Delay integral acquisition for spatial genomics imaging

Abstract

An imaging system for capturing a spatiomic image of a biological tissue sample may include an imaging chamber configured to secure a biological tissue sample placed in the imaging system; a delay and integration (TDI) imager including at least one scan line; a light source configured to illuminate a region on the biological tissue sample being captured by the TDI imager; and a controller configured to cause the TDI imager to scan a biological tissue sample using one or more TDI scans of the biological tissue sample.

Description

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS

[0002] This application claims priority to U.S. Provisional Patent Application No. 63 / 393,458, filed on July 29, 2022, entitled “TIME DELAY INTEGRATION ACQUISITION FOR SPATIAL GENOMICS IMAGING,” the entire disclosure of which is incorporated herein by reference for all purposes as if fully set forth herein. Technical Field

[0003] The present disclosure generally describes capturing multiplexed, spatial-omic images of biological tissue samples. More particularly, the present disclosure describes camera and light configurations that reduce capture time of biological imaging systems. Background Art

[0004] Spatial biology is the study of cellular and subcellular environments across multiple dimensions. Spatial biology tools can be used to determine what cells are present in a tissue sample, where these cells are located in the tissue sample, the biomarker co-expression pattern of these cells, and how these cells are organized to interact with each other in the tissue sample. Sample slides can be prepared with tissue samples in various imaging workflows to produce comprehensive images of tissues at the cellular and subcellular levels, thereby producing single-cell resolution to visualize and quantify biomarker expression. The resulting images can expose how cells interact and organize within a tissue sample.

[0005] Capturing these complex images of a cell's environment can be called spatial omics. High-resolution, highly multiplexed spatial omics is rapidly becoming an important tool for understanding disease and other biological conditions. Typically, this type of analysis involves hundreds of complex factors, variables, and processes. Integral Solutions can combine imaging and process control methods into a single machine for performing spatial omics. Summary of the invention

[0006] In some embodiments, an imaging system for capturing spatial omics images of a biological tissue sample may include: an imaging chamber configured to hold a biological tissue sample placed in the imaging system; a time delay and integration (TDI) imager comprising at least one scan line; a controller configured to illuminate an area on the biological tissue sample being captured by the TDI imager; and a controller configured to cause the TDI imager to scan the biological tissue sample using one or more TDI scans of the biological tissue sample.

[0007] In some embodiments, a method of capturing a spatial omics image of a biological tissue sample may include: fixing the biological tissue sample in an imaging chamber of an imaging system; directing light from a light source to illuminate an area on the biological tissue sample; and causing a time delay and integration (TDI) imager to scan the biological tissue sample using one or more TDI scans of the biological tissue sample, wherein the biological tissue sample may be illuminated by the light from the light source.

[0008] In some embodiments, an imaging system for capturing a spatial omics image of a biological tissue sample may include: an imaging chamber configured to fix a biological tissue sample placed in the imaging system; a time delay and integration (TDI) imager; a controller configured to illuminate an area on the biological tissue sample being captured by the TDI imager with a plurality of different wavelengths; and a controller configured to cause the TDI imager to simultaneously capture a plurality of different images of the biological tissue sample. Each of the plurality of different images may correspond to one of the plurality of different wavelengths.

[0009] In any embodiment, any or all of the following features may be implemented in any combination without limitation. The controller may cause the TDI imager to scan a biological sample without mechanical repositioning, stabilization, or focusing between scan lines in a capture column. The light source may deliver light to the biological tissue sample through an optical fiber line that may project light onto the biological tissue sample in an approximately circular illumination area. The light source may deliver light to the biological tissue sample through a hole having a rectangular shape. The hole may project a rectangular illumination area on the biological sample onto a rectangular field of view of the TDI imager. The light source may deliver light to the biological tissue sample through a hole having a shape that matches the shape of the field of view of the TDI imager. The light source may deliver light to the biological tissue sample through an optical fiber line having an elliptical shape. The system may include an optical fiber bundle including a plurality of optical fiber lines, each of the plurality of optical fiber lines being configured to deliver a different wavelength. Each of the plurality of optical fiber lines may have an elliptical or rectangular shape. Each of the plurality of optical fiber lines may be positioned to direct light at different portions of the biological tissue sample. The system / method may assemble equally spaced lines from a plurality of scan line columns from a TDI imager. The light from the light source may include multiple different wavelengths that can be projected onto the biological tissue sample simultaneously. The system / method may filter the multiple different wavelengths using multiple filters in front of the TDI imager. The system / method may filter the multiple different wavelengths using a beam splitter in front of the TDI imager. The TDI imager includes multiple scan lines, and each of the multiple scan lines can capture one of multiple different images. The TDI image may include multiple separate TDI cameras, and each of the multiple separate TDI cameras can be aimed at different positions of the biological tissue sample to capture the multiple different images simultaneously, and the system may include a filter wheel in front of the TDI imager. The filter wheel may include multiple filters corresponding to multiple different wavelengths. The light source may include a wide spectrum light source that combines the multiple different wavelengths onto a single illumination area. BRIEF DESCRIPTION OF THE DRAWINGS

[0010] A further understanding of the nature and advantages of the various embodiments may be realized by reference to the remainder of the specification and drawings, wherein the same reference numerals are used throughout the several drawings to represent the same elements. In some cases, a sub-label is associated with a reference numeral to represent one of multiple similar elements. When a reference numeral is referenced without specifying the presence of a sub-label, the reference numeral is intended to represent all such multiple similar elements.

[0011] Figure 1 Schematic diagram of a high-resolution biological imaging system according to some embodiments.

[0012] Figure 2 A flow chart illustrating a process for capturing multiple sets of images of a sample according to some embodiments.

[0013] Figure 3 A TDI camera that may be used in an imaging system according to some embodiments is illustrated.

[0014] Figure 4A Illustration of an illumination area for a conventional camera according to some embodiments.

[0015] Figure 4B Illustration of the illumination area of ​​a TDI camera according to some embodiments.

[0016] Figure 4C Illustrated is a lighting area where holes are utilized to minimize unused light areas according to some embodiments.

[0017] Figure 4D Illustrated is an illumination area where unused light area is minimized by the shape of the laser beam according to some embodiments.

[0018] Figure 5 Illustrate how a conventional camera system may capture multiple images illuminated by different wavelengths in accordance with some embodiments.

[0019] Figure 6 An imaging system configuration that allows for the capture of multiple wavelengths during a single pass through a tissue sample is illustrated in accordance with some embodiments.

[0020] Figure 7 Different configurations of filters for simultaneous wavelength capture are illustrated in accordance with some embodiments.

[0021] Figure 8 Illustration of a configuration using a beam splitter to separate images of different wavelengths.

[0022] Fig. 9 Flowchart illustrating a method of capturing a spatial omics image of a biological tissue sample.

[0023] Fig.10 An exemplary computer system is illustrated in which various embodiments may be implemented. DETAILED DESCRIPTION

[0024] Figure 1A high-resolution biological imaging system 100 is illustrated according to some embodiments. The imaging system 100 can be configured to combine multiple imaging workflows together into a single process to perform automated spatial analysis of tissue samples. The imaging system 100 can include multiple imaging chambers 108, 110, each of which can be configured to perform separate imaging operations on different tissue samples. The fluid system 102 can provide integrated fluid control to provide multiple fluorophores and / or other fluids to the imaging chambers 108, 110 during the imaging process. Different fluorophores and reagents can be loaded into containers in the fluid system 102 so that these fluids can be automatically provided to the imaging chambers 108, 110 when needed during the imaging process.

[0025] The imaging system 100 may include: a computer system including one or more processors; one or more memory devices; and instructions stored on the one or more memory devices, which instructions cause the imaging system 100 to perform imaging operations on tissue samples in the imaging chambers 108, 110. Therefore, each operation of the imaging process described herein may be represented by instructions stored on the one or more memory devices.

[0026] In an exemplary imaging workflow, a user or an automated process may load a tissue sample onto a slide and load the slide into the imaging chamber 108. After the tissue sample is secured in the imaging chamber 108, fluids may then be automatically pumped into the imaging chamber 108. For example, some fluid may be pumped into the imaging chamber 108 to clean the tissue and / or remove previous fluids or fluorophores that may be present within the imaging chamber 108. New fluids or fluorophores may be provided from the fluid system 102 in an automated manner as specified by instructions executed by the controller. During a typical cycle, one or more fluorophores may be pumped into the imaging chamber 108 that are configured to attach to cells in the tissue sample in order to visually highlight different features within the sample. Subsequently, the sample in the imaging chamber 108 may be illuminated using a corresponding laser wavelength, and a camera may capture an image of the illuminated sample. Fluorophores may be matched to different laser wavelengths that are configured to illuminate those specific fluorophores.

[0027] After the imaging process is complete, the controller can convert the raw images from the system into RNA spots. These RNA spots can be visualized as clusters of cell types highlighted by different fluorophores. Subsequently, multiple images can be merged for multi-omics analysis of tissue samples. The software tools provided by the controller of the imaging system 100 can provide different visualization, data filtering and analysis tools for viewing images.

[0028] Figure 2A flowchart 200 is illustrated of a process for capturing a multi-omics image of a sample according to some embodiments. As described above, the process may include loading a tissue sample on a substrate, such as a coverslip or a slide, and securing the slide within one of the imaging chambers of the imaging system (202). It should be noted that multiple imaging chambers may be operated independently and simultaneously in the same imaging system 100. For example, one imaging chamber 108 may capture an image of a sample while another imaging chamber 110 may exchange fluids in the imaging chamber 110. Subsequently, the imaging system 100 may provide fluids from the fluid system 102 to the imaging chamber 108 (204). Such fluids may include fluorophores that are configured to attach to specific cells or tissue types to be highlighted in the resulting image.

[0029] In order to capture images with a high enough resolution to visualize individual cells in detail, images of the sample can be captured in stages. For example, instead of capturing a single image of the sample, the field of view of the camera in the imaging system 100 can be reduced to increase the resolution. Subsequently, multiple images of the sample can be captured, and the multiple images can be stitched together to form an overall image. For example, the overall image 250 may include multiple sub-images that can be captured by the camera at a high resolution. Each of these images can correspond to the field of view of the image. Therefore, the process can include incrementally capturing field of view images using a camera (206), then moving the camera to a subsequent position with an adjacent field of view, and preparing the camera for a subsequent stage (208). The process can be repeated until the overall image 250 of the sample has been captured by a separate field of view image.

[0030] To capture the overall image 250, the camera may move its field of view in a pattern over the tissue sample. For example, a first field of view 252 (206) may be captured, and then the camera may move in a grid pattern to a second field of view 254 (208) that is optionally adjacent to the first field of view 252. This process may be repeated for each field of view in the sample until the overall image 250 has been captured. Note that Figure 2 The grid pattern shown in is provided by way of example only and is not meant to be limiting. Other embodiments may move horizontally, vertically, diagonally, and / or in any other pattern that may be used to capture individual field of view images that may be combined into overall image 250.

[0031] Multiple whole-body images 250 of a tissue sample may be captured in order to highlight different features in the tissue sample for overall multi-omics analysis. Thus, after capturing a whole-body image 250 for a particular fluorophore or set of fluorophores, the process may be repeated on the same tissue sample using another fluorophore or set of fluorophores. For example, the previous fluorophore may be pumped out of the imaging chamber 108, a cleaning agent or rinse may optionally be pumped through the imaging chamber 108 to clean the tissue sample, and a new set of fluorophores may be pumped into the imaging chamber 108 for the next image (204). Each whole-body image captured using different fluorophores to be combined in a multi-omics analysis may be referred to as an "imaging cycle" or "hyb". Typically, each sample may undergo multiple hybs using different fluorophores. For example, some embodiments may capture two, three, four, five, six, or more whole-body images of a sample, thereby repeating the cycle (204) multiple times. When the desired number of sample images have been captured, the sample may be removed from the imaging chamber 108 (210). Subsequently, a new sample may be added to the imaging chamber 108 (202), and the imaging process may be repeated.

[0032] Although the process does provide multiple sets of image data at high resolution, the process also takes a considerable amount of time. At each field of view image position, the sample can be illuminated by multiple different laser wavelengths (e.g., different colors configured to illuminate different fluorophores in the sample), and thus multiple images can be captured at different wavelengths at each position. Additionally, the sample itself can be adjusted axially to capture multiple images at different Z depth levels, thereby generating a three-dimensional image slice through the tissue sample. After capturing the image at each field of view position, the process may include moving the sample laterally so that the camera captures a new field of view position, which may require time to acquire a new image, physically move the sample using a piezoelectric motor or a segmented motor, configure the laser and corresponding filter for the desired wavelength, stabilize and focus the camera, and / or other operations. Combining these different factors together can result in a relatively large overall imaging time. For example, in an exemplary implementation using a camera with a 40x objective and a 30×30 field of view image grid to cover the sample, each hyb may take approximately 10 hours to complete. A typical four hyb sessions may take a total of 30 to 40 hours to complete. While reducing the resolution of the camera will increase the field of view and reduce the total number of field images required, doing so will also negatively impact the quality of the resulting image. This significant time requirement represents a technical problem in the field of biospatial omics.

[0033] To address these and other technical issues, embodiments described herein may use a number of technical improvements in an imaging system to greatly reduce overall imaging time. A Time Delay Integration (TDI) camera may be used to continuously scan a tissue sample in columns rather than moving between different fields of view. The laser beam projected onto the imaged sample may be shaped to approximately match a TDI image scan line. Some embodiments may simultaneously illuminate a tissue sample at different locations with different wavelengths captured by separate TDI cameras or portions of a single TDI camera. As described below, these improvements (whether alone or in combination) are used to significantly improve the imaging time of an imaging system.

[0034] Figure 3 FIG. 3 shows a TDI camera 302 that can be used in an imaging system according to some embodiments. The TDI camera 302 may include a charge-coupled device (CCD) as an image sensor for capturing an image. For example, the TDI camera 302 may include a CCD. Figure 3 304 of individual CCD pixels in a horizontal configuration. Note that for clarity, only a single row of pixels is shown in the scan line 304. However, this is not meant to be limiting. As discussed and illustrated below, the TDI camera 302 may include multiple horizontal rows of pixels.

[0035] The operation of the TDI camera 302 can be contrasted with the operation of the conventional camera described above. As described above, the conventional camera can capture a single field of view and then move to another non-overlapping field of view before capturing the next image. Simply return to Figure 2 , field of view 252 may include a horizontal grid of individual pixels within field of view 202, and when acquiring a camera shot, each of the individual pixels will capture an image simultaneously. In contrast, TDI camera 302 may use scan lines 304 of individual pixels. TDI camera 302 may sequentially and continuously scan an image in a vertical direction. The entire image may then be assembled from equally spaced lines through the linear field of view of scan lines 304. It should be noted that the terms "horizontal" and "vertical" are only used to indicate that the image is horizontally and vertically spaced. Figure 3 The orthogonal directions shown in are not meant to be limited to a particular direction.

[0036] The scan line 304 need not extend the entire horizontal length of the image. Instead, multiple vertical "columns" may be captured using multiple vertical sequential scans. For example, to capture the entire image 350, the scan line 352 may continuously scan down a first vertical column 362 of the imaging area. When the scan of the first vertical column 362 is complete, the scan line 352 of the TDI camera may be repositioned on a second vertical column 364, and the scan line 352 may then continuously scan down the second vertical column 364. These vertical columns may be stitched together to form the entire image 350 of the tissue sample.

[0037] The use of the TDI camera 302 represents a significant technical improvement over other cameras in scanning tissue samples. The TDI camera 302 can continuously capture each vertical scan column, thus eliminating the need to mechanically reposition the sample, stabilize, focus, and prepare for each individual field of view capture. Instead, the TDI camera 302 can move at a constant speed in the vertical direction and scan continuously to accumulate reflected light or fluorescent signals from the tissue sample. The only repositioning of the TDI camera 302 that needs to occur may be between each of the vertical column captures. The use of the TDI camera 302 thus increases the speed at which an image of a tissue sample can be fully captured, reducing the total time from more than 35 hours to approximately 5 hours in combination with the other improvements described below.

[0038] Figure 4A An illumination area 402 for a conventional camera according to some embodiments is illustrated. As described above, a conventional camera can capture a separate field of view 404 of a tissue sample. Within the field of view 404, a plurality of individual pixels arranged in a rectangular grid can each simultaneously capture a portion of the field of view 404. Prior to capturing an image, the tissue sample can be illuminated by using a laser of a specified wavelength. The wavelength can be configured to illuminate certain types of fluorophores present on the tissue sample. Light from the laser can be transmitted from the laser to an imaging chamber via an optical fiber line. Traditionally, the optical fiber line has a circular shape so that the output of the light directed onto the tissue sample can be in an approximately circular shape. As shown in FIG. 4 , this can produce an approximately circular illumination area 402 to surround the field of view 404 of the camera.

[0039] Because the circular shape of illumination area 402 does not exactly match the rectangular shape of field of view 404, there may be unused light areas 406 around the sides of field of view 404. However, compared to the size of field of view 404 and illumination area 402 as a whole, unused light areas 406 are relatively small.

[0040] Figure 4B 4 shows an illumination area 412 of a TDI camera according to some embodiments. As described above, the scan line 414 may include Figure 4B414. However, it should be noted that the shape of the illumination area 412 still corresponds to the circular output of a conventional fiber optic line that delivers laser light to a tissue sample. Because the shape of the illumination area 412 is quite large and different from the shape of the scan line 414, the unused light area 416 is much larger than when using a conventional camera. It has been found that in some cases, overexposing the tissue sample with excess light in the unused light area 416 may be harmful. For example, fluorophores in the unused light area 416 may be damaged, causing the image of those fluorophores to be washed out or have lower contrast. In addition, when the scan line 414 is used to capture an image, reflections of light from the unused light area 416 may be disruptive. Therefore, some embodiments may be further configured to minimize the unused light area 416 using the methods described below.

[0041] Figure 4C An illumination area 422 is illustrated in which an aperture is utilized to minimize unused light area 426 in accordance with some embodiments. This configuration continues to use the circular light output from a conventional fiber optic line. However, instead of outputting an entire circular light pattern onto a tissue sample, an aperture 428 may be used to shape the light output. In this example, the aperture may block the top and / or bottom of the output of the fiber optic line to make the illumination area 422 more rectangular in shape. In general, this embodiment may thus be characterized as providing light to a tissue sample in a shape that is substantially similar to the shape of a scan line 424 of a TDI camera, which in this particular example is rectangular in shape. It should be noted that other camera fields of view may use different shapes, so this example is not intended to be limiting.

[0042] While the general shape of the illumination area 422 on the tissue sample can be configured to generally match the shape of the scan line 424 of the TDI camera, some embodiments may include a buffer region of light around the scan line 424. Figure 4C In the example of FIG. 4 , an unused light region 426 still exists around the scan line 424. While some embodiments may minimize this unused light region 426, other embodiments may allow for the unused light region 426 so that the illumination area 422 is larger than the area of ​​the scan line 424. For example, the unused light region 426 may be larger than the area of ​​the scan line 424 by about 5% to 10%, about 10% to 15%, about 15% to 20%, about 20% to 25%, about 25% to 30%, about 30% to 35%, about 35% to 40%, about 40% to 45%, about 45% to 40%, about 50% to 55%, about 55% to 60%, about 60% to 65%, about 65% to 70%, about 70% to 75%, about 75% to 100%, or greater than 100%.

[0043] Figure 4DThe illustration shows an illumination area 432 in which the unused light area 436 is minimized by the shape of the fiber optic line according to some embodiments. As an alternative to using a circular fiber optic line, some embodiments may shape the fiber optic line to be more oval or rectangular to better match the shape of the scan line 334. In this example, as shown in FIG. Figure 4D As shown in , the elliptical fiber line can produce an illumination area 432 that more accurately matches the shape of the scan line 334. This configuration minimizes the unused light area 436 around the scan line 334, while also minimizing the wasted light output of the laser. Figure 4C Instead of blocking the light output through the holes shown in , shaping the fiber optic line can focus the light output onto the area of ​​scan line 334.

[0044] In addition to using a TDI camera and shaping the illumination area to match the scan lines of the TDI camera, some embodiments may further increase the throughput of the imaging system by optimizing how multiple images of the same area can be captured using different illumination wavelengths.

[0045] Figure 5 508. The conventional camera system according to some embodiments can capture multiple images illuminated by different wavelengths. In this example, each area of ​​the tissue sample can be illuminated by four different wavelengths. A first laser beam 502 can illuminate an area of ​​a tissue sample 504 with a first wavelength 503. Reflected light or fluorescence from the first laser beam 502 can pass through a first filter 506, which is configured to pass a first wavelength range 507, before being recorded by a camera 508. It should be noted that the first wavelength range 507 can be slightly higher than the first wavelength 503 from the first laser beam 502. Similarly, a second laser beam 512 can illuminate an area of ​​a tissue sample 504 with a second wavelength 513. Reflected light or fluorescence from the second laser beam 512 can pass through a second filter 516, which is configured to pass a second wavelength range 517, before being recorded by the camera 508. A third laser beam 522 can illuminate an area of ​​a tissue sample 504 with a third wavelength 523. Reflected light or fluorescence from the third laser beam 522 may pass through a third filter 526 configured to pass a third wavelength range 527 before being recorded by the camera 508. The fourth laser beam 532 may illuminate an area of ​​the tissue sample 504 with a fourth wavelength 533. Reflected light or fluorescence from the fourth laser beam 532 may pass through a fourth filter 536 configured to pass a fourth wavelength range 537 before being recorded by the camera 508. The images captured by the camera 508 by each laser beam may be stitched together to form a complete image of the tissue sample, each image illuminated by a different wavelength.

[0046] Typically, a complete set of field of view images is captured for each wavelength at each field of view position before moving to the next position. Between capturing images for each wavelength, time is required to change the filter wheel, fix the filter wheel, move the motor to account for wavelength-dependent focal plane shifts, etc. Therefore, each additional desired wavelength significantly increases the total time used to image the tissue sample. The embodiments described herein improve this process by using a camera and tissue sample to capture multiple wavelengths at once.

[0047] Figure 6 An imaging system configuration 600 is shown that allows for the capture of multiple wavelengths during a single pass through a tissue sample in accordance with some embodiments. Instead of a single fiber line that transmits a single wavelength, a fiber bundle 602 may transmit four separate fiber lines 603. Each of the separate fiber lines 603 may be shaped as an ellipse, rectangle, or other shape that is configured to match the size / shape of a corresponding scan line of a TDI camera. Figure 6 In the example of FIG. 5 , the individual fiber optic lines 603 may have an elliptical shape and may be stacked in a vertical configuration within the fiber optic bundle 602 so that the corresponding illumination areas on the tissue sample are vertically aligned, such as Figure 6 Alternatively, instead of shaping the fiber optic line itself, some embodiments may shape the output beam into a circular, square, or any other shape of the fiber optic line. For example, the output beam may be shaped into an elliptical shape before or after directing the light into or out of the fiber optic line.

[0048] Each of the fiber optic lines 603 can individually transmit a different wavelength corresponding to a different fluorophore in the tissue sample. Some embodiments can illuminate the tissue sample continuously during the TDI image acquisition process. For example, four illumination areas corresponding to the fiber optic lines 603 can be illuminated continuously and scanned vertically downwardly to the tissue sample 604 together with the corresponding scan lines of the TDI camera. The filter 606 can also be positioned and configured to filter out a portion of the reflected light or fluorescence of the tissue sample 604 based on the wavelength of the fiber optic line 603. An exemplary configuration of the filter 606 is described in detail below.

[0049] The system may also include a TDI camera 608 having four TDI sensors. For example, the TDI camera 608 may include Figure 6 608. Each of the scan lines can be configured to scan an area on the tissue sample 604 that is illuminated by a corresponding light from one of the fiber optic lines 603. Thus, each of the fiber optic lines 603 can be aligned with and correspond to a separate scan line in the scan lines 609 of the TDI camera 608. The filter 606 can also include a plurality of separate filters that are aligned with the optical path between the fiber optic lines 603 and the scan lines 609 of the TDI camera 608.

[0050] These embodiments allow for the simultaneous capture of multiple images, each with a different wavelength, during each vertical column scan of the tissue sample. As four separate passes through the tissue sample (one for each wavelength) are used, these embodiments can capture any or all desired wavelengths in a single pass. For four different wavelengths, this can reduce the total time by up to 4 times.

[0051] although Figure 6 Individual TDI camera scan lines are illustrated that may be physically and individually separated from one another, but other embodiments may use a single TDI scan line in which individual pixel rows are subdivided. For example, a single TDI imager may include a set of horizontal scan lines. However, the filter 606 and the fiber optic line 603 may be configured to reflect light onto different areas of a set pixel row in the scan line. For example, 3 to 4 rows of pixels on a TDI camera may be assigned to one wavelength, while the next 3 to 4 rows of pixels on the TDI camera may be assigned to the next wavelength, and so on. The software operating the TDI camera 608 may then subdivide these pixel rows when processing the image. Thus, as shown in FIG. Figure 6 Instead of physically separating the scan lines on the TDI camera 608 as shown in FIG, some embodiments may alternatively perform such separation of the different wavelength images in software.

[0052] Figure 7 Different configurations of filters for simultaneous wavelength capture according to some embodiments are illustrated. In a first configuration 704, individual filters may be placed on the scan lines of the TDI camera itself. As described above, the TDI camera may include a different scan line for each different wavelength. As part of the camera construction, for each wavelength, the filter may be placed adjacent to the individual scan line pixels. The filter may be integrated onto the camera using a custom CMOS image sensor.

[0053] In a second configuration 706, the individual filters may be fabricated together as a cover glass placed on the imager. For example, a cover may be placed on a TDI imager so that the area on the cover glass for each filter will be in front of a corresponding scan line of the TDI imager.

[0054] The third configuration 708 may place the filters together on a "butcher block" filter at a location corresponding to an intermediate focal plane of the TDI imager. In some embodiments, the third configuration 708 may be configured as a filter wheel.

[0055] Figure 8The configuration using a beam splitter to separate different wavelength images is shown. Instead of using four different laser wavelengths to illuminate different areas on the tissue sample, some embodiments can illuminate the tissue sample using a broad spectrum light that combines each desired wavelength into a single illumination area on the tissue sample. The reflected light or fluorescence 800 from the tissue sample can then pass through a beam splitter 802. The beam splitter 802 can then separate the different wavelengths in the reflected light or fluorescence 802 onto separate TDI imagers 806, 808, 810.

[0056] Fig. 9 Flowchart 900 illustrating a method of capturing a spatial omics image of a biological tissue sample. The method may include securing the biological tissue sample in an imaging chamber of an imaging system (902). The method may also include directing light from a light source to illuminate an area on the biological tissue sample (904). The method may further include causing a TDI imager to scan the biological tissue sample using one or more TDI scans of the biological tissue sample, wherein the biological tissue sample is illuminated by light from the light source (906). Each of these operations may be performed as described in detail above.

[0057] It should be understood that Fig. 9 The specific steps shown in provide specific methods for capturing spatial omics images of biological tissue samples according to various embodiments. According to alternative embodiments, other sequences of steps may also be performed. For example, alternative embodiments may perform the steps outlined above in a different order. In addition, Fig. 9 The individual steps shown in the embodiment may include multiple sub-steps, which may be performed in various orders suitable for the individual steps. In addition, additional steps may be added or removed depending on the specific application. Many variations, modifications and alternatives also fall within the scope of this case.

[0058] Each of the methods described herein can be implemented by a computer system. Each step of these methods can be automatically performed by a computer system, and / or can have input / output involving a user. For example, a user can provide input for each step in the method, and each of these inputs can be a response to a specific output requesting the input, wherein the output is generated by a computer system. Each input can be received in response to a corresponding request output. In addition, input can be received from a user, received as a data stream from another computer system, retrieved from a memory location, retrieved via a network, requested from a network service, and the like. Similarly, the output can be provided to a user as a data stream, provided to another computer system, stored in a memory location, sent via a network, provided to a network service, and the like. In short, each step of the method described herein can be performed by a computer system, and can involve any number of inputs, outputs, and / or requests from a computer system of a computer system, which can involve or not involve a user. Those steps that do not involve a user can be considered to be automatically performed by a computer system without human intervention. Therefore, it should be understood from the present disclosure that each step of each method described herein can be changed to include input and output to and from the user, or can be automatically completed by the computer system without human intervention, wherein any determination is made by the processor. In addition, some embodiments of each method described herein can be implemented as an instruction set stored on a tangible non-transitory storage medium to form a tangible software product.

[0059] Fig.10 An exemplary computer system 1000 in which various embodiments may be implemented is shown. System 1000 may be used to implement any of the above-described computer systems. As shown in the figure, computer system 1000 includes a processing unit 1004, which communicates with multiple peripheral subsystems via a bus subsystem 1002. These peripheral subsystems may include a processing acceleration unit 1006, an I / O subsystem 1008, a storage subsystem 1018, and a communication subsystem 1024. Storage subsystem 1018 includes a tangible computer-readable storage medium 1022 and a system memory 1010.

[0060] The bus subsystem 1002 provides a mechanism for enabling the various components and subsystems of the computer system 1000 to communicate with each other as expected. Although the bus subsystem 1002 is schematically shown as a single bus, alternative embodiments of the bus subsystem may utilize multiple buses. The bus subsystem 1002 may be any of several types of bus structures, including a memory bus or memory controller, a peripheral bus, and a local bus using any of a variety of bus architectures. For example, these architectures may include an Industry Standard Architecture (ISA) bus, a Micro Channel Architecture (MCA) bus, an Enhanced ISA (EISA) bus, a Video Electronics Standards Association (VESA) regional bus, and a Peripheral Component Interconnect (PCI) bus, each of which may be implemented as a mezzanine bus manufactured according to the IEEE P1386.1 standard.

[0061] The processing unit 1004 may be implemented as one or more integrated circuits (e.g., conventional microprocessors or microcontrollers) and controls the operation of the computer system 1000. One or more processors may be included in the processing unit 1004. These processors may include single-core or multi-core processors. In some embodiments, the processing unit 1004 may be configured as one or more independent processing units 1032 and / or 1034, each of which includes a single-core or multi-core processor. In other embodiments, the processing unit 1004 may also be implemented as a quad-core processing unit formed by integrating two dual-core processors into a single chip.

[0062] In various embodiments, the processing unit 1004 may execute various programs in response to the code, and may maintain multiple concurrently executing programs or processes. At any given time, some or all of the code to be executed may reside in the processor 1004 and / or the storage subsystem 1018. Through appropriate programming, the processor 1004 may provide various functions as described above. The computer system 1000 may additionally include a processing acceleration unit 1006, which may include a digital signal processor (DSP), a dedicated processor, etc.

[0063] I / O subsystem 1008 may include user interface input devices and user interface output devices. User interface input devices may include keyboards, pointing devices such as mice or trackballs, touch pads or touch screens incorporated into displays, scroll wheels, click wheels, dials, buttons, switches, keyboards, audio input devices with voice command recognition systems, microphones, and other types of input devices. User interface input devices may include, for example, motion sensing and / or gesture recognition devices, such as Microsoft Kinect® motion sensors, which enable users to control input devices such as Microsoft Xbox® 360 game controllers and interact with the input devices via a natural user interface using gestures and spoken commands. User interface input devices may also include eye movement recognition devices, such as Google Glass® blink detectors, which detect eye movements of the user (e.g., "blinking" when taking pictures and / or making menu selections) and convert eye movements as input to the input device (e.g., such as Google Glass®). In addition, user interface input devices may include voice recognition sensing devices, which enable users to interact with voice recognition systems (e.g., Siri® navigator) via voice commands.

[0064] User interface input devices may also include, but are not limited to, three-dimensional (3D) mice, joysticks or pointing sticks, game controllers, and drawing tablets, as well as audio / visual devices such as speakers, digital cameras, digital video recorders, portable media players, network monitors, image scanners, fingerprint scanners, barcode readers, 3D scanners, 3D printers, laser rangefinders, and eye gaze tracking devices. Additionally, user interface input devices may include, for example, medical imaging input devices such as computed tomography scanners, magnetic resonance imaging, positron emission tomography, and medical ultrasound devices. User interface input devices may also include, for example, audio input devices such as MIDI keyboards, digital musical instruments, and the like.

[0065] The user interface output device may include a display subsystem, indicator lights, or non-visual displays such as audio output devices. The display subsystem may be a cathode ray tube (CRT), a flat panel device such as a flat panel device using a liquid crystal display (LCD) or a plasma display, a projection device, a touch screen, etc. In general, the use of the term "output device" is intended to include all possible types of devices and mechanisms for outputting information from the computer system 1000 to a user or other computers. For example, the user interface output device may include, but is not limited to, various display devices that visually convey text, graphics, and audio / video information, such as monitors, printers, speakers, headphones, car navigation systems, plotters, voice output devices, and modems.

[0066] Computer system 1000 may include storage subsystem 1018, which includes software elements shown as currently located within system memory 1010. System memory 1010 may store program instructions that may be loaded and executed on processing unit 1004, as well as data generated during the execution of such programs.

[0067] Depending on the configuration and type of computer system 1000, system memory 1010 may be volatile (such as random access memory (RAM)) and / or non-volatile (such as read-only memory (ROM), flash memory, etc.). RAM typically contains data and / or program modules that are immediately accessible to and / or currently being operated on and executed by processing unit 1004. In some implementations, system memory 1010 may include a variety of different types of memory, such as static random access memory (SRAM) or dynamic random access memory (DRAM). In some implementations, a basic input / output system (BIOS) containing basic routines that help transfer information between elements within computer system 1000, such as during startup, may typically be stored in ROM. By way of example and not limitation, system memory 1010 also illustrates application programs 1012 (application programs 1012 may include client applications, web browsers, middle-tier applications, relational database management systems (RDBMS), etc.), program data 1014, and operating system 1016. For example, operating system 1016 may include various versions of Microsoft Windows®, Apple Macintosh®, and / or Linux operating systems, various commercially available UNIX® or UNIX-like operating systems (including, but not limited to, various GNU / Linux operating systems, Google Chrome® OS, etc.), and / or mobile operating systems such as iOS, Windows® Phone, Android® operating system, BlackBerry® 10 operating system, and Palm® operating system.

[0068] Storage subsystem 1018 may also provide a tangible computer-readable storage medium for storing basic programming and data structures that provide the functionality of some embodiments. Software (programs, code modules, instructions) that provide the above-mentioned functionality when executed by a processor may be stored in storage subsystem 1018. These software modules or instructions may be executed by processing unit 1004. Storage subsystem 1018 may also provide a repository for storing data used in accordance with some embodiments.

[0069] Storage subsystem 1000 may also include a computer-readable storage media reader 1020, which may be further connected to computer-readable storage media 1022. Computer-readable storage media 1022 (together with and optionally in conjunction with system memory 1010) may comprehensively represent remote, local, fixed and / or removable storage devices, as well as storage media for temporarily and / or more permanently containing, storing, transferring, and retrieving computer-readable information.

[0070] The computer-readable storage medium 1022 containing the code or portions of the code may also include any suitable media, including storage media and communication media, such as, but not limited to, volatile and non-volatile, removable and non-removable media implemented in any method or technology for storing and / or transmitting information. The media may include tangible computer-readable storage media, such as RAM, ROM, electronically erasable programmable ROM (EEPROM), flash memory or other memory technology, compact disk-read only memory (CD-ROM), digital versatile disk (DVD) or other optical storage, magnetic card, tape, magnetic disk storage or other magnetic storage device, or other tangible computer-readable medium. The media may also include non-tangible computer-readable media, such as data signals, data transmissions, or any other medium that can be used to transmit the desired information and can be accessed by the computing system 1000.

[0071] For example, the computer-readable storage medium 1022 may include a hard disk drive that reads from or writes to a non-removable, non-volatile magnetic medium; a magnetic disk drive that reads from or writes to a removable, non-volatile magnetic disk; and an optical disk drive that reads from or writes to a removable, non-volatile optical disk (such as a CD ROM, a DVD, and a Blu-Ray® disc) or other optical media. The computer-readable storage medium 1022 may include, but is not limited to, a Zip® drive, a flash memory card, a universal serial bus (USB) flash drive, a secure digital (SD) card, a DVD disk, a digital video tape, and the like. The computer-readable storage medium 1022 may also include a solid-state drive (SSD) based on non-volatile memory, such as a flash memory-based SSD, an enterprise flash drive, a solid-state ROM, and the like; a volatile memory-based SSD, such as a solid-state RAM, a dynamic RAM, a static RAM; a DRAM-based SSD, a magnetoresistive RAM (MRAM) SSD, and a hybrid SSD using a combination of DRAM and a flash memory-based SSD. The disk drives and their associated computer-readable media provide nonvolatile storage of computer-readable instructions, data structures, program modules and other data for the computer system 1000 .

[0072] The communication subsystem 1024 provides an interface to other computer systems and networks. The communication subsystem 1024 is used as an interface for receiving data from the computer system 1000 and sending data from the computer system to other systems. For example, the communication subsystem 1024 can enable the computer system 1000 to be connected to one or more devices via the Internet. In some embodiments, the communication subsystem 1024 may include a radio frequency (RF) transceiver element for accessing a wireless voice and / or data network (e.g., using cellular phone technology, such as 3G, 4G or EDGE (Enhanced Data Rates for Global Evolution) advanced data network technology), WiFi (IEEE 802.11 family standards, or other mobile communication technologies, or any combination thereof), a global positioning system (GPS) receiver element, and / or other elements. In some embodiments, in addition to or in lieu of a wireless interface, the communication subsystem 1024 may provide a wired network connection (e.g., Ethernet).

[0073] In some implementations, the communication subsystem 1024 may also receive incoming communications in the form of structured and / or unstructured data feeds 1026 , event streams 1028 , event updates 1030 , etc., on behalf of one or more users who may use the computer system 1000 .

[0074] For example, the communication subsystem 1024 may be configured to receive data feeds 1026 in real time from users of social networks and / or other communication services, such as Twitter® feeds, Facebook® updates, web feeds (such as Rich Site Summary (RSS) feeds), and / or real-time updates from one or more third-party information sources.

[0075] Additionally, the communication subsystem 1024 may also be configured to receive data in the form of a continuous data stream, which may include an event stream 1028 of real-time events and / or event updates 1030, which may be continuous or endless in nature and without a clear end. Examples of applications that generate continuous data may include, for example, sensor data applications, financial quoters, network performance measurement tools (e.g., network monitoring and traffic management applications), clickstream analysis tools, automobile traffic monitoring, etc.

[0076] The communication subsystem 1024 may also be configured to output structured and / or unstructured data feeds 1026 , event streams 1028 , event updates 1030 , etc. to one or more databases that may be in computer communication with one or more streaming data source computers coupled to the computer system 1000 .

[0077] Computer system 1000 can be one of various types, including a handheld portable device (e.g., an iPhone® cellular phone, an iPad® tablet computer, a PDA), a wearable device (e.g., a Google Glass® head-mounted display), a PC, a workstation, a mainframe, a kiosk, a server rack, or any other data processing system.

[0078] Due to the ever-changing nature of computers and networks, the description of the computer system 1000 shown in the figure is only used as a specific example. Many other configurations with more or fewer elements than the system shown in the figure are also possible. For example, customized hardware can also be used and / or specific elements can be implemented by hardware, firmware, software (including applets) or a combination thereof. In addition, connections with other computing devices such as network input / output devices can be adopted. Based on the disclosure and teachings provided herein, other ways and / or methods of implementing various embodiments should be obvious.

[0079] As used herein, the term "about" or "approximately" or "substantially" may be interpreted as being within the range expected by a person of ordinary skill in the art based on the specification.

[0080] In the following description, for the purpose of explanation, numerous specific details are set forth to provide a thorough understanding of various embodiments. However, it is apparent that some embodiments may be practiced without these specific details. In other cases, well-known structures and devices are shown in the form of block diagrams.

[0081] The foregoing description provides only exemplary embodiments and is not intended to limit the scope, applicability or configuration of the present disclosure. On the contrary, the foregoing description of various embodiments will provide a feasible disclosure for implementing at least one embodiment. It should be understood that various changes may be made to the function and arrangement of elements without departing from the spirit and scope of some embodiments set forth in the appended claims.

[0082] Specific details are given in the foregoing description to provide a thorough understanding of the embodiments. However, it should be understood that the embodiments may be practiced without these specific details. For example, circuits, systems, networks, processes, and other elements may have been shown as elements in block diagram form so as not to obscure the embodiments with unnecessary details. In other cases, well-known circuits, processes, algorithms, structures, and techniques may be shown without unnecessary details in order to avoid obscuring the embodiments.

[0083] In addition, it should be noted that various embodiments may be described as processes, which are shown as flow charts, flow charts, data flow diagrams, structure diagrams, or block diagrams. Although a flow chart may describe operations as a sequential process, many operations may be performed in parallel or concurrently. In addition, the order of the operations may be rearranged. A process terminates when its operations are completed, but may have other steps not included in the accompanying drawings. A process may correspond to a method, function, procedure, subroutine, subprogram, etc. When a process corresponds to a function, its termination may correspond to the function returning to the calling function or the main function.

[0084] The term "computer-readable medium" includes, but is not limited to, portable or fixed storage devices, optical storage devices, wireless channels, and various other media capable of storing, containing, or carrying instructions and / or data. A code segment or machine executable instruction may represent any combination of a process, function, subroutine, program, routine, subroutine, module, software package, class, or instruction, data structure, or program statement. A code segment may be coupled to another code segment or hardware circuit by passing and / or receiving information, data, actual parameters, parameters, or memory contents. Information, actual parameters, parameters, data, etc. may be passed, forwarded, or transmitted via any suitable means, including memory sharing, information passing, token passing, network transmission, etc.

[0085] In addition, the embodiments may be implemented by hardware, software, firmware, middleware, microcode, hardware description language, or any combination thereof. When implemented in software, firmware, middleware, or microcode, the code or code segments for performing the necessary tasks may be stored in a machine-readable medium. The processor may perform the necessary tasks.

[0086] In the foregoing description, features are described with reference to specific embodiments thereof, but it should be appreciated that not all embodiments are limited thereto. Various features and aspects of some embodiments may be used alone or in combination. Furthermore, the embodiments may be used in any number of environments and applications beyond those described herein without departing from the broader spirit and scope of the specification. Accordingly, the description and drawings are to be regarded as illustrative rather than restrictive.

[0087] In addition, for ease of explanation, the method is described in a specific order. It should be understood that in alternative embodiments, these methods can be performed in an order different from the described order. It should also be understood that the method described above can be formed by hardware elements, or can be implemented by a sequence of machine executable instructions, which can be used to make a machine (such as a general or special processor or logic circuit) programmed with instructions perform these methods. These machine executable instructions can be stored on one or more machine-readable media, such as CD-ROM or other types of optical disks, floppy disks, ROM, RAM, EPROM, EEPROM, magnetic cards or optical cards, flash memory or other types of machine-readable media suitable for storing electronic instructions. Alternatively, these methods can be performed by a combination of hardware and software.

Claims

1. An imaging system for capturing a spatial omics image of a biological tissue sample, the imaging system comprising: an imaging chamber configured to fix a biological tissue sample placed in the imaging system; a time delay and integration (TDI) imager, the TDI imager comprising at least one scan line; a light source configured to illuminate an area on the biological tissue sample being captured by the TDI imager; as well as A controller is configured to cause the TDI imager to scan the biological tissue sample using one or more TDI scans of the biological tissue sample.

2. The imaging system of claim 1, wherein the controller causes the TDI imager to scan the biological sample without requiring mechanical repositioning, stabilization, or focusing between capturing scan lines in a column.

3. The imaging system of claim 1, wherein the light source transmits light to the biological tissue sample via an optical fiber line, and the optical fiber line projects light onto the biological tissue sample in an approximately circular illumination area.

4. The imaging system of claim 1, wherein the light source delivers light to the biological tissue sample through an aperture having a rectangular shape.

5. The imaging system of claim 4, wherein the aperture projects a rectangular illumination area on the biological sample onto a rectangular field of view of the TDI imager.

6. The imaging system of claim 1, wherein the light source delivers light to the biological tissue sample through an aperture, the aperture having a shape that matches a shape of the field of view of the TDI imager.

7. The imaging system of claim 1, wherein the light source delivers light to the biological tissue sample through a fiber optic line having an elliptical shape.

8. The imaging system of claim 1, further comprising a fiber optic bundle comprising a plurality of fiber optic lines, each of the plurality of fiber optic lines being configured to transmit a different wavelength.

9. The imaging system of claim 8, wherein each of the plurality of optical fiber lines has an elliptical or rectangular shape.

10. The imaging system of claim 8, wherein each of the plurality of fiber optic lines is positioned to direct light at a different portion of the biological tissue sample.

11. A method for capturing a spatial omics image of a biological tissue sample, the method comprising the following steps: fixing the biological tissue sample in an imaging chamber of an imaging system; directing light from a light source to illuminate an area on the biological tissue sample; and A time delay and integration (TDI) imager is caused to scan the biological tissue sample using one or more TDI scans of the biological tissue sample, wherein the biological tissue sample is illuminated by the light from the light source.

12. The method of claim 11, further comprising the step of assembling equally spaced lines from a plurality of scan line columns from the TDI imager.

13. The method of claim 11, wherein the light in the light from the light source comprises a plurality of different wavelengths that are simultaneously projected onto the biological tissue sample.

14. The method of claim 13, further comprising the step of filtering the plurality of different wavelengths using a plurality of filters located in front of the TDI imager.

15. The method of claim 13, further comprising the step of filtering the plurality of different wavelengths using a beam splitter located in front of the TDI imager.

16. An imaging system for capturing a spatial omics image of a biological tissue sample, the imaging system comprising: an imaging chamber configured to fix a biological tissue sample placed in the imaging system; Time Delay and Integration (TDI) Imager; a light source configured to illuminate an area of ​​the biological tissue sample being captured by the TDI imager with a plurality of different wavelengths; as well as A controller is configured to cause the TDI imager to simultaneously capture a plurality of different images of the biological tissue sample, wherein each of the plurality of different images corresponds to one of the plurality of different wavelengths.

17. The imaging system of claim 16, wherein the TDI imager comprises a plurality of scan lines, and each of the plurality of scan lines captures one of a plurality of different images.

18. The imaging system of claim 16, wherein the TDI image comprises a plurality of separate TDI cameras, and each of the plurality of separate TDI cameras is aimed at a different location of the biological tissue sample, thereby capturing the plurality of different images simultaneously.

19. The imaging system of claim 16, further comprising a filter wheel in front of the TDI imager, the filter wheel comprising a plurality of filters corresponding to the plurality of different wavelengths.

20. The imaging system of claim 16, wherein the light source comprises a broad spectrum light source that combines the plurality of different wavelengths onto a single illumination area.

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