Cell image acquisition method and system

Abstract

The invention discloses a cell image acquisition method and system, and the method comprises the steps: determining the initial focal plane height of a three-dimensional control platform for carrying out the collection of a transparent carrier, and taking the initial focal plane height as a bearing position, and carrying out the traversal of an initial parameter, at least two laser focusing positions are determined according to focus measuring images of the bearing positions at a plurality of preset laser positions in the horizontal direction, and after the laser focusing positions of all the bearing positions are obtained, all the bearing positions are traversed again; and determining a target laser focusing position, and collecting a plurality of cell images of the bearing position based on the target laser focusing position. Therefore, the focal plane and the focusing position are rapidly determined through the bearing position fixed point position laser focusing, the rest laser focusing point position results are used as redundant standby, the corresponding laser focusing information can be automatically called during bright field photographing acquisition, and automatic and efficient focusing imaging is realized, so that the cell imaging efficiency is improved.

Description

Technical Field

[0001] This application relates to the field of biological image acquisition, and more specifically, to a method and system for acquiring cell images. Background Technology

[0002] With the rapid development of cell technology, cell imaging is being used more and more widely in fields such as biology and chemistry. As a core optical instrument, the microscope magnifies and images tiny cell samples through the objective lens, providing key support for operators to observe cell details.

[0003] Microscopes typically consist of one or more lenses, and their core function is to magnify tiny objects that are difficult to distinguish with the naked eye into the visible range. In cell image acquisition, transparent carriers are often used to hold cell samples. However, in practical applications, the thickness of the transparent carrier often varies, which places higher demands on the imaging quality and ease of operation of the microscope.

[0004] Traditional microscopes cannot automatically, quickly, and accurately determine the focal plane in the above scenarios. When it is necessary to observe cells at different thicknesses on a transparent carrier, the operator must manually and slowly adjust the focus after each sample movement. This not only increases the complexity of the operation but also leads to low overall efficiency in cell image acquisition, making it difficult to meet the needs of large-scale, high-efficiency cell imaging.

[0005] Therefore, how to design a cell image acquisition control scheme to quickly and accurately determine the focal plane of the microscope objective and achieve focusing and image acquisition during movement, thereby improving the efficiency of cell imaging, is a problem that needs attention. Summary of the Invention

[0006] In view of the above problems, this application provides a cell image acquisition method and system to quickly and accurately determine the focal plane of the microscope objective, and to achieve focusing and image acquisition during movement, thereby improving the efficiency of cell imaging.

[0007] To achieve the above objectives, the following specific solutions are proposed:

[0008] A cell image acquisition method is applied to a cell image acquisition system, the cell image acquisition system including a three-dimensional control platform, a bright field light source, a laser light source, an objective lens, a transparent carrier, and an imaging camera;

[0009] The three-dimensional control platform includes a horizontal motion platform and a vertical axis motor. The vertical axis motor is fixed with the objective lens. The transparent carrier is placed on the horizontal motion platform. The transparent carrier contains multiple support positions for culturing cells. The bright field light source emits a beam that passes sequentially through the transparent carrier and the objective lens and is then received by the imaging camera. The laser light source is located between the objective lens and the transparent carrier.

[0010] The method includes:

[0011] Determine the initial focal plane height of the three-dimensional control platform for data acquisition from the transparent carrier;

[0012] Using the initial focal plane height as the initial parameter for traversing the bearing positions, traverse each bearing position: based on the focusing images of each bearing position at multiple preset laser positions in the first horizontal direction, determine at least two laser focusing positions for that bearing position.

[0013] Traverse each bearing position: Based on the position of the horizontal motion platform in the first horizontal direction when traversing to each bearing position, determine the target laser focusing position among at least two laser focusing positions of that bearing position; based on the target laser focusing position of each bearing position, acquire multiple cell images of that bearing position.

[0014] Optionally, the cell image acquisition system further includes an imaging camera located vertically below the laser light source;

[0015] Determining the initial focal plane height for the 3D control platform to acquire data from the transparent carrier includes:

[0016] The initial bearing position of the transparent carrier is moved above the objective lens by the horizontal motion platform.

[0017] With both the bright field light source and the imaging camera turned on, the vertical axis motor is adjusted until the surface image sharpness of the transparent carrier is greater than the sharpness threshold, and then the current initial focal plane height of the vertical axis motor is determined.

[0018] Optionally, the method further includes:

[0019] When the vertical axis motor is at the initial focal plane height, the laser light source is turned on, and the laser spot center position information of the light spot image acquired by the imaging camera is recorded.

[0020] Optionally, using the initial focal plane height as the initial parameter for traversing the bearing positions, traverse each bearing position: based on the focusing images at multiple preset laser positions in the first-dimensional horizontal direction for each bearing position, determine at least two laser focusing positions for that bearing position, including:

[0021] Using the initial focal plane height as the initial parameter for traversing the bearing positions, traverse each bearing position:

[0022] For each traversed bearing position, when the horizontal motion platform moves along the first horizontal direction and moves to each preset laser position of the bearing position, the focus image at each preset laser position is acquired at the current height of the vertical axis motor;

[0023] For each traversed carrier position, at least two laser focusing positions of the carrier position are determined based on the spot information of the focusing image at each preset laser position of the carrier position.

[0024] Optionally, based on the spot information of the focusing image at each preset laser position of the bearing position, at least two laser focusing positions of the bearing position are determined, including:

[0025] If there is only one focusing image containing spot information among the focusing images of the bearing position, then at least two laser focusing positions of the bearing position are determined, which are the height of the vertical axis motor when the focusing image is acquired.

[0026] If none of the focusing images of the bearing position contain light spot information, then it is determined that at least two laser focusing positions of the bearing position are the current height of the vertical axis motor.

[0027] If there are two or more focusing images containing spot information in each focusing image of the bearing position, then the first laser focusing position of the bearing position is determined to be the height of the vertical axis motor when the focusing image containing spot information is acquired first, and the second laser focusing position of the bearing position is determined to be the height of the vertical axis motor when the focusing image containing spot information is acquired last.

[0028] Optionally, after determining at least two laser focusing positions of the carrier position based on the spot information of the focusing image at each preset laser position of the carrier position, the method further includes:

[0029] Adjust the vertical axis motor to the second laser focusing position.

[0030] Optionally, based on the position of the horizontal motion platform in the first horizontal direction when traversing to each bearing position, the target laser focusing position among at least two laser focusing positions of that bearing position is determined, including:

[0031] For each traversed bearing position, when the horizontal motion platform is about to move along the second horizontal direction, the target laser focusing position is determined from at least two laser focusing positions of the bearing position based on the current position of the horizontal motion platform in the first horizontal direction, wherein the second horizontal direction, the first horizontal direction, and the vertical direction are mutually perpendicular.

[0032] Optionally, based on the current position of the horizontal motion platform in the first horizontal direction, the target laser focusing position among at least two laser focusing positions of the bearing position is determined, including:

[0033] If the current position of the horizontal motion platform in the first horizontal direction is in the first half of the bearing position, then the target laser focusing position among the at least two laser focusing positions of the bearing position is determined as the first laser focusing position, wherein the first half of the bearing position is the half of the bearing position first passed through by the horizontal motion platform along the first horizontal direction through each preset laser position.

[0034] If the current position of the horizontal motion platform in the first horizontal direction is in the second half of the bearing position, then the target laser focusing position among the at least two laser focusing positions of the bearing position is determined as the second laser focusing position, wherein the second half of the bearing position is the half of the bearing position that is passed through last during the process of the horizontal motion platform passing through each preset laser position in the first horizontal direction.

[0035] Optionally, acquiring multiple cell images of each carrier position based on the target laser focusing position includes:

[0036] Adjust the vertical axis motor to the target laser focusing position;

[0037] For each carrier position traversed, when the vertical axis motor is in the target laser focusing position, and the horizontal motion platform moves along the second horizontal direction and moves to each preset image acquisition position of the carrier position, the actions of turning on the bright field light source, taking pictures with the imaging camera, and turning off the bright field light source are executed in sequence to acquire cell images of the carrier position at each preset image acquisition position.

[0038] A cell image acquisition system includes a three-dimensional control platform, a bright field light source, a laser light source, an objective lens, a transparent carrier, an imaging camera, and a controller;

[0039] The three-dimensional control platform includes a horizontal motion platform and a vertical axis motor. The vertical axis motor is fixed with the objective lens. The transparent carrier is placed on the horizontal motion platform. The transparent carrier contains multiple support positions for culturing cells. The bright field light source emits a beam that passes sequentially through the transparent carrier and the objective lens and is then received by the imaging camera. The laser light source is located between the objective lens and the transparent carrier.

[0040] The controller includes:

[0041] A focal plane initialization unit is used to determine the initial focal plane height for the three-dimensional control platform to collect data from the transparent carrier.

[0042] The laser focusing position determination unit is used to traverse each bearing position using the initial focal plane height as the initial parameter for bearing position traversal: based on the focusing images of each bearing position at multiple preset laser positions in the first horizontal direction, it determines at least two laser focusing positions for that bearing position.

[0043] The acquisition and control unit is used to traverse each carrier position: based on the position of the horizontal motion platform in the first horizontal direction when traversing to each carrier position, it determines the target laser focusing position among at least two laser focusing positions of that carrier position; based on the target laser focusing position of each carrier position, it acquires multiple cell images of that carrier position.

[0044] By employing the above technical solution, this application determines the initial focal plane height for the 3D control platform to acquire data from the transparent carrier. Using this initial focal plane height as the initial parameter for traversing the carrier positions, each carrier position is traversed. Based on the focusing images at multiple preset laser positions in the first-dimensional horizontal direction for each carrier position, at least two laser focusing positions for that carrier position are determined. After obtaining the laser focusing positions for all carrier positions, each carrier position is traversed again. Based on the position of the horizontal motion platform in the first-dimensional horizontal direction when traversing to each carrier position, the target laser focusing position among the at least two laser focusing positions for that carrier position is determined. Based on the target laser focusing position for each carrier position, multiple cell images of that carrier position are acquired. Therefore, by using fixed-point laser focusing on the carrier positions to quickly determine the focal plane and focusing position, while using the remaining laser focusing position results as redundancy, the corresponding laser focusing information can be automatically called during bright-field imaging, achieving automatic and efficient focusing and imaging, thereby improving cell imaging efficiency. Attached Figure Description

[0045] Various other advantages and benefits will become apparent to those skilled in the art upon reading the following detailed description of preferred embodiments. The accompanying drawings are for illustrative purposes only and are not intended to limit the scope of this application. Furthermore, the same reference numerals denote the same parts throughout the drawings. In the drawings:

[0046] Figure 1 This is a schematic diagram of a system structure for cell image acquisition provided in an embodiment of this application;

[0047] Figure 2 This is a schematic diagram of a process for implementing cell image acquisition provided in an embodiment of this application;

[0048] Figure 3 A schematic diagram of a process for obtaining an initial focal plane is provided for an embodiment of this application;

[0049] Figure 4 A layout diagram of a transparent carrier provided in an embodiment of this application;

[0050] Figure 5 A schematic diagram of the laser focusing position of a transparent carrier bearing position provided in an embodiment of this application;

[0051] Figure 6 A schematic diagram of a process for performing laser focusing is provided for an embodiment of this application;

[0052] Figure 7 This application provides a diagram of motor signal triggering during focusing and ranging.

[0053] Figure 8 This is a schematic diagram of a carrier cell image provided in an embodiment of this application;

[0054] Figure 9 This application provides a schematic diagram of a process for full-plate image acquisition on a transparent carrier.

[0055] Figure 10 This application provides a motor signal trigger diagram for full-board image acquisition.

[0056] Figure 11 This is a schematic diagram of the structure of a controller for a cell image acquisition system provided in an embodiment of this application, which implements cell image acquisition. Detailed Implementation

[0057] The technical solutions of the embodiments of this application will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of this application, and not all embodiments. Based on the embodiments of this application, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of this application.

[0058] This application provides an optional system architecture for cell image acquisition, which may include a three-dimensional control platform, a bright field light source, a laser light source, an objective lens, a transparent carrier, an imaging camera, a memory, and a controller.

[0059] The three-dimensional control platform may include a horizontal motion platform (XY motion platform), a horizontal axis motor (X motor), a vertical axis motor (Y motor), and a vertical axis motor (Z motor). The Z motor is fixed with an objective lens. A transparent carrier is placed on the horizontal motion platform. The transparent carrier contains multiple carrier sites for cell culture; different types of transparent carriers have different numbers of carrier sites, and the size of each carrier site also differs. For example, in a transparent carrier with 96 (12×8) carrier sites and a transparent carrier with 384 (24×16) carrier sites, the carrier site size of the 96-well plate is larger than that of the 384-well plate. The imaging camera is located vertically below the laser source. The bright-field light source emits a beam that passes sequentially through the transparent carrier and the objective lens before being received by the imaging camera. The laser source is located between the objective lens and the transparent carrier, and the imaging camera is located vertically below the objective lens.

[0060] Specifically, the X motor has its corresponding X motor encoder, the Y motor has its corresponding Y motor encoder, and the Z motor has its corresponding Z motor encoder. The signal flow and control method of this system can be found in [reference needed]. Figure 1 The controller can send corresponding motion commands to the laser light source, bright field light source, imaging camera, memory, and XYZ motors. The controller can also acquire position data stored in the memory and encoder data representing the movement of the XYZ motors. The XY motors control the movement of the XY motion platform in the horizontal two-dimensional direction, while the Z motors control the movement of the transparent carrier in the Z-axis direction.

[0061] Based on the above system architecture, system signal flow, and control method, Figure 2 This illustration shows a flowchart of a cell image acquisition method provided in an embodiment of this application. (Refer to...) Figure 2 The process may include:

[0062] Step S110: Determine the initial focal plane height for the 3D control platform to collect data from the transparent carrier.

[0063] Understandably, the initial focal plane height of the three-dimensional control platform is determined before focusing, so as to determine the initial focal plane at that height. This gives a rough position for focusing on the bearing position, which is convenient for subsequent focusing on each bearing position and avoids large adjustments to the Z motor when focusing on the bearing position.

[0064] Step S120: Using the initial focal plane height as the initial parameter for traversing the bearing positions, traverse each bearing position: Based on the focusing images of multiple preset laser positions in the first horizontal direction for each bearing position, determine at least two laser focusing positions for that bearing position.

[0065] The first horizontal dimension can be the direction of the X motor (X direction). The number of preset laser positions varies for transparent carriers with different numbers of holes, and the determined laser focusing positions also differ.

[0066] Understandably, focusing at multiple preset laser positions ensures the reliability of the focusing position. Furthermore, during laser focusing, the carrier position might fail to produce a laser ranging result due to other factors. Therefore, by using the laser ranging results from the remaining laser focusing positions as backups, the system compensates for situations where laser ranging fails, thus ensuring cell imaging efficiency.

[0067] Step S130: Traverse each bearing position: Based on the position of the horizontal motion platform in the first horizontal direction when traversing to each bearing position, determine the target laser focusing position among at least two laser focusing positions of that bearing position; based on the target laser focusing position of each bearing position, acquire multiple cell images of that bearing position.

[0068] It is understandable that at least two laser focusing positions of the carrier position are determined at multiple preset laser positions in the first-dimensional horizontal direction during the focusing stage. Therefore, when sampling the image at the corresponding carrier position, it is necessary to match the laser focusing position corresponding to or close to the preset laser position as the target focusing position as much as possible, so that the target focusing position can accurately adapt to the focal plane formed by the slight thickness deviation of the transparent carrier in this area.

[0069] The cell image acquisition method provided in this embodiment determines the initial focal plane height for the 3D control platform to acquire images from the transparent carrier. Using this initial focal plane height as the initial parameter for traversing the carrier positions, the method traverses each carrier position. Based on the focusing images at multiple preset laser positions in the first-dimensional horizontal direction for each carrier position, at least two laser focusing positions for that carrier position are determined. After obtaining the laser focusing positions for all carrier positions, the method traverses each carrier position again. Based on the position of the horizontal motion platform in the first-dimensional horizontal direction when traversing to each carrier position, the target laser focusing position among the at least two laser focusing positions for that carrier position is determined. Based on the target laser focusing position for each carrier position, multiple cell images of that carrier position are acquired. Therefore, by using fixed-point laser focusing on the carrier position to quickly determine the focal plane and focusing position, and using the remaining laser focusing position results as redundancy, the corresponding laser focusing information can be automatically called during bright-field imaging, achieving automatic and efficient focusing and imaging, thereby improving cell imaging efficiency.

[0070] In some embodiments of this application, the process of determining the initial focal plane height of the three-dimensional control platform for acquiring data from the transparent carrier in step S110 is described. This process may include:

[0071] S1. Move the initial bearing position of the transparent carrier to above the objective lens using a horizontal motion platform.

[0072] S2. With both the bright field light source and the imaging camera on, adjust the vertical axis motor until the surface image sharpness of the transparent carrier is greater than the sharpness threshold, and then determine the current initial focal plane height of the vertical axis motor.

[0073] Furthermore, by turning on the laser source when the vertical axis motor is at the initial focal plane height, the laser spot center position information of the laser spot image acquired by the imaging camera can be recorded. The laser spot center position information includes the initial focal plane height and the XY position of the laser spot center.

[0074] For example Figure 3 The initial focal plane acquisition process is as follows: the controller controls the XY motion platform to move to the A1 bearing position of the transparent carrier (e.g., Figure 4 (The A1 bearing position shown is the initial bearing position); turn on the bright field light source and imaging camera; adjust Z until a clear image of the transparent carrier surface is obtained and record the position as the initial focal plane height Z0; then turn off the bright field light source. Next, turn on the laser light source again, and the imaging camera can acquire a light spot image; record the XY position (X0, Y0) of the center of the light spot at this time; then turn off the laser light source.

[0075] In some embodiments of this application, the process of step S120 of the above embodiment, which involves traversing each carrier position using the initial focal plane height as the initial parameter, and determining at least two laser focusing positions for each carrier position based on the focusing images at multiple preset laser positions in the first-dimensional horizontal direction, is described. This process may include:

[0076] Using the initial focal plane height as the initial parameter, iterate through each bearing position.

[0077] For example Figure 4 As shown, during the focusing process, the traversal direction of the bearing positions can start from bearing position A1, with the X motor moving towards A12 until A12 is reached. After completing the traversal of bearing position A12 in row A, the Y motor can be driven to move to B12, and then the traversal can continue from B12 to B1. The entire board is traversed in this serpentine manner until all bearing positions have been traversed.

[0078] For each traversed bearing position, when the horizontal motion platform moves along the first horizontal direction and moves to each preset laser position of the bearing position, a focus image at that preset laser position is acquired at the current height of the vertical axis motor.

[0079] The first horizontal dimension can represent either the direction of motion of the X motor or the direction of motion of the Y motor. This example uses the direction of motion of the X motor as the first horizontal dimension, that is... Figure 4 The X direction. The preset laser position for each bearing position can be referenced. Figure 5Each bearing position can have three preset laser positions L1, L2, and L3 in the X direction. The laser imaging heights corresponding to the positions L1, L2, and L3 can be different, respectively Z L1 Z L2 and Z L3 That is, when the Z motor moves to the laser positions L1, L2, and L3, it needs to move the Z motor by the appropriate speed respectively. L1 Z L2 and Z L3 The image was acquired at the height of the laser light source, thus obtaining three focus images.

[0080] For each traversed carrier position, at least two laser focusing positions are determined based on the spot information of the focusing image at each preset laser position of the carrier position.

[0081] Specifically, the spot information of the focusing image at each preset laser position of the bearing position can include the following three cases.

[0082] The first method is to determine at least two laser focusing positions of the bearing position if there is only one focusing image containing light spot information among the focusing images of the bearing position. These positions are the height of the vertical axis motor when the focusing image is acquired.

[0083] For example Figure 5 For a 96-well plate, two laser focusing positions Z need to be determined. V1 and Z V2 If in L1, L2, or L3 there is only one focusing image containing spot information (assuming it is from Z...) L1 Collected from Z L1 The focus measurement image acquired at point Z has light spots, from Z L2 and Z L3 If the focus measurement image acquired at point Z has no light spot, then the two laser focusing positions Z are determined. V1 and Z V2 All equal to Z L1 .

[0084] The second method is to determine that if none of the focus images of the bearing position contain light spot information, then at least two laser focusing positions of the bearing position are the current height of the vertical axis motor.

[0085] For example Figure 5 If none of the three focus measurement images acquired in L1, L2, or L3 show a light spot, then Z is considered to be... L1 Z L2 and Z L3 If ineffective, then the Z motor remains stationary, and the two laser focusing positions Z... V1 and Z V2 All are equal to the current height Z of the Z motor. L0 .

[0086] After determining at least two laser focusing positions of the carrier position, the at least two laser focusing positions of the carrier position can be stored in the memory so that the laser focusing position information corresponding to the carrier position can be retrieved during cell image acquisition.

[0087] The third method is as follows: If there are two or more focusing images containing light spot information in each focusing image of the bearing position, then the first laser focusing position of the bearing position is determined to be the height of the vertical axis motor when the focusing image containing light spot information is acquired first, and the second laser focusing position of the bearing position is determined to be the height of the vertical axis motor when the focusing image containing light spot information is acquired last.

[0088] For example Figure 5 The sampling order is L1→L2→L3, in Z. L1 Z L2 and Z L3 In the middle, if Z L1 and Z L3 Contains light spot information, Z L2 If there is no light spot information, then Z V1 = Z L1 Z V2 = Z L3 If Z L2 and Z L3 Contains light spot information, Z L1 If there is no light spot information, then Z V1 =Z L2 Z V2 = Z L3 If Z L1 Z L2 and Z L3 Both contain light spot information, then Z V1 = Z L1 Z V2 = Z L3 .

[0089] Furthermore, after determining at least two laser focusing positions (such as Z) of the bearing position... V1 and Z V2 After that, the vertical axis motor can be adjusted to the second laser focusing position Z. V2 .

[0090] Understandably, in practical use, the flatness of transparent carriers can vary significantly due to differences in manufacturers and manufacturing processes, with the surface difference at the bottom of the holes at different carrier positions reaching up to 100µm. To ensure that some carrier positions can obtain laser spots and to avoid laser ranging failure, the Z motor needs to be moved to the Z... V2 .

[0091] More specifically, Figure 6The process of laser focusing across the entire plate carrier position is illustrated. The controller moves the XY platform to the starting position of carrier position A1 on the transparent carrier and turns on the laser source; for each row of Y, the X motor moves from X=1 to X=X. MAX During the X-axis movement, it will pass through preset laser positions L1, L2, and L3 within each support position. The camera will capture an image each time it passes a laser position. The acquired image will be analyzed by the controller of the control device to determine if there is a light spot. If a light spot is present, the Z-position Z of the current laser position will be calculated. L1 Z L2 and Z L3 After further processing and judgment, two of the Z's were removed. V1 and Z V2 Write the data into the memory of the control device; after X has passed through the three laser positions within each bearing position, the Z motor moves to Z. V2 After X has passed through all laser positions within all bearing positions, the Y motor moves to the next row until Y = Y. MAX When the laser is turned off, laser focusing ends. The timing signal reference for the entire laser focusing and ranging process is as follows: Figure 7 When the Y motor is stationary, the X motor moves left and right to perform laser ranging.

[0092] In some embodiments of this application, the process of determining the target laser focusing position among at least two laser focusing positions of a bearing position based on the position of the horizontal motion platform in the first horizontal direction when traversing each bearing position, as mentioned in the above embodiments, is described. Specifically, it may include:

[0093] For each traversed bearing position, when the horizontal motion platform is about to move along the second horizontal direction, the target laser focusing position is determined from at least two laser focusing positions of the bearing position based on the current position of the horizontal motion platform in the first horizontal direction.

[0094] The second horizontal dimension represents the direction of the Y motor. The second horizontal dimension, the first horizontal dimension, and the vertical direction are all perpendicular to each other.

[0095] Specifically, the process of determining the target laser focusing position of the bearing position can include the following two situations.

[0096] The first method is to determine the target laser focusing position among at least two laser focusing positions of the bearing position if the current position of the horizontal motion platform is in the first half of the bearing position.

[0097] The first half-zone refers to the half-zone that is traversed first as the bearing position moves along the first horizontal dimension through various preset laser positions by the horizontal motion platform. (Reference) Figure 8 , Figure 8 This indicates that a data bit is divided into 4 columns from left to right. The order from left to right here corresponds to the data bit's position in the image. Figure 5 The order of the captured focusing images is consistent. Therefore, during the process of the horizontal motion platform / X motor passing through the preset laser position of the bearing position along the X direction, the first half-zone it passes through is the 1st and 2nd columns. If the X position of the X motor at this time falls in the first half-zone (or the 1st and 2nd columns), then the target laser focusing position of the bearing position is determined to be the first laser focusing position Z of the two laser focusing positions. V1 .

[0098] The second method is to determine the target laser focusing position among at least two laser focusing positions of the bearing position if the current position of the horizontal motion platform in the first horizontal direction is in the second half of the bearing position.

[0099] The second half-zone refers to the half-zone traversed last during the process of the bearing position being moved by the horizontal motion platform along the first horizontal direction through various preset laser positions. (Reference) Figure 8 , Figure 8 This indicates that a data bit is divided into 4 columns from left to right. The order from left to right here corresponds to the data bit's position in the image. Figure 5 The order of the acquired focusing images is consistent. Therefore, during the process of the horizontal motion platform / X motor passing through the preset laser position of the bearing position along the X direction, the latter half of the zone is the 3rd and 4th columns. If the X position of the X motor at this time falls in the second half of the zone (or the 3rd and 4th columns), then the target laser focusing position of the bearing position is determined to be the first laser focusing position Z of the two laser focusing positions. V2 .

[0100] Understandably, during laser focusing, three ranging positions, L1, L2, and L3, are preset at each bearing position. V1 Z V2 The effective focal plane parameters are selected from these parameters, and the sampling positions C1 to C4 are distributed in columns within the bearing position. X is closer to the generation of Z in the first and second sampling positions. V1 In the laser point area, X is closer to the generated Z in the 3rd and 4th columns. V2 For the laser point area, select the corresponding Z V The value can precisely adapt to the focal plane formed by the minute thickness deviation of the transparent carrier in this area. This allows for the selection of the target laser focusing position Z. V This approach not only maintains the high efficiency of fixed-point laser focusing instead of point-by-point focusing, but also avoids image defocusing caused by local thickness differences within the bearing position by calling matching preset focal plane parameters in different regions, thus ensuring the clarity of the image across the entire bearing position. At the same time, combined with the redundant design of laser focusing, it further reduces the impact of a single ranging anomaly on the image quality, achieving a balance between efficiency and imaging quality.

[0101] In addition, for some types of transparent carriers with only one preset laser position, there is only one identifiable laser focusing position. During full-plate image acquisition on these transparent carriers, since there is only one laser focusing position, the process of determining the laser focusing position based on the X-motor position can be skipped (or this single laser focusing position can be directly determined as the target laser focusing position), and cell image acquisition can be performed directly according to the laser focusing position.

[0102] Based on this, the process of acquiring multiple cell images of each carrier position based on the target laser focusing position of each carrier position, as mentioned in the foregoing embodiments, will be introduced.

[0103] Specifically, after determining the target laser focusing position, first adjust the vertical axis motor / Z motor to the target laser focusing position to fix the focus position.

[0104] Furthermore, for each carrier position traversed, with the vertical axis motor in the target laser focusing position, when the horizontal motion platform moves along the second horizontal direction and moves to each preset image acquisition position of the carrier position, the actions of turning on the bright field light source, taking pictures with the imaging camera, and turning off the bright field light source are executed in sequence to acquire cell images of the carrier position at each preset image acquisition position.

[0105] For details, please refer to Figure 8 If motor X is in column 1, then motor Z is in column Z. V1 Below, the Y motor acquires four cell images in the order of C1→C2→C3→C4 from top to bottom in the first column; if the X motor is located in the second column, then the Z motor is located in the Z... V1 Below, the Y motor acquires four cell images in the order of C1→C2→C3→C4 from bottom to top in the second column; if the X motor is located in the third column, then the Z motor is located in the Z... V2 Below, the Y motor acquires four cell images in the order of C1→C2→C3→C4 from top to bottom in the 3rd column; if the X motor is located in the 4th column, then the Z motor is in the Z position. V2 The Y-motor acquires four cell images in the order of C1→C2→C3→C4 from top to bottom in the fourth column. Each time it passes one of the acquisition positions, it sequentially performs the actions of turning on the bright-field light source, taking a picture with the imaging camera, and turning off the bright-field light source, thereby obtaining the cell image at that acquisition position.

[0106] For a more detailed process of full-board image acquisition, please refer to [link / reference]. Figure 9 and combined Figure 4The XY platform first moves to the starting position of the A1 bearing position and begins acquiring data from the first column of A1 towards H1. After acquiring data from the first column of H1, the X motor moves to the second column of H1 and begins acquiring data from the second column of H1 towards A1, continuing this process until the second column of A1 is acquired, and so on. After acquiring data from the fourth column of A1, the X motor moves to the first column of A2 and acquires data from A2 to H2 using the serpentine movement pattern described above, until the entire board is acquired. The timing signals for full-board acquisition are referenced below. Figure 10 When the X motor is stationary, the Y motor moves back and forth to take open-field photos.

[0107] The controller for cell image acquisition provided in the embodiments of this application is described below. The controller for cell image acquisition described below can be referred to in correspondence with the method for cell image acquisition described above.

[0108] See Figure 11 , Figure 11 This is a schematic diagram of a controller structure for acquiring cell images, as disclosed in an embodiment of this application.

[0109] like Figure 11 As shown, the controller may include:

[0110] The focal plane initialization unit 100 is used to determine the initial focal plane height for the three-dimensional control platform to acquire data from the transparent carrier.

[0111] The laser focusing position determination unit 200 is used to traverse each bearing position using the initial focal plane height as the initial parameter for bearing position traversal: based on the focusing image 300 at multiple preset laser positions in the first horizontal direction of each bearing position, at least two laser focusing positions of that bearing position are determined.

[0112] The acquisition and control unit is used to traverse each carrier position: based on the position of the horizontal motion platform in the first horizontal direction when traversing to each carrier position, it determines the target laser focusing position among at least two laser focusing positions of that carrier position; based on the target laser focusing position of each carrier position, it acquires multiple cell images of that carrier position.

[0113] Optionally, the specific control logic implemented by other units of the controller corresponds one-to-one with the cell image acquisition scheme of the aforementioned embodiments, as detailed in the aforementioned embodiments, and will not be repeated here.

[0114] Finally, it should be noted that in this document, relational terms such as "first" and "second" are used only to distinguish one entity or operation from another, and do not necessarily require or imply any such actual relationship or order between these entities or operations. Furthermore, the terms "comprising," "including," or any other variations thereof are intended to cover non-exclusive inclusion, such that a process, method, article, or apparatus that comprises a list of elements includes not only those elements but also other elements not expressly listed, or elements inherent to such a process, method, article, or apparatus. Without further limitations, an element defined by the phrase "comprising one..." does not exclude the presence of other identical elements in the process, method, article, or apparatus that includes said element.

[0115] The various embodiments in this specification are described in a progressive manner. Each embodiment focuses on the differences from other embodiments. The various embodiments can be combined as needed, and the same or similar parts can be referred to each other.

[0116] The above description of the disclosed embodiments enables those skilled in the art to make or use this application. Various modifications to these embodiments will be readily apparent to those skilled in the art, and the general principles defined herein may be implemented in other embodiments without departing from the spirit or scope of this application. Therefore, this application is not to be limited to the embodiments shown herein, but is to be accorded the widest scope consistent with the principles and novel features disclosed herein.

Claims

1. A method for acquiring cell images, characterized in that, This is applied to a cell image acquisition system, which includes a three-dimensional control platform, a bright field light source, a laser light source, an objective lens, a transparent carrier, and an imaging camera. The three-dimensional control platform includes a horizontal motion platform and a vertical axis motor. The vertical axis motor is fixed with the objective lens. The transparent carrier is placed on the horizontal motion platform. The transparent carrier contains multiple support positions for culturing cells. The bright field light source emits a beam that passes through the transparent carrier and the objective lens in sequence and is then received by the imaging camera. The laser light source is located between the objective lens and the transparent carrier. The method includes: Determine the initial focal plane height of the three-dimensional control platform for data acquisition from the transparent carrier; Using the initial focal plane height as the initial parameter for traversing the bearing positions, traverse each bearing position: based on the focusing images of each bearing position at multiple preset laser positions in the first horizontal direction, determine at least two laser focusing positions for that bearing position. Traverse each bearing position: Based on the position of the horizontal motion platform in the first horizontal direction when traversing to each bearing position, determine the target laser focusing position among at least two laser focusing positions of that bearing position; based on the target laser focusing position of each bearing position, acquire multiple cell images of that bearing position.

2. The method according to claim 1, characterized in that, The imaging camera is located vertically below the objective lens; Determining the initial focal plane height for the 3D control platform to acquire data from the transparent carrier includes: The initial bearing position of the transparent carrier is moved above the objective lens by the horizontal motion platform. With both the bright field light source and the imaging camera turned on, the vertical axis motor is adjusted until the surface image sharpness of the transparent carrier is greater than the sharpness threshold, and then the current initial focal plane height of the vertical axis motor is determined.

3. The method according to claim 2, characterized in that, Also includes: When the vertical axis motor is at the initial focal plane height, the laser light source is turned on, and the laser spot center position information of the light spot image acquired by the imaging camera is recorded.

4. The method according to claim 1, characterized in that, Using the initial focal plane height as the initial parameter for traversing the bearing positions, traverse each bearing position: based on the focusing images of multiple preset laser positions at each bearing position in the first-dimensional horizontal direction, determine at least two laser focusing positions for that bearing position, including: Using the initial focal plane height as the initial parameter for traversing the bearing positions, traverse each bearing position: For each traversed bearing position, when the horizontal motion platform moves along the first horizontal direction and moves to each preset laser position of the bearing position, the focus image at each preset laser position is acquired at the current height of the vertical axis motor; For each traversed carrier position, at least two laser focusing positions of the carrier position are determined based on the spot information of the focusing image at each preset laser position of the carrier position.

5. The method according to claim 4, characterized in that, Based on the spot information of the focusing image at each preset laser position of the bearing position, at least two laser focusing positions of the bearing position are determined, including: If there is only one focusing image containing spot information among the focusing images of the bearing position, then at least two laser focusing positions of the bearing position are determined, which are the height of the vertical axis motor when the focusing image is acquired. If none of the focusing images of the bearing position contain light spot information, then it is determined that at least two laser focusing positions of the bearing position are the current height of the vertical axis motor. If there are two or more focusing images containing spot information in each focusing image of the bearing position, then the first laser focusing position of the bearing position is determined to be the height of the vertical axis motor when the focusing image containing spot information is acquired first, and the second laser focusing position of the bearing position is determined to be the height of the vertical axis motor when the focusing image containing spot information is acquired last.

6. The method according to claim 5, characterized in that, After determining at least two laser focusing positions of the carrier position based on the spot information of the focusing image at each preset laser position of the carrier position, the method further includes: Adjust the vertical axis motor to the second laser focusing position.

7. The method according to claim 5, characterized in that, Based on the position of the horizontal motion platform in the first horizontal direction when traversing to each bearing position, determine the target laser focusing position among at least two laser focusing positions for that bearing position, including: For each traversed bearing position, when the horizontal motion platform is about to move along the second horizontal direction, the target laser focusing position is determined from at least two laser focusing positions of the bearing position based on the current position of the horizontal motion platform in the first horizontal direction, wherein the second horizontal direction, the first horizontal direction, and the vertical direction are mutually perpendicular.

8. The method according to claim 7, characterized in that, Based on the current position of the horizontal motion platform in the first horizontal direction, determine the target laser focusing position among at least two laser focusing positions of the bearing position, including: If the current position of the horizontal motion platform in the first horizontal direction is in the first half of the bearing position, then the target laser focusing position among the at least two laser focusing positions of the bearing position is determined as the first laser focusing position, wherein the first half of the bearing position is the half of the bearing position first passed through by the horizontal motion platform along the first horizontal direction through each preset laser position. If the current position of the horizontal motion platform in the first horizontal direction is in the second half of the bearing position, then the target laser focusing position among the at least two laser focusing positions of the bearing position is determined as the second laser focusing position, wherein the second half of the bearing position is the half of the bearing position that is passed through last during the process of the horizontal motion platform passing through each preset laser position in the first horizontal direction.

9. The method according to claim 5, characterized in that, The acquisition of multiple cell images for each carrier position based on the target laser focusing position includes: Adjust the vertical axis motor to the target laser focusing position; For each carrier position traversed, when the vertical axis motor is in the target laser focusing position, and the horizontal motion platform moves along the second horizontal direction and moves to each preset image acquisition position of the carrier position, the actions of turning on the bright field light source, taking pictures with the imaging camera, and turning off the bright field light source are executed in sequence to acquire cell images of the carrier position at each preset image acquisition position.

10. A cell image acquisition system, characterized in that, It includes a 3D control platform, a bright field light source, a laser light source, objective lenses, a transparent carrier, an imaging camera, and a controller; The three-dimensional control platform includes a horizontal motion platform and a vertical axis motor. The vertical axis motor is fixed with the objective lens. The transparent carrier is placed on the horizontal motion platform. The transparent carrier contains multiple support positions for culturing cells. The bright field light source emits a beam that passes through the transparent carrier and the objective lens in sequence and is then received by the imaging camera. The laser light source is located between the objective lens and the transparent carrier. The controller includes: A focal plane initialization unit is used to determine the initial focal plane height for the three-dimensional control platform to collect data from the transparent carrier. The laser focusing position determination unit is used to traverse each bearing position using the initial focal plane height as the initial parameter for bearing position traversal: based on the focusing images of at least two preset laser positions of each bearing position in the first horizontal direction, at least two laser focusing positions of that bearing position are determined. The acquisition and control unit is used to traverse each carrier position: based on the position of the horizontal motion platform in the first horizontal direction when traversing to each carrier position, it determines the target laser focusing position among at least two laser focusing positions of that carrier position; based on the target laser focusing position of each carrier position, it acquires multiple cell images of that carrier position.

Images