Apparatus and method for obtaining image of sample during non-uniform motion

Abstract

A method for imaging a sample as it experiences non-uniform motion. A field of view of a camera for imaging the sample is illuminated as the sample moves. The intensity of the illumination may be modulated such that a dose per unit length of a sample container may remain constant even if a period during which any particular feature of the sample container is within the field of view of the camera may vary due to motion inhomogeneity.

Description

Background Art

[0001] The subject matter discussed in this section should not be considered to be prior art simply because it is mentioned in this section. Similarly, problems mentioned in this section or associated with the subject matter provided as background technology should not be considered to have been previously recognized in the prior art. The subject matter in this section merely represents different approaches, which themselves may also correspond to specific implementations of the technology protected by the claims.

[0002] For objects to be imaged, photons must be collected while they are in the field of view of the imaging device. This in turn requires illumination of the objects. To ensure consistent data collection, the energy applied by illumination should be uniform during the time each object is in the field of view. High-precision motion sample stages, time delay integration (TDI) cameras, and diode pumped solid-state (DPSS) lasers are among the components used to achieve this goal. Summary of the invention

[0003] Examples disclosed herein relate to techniques for illuminating an object, and specifically focus on techniques for illuminating a sample of genetic material to be sequenced.

[0004] A specific implementation relates to a machine comprising: a sample stage; a camera for capturing an image; and an actuation system for moving the sample stage relative to a field of view of the camera overlapping the sample stage. Such a machine may also include a controller for generating a set of analysis signals of features of a sample container on the sample stage by performing actions including a set of non-uniform motion imaging actions, wherein each of the analysis signals has a uniform response trajectory. The set of non-uniform motion imaging actions may include: during a first period of time, accelerating the sample stage from a first speed to a second speed relative to the field of view of the camera, and illuminating the field of view of the camera at a first average intensity. The set of non-uniform motion imaging actions may include: during a second period of time after the first period of time, moving the sample stage relative to the field of view of the camera at a predetermined, substantially constant scanning speed, and illuminating the field of view of the camera at a second intensity, the second intensity being substantially constant, wherein the predetermined scanning speed is greater than or equal to the second speed. The set of non-uniform motion imaging actions may include: during a third time period after the second time period, decelerating the sample stage from a third speed to a fourth speed relative to the field of view of the camera and illuminating the field of view of the camera at a third average intensity, wherein the third speed is less than or equal to the predetermined scanning speed. The set of non-uniform motion imaging actions may include: during the first time period, the second time period, and the third time period, capturing exposure of a feature of the sample container on the sample stage.

[0005] In some specific implementations of the machine such as described in the preceding paragraphs of the present disclosure, the controller illuminates the field of view of the camera with the first average intensity during the first time period, with the second intensity during the second time period, and with the third average intensity during the third time period by performing actions including providing a motion trajectory of the sample stage to an illumination controller, wherein the illumination controller provides the uniform response trajectory of each of the analysis signals by continuously modulating the intensity of the illumination source based on the speed of the sample stage when the sample stage accelerates relative to the field of view of the camera and when the sample stage decelerates relative to the field of view of the camera.

[0006] In some implementations of a machine such as described in the second paragraph of the present disclosure, the machine may include: an illumination controller for controlling the illumination source; and an encoder for providing a measurement of the position of the sample stage. In some implementations of such a machine, the controller illuminates the field of view of the camera at the first average intensity during the first period by performing actions including synchronizing the measurement of the position of the sample stage from the encoder with the illumination controller.

[0007] In some implementations of the machine, such as described in the second paragraph of the present disclosure, the first average intensity, the second intensity, and the third average intensity may all be equal. In some implementations of such a machine, the controller generates the set of analysis signals by performing actions including, for each exposure of the feature of the sample container on the sample stage, normalizing the data of the exposure based on the speed of the sample stage when the exposure was captured.

[0008] In some implementations of a machine such as any of the second to fifth paragraphs of the present disclosure, the actuation system may be a motor and the camera may be a time delay integration (TDI) line scan camera. In some implementations of such a machine, the illumination source may be selected from the group consisting of a diode laser and a light emitting diode.

[0009] In some specific implementations of a machine such as any one of the second to sixth paragraphs of the present invention, the first period may include an acceleration period during which the controller accelerates the sample stage from motionless to the predetermined scanning speed relative to the field of view of the camera. In some specific implementations of such a machine, the third period may include a deceleration period during which the controller decelerates the sample stage from the predetermined scanning speed to motionless relative to the field of view of the camera. In some specific implementations of such a machine, the controller may perform the set of non-uniform motion imaging actions multiple times, wherein each execution of the set of non-uniform motion imaging actions corresponds to a strip of features of the sample container. In some specific implementations of such a machine, during each execution of the set of non-uniform motion imaging actions, the controller moves the sample stage along a scanning direction relative to the field of view of the camera. In some specific implementations of such a machine, the controller moves from a strip corresponding to the first of the two consecutive executions to a strip corresponding to the second of the two consecutive executions by moving the sample stage in a direction oblique to the scanning direction between any two consecutive executions of the set of non-uniform imaging actions.

[0010] In some specific implementations of the machine such as any of the second to seventh paragraphs of the present disclosure, the controller may focus the camera during the second period based on an exposure of the feature of the sample container on the sample stage captured at the end of the first period.

[0011] In some specific implementations of the machine such as any one of the second to eighth paragraphs of the present disclosure, the product of the first average intensity and the average speed during the first period is equal to the product of the predetermined scanning speed and the second average intensity. In some such specific implementations, the product of the third average intensity and the average speed during the third period is equal to the product of the predetermined scanning speed and the second average intensity.

[0012] In some implementations of the machine, such as any of the second to ninth paragraphs of the present disclosure, the first period consists of an acceleration period during which the controller accelerates the sample stage from motionless to the predetermined scanning speed relative to the field of view of the camera. In some such implementations, the set of non-uniform motion imaging actions includes illuminating the field of view of the camera at the first average intensity during the first period by continuously providing the illumination source current during the acceleration period at a level that varies with the speed of the sample stage during the acceleration period.

[0013] In some specific implementations of the machine such as any of the second to ninth paragraphs of the present disclosure, the first period consists of an acceleration period during which the controller accelerates the sample stage from motionless to the predetermined scanning speed relative to the field of view of the camera. In some such specific implementations, the set of non-uniform motion imaging actions includes: illuminating the field of view of the camera at the first average intensity during the first period by pulsing the illumination source current at a rate that varies with the speed of the sample stage during the acceleration period.

[0014] Another implementation relates to a method, the method comprising: generating a set of analysis signals of features of a sample container by performing a set of non-uniform motion imaging actions, wherein each of the analysis signals has a uniform response trajectory. In some such implementations, the set of non-uniform motion imaging actions may include: during a first time period, accelerating a sample stage from a first speed to a second speed relative to a field of view of a camera, and illuminating the field of view of the camera at a first average intensity. In some implementations of such methods, the set of non-uniform motion imaging actions further comprises: during a second time period after the first time period, moving the sample stage relative to the field of view of the camera at a predetermined scanning speed, and illuminating the field of view of the camera at a second average intensity, wherein the predetermined scanning speed is greater than or equal to the second speed. In some implementations of such methods, the set of non-uniform motion imaging actions further comprises: during a third time period after the second time period, decelerating the sample stage from a third speed to a fourth speed relative to the field of view of the camera, and illuminating the field of view of the camera at a third average intensity, wherein the third speed is less than or equal to the predetermined scanning speed. In some implementations of such methods, the set of non-uniform motion imaging actions further includes capturing exposure of a feature of the sample container on the sample stage during the first time period, the second time period, and the third time period.

[0015] In some implementations of the method such as described in the preceding paragraphs, the method further includes: providing the motion trajectory of the sample stage to a controller, the controller controlling an illumination source to illuminate the field of view of the camera. In some such implementations, the method may further include: the controller controlling the illumination source to provide the uniform response trajectory of each of the analysis signals by continuously modulating the intensity of the illumination source based on the speed of the sample stage when the sample stage is accelerated relative to the field of view of the camera and when the sample stage is decelerated relative to the field of view of the camera.

[0016] In some implementations of the method such as described in paragraph 12 of the present disclosure, the method further includes: measuring the position of the sample stage using an encoder during a period including the first period and the third period. In some implementations of such methods, illuminating the field of view of the camera with the first intensity and the third intensity during the first period and the third period is based on providing the measurement result of the position of the sample stage from the encoder to a controller, and the controller controls the illumination source and modulates the illumination intensity based on the measurement result of the position of the sample stage from the encoder.

[0017] In some implementations of the method, such as described in paragraph 12 of the present disclosure, the first average intensity, the second average intensity, and the third average intensity are all equal. In some such implementations, the method includes generating the set of analysis signals by performing an action comprising: for each exposure of the feature of the sample container on the sample stage, normalizing the data of the exposure based on the speed of the sample stage when the exposure was captured.

[0018] In some implementations of the method of any of paragraphs 12 to 15 of the present disclosure, the camera is a time delay integration (TDI) line scan camera. In some implementations of such methods, illuminating the field of view of the camera during the first time period, the second time period, and the third time period is performed using an illumination source selected from the group consisting of: a diode laser and a light emitting diode.

[0019] In some specific implementations of the method such as any one of the twelfth to sixteenth paragraphs of the present invention, the first period consists of an acceleration period, during which the sample stage is accelerated from motionless to the predetermined scanning speed relative to the field of view of the camera. In some such specific implementations, the third period consists of a deceleration period, during which the sample stage is decelerated from the predetermined scanning speed to motionless relative to the field of view of the camera. In some such specific implementations, the method includes performing the set of non-uniform motion imaging actions multiple times, wherein each execution of the set of non-uniform motion imaging actions corresponds to a strip of features on the sample container. In some specific implementations of such methods, during each execution of the set of non-uniform motion imaging actions, the sample stage moves relative to the field of view of the camera along a scanning direction. In some specific implementations of such methods, the method includes moving from a strip corresponding to the first of the two consecutive executions to a strip corresponding to the second of the two consecutive executions by moving the sample stage in a direction oblique to the scanning direction between any two consecutive executions of the set of non-uniform motion imaging actions.

[0020] In some specific implementations of the method such as any of paragraphs 12 to 17 of the present disclosure, the method includes focusing the camera based on an exposure of a feature of the sample container on the sample stage captured during the first period of time.

[0021] In some specific implementations of the method such as any one of paragraphs 12 to 18 of the present disclosure, the product of the first average intensity and the average speed during the first time period is equal to the product of the predetermined scanning speed and the second average intensity. In some specific implementations of such methods, the product of the third average intensity and the average speed during the third time period is equal to the product of the predetermined scanning speed and the second average intensity.

[0022] In some specific implementations of the method such as any of paragraphs 12 to 19 of the present disclosure, the first period consists of an acceleration period during which the sample stage is accelerated from motionless to the predetermined scanning speed relative to the field of view of the camera. In some such specific implementations, the set of non-uniform motion imaging actions includes: illuminating the field of view of the camera at the first average intensity during the first period by continuously providing an illumination source current that illuminates the field of view of the camera during the acceleration period at a level that varies with the speed of the sample stage during the acceleration period.

[0023] In some specific implementations of the method such as any of paragraphs 12 to 19 of the present disclosure, the first period consists of an acceleration period during which the sample stage is accelerated from motionless to the predetermined scanning speed relative to the field of view of the camera. In some such specific implementations, the set of non-uniform motion imaging actions includes: illuminating the field of view of the camera at the first average intensity during the first period by pulsing an illumination source current that illuminates the field of view of the camera at a rate that varies with the speed of the sample stage during the acceleration period.

[0024] Another specific implementation involves a machine-readable medium storing instructions for executing a method such as any one of the methods described in paragraphs 12 to 21 of the present disclosure.

[0025] Other features and aspects of the disclosed technology will become apparent from the following detailed description in conjunction with the accompanying drawings, which illustrate features of examples according to the disclosed technology by way of example. This summary is not intended to limit any scope of protection provided by this document or any related document, which is defined by the claims and equivalents of the corresponding documents.

[0026] It should be understood that all combinations of the aforementioned concepts (assuming such concepts are not mutually contradictory) are contemplated as being part of the inventive subject matter disclosed herein. Specifically, all combinations of the claimed subject matter appearing at the end of this disclosure are contemplated as being part of the inventive subject matter disclosed herein. BRIEF DESCRIPTION OF THE DRAWINGS

[0027] The present disclosure according to one or more various examples is described in detail with reference to the following drawings. These drawings are provided for illustration purposes only and merely depict typical or example implementations.

[0028] Figure 1 In one example, a generalized block diagram of an example image scanning system with which the systems and methods disclosed herein may be implemented is illustrated.

[0029] Figure 2 is a block diagram illustrating an example dual-channel line-scan modular optical imaging system that may be implemented in certain implementations.

[0030] Figure 3 Example configurations of patterned samples that may be imaged according to implementations disclosed herein are illustrated.

[0031] FIG. 4A to FIG. 4B Motion trajectories illustrating the potential impact of allowing imaging during periods of non-uniform motion are depicted.

[0032] Figure 5 An example process for modulating illumination during periods of non-uniform motion is illustrated.

[0033] Figure 6 Example computing modules that can be used to implement various features of the implementations described in this disclosure are illustrated.

[0034] The drawings are not exhaustive and do not limit the disclosure to the precise forms disclosed. DETAILED DESCRIPTION

[0035] As used herein to refer to a sample, the term "spot" or "feature" is intended to mean a point or area in a pattern that can be distinguished from other points or areas based on relative position. A single spot may include one or more molecules of a particular type. For example, a spot may contain a single target nucleic acid molecule having a particular sequence, or a spot may contain several nucleic acid molecules having the same sequence (and / or its complement).

[0036] As used herein to refer to a spot or feature in reference to a binding direction, the term "pitch" is intended to refer to the separation of that spot or feature from other spots or features in that direction. For example, if a sample container has an array of features that are separated by 650 nm from one another in the direction in which the container will move during imaging, the "pitch" of the features in that direction may be referred to as 650 nm.

[0037] As used herein, the term "xy plane" is intended to refer to a 2-dimensional region defined by linear axes x and y in a Cartesian coordinate system. When used with reference to a detector and an object observed by the detector, the region may be further specified as being orthogonal to the viewing direction between the detector and the object being detected. When used herein to refer to a line scanner, the term "y direction" refers to the scanning direction.

[0038] As used herein, the term "z-coordinate" is intended to mean information specifying the position of a point, line, or area along an axis orthogonal to the xy plane. In a particular implementation, the z-axis is orthogonal to the area of ​​the object observed by the detector. For example, the focus direction of an optical system may be specified along the z-axis.

[0039] As used herein, the term "scan line" is intended to mean a 2-dimensional cross-section of a detection object in the xy plane, which cross-section is rectangular or oblong and causes relative movement between the cross-section and the object. For example, in the case of fluorescence imaging, a region of an object having a rectangular or oblong shape can be specifically excited (to the exclusion of other regions) and / or emission from this region can be specifically acquired (to the exclusion of other regions) at a given time point in a scan.

[0040] Specific implementations disclosed herein relate to illumination of an object to be imaged while in motion. Illumination may be provided for one or more transient intervals, and data corresponding to multiple illumination transient intervals may be combined to generate an image.

[0041] Figure 1 is an example imaging system 100 in which the techniques disclosed herein may be implemented. The example imaging system 100 may include a device for obtaining or generating an image of a sample. Figure 1 The examples outlined in show example imaging configurations of specific implementations of backlight design. It should be noted that although systems and methods may be described herein from time to time in the context of example imaging system 100, these are merely examples of specific implementations that may implement the illumination and imaging techniques disclosed herein.

[0042] As in Figure 1As can be seen in the example of , a subject sample is located on a sample container 110 (e.g., a flow cell as described herein), which is positioned on a sample stage 170, which is mounted on a frame 190 below an objective lens 142. A light source 160 and associated optical devices direct a light beam (such as a laser) to a selected sample position on the sample container 110. The sample emits fluorescence, and the resulting light is collected by the objective lens 142 and directed to the image sensor of the camera system 140 to detect the fluorescence. The sample stage 170 moves relative to the objective lens 142 to position the next sample position on the sample container 110 at the focus of the objective lens 142. Movement of the sample stage 170 relative to the objective lens 142 can be achieved by moving the sample stage itself, the objective lens, some other component of the imaging system, or any combination of the foregoing. Additional specific implementations may also include moving the entire imaging system onto a fixed sample.

[0043] The fluid delivery module or device 180 can direct the flow of reagents (e.g., fluorescently labeled nucleotides, buffers, enzymes, cleavage reagents, etc.) to (and through) the sample container 110 and the waste valve 120. The sample container 110 may include one or more substrates on which the sample is provided. For example, in the case of a system for analyzing a large number of different nucleic acid sequences, the sample container 110 may include one or more substrates on which the nucleic acid to be sequenced is bound, attached or associated. In various specific implementations, the substrate may include any inert substrate or matrix to which the nucleic acid can be attached, such as a glass surface, a plastic surface, latex, dextran, a polystyrene surface, a polypropylene surface, a polyacrylamide gel, a gold surface, and a silicon wafer. In some applications, the substrate is in a channel or other region located at multiple positions formed in a matrix or array on the sample container 110.

[0044] In some implementations, the sample container 110 may include a biological sample imaged using one or more fluorescent dyes. For example, in a specific implementation, the sample container 110 may be implemented as a patterned flow cell, including a translucent cover, a substrate, and a liquid sandwiched therebetween, and the biological sample may be located at the inner surface of the translucent cover or the inner surface of the substrate. The flow cell may include a large number (e.g., thousands, millions, or billions) of holes or other types of spots (e.g., pads, turfs) that are designed to define arrays (e.g., hexagonal arrays, rectangular arrays, etc.) on the substrate. Each spot may form a cluster (e.g., a monoclonal cluster) of a biological sample (such as DNA, RNA, or another genomic material) that can be sequenced, for example, using sequencing by synthesis. The flow cell may be further divided into a plurality of spaced-apart lanes (e.g., eight lanes), each of which includes a cluster of a hexagonal array. An example flow cell that can be used in the implementation disclosed herein is described in U.S. Pat. No. 8,778,848.

[0045] The system also includes a temperature station actuator 130 and a heater / cooler 135 that can optionally adjust the temperature of the fluid conditions within the sample container 110. A camera system 140 can be included to monitor and track the sequencing of the sample container 110. The camera system 140 can be implemented, for example, as a charge coupled device (CCD) camera (e.g., a time delay integration (TDI) CCD camera) that can interact with various filters within the filter switching assembly 145, the objective lens 142, and the focusing laser / focusing laser assembly 150. The camera system 140 is not limited to a CCD camera, and other camera and image sensor technologies can be used. In a specific implementation, the camera sensor can have a pixel size between about 5 μm and about 15 μm, but other pixel sizes, such as 2.4 μm, can also be used in some cases.

[0046] Output data from the sensors of camera system 140 may be transmitted to a real-time analysis module (not shown), which may be implemented as a software application that analyzes the image data (e.g., image quality scoring), reports or displays characteristics of the laser beam (e.g., focus, shape, intensity, power, brightness, position) to a graphical user interface (GUI), and, as further described below, dynamically corrects distortions in the image data.

[0047] A light source 160 (e.g., an excitation laser within an assembly that optionally includes multiple lasers) or other light source may be included to illuminate a fluorescent sequencing reaction within a sample via illumination through a fiber optic interface (which may optionally include one or more re-imaging lenses, fiber optic mounts, etc.). A low-watt lamp 165 and a focusing laser 150 are also presented in the illustrated example. In some implementations, the focusing laser 150 may be turned off during imaging. In other implementations, an alternative focusing configuration may include a second focusing camera (not shown), which may be a quadrant detector, a position sensitive detector (PSD), or a similar detector to measure the position of a scattered light beam reflected from a surface while data is collected.

[0048] Although illustrated as a backlight device, other examples may include light from a laser or other light source that is directed through the objective lens 142 onto a sample on the sample container 110. The sample container 110 may ultimately be mounted on a sample stage 170 to provide movement and alignment of the sample container 110 relative to the objective lens 142. The sample stage may have one or more actuators to allow it to move in any of three dimensions. For example, in terms of a Cartesian coordinate system, actuators may be provided to allow the sample stage to move in the X, Y, and Z directions relative to the objective lens. This may allow one or more sample locations on the sample container 110 to be positioned to be optically aligned with the objective lens 142.

[0049] A focusing (z-axis) component 175 is shown in this example as being included to control the positioning of the optical components relative to the sample container 110 in a focusing direction (commonly referred to as the z-axis or z-direction). The focusing component 175 may include one or more actuators physically coupled to the optical sample stage or the sample stage, or both, to move the sample container 110 on the sample stage 170 relative to the optical components (e.g., the objective lens 142) to provide appropriate focus for the imaging operation. For example, the actuator may be physically coupled to the corresponding sample stage, such as by mechanical, magnetic, fluid or other attachment or direct or indirect contact with the sample stage or with it. The one or more actuators may move the sample stage in the z-direction while maintaining the sample stage in the same plane (e.g., maintaining a horizontal or horizontal attitude perpendicular to the optical axis). The one or more actuators may also tilt the sample stage. For example, this may be done so that the sample container 110 can be dynamically leveled to account for any slope on its surface.

[0050] Focusing of the system generally refers to aligning the focal plane of the objective lens with the sample to be imaged at a selected sample location. However, focusing may also refer to adjustment of the system to obtain desired characteristics of the sample representation, such as, for example, a desired clarity or contrast level of the test sample image. Because the available depth of field of the focal plane of the objective lens may be small (sometimes on the order of 1 μm or less), the focusing component 175 closely follows the surface being imaged. Because the sample container is not completely flat as it is fixed in the instrument, the focusing component 175 can be arranged to follow this contour when moving along the scanning direction (referred to herein as the y-axis).

[0051] Light emitted from the test sample at the sample location being imaged may be directed to one or more detectors of the camera system 140. An aperture may be included and positioned to allow only light emitted from the focus area to pass to the detector. An aperture may be included to improve image quality by filtering out components of light emitted from areas outside the focus area. An emission filter may be included in the filter switching assembly 145, which may be selected to record a determined emission wavelength and cut off any stray laser light.

[0052] Although not illustrated, a controller may be provided to control the operation of the scanning system, and the controller may be implemented as described below. Figure 6The computing module discussed in the context of . The controller can be implemented to control various aspects of the system operation, such as, for example, focusing, sample stage movement, and imaging operations. In various specific implementations, the controller can be implemented using hardware, algorithms (e.g., machine executable instructions), or a combination of the foregoing. For example, in some specific implementations, the controller may include one or more CPUs or processors with associated memory. As another example, the controller may include hardware or other circuit systems to control operations, such as a computer processor and a non-transitory computer-readable medium in which machine-readable instructions are stored. For example, the circuit system may include one or more of the following: a field programmable gate array (FPGA), an application specific integrated circuit (ASIC), a programmable logic device (PLD), a complex programmable logic device (CPLD), a programmable logic array (PLA), a programmable array logic (PAL), or other similar processing devices or circuit systems. As another example, the controller may include a combination of the circuit system and one or more processors.

[0053] Other imaging systems may also be used when implementing the disclosed techniques. For example, Figure 2 is a block diagram illustrating an example dual-channel line scanning modular optical imaging system 200 in which various aspects of the disclosed technology can be implemented. In some specific implementations, the system 200 can be used for sequencing of nucleic acids. Applicable techniques include techniques in which nucleic acids are attached to fixed positions in an array (e.g., holes in a flow cell) and the array is repeatedly imaged while the array moves relative to the field of view of a camera in the imaging system 200. In such specific implementations, the system 200 can obtain images in two different color channels that can be used to distinguish a specific nucleotide base type from another base type. More specifically, the system 200 can implement a process known as "base calling", which generally refers to the process of determining a base call (e.g., adenine (A), cytosine (C), guanine (G), or thymine (T)) for a given spot position of an image in an imaging cycle. During dual-channel base calling, image data extracted from two images can be used to determine the presence of one of the four base types by encoding the base identity as a combination of the two image intensities. For a given spot or location in each of the two images, the base identity can be determined based on whether the combination of signal signatures is [on, on], [on, off], [off, on], or [off, off].

[0054] Referring again to the imaging system 200, the system includes a line generation module (LGM) 210 in which two light sources 211 and 212 are disposed. The light sources 211 and 212 may be coherent light sources, such as laser diodes that output laser beams. The light source 211 may emit light of a first wavelength (e.g., a red wavelength), and the light source 212 may emit light of a second wavelength (e.g., a green wavelength). The light beams output from the laser sources 211 and 212 may be directed through one or more beam shaping lenses 213. In some implementations, a single light shaping lens may be used to shape the light beams output from the two light sources. In other implementations, a separate beam shaping lens may be used for each light beam. In some examples, the beam shaping lens is a Powell lens, so that the light beam is shaped into a line pattern. The beam shaping lens of the LGM 210 or other optical component imaging system may shape the light emitted by the light sources 211 and 212 into a line pattern (e.g., by using one or more Powell lenses or other beam shaping lenses, diffraction or scattering components).

[0055] The LGM 210 may also include a reflective mirror 214 and a semi-reflective mirror 215 to direct the light beam through a single interface port to an emission optics module (EOM) 230. The light beam may pass through a shutter element 216. The EOM 230 may include an objective lens 235 and a z-sample stage 236 that moves the objective lens 235 longitudinally closer to or farther from a target 250. For example, the target 250 may include a liquid layer 252 and a translucent cover 251, and the biological sample may be located at the inner surface of the translucent cover and at the inner surface of a substrate layer located below the liquid layer. The z-sample stage 236 may then move the objective lens to focus the light beam onto any inner surface of the flow cell (e.g., onto the biological sample). Similarly, in some implementations, the target 250 may be mounted on or may include a sample stage that is movable relative to the objective lens 235 in an xy plane. The biological sample may be DNA, RNA, protein, or other biological material known in the art to be responsive to optical sequencing.

[0056] The EOM 230 may include a semi-reflective mirror 233 to reflect a focus tracking beam emitted from a focus tracking module (FTM) 240 onto a target 250 and then reflect light returned from the target 250 back into the FTM 240. The FTM 240 may include a focus tracking optical sensor to detect characteristics of the returned focus tracking beam and generate a feedback signal to optimize the focus of the objective lens 235 on the target 250.

[0057] The EOM 230 may also include a semi-reflective mirror 234 to direct light through the objective lens 235 while allowing light returned from the target 250 to pass through. In some implementations, the EOM 230 may include a tube lens 232. Light transmitted through the tube lens 232 may pass through a filter element 231 and enter a camera module (CAM) 220. The CAM 220 may include one or more optical sensors 221 to detect light emitted from the biological sample in response to an incident light beam (e.g., fluorescence in response to red and green light received from the light sources 211 and 212).

[0058] Output data from the sensor of CAM 220 may be transmitted to real-time analysis module 225. In various implementations, the real-time analysis module executes computer-readable instructions for analyzing image data (e.g., image quality scores, base calls, etc.), reporting or displaying characteristics of the beam (e.g., focus, shape, intensity, power, brightness, position), etc. to a graphical user interface (GUI). These operations may be performed in real time during the imaging cycle to minimize downstream analysis time and provide real-time feedback and troubleshooting during the imaging run. In a specific implementation, the real-time analysis module may be a computing device (e.g., computing device 1000) that is communicatively coupled to and controls the imaging system 200. In a specific implementation further described below, the real-time analysis module 225 may additionally execute computer-readable instructions for controlling the illumination of the target 250 and optionally for integrating data collected during multiple exposures of the optical sensor 221 into an image.

[0059] Figure 3 An example configuration of a sample container 300 that can be imaged according to an implementation disclosed herein is illustrated. In this example, the sample container 300 is patterned into a hexagonal array with ordered spots 310 that can be imaged simultaneously during an imaging run. Although a hexagonal array is illustrated in this example, in other implementations, a linear array, a circular array, an octagonal array, or some other array pattern can be used to pattern the sample container. For ease of illustration, the sample container 300 is illustrated as having tens to hundreds of spots 310. However, as can be appreciated by those skilled in the art, the sample container 300 can have thousands, millions, or billions of spots 310 that are imaged. In addition, in some cases, the sample container 300 can be a multi-plane sample that includes multiple planes (perpendicular to the focus direction) of spots 310 sampled during an imaging run.

[0060] In one particular implementation, the sample container 300 may be a flow cell patterned with millions or billions of wells divided into lanes. In this particular implementation, each well of the flow cell may contain biological material that is sequenced using sequencing by synthesis.

[0061] As discussed above, illumination and imaging of objects that move relative to the field of view of an imaging device has been achieved through a high precision motion sample stage, a time delay integration (TDI) camera, and a diode pumped solid state laser. To achieve consistent data collection, imaging is performed during periods of uniform motion of the objects to be imaged relative to the device used to image these objects (e.g., a TDI camera). However, specific implementations of the disclosed technology can achieve the same goals while allowing imaging during periods of acceleration or deceleration of the objects to be imaged relative to the device used to image them. This can include imaging during acceleration and deceleration periods at the beginning and end of sample imaging, and can also include other periods of non-uniform motion during sample imaging. For example, in some cases, after scanning a swath, during a period such as Figure 1 to Figure 2 When the illustrated imaging system decelerates and changes to the next strip, there will be a period of deceleration and then re-acceleration to the uniform scanning speed to image the next strip. In some cases, specific implementations of the disclosed technology can allow imaging during these strip transition periods, in addition to or instead of allowing imaging at the beginning and end of the sample scan.

[0062] To illustrate the potential impact of allowing imaging during periods of non-uniform motion, consider FIG. 4A to FIG. 4B . Figure 4A Three motion trajectories are provided that depict the velocity of the sample on the sample stage relative to the imaging device as a function of the position of the sample relative to the imaging device. First, Figure 4A A first motion trajectory 401 of a scenario where non-uniform motion is minimized is depicted. For example, this can be done in an imaging system designed to accelerate and decelerate the sample as quickly as possible, since imaging will only occur when the sample moves at a predefined uniform scanning speed. Figure 4A Also depicted are a second motion trajectory 402 and a third motion trajectory 403, which correspond to scenarios where the acceleration and deceleration are 50% and 75% of the acceleration and deceleration, respectively, of the first motion trajectory 401. In motion trajectory 402 and motion trajectory 403, the area 404 between the solid motion trajectory line and the outer dashed line represents the reduction in the amount of space that can be imaged when the sample moves at a uniform scan speed under the lower acceleration / deceleration motion trajectory 402 and acceleration / deceleration motion trajectory 403 relative to the first motion trajectory 401. Similarly, the area 405 between the solid motion trajectory line and the inner dashed line in the third motion trajectory 403 represents the increase in the amount of space that can be imaged when the sample moves at a uniform scan speed under the third motion trajectory 403 relative to the second motion trajectory 402, thereby demonstrating the effect of acceleration / deceleration on potential uniform motion scan time in terms of position.

[0063] and Figure 4A similar, Figure 4B Three motion trajectories are provided. However, Figure 4B The trace of illustrates velocity with respect to time, rather than with respect to position, and this velocity plot with respect to time is used to show the effect of allowing imaging during periods of non-uniform motion on the total time of a strip, where the total time of a strip includes the time to image the strip and the turnaround time between the end of imaging of one strip and the beginning of imaging of the next strip. Figure 4A Similar to the first motion trajectory 401 in, Figure 4B The first motion trajectory 411 in FIG. 4 shows the motion trajectory for a case where the imaging system is designed to accelerate and decelerate as quickly as possible, wherein Figure 4B The data acquisition time (t dat_0 ). Figure 4B The second motion trajectory 412 and the third motion trajectory 413 depict the speed of the sample over time, and the two speeds are respectively Figure 4B The acceleration and deceleration are performed at 50% and 75% of the acceleration and deceleration of the first motion trajectory 411 in FIG. Figure 4B As shown, where images are captured during acceleration and deceleration periods, the total time of the strip accelerating and decelerating at less than the depicted maximum rate can be equal to (in the case of the illustrated 50% acceleration and deceleration) or less than (in the case of the illustrated 75% acceleration and deceleration) the total time of the strip accelerating and decelerating at the indicated maximum rate.

[0064] Table 1 below provides further illustration of the potential effects of varying acceleration and deceleration in situations where images are captured while the sample is not at its uniform scan speed. The table illustrates exemplary values ​​of acceleration, scan speed, turnaround time, and other parameters that may affect compliance with Figure 4A and Figure 4B Image acquisition in the system of motion trajectory.

[0065]

[0066] Table 1

[0067] Although Figure 4A and Figure 4B and Table 1 illustrate how imaging during non-uniform motion can allow for comparable or better performance than can be achieved by a system having higher acceleration but capturing images only when the sample is at a constant velocity, but it should be understood that these figures and Table 1 are for illustrative purposes only and are not intended to represent the precise motion profiles that will necessarily exist in a system based on implementations of the present disclosure. For example, in a system based on the present disclosure, the acceleration and deceleration periods in the motion profile will most likely have an S-curve shape rather than having a Figure 4Aand Figure 4B to reflect that the acceleration will vary over time rather than being constant. Similarly, while potential relative turnaround times for strips are illustrated, turnaround times for different systems can vary greatly and may be greater (e.g., in cases where cheaper components are used to achieve lower maximum acceleration and deceleration rates for the system, and these cheaper components also increase the time required to switch between strips) or less than (e.g., in a system where the sample switches from one strip to another by following a curve rather than stopping, switching, and restarting, in which case imaging can run continuously while switching from one strip to another) the times illustrated in these figures and Table 1. Therefore, Figure 4A , Figure 4B The traces of Table 1 and their accompanying discussion should be understood to be illustrative only and should not be taken as implying limitations on the scope of the claims provided in this or any related document.

[0068] Now turn to Figure 5 , which illustrates a method that can be used to support imaging during periods of non-uniform motion, such as in Figure 4A The second motion trajectory 402 and the third motion trajectory 403 and Figure 4B The second motion trajectory 412 and the third motion trajectory 413 are described in the context of FIG. Figure 5 The method begins by initiating acceleration of the sample relative to the imaging device in box 501. As the sample accelerates, it can be illuminated at an intensity that matches its current speed, as shown in box 502. To support this, in some embodiments, a motion trajectory of the sample can be provided to a controller of a laser diode, light emitting diode, or other illumination source. Then, based on the longer distance of the sample within the field of view of the imaging device (e.g., a TDI camera) while accelerating, the illumination controller can provide current at a level that is proportionally lower than the current used when the system is at maximum sample speed. In this way, it is implemented to perform operations such as Figure 5 The system of the method shown can modulate the illumination when the sample undergoes non-uniform motion so that the product of the illumination and the average velocity remains constant over all imaging periods. For example, the relationship between the illumination provided during the sample acceleration period and the illumination provided when the sample moves at its maximum velocity (e.g., scanning velocity) can be expressed by the following formula 1:

[0069] I 01 *(V 0 +V 1 ) / 2=I s *V s

[0070] Formula 1

[0071] In the above formula, I01 Refers to the irradiation provided from time 0 to time 1 (e.g., from the beginning to the end of the acceleration period, from the beginning of acceleration to the end of the first 0.1 second acceleration time increment, etc.). s V is the illumination provided to the sample when the sample is scanned at the maximum speed of the system (e.g., the illumination provided when the sample moves at 48.0 mm / s in a system having the scanning speeds listed in Table 1). 0 and V 1 are the speeds of the sample at time 0 and time 1 respectively. V s is the speed at which the sample moves when it is scanned at the maximum speed of the system (e.g., 48.0 mm / s in a system having the scanning speeds listed in Table 1). Therefore, as shown in block 502, by using an intensity I satisfying a formula such as Equation 1 01 The sample may be illuminated at an intensity that matches the sample velocity.

[0072] exist Figure 5 In the method of , once acceleration is complete and the sample reaches its uniform scan speed, it can continue to be illuminated at the system's scan intensity (e.g., 0.5 Watt), as shown in block 503. This can continue until the sample begins to decelerate (e.g., when the end of the strip is reached and the system decelerates to move to the next strip or load another sample). As the sample decelerates, in block 504 it can be illuminated at an intensity that matches its current speed, such as by using current modification as described above. When deceleration is complete, if there are more strips, then in block 505 the system can move to the next strip. For example, in a method such as Figure 3 In the case where the sample container 300 shown is moved horizontally relative to the imaging device during scanning, the movement to the next strip in block 505 can be achieved by moving vertically so that the next set of horizontal spots is then within the field of view of the imaging device. During this period, the illumination and imaging of the sample can be paused while the sample is stationary in the direction in which it was moving while being scanned. Figure 5 After the system of the method shown moves to the next strip in block 505, it can then return to block 501 to accelerate the next strip to the scanning speed and image it at the scanning speed during the acceleration period as described above. Alternatively, if there are no further strips, then Figure 5 The process may terminate at block 506 (eg, in order to load the next sample so that it may be imaged as described).

[0073] It should be understood that the above methods and examples of how they may be applied are intended to be illustrative only, and that other methods and variations of the described methods are possible and may be applied in some implementations. For illustration, consider a method for modulating the intensity of the illumination provided to a sample during a period of non-uniform motion (e.g., acceleration and deceleration). While some implementations may control the intensity of the illumination by increasing or decreasing the current provided to the illumination source, another method that may be used in some cases is to illuminate the sample with high frequency pulses, setting the duty cycle of the illumination source so that the total illumination provided to the feature when the feature is within the field of view of the imaging device is the same as the illumination provided when the sample is scanned at the scanning speed of the system. Similarly, while some implementations may set the intensity of the illumination by providing a motion trajectory to a controller of the illumination source, another method is to synchronize the illumination with the pulses of an encoder used to measure the position of the sample (e.g., by pulse width modulation). For example, some such versions may provide pulse width modulation (PWM). However, in versions using PWM, the pulses of the PWM do not necessarily have to be synchronized with the pulses of the encoder.

[0074] In some implementations, it is also possible to support imaging during periods of non-uniform motion without requiring a 3D imager such as described above. Figure 5 For example, in some cases, the sample can be illuminated at a constant intensity (e.g., 0.5 Watt) throughout the imaging process, and a normalization process can be applied to the captured images to account for the larger illumination dose that will be provided when imaging features during relatively low speed periods (e.g., when the sample is accelerated to its scanning speed). For example, in some systems, the signal of a feature in an exposure (e.g., light emitted by a sample in a well in response to incident light) can be used to derive a device analysis signal by multiplying the signal in the exposure by the instantaneous speed of the feature when the exposure was captured (e.g., as determined using encoder measurements, or by matching the time of the captured exposure to a motion trajectory). Using this approach, the signal of a feature imaged during an acceleration or deceleration period will be multiplied by a lower normalization factor (e.g., lower speed) than the signal of a feature imaged when the sample is moving at its scanning speed. This will provide an analysis signal with a uniform response trajectory (i.e., the same signal can be considered to have the same meaning between analysis images), which can be used for purposes such as base calling.

[0075] Variations are also possible in the potential uses of imaging during periods of non-uniform motion. For example, in some cases, periods of non-uniform motion may be used to collect information about a sample that will be used for purposes such as base calling during nucleotide sequencing. However, in other cases, images captured during periods of non-uniform motion may be used for purposes such as focusing an imaging device. Combination approaches (such as using an initial acceleration period for focusing and then using subsequent acceleration and deceleration periods to capture images of the sample for base calling) and other types of variations (such as continuing to illuminate and capture images of the sample while moving to a new strip in box 505) are also possible and will be immediately apparent to those of ordinary skill in the art in light of this disclosure and may be implemented without undue experimentation. Thus, the above descriptions of potential variations (such as FIG. 4A to FIG. 4B and Figure 5 The description of ) should be understood as merely illustrative and should not be regarded as restrictive.

[0076] Now turn to Figure 6 , which illustrates an example computing component that can be used to implement various features of the systems and methods disclosed herein, such as the aforementioned features and functions of one or more aspects of methods 400 and 450. For example, the computing component can be implemented as real-time analysis module 225.

[0077] As used herein, the term module may describe a given functional unit that can be performed according to one or more specific implementations of the present application. As used herein, a module may be implemented using any form of hardware, software, or a combination thereof. For example, one or more processors, controllers, ASICs, PLAs, PALs, CPLDs, FPGAs, logic components, software routines, or other mechanisms may be implemented to form a module. In a specific implementation, the various modules described herein may be implemented as discrete modules, or the functions and features described may be partially or completely shared between one or more modules. In other words, as may be apparent to those of ordinary skill in the art after reading this specification, the various features and functions described herein may be implemented in any given application, and may be implemented in one or more separate or shared modules in various combinations and arrangements. Even if various features or functional elements may be individually described or protected by claims as separate modules, those of ordinary skill in the art will understand that these features and functions may be shared between one or more common software and hardware elements, and such descriptions should not require or imply the use of separate hardware or software components to implement such features or functions.

[0078] Where components or modules of an application are implemented in whole or in part using software, in one implementation, these software elements may be implemented to operate in conjunction with a computing or processing module capable of performing the functions described with respect thereto. Figure 61000. Various implementations are described based on the example computing module 1000. After reading this specification, it will become apparent to those skilled in the relevant art how to implement applications using other computing modules or architectures.

[0079] Reference now Figure 6 , computing module 1000 may represent computing or processing capabilities such as found in: sample desktop computers, laptop computers, notebook computers, and tablet computers; handheld computing devices (tablet computers, PDAs, smart phones, cell phones, palmtops, etc.); mainframes, supercomputers, workstations, or servers; or any other type of special-purpose or general-purpose computing device that may be desirable or appropriate for a given application or environment. Computing module 1000 may also represent computing capabilities embedded within or otherwise available to a given device. For example, computing modules may be found in other electronic devices such as, for example, digital cameras, navigation systems, cellular phones, portable computing devices, modems, routers, WAPs, terminals, and other electronic devices that may include some form of processing capabilities.

[0080] The computing module 1000 may include, for example, one or more processors, controllers, control modules, or other processing devices such as a processor 1004. The processor 1004 may be implemented using a general or special purpose processing engine such as, for example, a microprocessor, controller, or other control logic. In the illustrated example, the processor 1004 is connected to a bus 1002, but any communication medium may be used to facilitate interaction with other components of the computing module 1000 or external communication.

[0081] The computing module 1000 may also include one or more memory modules, referred to herein as main memory 1008. For example, preferably, a random access memory (RAM) or other dynamic memory may be used to store information and instructions to be executed by the processor 1004. The main memory 1008 may also be used to store temporary variables or other intermediate information during the execution of instructions to be executed by the processor 1004. The computing module 1000 may also include a read-only memory ("ROM") or other static storage device coupled to the bus 1002 for storing static information and instructions for the processor 1004.

[0082] The computing module 1000 may also include one or more information storage mechanisms 1010 in various forms, which may include, for example, a media drive 1012 and a storage unit interface 1020. The media drive 1012 may include a drive or other mechanism for supporting fixed or removable storage media 1014. For example, a hard disk drive, a solid state drive, a tape drive, an optical drive, a CD or DVD drive (R or RW), or other removable or fixed media drive may be provided. Thus, the storage medium 1014 may include, for example, a hard disk, a solid state drive, a magnetic tape, a cassette, an optical disk, a CD, a DVD, or a Blu-ray, or other fixed or removable media that is read, written, or accessed by the media drive 1012. As illustrated by these examples, the storage medium 1014 may include a computer usable storage medium having computer software or data stored therein.

[0083] In alternative implementations, the information storage mechanism 1010 may include other similar means for allowing computer programs or other instructions or data to be loaded into the computing module 1000. Such means may include, for example, fixed or removable storage units 1022 and interfaces 1020. Examples of such storage units 1022 and interfaces 1020 may include program cartridges and cartridge interfaces, removable memory (e.g., flash memory or other removable memory modules) and memory slots, PCMCIA slots and cards, and other fixed or removable storage units 1022 and interfaces 1020 that allow software and data to be transferred from the storage unit 1022 to the computing module 1000.

[0084] The computing module 1000 may also include a communication interface 1024. The communication interface 1024 may be used to allow software and data to be transferred between the computing module 1000 and external devices. Examples of the communication interface 1024 may include a modem or soft modem, a network interface (such as Ethernet, a network interface card, WiMedia, IEEE 802.XX or other interface), a communication port (such as, for example, a USB port, an IR port, an RS232 port, The software and data transmitted via the communication interface 1024 may generally be carried on signals, which may be electronic signals, electromagnetic signals (including optical signals), or other signals capable of being exchanged by a given communication interface 1024. These signals may be provided to the communication interface 1024 via a channel 1028. The channel 1028 may carry signals and may be implemented using a wired or wireless communication medium. Some examples of a channel may include a telephone line, a cellular link, an RF link, an optical link, a network interface, a local area network or a wide area network, and other wired or wireless communication channels.

[0085] In this document, the terms "computer-readable medium," "computer-usable medium," and "computer program medium" are generally used to refer to non-transitory media, whether volatile or non-volatile, such as, for example, memory 1008, storage unit 1022, and medium 1014. These and other various forms of computer program media or computer-usable media may involve carrying one or more sequences of one or more instructions to a processing device for execution. Such instructions embodied on the medium are generally referred to as "computer program code" or "computer program products" (which may be grouped in the form of a computer program or other grouping). When executed, such instructions may enable computing module 1000 to perform features or functions of the present application as discussed herein.

[0086] Although described above in terms of various implementations, it should be understood that the various features, aspects, and functions described in one or more of the individual implementations are not limited in their applicability to the specific implementations in which they are described, but may be applied alone or in various combinations to one or more of the other implementations of the present application, regardless of whether such implementations are described and whether such features are present as part of the described implementations. Therefore, the breadth and scope of the claims provided by this document or any related documents should not be limited by any of the above-mentioned implementations.

[0087] It should be understood that all combinations of the aforementioned concepts (assuming such concepts are not mutually contradictory) are contemplated as being part of the inventive subject matter disclosed herein. Specifically, all combinations of the claimed subject matter appearing at the end of this disclosure are contemplated as being part of the inventive subject matter disclosed herein.

[0088] The terms "substantially" and "approximately" used throughout this disclosure (including claims) are used to describe and illustrate small fluctuations, such as those caused by variations in processing. For example, they can mean less than or equal to ±5%, such as less than or equal to ±2%, such as less than or equal to ±1%, such as less than or equal to ±0.5%, such as less than or equal to ±0.2%, such as less than or equal to ±0.1%, such as less than or equal to ±0.05%.

[0089] To the extent applicable, the terms "first", "second", "third", etc. herein are used merely to show the respective objects described by these terms as separate entities and are not meant to imply a time sequence unless otherwise expressly stated herein.

[0090] Unless expressly stated otherwise, the terms and phrases used in this document and their variations should be interpreted as open-ended and not limiting. As an example of the foregoing: the term "including" should be understood to mean "including but not limited to", etc.; the term "example" is used to provide an example of the items discussed, rather than an exhaustive or limiting list thereof; the term "one" or "an" should be understood to mean "at least one", "one or more", etc.; and adjectives such as "pre-existing", "traditional", "normal", "standard", "known", and terms of similar meaning should not be interpreted as limiting the items described to a given time period or to items available within a given time period, but should be understood to include pre-existing, traditional, normal, or standard technologies available or known at any time now or in the future. Similarly, where this document refers to technologies that may be obvious or known to a person of ordinary skill in the art, such technologies include those technologies that are obvious or known to a person of ordinary skill in the art at any time now or in the future.

[0091] In some cases, the presence of broadening words and phrases such as "one or more," "at least," "but not limited to," or other similar phrases should not be understood to mean that a narrower case is intended or required where such broadening phrases may not be present. The use of the term "module" does not mean that the components or functions described or claimed as part of a module are all configured in a common package. In fact, any or all of the various components of a module, whether control logic or other components, may be combined in a single package or maintained separately, and may also be distributed in multiple groupings or packages or across multiple locations.

[0092] In addition, various implementations described herein are described in terms of block diagrams, flow charts, and other illustrations. As will be apparent to one of ordinary skill in the art after reading this document, the illustrated implementations and their various alternatives may be implemented without limitation to the illustrated examples. For example, the block diagrams and their accompanying descriptions should not be construed as mandatory requirements for a particular architecture or configuration.

[0093] Although various specific implementations of the present disclosure have been described above, it should be understood that they are presented only by way of example and not limitation. Similarly, each figure may depict an example architecture or other configuration of the present disclosure, which is done to help understand the features and functions that may be included in the present disclosure. The present disclosure is not limited to the illustrated example architecture or configuration, but various alternative architectures and configurations may be used to achieve the desired features. In fact, it is obvious to those skilled in the art how to implement alternative functions, logical or physical partitions and configurations to achieve the desired features of the present disclosure. In addition, many different component module names other than those described herein may be applied to various partitions. In addition, with respect to flow charts, operational descriptions, and method claims, unless the context otherwise dictates, the order of the actions presented herein should not enforce that various specific implementations perform the functions in the same order.

Claims

1. A machine, comprising: Sample table; a camera for capturing images; an actuation system for moving the sample stage relative to a field of view of the camera overlapping the sample stage; an illumination source, the illumination source being used to illuminate the field of view of the camera; and A controller configured to generate a set of analysis signals of characteristics of a sample container on the sample stage by performing actions including a set of non-uniform motion imaging actions, wherein each of the analysis signals has a uniform response trajectory, the set of non-uniform motion imaging actions comprising: During a first period, accelerating the sample stage from a first speed to a second speed relative to the field of view of the camera and illuminating the field of view of the camera at a first average intensity; during a second time period after the first time period, moving the sample stage relative to the field of view of the camera at a predetermined, substantially constant scan speed and illuminating the field of view of the camera at a second intensity, the second intensity being substantially constant, wherein the predetermined scan speed is greater than or equal to the second speed; During a third time period after the second time period, decelerating the sample stage from a third speed to a fourth speed relative to the field of view of the camera and illuminating the field of view of the camera at a third average intensity, wherein the third speed is less than or equal to the predetermined scanning speed; and During the first time period, the second time period, and the third time period, exposures of features of the sample container on the sample stage are captured.

2. The machine of claim 1 , wherein the controller illuminates the field of view of the camera at the first average intensity during the first time period, at the second intensity during the second time period, and at the third average intensity during the third time period by performing actions including providing a motion trajectory of the sample stage to an illumination controller, wherein the illumination controller provides the uniform response trajectory for each of the analytical signals by continuously modulating the intensity of the illumination source based on the speed of the sample stage when the sample stage accelerates relative to the field of view of the camera and when the sample stage decelerates relative to the field of view of the camera.

3. The machine of claim 1, wherein the machine comprises: an illumination controller for controlling the illumination source; and an encoder for providing measurements of the position of the sample stage, and wherein the controller illuminates the field of view of the camera at the first average intensity during the first time period by performing actions including synchronizing the measurements of the position of the sample stage from the encoder with the illumination controller.

4. The machine according to claim 1, wherein: The first average intensity, the second intensity, and the third average intensity are all equal; and The controller generates the set of analysis signals by performing actions including, for each exposure of a feature of the sample container on the sample stage, normalizing data of the exposure based on a velocity of the sample stage when the exposure was captured.

5. A machine according to any one of claims 1 to 4, wherein: The actuation system is a motor; The camera is a time delay integration (TDI) line scan camera; and The radiation source is selected from the group consisting of: Diode lasers; and Light Emitting Diode.

6. A machine according to any one of claims 1 to 5, wherein: The first period includes an acceleration period during which the controller accelerates the sample stage from motionless to the predetermined scanning speed relative to the field of view of the camera; The third period includes a deceleration period during which the controller decelerates the sample stage from the predetermined scanning speed to a stationary state relative to the field of view of the camera; the controller performing the set of non-uniform motion imaging actions a plurality of times, wherein each execution of the set of non-uniform motion imaging actions corresponds to a strip of features of the sample container; During each execution of the set of non-uniform motion imaging actions, the controller moves the sample stage relative to the field of view of the camera along a scanning direction; and The controller moves the sample stage in a direction oblique to the scanning direction between any two consecutive executions of the group of non-uniform motion imaging actions, so as to move from a strip corresponding to a first execution of the two consecutive executions to a strip corresponding to a second execution of the two consecutive executions.

7. The machine of any one of claims 1 to 6, wherein the controller focuses the camera during the second period based on an exposure of a feature of the sample container on the sample stage captured at the end of the first period.

8. A machine according to any one of claims 1 to 7, wherein: The product of the first average intensity and the average speed during the first period is equal to the product of the predetermined scanning speed and the second intensity; and The product of the third average intensity and the average speed during the third period is equal to the product of the predetermined scanning speed and the second intensity.

9. A machine according to any one of claims 1 to 8, wherein: The first period consists of an acceleration period during which the controller accelerates the sample stage from motionless to the predetermined scanning speed relative to the field of view of the camera; and The set of non-uniform motion imaging actions includes illuminating the field of view of the camera at the first average intensity during the first period by continuously providing the illumination source current during the acceleration period at a level that varies with a speed of the sample stage during the acceleration period.

10. A machine according to any one of claims 1 to 8, wherein: The first period consists of an acceleration period during which the controller accelerates the sample stage from motionless to the predetermined scanning speed relative to the field of view of the camera; and The set of non-uniform motion imaging actions includes illuminating the field of view of the camera at the first average intensity during the first period by pulsing the illumination source current at a rate that varies with the speed of the sample stage during the acceleration period.

11. A method comprising: Generating a set of analysis signals characteristic of the sample container by performing actions including performing a set of non-uniform motion imaging actions, wherein each of the analysis signals has a uniform response trajectory, the set of non-uniform motion imaging actions including: During a first period, accelerating the sample stage from a first speed to a second speed relative to a field of view of a camera and illuminating the field of view of the camera at a first average intensity; During a second time period after the first time period, moving the sample stage relative to the field of view of the camera at a predetermined scanning speed and illuminating the field of view of the camera at a second average intensity, the predetermined scanning speed being greater than or equal to the second speed; During a third time period after the second time period, decelerating the sample stage from a third speed to a fourth speed relative to the field of view of the camera and illuminating the field of view of the camera at a third average intensity, wherein the third speed is less than or equal to the predetermined scanning speed; and During the first time period, the second time period, and the third time period, exposures of features of the sample container on the sample stage are captured.

12. The method according to claim 11, wherein the method comprises: providing a motion trajectory of the sample stage to a controller, the controller controlling an illumination source to illuminate the field of view of the camera; as well as The controller that controls the illumination source provides the uniform response trajectory for each of the analysis signals by continuously modulating the intensity of the illumination source based on the speed of the sample stage as the sample stage accelerates relative to the field of view of the camera and as the sample stage decelerates relative to the field of view of the camera.

13. The method according to claim 11, wherein: The method includes measuring the position of the sample stage using an encoder during a period including the first period and the third period; and Illuminating the field of view of the camera with the first average intensity and the third average intensity during the first time period and the third time period is based on providing measurements of the position of the sample stage from the encoder to a controller, the controller controlling an illumination source and modulating illumination intensity based on the measurements of the position of the sample stage from the encoder.

14. The method according to claim 11, wherein: The first average intensity, the second average intensity, and the third average intensity are all equal; and The method includes generating the set of analysis signals by performing acts including, for each exposure of a feature of the sample container on the sample stage, normalizing data of the exposure based on a velocity of the sample stage when the exposure was captured.

15. The method according to any one of claims 11 to 14, wherein: The camera is a time delay integration (TDI) line scan camera; and Illuminating the field of view of the camera during the first, second, and third time periods is performed using an illumination source selected from the group consisting of: Diode lasers; and Light Emitting Diode.

16. A method according to any one of claims 11 to 15, wherein: The first period consists of an acceleration period during which the sample stage is accelerated from motionless to the predetermined scanning speed relative to the field of view of the camera; The third period consists of a deceleration period during which the sample stage is decelerated from the predetermined scanning speed to a stationary state relative to the field of view of the camera; The method includes performing the set of non-uniform motion imaging acts a plurality of times, wherein each performance of the set of non-uniform motion imaging acts corresponds to a strip of features on the sample container; During each execution of the set of non-uniform motion imaging actions, the sample stage moves relative to the field of view of the camera along a scanning direction; and The method includes moving the sample stage in a direction oblique to the scanning direction between any two consecutive executions of the set of non-uniform motion imaging actions, from a strip corresponding to a first execution of the two consecutive executions to a strip corresponding to a second execution of the two consecutive executions.

17. The method of any one of claims 11 to 16, wherein the method comprises focusing the camera based on an exposure of a feature of the sample container on the sample stage captured during the first period of time.

18. A method according to any one of claims 11 to 17, wherein: The product of the first average intensity and the average speed during the first period is equal to the product of the predetermined scanning speed and the second average intensity; and The product of the third average intensity and the average speed during the third period is equal to the product of the predetermined scanning speed and the second average intensity.

19. The method according to any one of claims 11 to 18, wherein: The first period consists of an acceleration period during which the sample stage is accelerated from motionless to the predetermined scanning speed relative to the field of view of the camera; and The set of non-uniform motion imaging actions includes illuminating the field of view of the camera at the first average intensity during the first period by continuously providing an illumination source current that illuminates the field of view of the camera during the acceleration period at a level that varies with the speed of the sample stage during the acceleration period.

20. The method according to any one of claims 11 to 18, wherein: The first period consists of an acceleration period during which the sample stage is accelerated from motionless to the predetermined scanning speed relative to the field of view of the camera; and The set of non-uniform motion imaging actions includes illuminating the field of view of the camera at the first average intensity during the first period by pulsing an illumination source current that illuminates the field of view of the camera at a rate that varies with the speed of the sample stage during the acceleration period.

Images