal gland tissue includes the seminal gland tissue of avian animals; the avian animals include chickens, ducks, geese, pigeons, or turkeys.

3. The method according to claim 1, characterized in that, The seminal gland tissue includes the reproductive gland tissue sites at the uterovaginal junction or sperm storage.

4. The method according to claim 1, characterized in that, The section of the seminal gland tissue is a paraffin section.

5. The method according to claim 4, wherein The preparation method of the paraffin section includes the following steps: Fix the seminal gland tissue successively with a first fixing solution, trim the block, and then fix it with a second fixing solution to obtain a fixed sample; Subject the fixed sample to dehydration, embedding, sectioning, and spreading successively to obtain a paraffin section of the seminal gland tissue.

6. The method according to claim 5, wherein The thickness of the cross-section of the trimmed seminal gland block is 1 - 4 mm.

7. The method according to claim 5, wherein The thickness of the section is 1 - 4 μm.

8. The method according to claim 1, characterized in that, The staining method of the section of the seminal gland tissue includes H&E staining, Masson staining, or safranin fast green staining.

9. The method according to claim 1, wherein The magnification of the digital panoramic section is 1× - 10×.

10. Application of the method according to any one of claims 1 - 9 in the research of avian reproductive biology and reproductive physiology, the evaluation of avian reproductive ability, and / or the optimization of livestock and poultry breeding.

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