A method for field of view leveling of an ultra-large field of view microscope driven by an electric cylinder

Abstract

The present invention discloses a method for field leveling of a super-large field-of-view microscope driven by an electric cylinder, including an electric cylinder, a lens and a microscope stage. The microscope stage includes an X-axis and a Y-axis for controlling the deflection of the stage in the X-axis and Y-axis directions. A sample to be photographed is provided on the microscope stage. The method comprises the following steps: S1: Collecting a microscopic image of the sample to be photographed at the starting angle through the lens to obtain a starting microscopic image; S2: Starting from the starting angle, collecting microscopic images at several angles within a set angle deflection range on the X-axis to obtain several microscopic images with angle deflections; S3: After each angle deflection, performing microscopic image registration and cropping on the microscopic images with angle deflections according to the starting microscopic image; S4: Calculating and obtaining the sharpness of the microscopic images at the starting angle and several angles; S5: Taking the best angle of the X-axis as a reference, repeating S2 to S4 to adjust the best angle of the Y-axis, and completing the global leveling of the XY double axes.

Description

Technical Field

[0001] The present invention relates to the field of microscopic image technology, and particularly to a method for leveling the field of view of an ultra-large field-of-view microscope driven by an electric cylinder. Background Art

[0002] In modern scientific research and technological applications, large field-of-view microscopes are widely welcomed due to their unique advantages. Such microscopes are mainly used for macroscopic to microscopic structure analysis in fields such as materials science, life science, and medical research. Compared with traditional microscopes, large field-of-view microscopes can provide a wider field of view without sacrificing resolution, enabling observers to obtain information on a larger sample area at one time, which is of great significance for understanding complex biological tissue structures or material surface features. In addition, it also has the characteristics of high throughput, capable of quickly scanning large sample areas, greatly improving the experimental efficiency and data acquisition speed, which is particularly important for applications that require a large number of sample detections.

[0003] When using a large field-of-view microscope for observation, leveling the field of view is a crucial step. Due to the limited working distance of the microscope, when the magnification increases, the depth of focus will correspondingly decrease, which means that only a very thin layer of the sample is in the focal position. Therefore, in order to ensure that all regions of interest in the entire field of view can be clearly imaged, the sample must be precisely leveled. Good leveling can not only improve the image quality, reduce edge blurring, but also avoid the loss of important information caused by local defocus, thus ensuring the accuracy and reliability of the data. Especially when performing quantitative analysis, such as particle size measurement or cell counting, the accuracy of field-of-view leveling is directly related to the effectiveness of the final result.

[0004] Traditional manual leveling methods, although capable of performing the work in some simple cases, have many limitations.

[0005] Firstly, manual operation is time-consuming, especially when dealing with complex or large-sized samples, and it is necessary to repeatedly adjust until satisfactory; secondly, human eye judgment is easily affected by subjective factors, and different operators may obtain different leveling results, resulting in poor repeatability; furthermore, under long-term high-intensity work, operators are prone to fatigue, which in turn affects the leveling accuracy.

[0006] Therefore, improvements are needed, and a method for leveling the field of view of an ultra-large field-of-view microscope driven by an electric cylinder is proposed. Summary of the Invention

[0007] The purpose of the present invention is to provide a method for leveling the field of view of an ultra-large field-of-view microscope driven by an electric cylinder to overcome the deficiencies in the prior art.

[0008] To achieve the above object, the present invention provides the following technical solutions:

[0009] The present application discloses a method for leveling the field of view of an ultra-large field of view microscope driven by an electric cylinder, including an electric cylinder, a lens, and a microscope stage. The microscope stage includes an X-axis and a Y-axis for controlling the deflection of the stage on the X-axis and Y-axis. A sample to be photographed is provided on the microscope stage, and the method includes the following steps:

[0010] S1: Collect a microscopic image of the sample to be photographed at the starting angle through the lens to obtain the starting microscopic image;

[0011] S2: Starting from the starting angle, collect microscopic images at several angles within a set deflection angle range on the X-axis to obtain several microscopic images at deflection angles;

[0012] S3: After each angle deflection, perform microscopic image registration and cropping on the microscopic images at deflection angles based on the starting microscopic image to ensure the accuracy of subsequent images;

[0013] S4: Calculate and obtain the clarity of the microscopic images at the starting angle and several angles;

[0014] S5: Taking the best angle on the X-axis as a reference, repeat S2~S4 to adjust the best angle on the Y-axis to complete the global leveling of the XY two axes.

[0015] Preferably, in S2, the collection is performed on either the X-axis or the Y-axis, and when returning to S2 in S5, it is performed on the other one of the X-axis and the Y-axis.

[0016] Preferably, S1 includes the following content: Obtain the region of interest of the sample to be photographed, and take the current position as the starting angle to collect the microscopic image at the current angle.

[0017] Preferably, S2 includes the following sub-steps:

[0018] S21: Obtain several collection angles within a preset deflection angle range according to the starting angle;

[0019] S22: Collect microscopic images at several collection angles;

[0020] S23: Obtain an image sequence based on the microscopic images at several angles.

[0021] Preferably, S3 includes the following content: Since the sample to be photographed in the field of view moves during the angle deflection, each image needs to be translated and registered based on the starting angle as a reference, and the registered image is cropped to retain the common area.

[0022] Preferably, the translational registration in S3 includes the following: registering the microscopic images through the matchTemplate function in OpenCV.

[0023] Preferably, S4 includes the following: for the starting microscopic image and the angular microscopic image cropped in S3, calculate their total variation respectively through the height, width, row and column indexes of their images; on the premise of having the same foreground target, the greater the total variation, the higher the clarity.

[0024] Preferably, S5 includes the following: according to the calculation result of S4, extract the angular microscopic image corresponding to the clearest image, and deflect the microscope stage to this angle; then return to S2, repeat the content of S2 - S4 on the Y-axis, obtain the clearest angles on the X-axis and Y-axis, and thus realize the global field of view leveling in two dimensions of XY.

[0025] This application also discloses a field of view leveling device for an ultra-large field of view microscope driven by an electric cylinder, including a memory and one or more processors. An executable code is stored in the memory. When the one or more processors execute the executable code, it is used to implement the above-mentioned field of view leveling method for an ultra-large field of view microscope driven by an electric cylinder.

[0026] This application also discloses a computer-readable storage medium, on which a program is stored. When the program is executed by a processor, it implements the above-mentioned field of view leveling method for an ultra-large field of view microscope driven by an electric cylinder.

[0027] Advantages of the present invention:

[0028] (1) The field of view of a large field of view microscope is very large, and it is difficult to ensure an ideal focal plane throughout the field of view by manual focusing; the present invention adopts an automated electric cylinder leveling system, which can accurately level the entire large field of view and ensure the consistency of imaging quality from the center to the edge.

[0029] (2) In biological research, it is often necessary to perform imaging analysis on large-area tissue sections or cell samples, which needs to be completed under a large field of view; the automated leveling in the present invention can greatly improve the efficiency of this process, realize true high-throughput and unattended processing, and accelerate the analysis process of large field of view samples.

[0030] (3) For large field of view imaging, manual leveling is a very heavy task, and the operator is prone to visual fatigue, thus increasing the risk of misoperation; while the automated leveling in the present invention can reduce this burden and improve the reliability of experimental data.

[0031] The features and advantages of the present invention will be described in detail through embodiments in conjunction with the drawings. Brief Description of the Drawings

[0032] Figure 1 is a flowchart of the steps of a method for leveling the field of view of a super-large field-of-view microscope driven by an electric cylinder according to the present invention;

[0033] Figure 2 is a schematic diagram of the single-dimensional leveling process of the present invention;

[0034] Figure 3 are schematic diagrams before and after registration and cutting of the present invention;

[0035] Figure 4 is a schematic diagram of the device of the present invention. Detailed Description of the Invention

[0036] To make the objectives, technical solutions and advantages of the present invention clearer, the present invention will be further described in detail below with reference to the drawings and embodiments. However, it should be understood that the specific embodiments described herein are only used to explain the present invention and are not used to limit the scope of the present invention. In addition, in the following description, the descriptions of well-known structures and technologies are omitted to avoid unnecessarily confusing the concepts of the present invention.

[0037] By integrating advanced sensor technologies or computer vision algorithms, the automated system can achieve a fast and accurate leveling process. It not only greatly shortens the preparation time, improves work efficiency, but also significantly reduces human errors, ensuring that each leveling reaches the best state. Therefore, automated leveling is not only an inevitable trend of technological progress but also the key to improving scientific research quality and production efficiency.

[0038] Refer to Figure 1 , an embodiment of the present invention provides a method for leveling the field of view of a super-large field-of-view microscope driven by an electric cylinder. Using multiple real-time captured images of the object to be photographed as the data source, the globally clearest image is found through the optimal search idea to achieve the purpose of leveling.

[0039] The embodiments of the present invention include the following parts:

[0040] The deflection of the microscope stage driven by the cylinder around the X-axis and the deflection around the Y-axis can be simplified. The following steps take the X-axis as an example, and the schematic diagram of the single-dimensional leveling process is as Figure 2 shown.

[0041] S1. Acquisition of microscopic images at the starting point: Find the ROI (region of interest) of the object to be photographed, and use the angle of the electric cylinder driving platform in the x direction at this time as the starting angle , and acquire the microscopic image at this angle ;

[0042] S2. Acquisition of microscopic images at multiple angles: Send instructions to the electric cylinder driver through the program to make it at is the starting angle, in the setting angle range In preset angle steps Perform multi-angle microscopic image acquisition to obtain image sequences ,in , This is because the starting angle has been collected in S1 The microscopic image does not need to be collected again at this step. Set the deflection angle range to 1.5° is (-1°, 1°), angle step The configuration of 0.2° is used as an example case. The image sequence obtained at this time is n=(2*1 / 0.2+1)-1=10;

[0043] S3. Multi-angle microscopic image registration: Since the sample in the field of view will move slightly during the angle deflection process, the image sequence needs to be Each image in the image is a microscopic image with the starting point As a benchmark, image translation and registration are performed to ensure that subsequent calculations are based on and The registration is performed in the public field of view. The matchTemplate function in OpenCV can be used for registration. All registered images are cropped and only the public area is retained for subsequent calculations. The schematic diagram of the effect before and after registration and cropping is shown in the figure. Figure 3 As shown;

[0044] In a feasible embodiment, after completing the registration and cropping, the image is processed with a uniform resolution, so as to reduce the time and resources required for processing in the subsequent processing and calculation process.

[0045] S4. Global clarity calculation: For the cropped and , respectively calculate and TV (total variation), under the premise of having the same foreground target, the larger the TV value, the more drastic the pixel change between adjacent pixels. In terms of visual perception, the more obvious the distinction between tissues is, the clearer it is. Therefore, the TV value can be used as the basis for its clarity. The TV calculation process can be expressed as formula (1):

[0046] (1)

[0047] Among them, h and w represent the pixel height and width of the cropped image, respectively, and i and j represent the row and column indexes of the image, respectively;

[0048] S5. Calculation result feedback: After obtaining the TV calculation result in S4, the corresponding deflection angle can be obtained according to the position index with the largest TV value. , and feedback this deflection angle value to the electric cylinder driver to drive the stage to deflect to the specified angle. , and capture the microscopic image corresponding to this angle as the starting image. , then execute the above steps S2 to S4 on the Y-axis, and finally feedback the Y-axis to the electric cylinder driver to deflect it to the specified angle, then the global vision leveling of the sample in the XY two dimensions can be completed.

[0049] An embodiment of the vision leveling device for an ultra-large field-of-view microscope driven by an electric cylinder according to the present invention can be applied to any device with data processing capabilities, and the any device with data processing capabilities can be a device or apparatus such as a computer. The device embodiment can be implemented by software, or by hardware or a combination of software and hardware. Taking software implementation as an example, as a logically meaningful device, it is formed by the processor of any device with data processing capabilities where it is located reading the corresponding computer program instructions in the non-volatile memory into the memory and running. From the hardware level, as Figure 4 shown, it is a hardware structure diagram of any device with data processing capabilities where the vision leveling device for an ultra-large field-of-view microscope driven by an electric cylinder according to the present invention is located. Except for Figure 4 the shown processor, memory, network interface, and non-volatile memory, the any device with data processing capabilities where the device is located in the embodiment usually also includes other hardware according to the actual functions of the any device with data processing capabilities, which will not be elaborated here. The implementation processes of the functions and roles of each unit in the above device are specifically detailed in the implementation processes of the corresponding steps in the above method, which will not be elaborated here.

[0050] For the device embodiment, since it basically corresponds to the method embodiment, the relevant parts can refer to the partial description of the method embodiment. The device embodiments described above are only illustrative, where the units described as separate components may or may not be physically separated, and the components shown as units may or may not be physical units, that is, they can be located in one place, or distributed to multiple network units. Some or all of the modules can be selected according to actual needs to achieve the purpose of the solution of the present invention. Those of ordinary skill in the art can understand and implement it without creative work.

[0051] The embodiment of the present invention also provides a computer-readable storage medium, on which a program is stored, and when the program is executed by a processor, it implements a vision leveling device for an ultra-large field-of-view microscope driven by an electric cylinder in the above embodiment.

[0052] The computer-readable storage medium may be an internal storage unit of any device with data processing capabilities described in any of the foregoing embodiments, such as a hard disk or memory. The computer-readable storage medium may also be an external storage device of any device with data processing capabilities, such as a plug-in hard disk, a Smart Media Card (SMC), an SD card, a Flash Card, etc. equipped on the device. Further, the computer-readable storage medium may also include both an internal storage unit and an external storage device of any device with data processing capabilities. The computer-readable storage medium is used to store the computer program and other programs and data required by any device with data processing capabilities, and may also be used to temporarily store the data that has been output or will be output.

[0053] The foregoing is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent replacements, or improvements made within the spirit and principle of the present invention shall be included within the protection scope of the present invention.

Claims

1. A method for leveling the field of view of an ultra-large field of view microscope driven by an electric cylinder, characterized in that: The method comprises an electric cylinder, a lens and a microscope stage, wherein the microscope stage comprises an X-axis and a Y-axis, and the electric cylinder is used to control the deflection of the stage on the X-axis and the Y-axis. A sample to be photographed is arranged on the microscope stage, and the method comprises the following steps: S1: Capture a microscopic image of the sample at the starting angle through the lens to obtain a starting microscopic image; S2: Sending instructions to the electric cylinder driver through the program, so that it starts from the starting angle and performs multi-angle microscopic image acquisition within the set deflection angle range on the X-axis with a preset angle step length to obtain a number of deflection microscopic images; S3: After each angle deflection, the deflected microscopic image is registered and cropped according to the starting microscopic image, and the common field of view area is retained to ensure the accuracy of subsequent images; S4: Calculate the clarity of the microscopic images at the starting angle and several angles, and calculate the TV total variation of the starting microscopic image; S5: After obtaining the TV total variation calculation result of S4, the X-axis angle with the highest clarity corresponding to the position index with the largest TV value can be obtained, and this deflection angle value is fed back to the electric cylinder driver to drive the stage to deflect to the specified angle, and the microscopic image corresponding to this angle is taken as the starting image, and then the steps S2 to S4 are performed on the Y axis, and finally the Y axis is fed back to the electric cylinder driver to deflect it to the specified angle to complete the global field of view leveling of the sample in the XY two dimensions.

2. The method for leveling the field of view of a super-large field of view microscope driven by an electric cylinder as claimed in claim 1, characterized in that: The global field of view leveling in S5 is achieved through iterative optimization, and the order of leveling the X-axis and the Y-axis is interchangeable; S2 includes collecting from any one of the X-axis and the Y-axis, and returning to S2 in S5 is executed from the other one of the X-axis and the Y-axis.

3. The method for leveling the field of view of a super-large field of view microscope driven by an electric cylinder as claimed in claim 1, characterized in that: The S1 includes the following contents: obtaining the region of interest of the sample being photographed, and taking the current position as the starting angle, collecting the microscopic image at the current angle.

4. The method for leveling the field of view of a super-large field of view microscope driven by an electric cylinder as claimed in claim 1, characterized in that: The S2 comprises the following sub-steps: S21: acquiring a plurality of acquisition angles within a preset deflection angle range according to the starting angle; S22: collecting microscopic images at several collection angles; S23: Acquire an image sequence based on the microscopic images at several angles.

5. The method for leveling the field of view of an ultra-large field of view microscope driven by an electric cylinder as claimed in claim 1, characterized in that: S3 includes the following contents: since the photographed samples in the field of view move during the angle deflection process, each image needs to be translated and registered based on the starting angle as a reference, and the registered images are cropped to retain the common area.

6. A method for leveling the field of view of an ultra-large field of view microscope driven by an electric cylinder as claimed in claim 5, characterized in that: The translation registration in S3 includes the following contents: registering the microscopic image by using the matchTemplate function in OpenCV.

7. The method for leveling the field of view of an ultra-large field of view microscope driven by an electric cylinder as claimed in claim 1, characterized in that: S4 includes the following contents: for the initial microscopic image and the deflected microscopic image cropped in S3, the total variation is calculated by using the height, width and row and column indexes of the images respectively; under the premise of having the same foreground target, the larger the total variation, the higher the clarity.

8. The method for leveling the field of view of a super-large field of view microscope driven by an electric cylinder as claimed in claim 1, characterized in that: S5 includes the following contents: according to the calculation result of S4, extract the deflection microscopic image corresponding to the clearest image, and deflect the microscope stage to the angle; then return to S2, repeat the contents of S2 to S4 on the Y axis, obtain the clearest angles on the X axis and the Y axis, and then realize the global field of view leveling in the XY two-dimensional.

9. A field of view leveling device for an ultra-large field of view microscope driven by an electric cylinder, characterized in that: It comprises a memory and one or more processors, wherein the memory stores executable codes, and when the one or more processors execute the executable codes, they are used to implement the field of view leveling method for an ultra-large field of view microscope driven by an electric cylinder as described in any one of claims 1 to 8.

10. A computer-readable storage medium, characterized in that: A program is stored thereon, and when the program is executed by a processor, a method for leveling the field of view of an ultra-large field of view microscope driven by an electric cylinder as described in any one of claims 1 to 8 is implemented.

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