Multispectral imaging device

Abstract

The apparatus includes a sample stage area, an optical assembly, and a camera assembly. The optical assembly includes an objective element, an imaging lens, a first dichroic element, and a corrector element. The objective element provides a field of view including an object plane. The imaging lens is configured to receive light transmitted through the objective element. The optical assembly is configured to transmit light from at least a first color channel, a second color channel, a third color channel, and a fourth color channel from the imaging lens toward the first dichroic element. The first dichroic element is configured to transmit light from the first and third color channels, while reflecting light from the second and fourth color channels. The corrector element is configured to induce astigmatism in the first and third color channels to offset the astigmatism induced by the dichroic element.

Description

Technical Field

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[0001] (Priority) This application claims priority to U.S. Provisional Patent Application No. 63 / 469,120, filed May 26, 2023, entitled "Multispectral Imager", the entire disclosure of which is incorporated herein by reference.

Background Art

[0002] Aspects of the present disclosure generally relate to devices, systems, and methods for providing biological or chemical analysis. Various protocols in biological or chemical research involve performing a number of controlled reactions on a local support surface or within a defined reaction chamber. Subsequently, a specified reaction can be observed or detected, and the subsequent analysis may serve to identify or clarify the properties of the chemical substances involved in the reaction. For example, in some multiplex assays, an unknown analyte having an identifiable label (e.g., a fluorescent label) may be exposed to thousands of well-known probes under controlled conditions. Each well-known probe may be deposited in a corresponding well of a flow cell channel. Observing any chemical reaction that occurs between the well-known probe and the unknown analyte in the well may serve to identify or clarify the properties of the analyte. Other examples of such protocols include well-known DNA sequencing processes such as Sequencing-By-Synthesis (SBS) or circular array sequencing.

[0003] Various devices, systems, and methods have been made and used to perform biological or chemical analysis, but prior to the present inventor(s), no one is believed to have made or used the devices and techniques described herein.

Brief Description of the Drawings

[0004] [Figure 1]A schematic diagram of one embodiment of a system that may be used to provide biological or chemical analysis is shown. [Figure 2] A schematic diagram of one embodiment of a set of components that can cooperate to provide a fluid path in the system shown in Figure 1 is depicted. [Figure 3] A schematic diagram of another embodiment of a system that may be used to provide biological or chemical analysis is depicted. [Figure 4] A cross-sectional view of one embodiment of a flow cell that may be used in the system shown in Figure 1 is depicted. [Figure 5] A cross-sectional view of another embodiment of a flow cell that may be used in the system shown in Figure 1 is depicted. [Figure 6] Figure 5 depicts a top view of the flow cell, with the upper wafer omitted to reveal the lower wafer. [Figure 7] A schematic diagram of another embodiment of a system that may be used to provide biological or chemical analysis is depicted. [Figure 8] A schematic diagram of one embodiment of an imaging element that can be integrated into the system shown in Figure 7 is depicted. [Figure 9] A schematic diagram of another embodiment of the imaging element that can be integrated into the system shown in Figure 7 is depicted. [Figure 10] A schematic diagram of one embodiment of an imaging element that can be integrated into the system shown in Figure 7 is depicted. [Figure 11] A schematic diagram illustrating an example of a color separation assembly to either the imaging system or imaging assembly described herein is shown. [Figure 12] A schematic diagram of an embodiment of a color separation assembly positioned in a collimated space between the objective lens assembly and the imaging lens assembly is shown. [Figure 13] This diagram illustrates a schematic representation of one embodiment of the arrangement of imaging elements, including a color separation assembly, and is provided along the xz plane. [Figure 14] Figure 13 shows a schematic diagram of the imaging element, which is provided along the xy plane. [Figure 15]Figure 13 shows a schematic perspective view of the imaging element. [Figure 16] This diagram illustrates an example of another arrangement of imaging elements, including a color separation assembly, and is provided along the xz plane. [Figure 17] Figure 16 shows a schematic diagram of the imaging element, which is provided along the xy plane. [Figure 18] A schematic diagram of the arrangement of imaging components, including the beam separation element, is shown. [Modes for carrying out the invention]

[0005] The following detailed descriptions of specific embodiments will be better understood when read in conjunction with the accompanying drawings. To the extent that the drawings illustrate functional blocks of various embodiments, functional blocks do not necessarily represent divisions between hardware components. Therefore, for example, one or more functional blocks (e.g., processors or memory) may be implemented on a single piece of hardware (e.g., a general-purpose signal processor or random-access memory, a hard disk, etc.). Similarly, a program may be a standalone program, incorporated as a subroutine within an operating system, or a function within an installed software package, and so on. It should be understood that the various embodiments are not limited to the arrangements and means shown in the drawings.

[0006] I. Overview of Systems for Biological or Chemical Analysis The examples described herein may be used in a variety of biological or chemical processes and systems for scientific, commercial, or other analytical purposes. More specifically, the examples described herein may be used in a variety of processes and systems where it is desirable to detect events, properties, qualities, or characteristics that exhibit a specified reaction. A bioassay system as described herein may be configured to perform a number of specified reactions that can be detected individually or collectively. For example, a bioassay system may be used to sequence a high-density array of nucleic acid features through repeated cycles of enzymatic manipulation and image acquisition. In some embodiments, nucleic acids may be attached to a surface and amplified. Examples of such amplification are described in U.S. Patent No. 7,741,463, published June 22, 2010, titled "Method of Preparing Libraries of Template Polynucleotides" (the entire disclosure of which is incorporated herein by reference), and / or U.S. Patent No. 7,270,981, published September 18, 2007, titled "Recombinase Polymerase Amplification" (the entire disclosure of which is incorporated herein by reference).

[0007] Components used in a bioassay system may include one or more microfluidic channels for delivering reagents or other reactants to reaction sites. Reaction sites may be randomly distributed across a substantially flat surface, or they may be patterned across a substantially flat surface. Each reaction site may be imaged to detect light emanating from it. Signals indicating photons emitted from the reaction sites and detected by the imaging sensor may provide illumination values. These illumination values ​​may be coupled to images showing the photons detected from the reaction sites. These images may be further analyzed to identify the composition, reaction, conditions, etc., at each reaction site.

[0008] II. Examples of Fluid Element Engineering Devices and Fluid Flow Channels A. Examples of systems with higher volume throughput Figure 1 shows a schematic diagram of one embodiment of a system (100) that may be used to perform analysis on one or more samples of interest. In some implementations, the sample may consist of one or more clusters of linearized nucleotides (e.g., DNA) that form single-stranded DNA (sstDNA). In the shown implementation, the system (100) is configured to receive a flow cell cartridge assembly (102) containing a flow cell assembly (103) and a sample cartridge (104). The system (100) includes a flow cell receptacle (122) that receives the flow cell cartridge assembly (102), a vacuum chuck (124) that supports the flow cell assembly (103), and a flow cell interface (126) used to establish a fluid bond between the system (100) and the flow cell assembly (103). The flow cell interface 126 may include one or more manifolds. The system (100) further includes a shipper manifold assembly (106), a sample loading manifold assembly (108), and a pump manifold assembly (110). The system 100 also includes a drive assembly 112, a controller 114, an imaging system 116, and a waste reservoir 118. The controller (114) is electrically and / or communicatively coupled to the drive assembly (112) and the imaging system (116) and is configured to cause the drive assembly (112) and / or the imaging system (116) to perform various functions as disclosed herein.

[0009] In this embodiment, the flow cell assembly (103) includes a flow cell (128) having a channel (130), fluid-coupled to the channel (130), and defining a plurality of first openings (132) located on a first side (134) of the channel (130). The flow cell (128) further includes a plurality of second openings (136) fluid-coupled to the channel (130) and located on a second side (138) of the channel (130). Thus, fluid may flow through the channel and through the flow cell (128). Although the flow cell (128) is shown to include one channel (130), the flow cell (128) may include two or more channels (130). The flow cell assembly (103) also includes a flow cell manifold assembly (140) connected to the flow cell (128) and having a first manifold fluid line (142) and a second manifold fluid line (144). The flow cell manifold assembly (140) may be in the form of a laminate containing multiple layers, as will be discussed in more detail below.

[0010] In the shown implementation, a first manifold fluid line (142) has a first fluid line opening (146) and is fluid-coupled to each of the first openings (132) of the flow cell (128), and a second manifold fluid line (144) has a second fluid line opening (148) and is fluid-coupled to each of the second openings (136). As shown, the flow cell assembly (103) is coupled to the flow cell manifold assembly (140) and includes a gasket (150) fluid-coupled to the fluid line openings (146, 148). In some implementations in which the flow cell (128) includes multiple channels (130), the flow cell manifold assembly (140) may include an additional fluid line (152) that connects the first fluid line openings (146) to a single manifold port (154). In this configuration, a single gasket (150) may be coupled to a flow cell manifold assembly (140) that surrounds a manifold port (154) and has fluid communication with multiple channels (130). During operation, the flow cell interface (126) engages with the corresponding gasket (150) to establish a fluid coupling between the system (100) and the flow cell (128). The engagement between the flow cell interface (126) and the gasket (150) reduces or eliminates fluid leakage between the flow cell interface (126) and the flow cell (128).

[0011] In the provided implementation, the first manifold fluid line (142) has a portion (156) substantially parallel to the longitudinal axis (158) of the channel (130), and the second manifold fluid line (144) has a portion (160) substantially parallel to the longitudinal axis (158) of the channel (130). The first manifold fluid line (142) is shown at least partially adjacent to the first end (162) of the flow cell (128) and spaced apart from the second end (164) of the flow cell (128), and the second manifold fluid line (144) is shown at least partially adjacent to the second end (164) of the flow cell (128) and spaced apart from the first end (162). However, other arrangements of the manifold fluid lines (142, 144) may prove to be preferable.

[0012] In the shown implementation, the system (100) includes a sample cartridge receptacle (166) that receives a sample cartridge (104) carrying one or more target samples (e.g., analytes). The system (100) also includes a sample cartridge interface (168) that establishes a fluid connection with the sample cartridge (104). The sample loading manifold assembly (108) includes one or more sample valves (170). The pump manifold assembly (110) includes one or more pumps (172), one or more pump valves (174), and a cache (176). The valves (170, 174) and pumps (172) may take any preferred form. The cache (176) may include a meandering cache, which may temporarily store one or more reaction components during bypass operation of the system (100). Although the cache (176) is shown to be contained within the pump manifold assembly (110), the cache (176) may alternatively be located elsewhere (for example, within the shipper manifold assembly (106) or in another manifold downstream of the bypass fluid line (178)).

[0013] The sample loading manifold assembly (108) and the pump manifold assembly (110) transport one or more samples of interest from the sample cartridge (104) to the flow cell cartridge assembly (102) through the fluid line (180). In some implementations, the sample loading manifold assembly (108) may load or handle each of the samples of interest individually into each channel (130) of the flow cell (128). The process of loading the samples of interest into the channels (130) may be performed automatically using a system (100). As shown in Figure 1, the sample cartridge (104) and the sample loading manifold assembly (108) are located downstream of the flow cell cartridge assembly (102). In the implementation shown, the sample loading manifold assembly (108) is coupled between the flow cell cartridge assembly (102) and the pump manifold assembly (110). To draw the target sample from the sample cartridge (104) toward the pump manifold assembly (110), the sample valve (170), the pump valve (174), and / or the pump (172) may be selectively actuated to bias the target sample toward the pump manifold assembly (110). The sample cartridge (104) may include multiple sample reservoirs that are selectively fluid-accessible via the corresponding sample valves (170). To individually flow the target sample toward the channels (130) of the flow cell 128 and away from the pump manifold assembly (110), the sample valves (170), the pump valves (174), and / or the pump (172) may be selectively actuated to bias the target sample toward each channel (130) of the flow cell (128) toward the flow cell cartridge assembly (102).

[0014] The drive assembly (112) interfaces with the sipper manifold assembly (106) and the pump manifold assembly (110) to flow one or more reagents that interact with the sample within flow cell (128). In some scenarios, reversible terminators are attached to the reagents to enable the incorporation of a single nucleotide onto the elongating DNA strand. In some such implementations, one or more of the nucleotides have unique fluorescent labels that emit color when excited. The color (or its absence) is used to detect the corresponding nucleotide. In the illustrated implementation, imaging system (116) excites one or more of the distinguishable labels (e.g., fluorescent labels) and then acquires image data of the distinguishable labels. The labels can be excited by incident light and / or a laser, and the image data may include one or more colors emitted by each label in response to the excitation. The image data (e.g., detection data) may be analyzed by system (100). Examples of features and functions that can be incorporated into imaging system (116) are described in more detail below.

[0015] After the image data is obtained, drive assembly (112) interfaces with sipper manifold assembly (106) and pump manifold assembly (110) to flow another reaction component (e.g., reagent) through flow cell (128), which is then received by waste reservoir (118) via main waste line (182) and / or otherwise discharged by system (100). Some reaction components may perform a flushing operation that chemically cleaves the fluorescent label and reversible terminator from the sstDNA. Next, the sstDNA may be ready for another cycle.

[0016] The main waste liquid line (182) is coupled between the pump manifold assembly (110) and the waste reservoir (118). In some implementations, the pump (172) and / or the pump valve (174) of the pump manifold assembly (110) selectively flow the reaction components from the flow cell cartridge assembly (102) through the fluid line (180) and the sample loading manifold assembly (108) to the main waste liquid line (182). The flow cell cartridge assembly (102) is coupled to the central valve (184) via the flow cell interface (126). The central valve (184) is connected to the flow cell interface (126) via the fluid line (185). The auxiliary waste liquid line (186) is connected to the central valve (184) and the waste reservoir (118). In some implementations, the auxiliary waste liquid line (186) receives the excess fluid of the target sample from the flow cell cartridge assembly (102) via the central valve (184) and flows the excess fluid of the target sample to the waste reservoir (118) when loading the target sample back into the flow cell (128) as described herein.

[0017] The shipper manifold assembly (106) includes a shared line valve (188) and a bypass valve (190). The shared line valve (188) may be referred to as a reagent selector valve. The central valve (184) and the valves (188, 190) of the shipper manifold assembly (106) may be selectively actuated to control the flow of fluid through the fluid lines (192, 194, 196). The shipper manifold assembly (106) may be coupled to a corresponding number of reagent reservoirs (198) via reagent shippers (200). The reagent reservoirs (198) may contain fluid (e.g., reagents and / or other reactive components). In some implementations, the shipper manifold assembly (106) includes multiple ports. Each port of the shipper manifold assembly (106) may accept one of the reagent shippers (200). The reagent shippers (200) may be referred to as fluid lines. Some forms of the reagent shipper (200) may include an array of shipper tubes extending downward along the z-dimension from a port in the body of the shipper manifold assembly (106). Reagent reservoirs (198) may be provided in a cartridge, and the tubes of the reagent shipper (200) may be configured to be inserted into the corresponding reagent reservoirs (198) in the reagent cartridge so that liquid reagents can be drawn from each reagent reservoir (198) into the shipper manifold assembly (106).

[0018] The shared line valve (188) of the sipper manifold assembly (106) is connected to the central valve (184) via the shared reagent fluid line (196). Different reagents may flow through the shared reagent fluid line (196) at different times. In some variations, when performing a flushing operation before changing one reagent with another, the pump manifold assembly (110) may draw wash buffer through the shared reagent fluid line (196), the central valve (184), and the flow cell cartridge assembly (102).

[0019] The bypass valve (190) of the sipper manifold assembly (106) is connected to the central valve (184) via dedicated reagent fluid lines (194, 196). Each of the dedicated reagent fluid lines (194, 196) may be associated with a single reagent. Fluids that may flow through the dedicated reagent fluid lines (194, 196) may be used during the sequencing operation and may include cutting reagents, integration reagents, scanning reagents, cutting wash solutions, and / or wash buffers.

[0020] The bypass valve (190) is also coupled to the cache (176) of the pump manifold assembly (110) via the bypass fluid line (178). One or more reagent priming, hydration, mixing, and / or transfer operations may be performed using the bypass fluid line (178). The priming, hydration, mixing, and / or transfer operations may be performed independently of the flow cell cartridge assembly (102). Thus, operations using the bypass fluid line (178) may be performed, for example, during the incubation of one or more target samples in the flow cell cartridge assembly (102). That is, the shared line valve (188) may be used independently of the bypass valve (190), so that while the shared line valve (188) and / or the central valve (184) are performing other operations simultaneously, substantially simultaneously, or in offset synchronization, the bypass valve (190) may use the bypass fluid line (178) and / or the cache (176) to perform one or more operations.

[0021] The drive assembly (112) includes a pump drive assembly (202) and a valve drive assembly (204). The pump drive assembly (202) may interface with one or more pumps (172) to pump fluid through a flow cell (128) and / or load one or more samples of interest into the flow cell (128). The valve drive assembly (204) may interface with one or more valves (170, 174, 184, 188, 190) to control the position of the corresponding valves (170, 174, 184, 188, 190).

[0022] Figure 2 shows an embodiment of a fluid device (220) that may be incorporated into a modified version of system (100). The fluid device (220) in this embodiment includes a pump manifold assembly (222) which can operate similarly to the pump manifold assembly (110) described above, a sample loading manifold assembly (228) which can operate similarly to the sample loading manifold assembly (108) described above, a flow cell interface (240) which can operate similarly to the flow cell interface (126) described above, a shipper manifold assembly (250) which can operate similarly to the shipper manifold assembly (106) described above, and a waste reservoir (270) which can operate similarly to the waste reservoir (118) described above. The pump manifold assembly (222) is connected to the port assembly (258) of the shipper manifold assembly (250) via a fluid line (224) which may be similar to the fluid line (178), and is connected to the sample loading manifold assembly (228) via a fluid line (226). The sample loading manifold assembly (228) is connected to a flow cell interface (240) via a fluid line (230) which may be similar to fluid line (180), and to a port assembly (258) via fluid lines (232, 234). The flow cell interface (240) is connected to a shipper manifold assembly (250) via a fluid line (242) which may be similar to fluid line (185). The shipper manifold assembly (250) includes a manifold body (252) and a common output port (256) which provides fluid communication via fluid line (185). A valve assembly (254) controls the flow of fluid through the common output port (256) and may operate similarly to a central valve (184). The port assembly (258) of the shipper manifold assembly (250) is connected to a waste reservoir (270) via a fluid line (272) which may be similar to fluid line (186).

[0023] Multiple reagent shippers (260) extend from the manifold body (252) and are fluid-coupled to a valve assembly (254) via their respective fluid channels (262) within the manifold body (252). The reagent shippers (260) may operate similarly to the reagent shipper (200). The valve assembly (254) can selectively connect the fluid channels (262) to a flow cell interface (240) via a common output port (256) and a fluid line (230), thereby enabling it to selectively supply different reagents to the flow cell interface (240). In other words, if each reagent shipper (260) is located within a different reagent (e.g., within each reagent reservoir (198)), a flow cell (e.g., a flow cell (128)) connected to the flow cell interface (240) may selectively accept these different reagents based on the control of the valve assembly (254).

[0024] The port assembly (258) may also provide a fluid interface between the pump manifold assembly (222) and the shipper manifold assembly (250), thereby enabling the shipper manifold assembly (250) to receive pressurized fluid from the pump manifold assembly (222). The port assembly (258) may also provide a fluid interface between the sample loading manifold assembly (228) and the shipper manifold assembly (250), thereby enabling the shipper manifold assembly (250) to receive sample fluid from the sample loading manifold assembly (228). The port assembly (258) may also provide a fluid interface between the waste reservoir (270) and the shipper manifold assembly (250), thereby enabling the shipper manifold assembly (250) to communicate waste liquid with the waste reservoir (270). Fluid communication through the port assembly (258) may be at least partially regulated by the valve assembly (254).

[0025] Referring back to Figure 1, the controller (114) of this embodiment includes a user interface (206), a communication interface (208), one or more processors (210), and a memory (212) that stores instructions executable by one or more processors (210) for performing various functions, including the disclosed implementation. The user interface (206), the communication interface (133), and the memory (212) are electrically and / or communicatively coupled to one or more processors (210). The user interface (206) may be adapted to receive input from a user and provide the user with information associated with the operation of the system (100) and / or analysis performed. The user interface (206) may include a touchscreen, a display, a keyboard, a speaker, a mouse, a trackball, and / or a voice recognition system.

[0026] The communication interface (208) is adapted to enable communication between the system (100) and a remote system (e.g., a computer) via a network (e.g., the Internet, an intranet, a Local-Area Network (LAN), a Wide-Area Network (WAN), a coaxial cable network, a wireless network, a wired network, a satellite network, a Digital Subscriber Line (DSL) network, a cellular network, a Bluetooth connection, a Near Field Communication (NFC) connection, etc.). Some of the communication provided to the remote system may be associated with analysis results, imaging data, etc., generated or otherwise acquired by the system (100). Some of the communication provided to the system (100) may be associated with fluid analysis operations, patient records, and / or protocols performed by the system (100).

[0027] One or more processors (210) and / or systems (100) may include one or more processor-based systems or microprocessor-based systems. In some implementations, one or more processors (210) and / or systems (100) may include one or more programmable processors, programmable controllers, microprocessors, microcontrollers, graphics processing units (GPUs), digital signal processors (DSPs), reduced-instruction set computers (RISCs), application-specific integrated circuits (ASICs), field programmable gate arrays (FPGAs), field programmable logic devices (FPLDs), logic circuits, and / or other logic-based devices that perform various functions, including those described herein.

[0028] Memory (212) includes semiconductor memory, magnetically readable memory, optical memory, hard disk drives (HDDs), optical storage drives, solid-state storage devices, solid-state drives (SSDs), flash memory, read-only memory (ROM), erasable programmable read-only memory (EPROM), electrically erasable programmable read-only memory (EEPROM), random-access memory (RAM), non-volatile RAM (NVRAM) memory, compact discs (CDs), compact disc read-only memory (CD-ROMs), digital versatile discs (DVDs), Blu-ray discs, and redundant arrays of independent discs. It may include one or more of the following storage devices or storage disks: a disk, RAID system, cache, and / or any other storage device or storage disk in which information is stored over any duration (e.g., permanently, temporarily, over a long period, for buffering, for caching).

[0029] B. Examples of systems with lower volume throughput Figure 3 shows a schematic diagram of another embodiment of system (300) that may be used to perform analysis on one or more samples. Unless otherwise described below, system (300) in this embodiment may be configured and operable in the same way as system (100) described above with reference to Figure 1. In some embodiments, system (100) is used to provide a higher volume throughput, while system (300) is used to provide a lower volume throughput. Alternatively, systems (100, 300) may provide any other suitable volume or degree of throughput. System (300) in this embodiment receives a reagent cartridge (302) and includes, in part, a gas source (304), a drive assembly (306), a controller (308), an imaging system (310), and a waste reservoir (312). The reagent cartridge (302) may be referred to as a consumable, a reagent reservoir, or a reagent assembly. The controller (308) is electrically and / or communicatively coupled to the drive assembly (306) and the imaging system (310), and causes the drive assembly (306) and / or the imaging system (310) to perform various functions as disclosed herein.

[0030] The reagent cartridge (302) in the shown configuration includes a well assembly (314) having a body (316). The body (316) has a first wall (318) defining a well (320) having a port (322). The first wall (318) has a distal end (324) defining an opening (326) having an opening periphery (328). The second wall (330) surrounds the first wall (318) and has a distal end (332). The distal end (332) may be referred to as the edge or outer edge. A cover (334) is connected to the distal end (324) of the first wall (318), covering the opening (326) along the opening periphery (328) at the connection portion (336), and is disconnected from the distal end (324) of the first wall (318) at the disconnected portion (338). The connection portion (336) may be referred to as a connection section or connection segment, and the disconnected portion (338) may be referred to as a disconnection section or disconnection segment. The first wall (318) has a height, and the second wall (330) has a height greater than the height of the first wall (318). Alternatively, the first well (318) and the second well (330) may be the same or similar height. An impermeable barrier (340) is connected to the distal end (332) of the second wall (330) and covers the well (320). The impermeable barrier (340) may be foil, plastic, or the like, and may prevent or block moisture from penetrating into the wells (320) of the reagent cartridge (302).

[0031] The unconnected portion (338) of the cover (334) forms a vent (342) that allows airflow from the well (320). The dry reagent (348) is contained within the well (320), and the vent (342) is sized to substantially hold the dry reagent (348) within the well (320). The body (316) may contain multiple wells (320), while Figure 3 shows one well (320). The liquid (346) may actually flow into the well (320) via the port (322) to rehydrate the dry reagent (348). The vent (342) may allow gas to escape from the well (320) as the liquid (346) flows into the well (320), and the cover (334) prevents or prevents the reagent (348) and / or liquid (346) from leaking out of the well (320). In other words, the vent (342) holds the reagent (348) and / or liquid (346) within the well (320) and prevents or blocks the reagent (348) and / or liquid (346) from moving out of the well (320). The vent (342) and cover (334) prevent or block secondary contamination between reagents when the reagent cartridge (302) contains two or more wells (320). The liquid (346) and dry reagent (348) may flow into and out of the well (320) to mix the liquid (346) and dry reagent (348) from the liquid reservoir (362). The system (300) and / or reagent cartridge (302) may include a mixing chamber used for mixing the liquid (346) and dry reagent (348) in some implementations. The impermeable barrier (340) may be penetrated before the liquid (346) flows into the well (320).

[0032] A gas source (304) may be used to pressurize a liquid reservoir (362) to draw liquid (346) into a well (320), and / or a pump (350) may draw liquid (346) from the liquid reservoir (362) and flow liquid (346) into the well (320) to rehydrate the reagent (348). The gas source (304) may be provided by a system (300) and / or supported by a reagent cartridge (302). The gas source (304) may be omitted as an alternative. The pump (350) may be implemented by a syringe pump, a peristaltic pump, a diaphragm pump, etc. The pump (350) may be located downstream of the flow cell (368) as shown, the pump (350) may be located upstream of the flow cell (368), or may be omitted entirely.

[0033] The reagent cartridge (302) and / or system (300) includes a valve (352) which may be selectively actuated to control the flow of fluid through a fluid line (356). Such a valve (352) may be implemented by a valve manifold, rotary valve, switching valve, pinch valve, flat valve, solenoid valve, check valve, piezo valve, etc. A regulator (354) may be positioned between the gas source (304) and the valve (352) and regulate the pressure of the gas supplied to the valve (352). The regulator (354) may include a valve that controls the flow of gas from the gas source (304).

[0034] The body (316) of the well assembly (314) has a rim (364), and the impermeable barrier (340) may be hermetically connected to the body (316) along the rim (364). The impermeable barrier (340) may include foil, plastic, and / or any other suitable material. The system (300) may penetrate the impermeable barrier (340), the impermeable barrier (340) may be penetrated individually before use, or the impermeable barrier (340) may be penetrated by some other structure or method. The system (300) includes an actuator assembly (360) in the shown configuration that interfaces with the impermeable barrier (340) to penetrate the impermeable barrier (340). The system (300) may include projections such as struts having blunt or sharp ends that are movable by the actuator assembly (360) to penetrate the impermeable barrier (340). Alternatively, the impermeable barrier (340) may be punctured by an operator before the reagent cartridge (302) is positioned within the system (300). The system (300) also includes a liquid reservoir (362) containing liquid (346). Liquid (346) may include a rehydration solution, a washing buffer, and / or any other suitable type of liquid.

[0035] The system (300) further includes a flow cell receptacle (366) that receives a flow cell (368). The flow cell (368) may be configured and operable in the same way as the flow cell (128). In some modifications, the flow cell (368) is supported by and / or incorporated into a reagent cartridge (302). The flow cell 368 may support the sample of interest. A gas source (304) and / or a pump (350) may flow one or more liquid reagents through a reagent cartridge (302) that interacts with the sample, by flowing liquid (346) to rehydrate the dry reagent (348). The imaging system (310) may be configured and operable in the same way as the imaging system (116) so that the imaging system (310) can be used to acquire image data from the flow cell (368). After image data is obtained, the drive assembly (306) may interface with the reagent cartridge (302) to flow another reaction component (e.g., a reagent) through the flow cell (368), which is then received by the waste reservoir (312) and / or discharged by the reagent cartridge (302) in another manner. In this embodiment, the drive assembly (306) includes a pump drive assembly (370), a valve drive assembly (372), and an actuator assembly (360). The pump drive assembly (370) interfaces with the pump (350) to pump fluid through the reagent cartridge (302) and / or the flow cell (368), and the valve drive assembly (372) interfaces with the valve (352) to control the position of the valve (352).

[0036] The controller (308) in this embodiment includes a user interface (374), a communication interface (376), a processor (378), and memory (380). The user interface (374) may be configured and operable as the user interface (206) of the system (100). The communication interface (376) may be configured and operable as the communication interface (208) of the system (100). The processor (378) may be configured and operable as the processor (210) of the system (100). The memory (380) may be configured and operable as the memory (212) of the system (100).

[0037] Further examples and details of how the various features of each system (100, 300) can be configured and operated are described below. As merely a further example, various features of the system (100, 300) may be configured and operable in accordance with at least some of the teachings of International Publication No. 2023 / 055873, title of the invention "Flow Cells and Related Flow Cell Manifold Assemblies and Methods," published on April 6, 2023 (the entire disclosure of which is incorporated herein by reference), U.S. Patent No. 9,958,465, title of the invention "Detection Apparatus having a Microfluorometer, a Fluidic System, and a Flow Cell Latch Clamp Module," issued on May 1, 2018 (the entire disclosure of which is incorporated herein by reference), and / or U.S. Patent Application Publication No. 63 / 325,462, title of the invention "Well Assemblies and Related Systems and Methods," filed on March 30, 2022 (the entire disclosure of which is incorporated herein by reference).

[0038] III. Examples of Flow Cell Structures As described above, the system (100, 300) may carry out a reaction within a flow cell (128, 368) and / or perform analysis on one or more target samples within the flow cell (128, 368). Examples of possible forms of such a flow cell (128, 368) are described below, but it should be understood that the flow cell (128, 368) may take various other forms and may have various other features in addition to, or instead of, those described below.

[0039] A. Examples of single-surface patterned flow cells Figure 4 shows an embodiment of a flow cell (400) comprising a patterned substrate (402) including recesses (404) separated by interstitial regions (406) and surface chemistry (410, 412) positioned within the recesses (404). The recesses (404) may be in the form of microwells or nanowells. The recesses (404) may contain nucleic acid chains or other oligonucleotides and thereby be configured to provide reaction sites for SBS and / or other types of processes. In some variants, each recess (404) has a cylindrical configuration with a generally circular cross-sectional profile. In some other variants, each recess (404) has a polygonal (e.g., hexagonal, octagonal, square, rectangular, elliptical, etc.) cross-sectional profile. Alternatively, the recesses (404) may have any other preferred configuration. It should also be understood that the recesses (404) may be arranged in any preferred pattern, including but not limited to a grid pattern.

[0040] The surface chemistry (410, 412) in this embodiment includes a functionalized coating layer (410) and a primer (412). Although not shown, the recess (404) may also have a surface preparation or treatment chemical (e.g., silane or silane derivative) positioned between the substrate (402) and the functionalized coating layer (410). This same surface preparation or treatment chemical may also be positioned on the gap region (406). In this embodiment, the hydrogel (440) is applied before the lid (420) is bonded to the substrate (402). The hydrogel (440) covers the surface chemistry (410, 412) in the recess (404) and at least a portion of the patterned substrate (402) (e.g., the gap region (406) which is not the bonding region (422)). As just one example, the hydrogel (440) may include PAZAM, crosslinked polyacrylamide, agarose gel, etc.

[0041] The flow cell (400) in this embodiment further includes a lid (420) bonded to a bonding region (422) of a patterned substrate (402). In the embodiment shown in Figure 4, the lid (420) includes a top (424) connected to several side walls (426), these components (424, 426) define a portion of each of the six channels (430A, 430B, 430C, 430D, 430E, 430F). Each side wall (426) isolates one channel (430A, 430B, 430C, 430D, 430E, 430F) from each adjacent channel (430A, 430B, 430C, 430D, 430E, 430F). Each flow path (430A, 430B, 430C, 430D, 430E, 430F) is selectively in fluid communication with each set of recesses (404).

[0042] The lid (420) may be bonded to the bonding region (422) of the substrate (402) using any preferred technique such as laser bonding, diffusion bonding, anodic bonding, eutectic bonding, plasma-activated bonding, glass frit bonding, or other methods well known in the art. In some variations, a spacer layer (428) may be used to bond the lid (420) to the bonding region (422). The spacer layer (428) may include any material that seals together at least a portion of the gap region (404) (e.g., the bonding region (422)) between the substrate (402) and the lid (420). Although not shown, the lid (420) or patterned substrate (402) may include inlet and outlet ports that fluidically engage with other ports (not shown), such as those of the sample cartridge interface (168), to direct fluid into each channel (430A, 430B, 430C, 430D, 430E, 430F) (e.g., from a reagent cartridge or other fluid storage system) and out of the channels (e.g., to a waste reservoir (118) or another waste removal system). The channels (430A, 430B, 430C, 430D, 430E, 430F) may function to selectively introduce reactants or reactants into the hydrogel (440) and the underlying surface chemistry (410, 412) to initiate a specified reaction, for example, within a recess (404).

[0043] The flow cell (400) includes a pattern of recesses (404) for providing an array of reaction sites, but other modifications may provide reaction sites on or thereon on a variety of other types of structural features, including but not limited to continuously flat surfaces and / or protruding surfaces. As just a further example, the flow cell (400) may be constructed and operable in accordance with at least part of the teachings of U.S. Patent No. 10,919,033, issued February 16, 2021, titled “Flow Cells with Hydrogel Coating,” the entire disclosure of which is incorporated herein by reference.

[0044] B. Examples of dual-surface patterned flow cells Figure 4 shows an example of a flow cell (400) having a single surface patterned with reaction sites (i.e., recesses (404) formed in the substrate (402)). However, in some cases, it may be desirable to provide a variation of the flow cell (400) that provides two surfaces patterned with reaction sites. An example of a double-surface patterned flow cell (450) is shown in Figures 5-6. In this example, the flow cell (450) includes a pair of wafers (452, 454) joined together, with a spacer layer (456) inserted between the wafers (452, 454). Each wafer (452, 454) is patterned to provide a number of recesses (462, 464) such that when the flow cell (450) is assembled, the recesses (462) of wafer (452) align with the recesses (464) of wafer (454). The recesses (462) are separated from each other by gap regions (466), and the recesses (464) are separated from each other by gap regions (468). In this embodiment, the spacer layer (456) does not come into contact with the gap regions (466, 468).

[0045] The recesses (462, 464) of the flow cell (450) are configured and may be operable in the same manner as the recess (404) of the flow cell (400) described above. Each recess (462, 464) in this embodiment includes a grafted coating (470) which may be similar to the functionalized coating layer (410), and a primer (472) which may be similar to the primer (412) described above. Each recess (462, 464) may further include a hydrogel such as a hydrogel (440), and / or any other preferred features. As shown in Figures 5-6, the recesses (462, 464) are located within a plurality of channels (480A, 480B, 480C, 480D). The channels (480A, 480B, 480C, 480D) are separated from each other by walls (458) and ends (459) formed by a spacer layer (456). In the embodiment shown in Figure 6, the flow cell (450) provides four channels (480A, 480B, 480C, 480D), each channel (480A, 480B, 480C, 480D) containing several rows of recesses (462, 464). When the flow cell (450) is used in a system (100, 300), the channels (480A, 480B, 480C, 480D) may function to selectively introduce reactants or reactant surface chemistry (470, 472) to initiate a specified reaction in / at the recesses (462, 464), for example. In some cases, each wafer (452, 454) has its own set of recesses (462, 464) that provide corresponding reaction sites, so that the flow cell (450) can provide twice the reaction of a flow cell (400) of similar size over a given period of time.

[0046] The dashed lines in Figure 6 show how the flow cell (450) can be diced to effectively form smaller flow cells (450A, 450A), each having its own pair of channels (480A, 480B, 480C, 480D). However, in this embodiment, a single flow cell (450) has three or more channels (480A, 480B, 480C, 480D). The flow cell (450) includes a pattern of recesses (462, 464) for providing an array of reaction sites, but other modifications may provide reaction sites on or thereon on a variety of other types of structural features, including but not limited to continuously flat surfaces and / or protruding surfaces. As merely a further example, a flow cell (450) may be constructed and operational in accordance with at least part of the teachings of U.S. Patent No. 10,955,332, titled "Flow Cell Package and Method for Making the Same," issued March 23, 2021, the entire disclosure of which is incorporated herein by reference.

[0047] IV. Examples of Imaging System Features As described above, the system (100, 300) includes an imaging system (116, 310) that excites one or more identifiable labels (e.g., fluorescent labels) in a sample within a reaction site provided by recesses (404, 462, 464) of flow cells (128, 368, 400, 450), and subsequently acquires image data of the identifiable labels. This image data is used to identify nucleotides as part of a nucleic acid sequencing process. Alternatively, the image data may be used for various other purposes. The following description provides details on how several variations of the imaging system (116, 310) may be configured and operational.

[0048] Figure 7 shows a schematic diagram of another embodiment of system (500) that may be used to perform analysis on one or more samples of interest. Unless otherwise described below, system (500) in this embodiment may be configured and operable in the same way as systems (100, 300) described above. System (500) is configured to perform a number of parallel reactions within a flow cell (510). Flow cell (510) may be configured and operable in the same way as flow cells (400, 450) described above, or may have any other preferred configuration. Flow cell (510) may therefore include one or more channels that receive the solution from system (500) and direct the solution toward the reaction sites in flow cell (510).

[0049] The system (500) includes a system controller (520) capable of communicating with various components, assemblies, and subsystems of the system (500). The controller (520) may be configured and operable in the same manner as the controllers (114, 308) described above. The imaging assembly (522) of the system (500) includes a light-emitting assembly (550) that emits light that reaches the reaction site on the flow cell (510). The light-emitting assembly (550) may include an incoherent light emitter (e.g., emitting a light beam output by one or more excitation diodes) or a coherent light emitter such as a light emitter output by one or more lasers or laser diodes. In some implementations, the light-emitting assembly (550) may include several different light sources (not shown), each emitting light in a different wavelength range. Some variations of the light emission assembly (550) may also include one or more collimating lenses (not shown), a light-structured optical assembly (not shown), a projection lens (not shown) that can be operated to adjust the structured beam shape and path, an epifluorescence microscope component, and / or other components. Although the system (500) is shown as having a single light emission assembly (550), some other implementations may include multiple light emission assemblies (550).

[0050] In this embodiment, light from the light-emitting assembly (550) is directed by the dichroic mirror assembly (546) through the objective lens assembly (542) onto the sample in the flow cell (510) positioned on the mobile stage (570). In fluorescence microscopy of the sample, the fluorescent element associated with the sample emits fluorescence in response to excitation light, and the resulting light is collected by the objective lens assembly (542) and directed to the imaging sensor of the camera system (540) to detect the emitted fluorescence. In some implementations, the imaging lens assembly may be positioned between the objective lens assembly (542) and the dichroic mirror assembly (546), or between the dichroic mirror (546) and the imaging sensor of the camera system (540). The movable lens element may be able to translate along the longitudinal axis of the imaging lens assembly, taking into account spherical aberration introduced by focusing onto the upper or lower inner surface of the flow cell (510) and / or by the movement of the objective lens assembly (542).

[0051] In this embodiment, a filter switching assembly (544) is inserted between the dichroic mirror assembly (546) and the camera system (540). The filter switching assembly (544) includes one or more emission filters that can be used to allow emission wavelengths within a specific range to pass through and to block (or reflect) emission wavelengths within other ranges. For example, emission filters may be used to direct different wavelength ranges of emitted light to different imaging sensors of the camera system (540) of the imaging assembly (522). For example, the emission filters may be implemented as dichroic mirrors that direct emitted light of different wavelengths from the flow cell (510) to different imaging sensors of the camera system (540). In some modifications, a projection lens is inserted between the filter switching assembly (544) and the camera system (540). The filter switching assembly (544) may be omitted in some variations.

[0052] The system (500) further includes a fluid delivery assembly (590) capable of directing the flow of reagents (e.g., fluorescently labeled nucleotides, buffers, enzymes, dissection reagents, etc.) to a flow cell (510) and a waste valve (580) (and through the flow cell (510) and the waste valve (580)). The fluid delivery assembly (590) may be configured and operable as the various fluid delivery components described above in the context of Figures 1 to 3. The system (500) of this embodiment also includes a temperature station actuator (530) and a heater / cooler (532) capable of optionally adjusting the temperature conditions of the fluid in the flow cell (510). In some implementations, the heater / cooler (532) may be fixed to and / or integrated into the sample stage (570) on which the flow cell (510) is mounted.

[0053] The flow cell (510) may be detachably mounted on a sample stage (570) which can provide movement and alignment of the flow cell (510) relative to the objective lens assembly (542). The sample stage (570) may have one or more actuators to enable movement of the sample stage (570) in any of three dimensions. For example, actuators may be provided to enable the sample stage (570) to move in the x, y, and z directions relative to the objective lens assembly (542), to tilt relative to the objective lens assembly (542), and / or to move relative to the objective lens assembly (542). Movement of the sample stage (570) may enable one or more sample positions on the flow cell (510) to be optically aligned and positioned with respect to the objective lens assembly (542). Movement of the sample stage (570) relative to the objective lens assembly (542) may be achieved by moving the sample stage (570) itself, by moving the objective lens assembly (542), by moving some other components of the imaging assembly (522), by moving some other components of the system (500), or by any combination thereof. For example, in some implementations, the sample stage (570) may be operable in the x and y directions relative to the objective lens assembly (542), while the focal component (562) or z-stage may move the objective lens assembly (542) along the z-direction relative to the sample stage (570).

[0054] In some implementations, a focusing component (562) may be included to control the positioning of one or more elements of the objective lens assembly (542) relative to the flow cell (510) in the focal direction (e.g., along the z-axis or z-dimension). The focusing component (562) may include one or more actuators physically coupled to the objective lens assembly (542), an optical stage, a sample stage (570), or a combination thereof, to move the flow cell (510) on the sample stage (570) relative to the objective lens assembly (542) and provide proper focusing for the imaging operation. In this embodiment, the focal component (562) is configured to detect the displacement of the objective lens assembly (542) relative to a portion of the flow cell (510) and output data indicating the focal position to the focal component (562) or to its components, or a focus tracking module (560) is used which can control the focal component (562), such as a controller (520), to move the objective lens assembly (542) and position the corresponding portion of the flow cell (510) at the focal point of the objective lens assembly (542).

[0055] In some implementations, actuators for the focal component (562) or sample stage (570) may be physically coupled to the objective lens assembly (542), optical stage, sample stage (570), or a combination thereof, for example, by direct or indirect mechanical, magnetic, fluid, or other mounting or contact to or with the stage or its components. The actuator for the focal component (562) may be configured to move the objective lens assembly (542) in the z-direction while keeping the sample stage (570) in the same plane (for example, while maintaining a level or horizontal orientation perpendicular to the optical axis). In some implementations, the sample stage (570) includes x-direction actuators and y-direction actuators to form an xy-stage. The sample stage (570) may also be configured to include one or more tilt or inclination actuators to tilt or incline the sample stage (570) and / or a portion thereof, taking into account the gradient of its surface.

[0056] The camera system (540) may include one or more imaging sensors for monitoring and tracking the imaging (e.g., sequencing) of the flow cell (510). The camera system (540) may be implemented, for example, as a CCD or CMOS imaging sensor camera, but other imaging sensor technologies (e.g., active pixel sensors) may be used. As just further examples, the camera system (540) may include a dual-sensor time delay integration (TDI) camera, a single-sensor camera, a camera having one or more two-dimensional imaging sensors, and / or other types of camera technologies. Although the camera system (540) and associated optical components are shown positioned above the flow cell (510) in Figure 7, one or more imaging sensors or other camera components may be incorporated into the system (500) in a number of other ways, as will be apparent to those skilled in the art by considering the teachings herein. For example, one or more imaging sensors may be positioned below the flow cell (510), such as within or below the sample stage (570), or they may be incorporated into the flow cell (510).

[0057] Figure 8 shows embodiments of various components that may be incorporated into the imaging assembly (522) of the system (500). In particular, the configuration shown in Figure 8 may represent a variation of the light emission assembly (550). The configuration shown in Figure 8 may be particularly useful in a scenario in which the camera system (540) includes a TDI camera. In the configuration shown in Figure 8, the Line Generation Module (LGM) (602) and the Emission Optics Module (EOM) (604) are mechanically coupled to and aligned with a precision mounting plate (610). The EOM (604) includes an objective lens assembly (606) aligned with an imaging lens (620) via a mirror (608), and the imaging lens is optically coupled to the LGM (602). The LGM (602) may include one or more light sources (e.g., coherent light sources such as laser diodes). In some embodiments, the LGM(602) may include a first light source configured to emit red wavelength light and a second light source configured to emit green wavelength light. The LGM(602) may further include optical components such as focal planes, lenses, reflectors, or mirrors. The optical components may be located within the housing of the LGM(602) and may focus the light emitted from one or more light sources by directing it into adjacent module subassemblies. One or more of the optical components of the LGM(602) may also be configured to shape the light emitted from one or more light sources into a desired pattern. For example, in some implementations, the optical components may shape the light into a line pattern (e.g., by using one or more Powell lenses or other beam-shaping lenses, diffraction or scattering components). In some modifications, the LGM(602) may include one or more laser modules that can be individually removed from and replaced from the LGM(602).

[0058] The light beam generated by the LGM(602) passes through an interface baffle between the LGM(602) and the EOM(604), through the objective lens assembly(606), and strikes an optical target (e.g., a flow cell(510)). In some variations, the interface baffle includes an aperture shaped to allow light to pass through its center while obscuring interference from an external light source. The response light emission from the target can pass back through the objective lens assembly(606) and enter the imaging lens(622). The lens element(622), which may form part of the imaging lens(620), is configured to articulate along an axis (e.g., the z-axis) to correct spherical aberration artifacts introduced by the objective lens assembly(606) as it images through varying thicknesses of the flow cell(510) components. As shown in the figure, the lens element (622) may articulate to move closer to or further away from the objective lens assembly (606) to adjust the beam shape and path. The objective lens assembly (606) may emit excitation light toward an optical target (e.g., a flow cell (510)) and receive fluorescence emission from the optical target. The actuator may be configured to position the objective lens assembly (606) in a target region close to the optical target. Next, the processor of the controller (520) may execute program instructions to detect fluorescence emission from the optical target.

[0059] Figure 9 shows an embodiment of another configuration that may be provided within the imaging assembly (522). In particular, Figure 9 shows the imaging assembly (650) positioned relative to a flow cell (670). The flow cell (670) may represent any of the variations of the flow cells (128, 368, 400, 450, 510) described herein. The flow cell (670) has an upper layer (671) and a lower layer (673) separated by a fluid-filled channel (675). In the shown configuration, the upper layer (671) is optically transparent, and the imaging assembly (650) is focused on a region (676) on the inner surface (672) of the upper layer (671). In other variations, the imaging assembly (650) may be focused on the inner surface (674) of the lower layer (673). One or both of the surfaces (672, 674) may include array feature regions detected by the imaging assembly (650).

[0060] The imaging assembly (650) includes an objective lens assembly (666) configured to direct excitation radiation from the light-emitting assembly (652) to the flow cell (670), and radiation from the flow cell (670) to the detector (664). In the illustrated configuration, excitation radiation from the light-emitting assembly (652) passes through a lens (658), a beam splitter (660), and the objective lens assembly (666) to reach the flow cell (670). In this embodiment, the light-emitting assembly (652) includes two light-emitting diodes (LEDs) (656, 654) that produce radiation of different wavelengths. The radiation emitted from the flow cell (670) is captured by the objective lens assembly (666), reflected by the beam splitter (660), and passes through the tuner optical system (662) to the detector (664) (e.g., a CMOS sensor). The beam splitter (660) serves to direct the emitted radiation in a direction perpendicular to the path of the excitation radiation. The position of the objective lens assembly (666) may be moved in the z dimension to change the focus of the imaging assembly (650). The imaging assembly (650) may be moved back and forth in the y direction to capture images of several regions of at least one inner surface (672, 674) of the flow cell (670).

[0061] In this embodiment, a single imaging assembly (650) includes two LEDs (656, 654) that emit light at two different wavelengths, and a single detector (664) detects the light emitted from the fluorophore label in the flow cell (670) in response to irradiation at these two different wavelengths. In some other variations, there are two or more imaging assemblies (650), each imaging assembly (650) including a single LED (656, 654) and a single detector (664), and each imaging assembly (650) provides irradiation at only one single wavelength. In another variation, two or more detectors (664) may receive excitation radiation from a common emission assembly (652).

[0062] Figure 10 shows an embodiment of another configuration that may be provided within the imaging assembly (522). In particular, Figure 10 shows the imaging assembly (700) positioned relative to a flow cell (774). The flow cell (774) may represent any of the variations of the flow cells (128, 368, 400, 450, 510) described herein. The flow cell (770) has a translucent cover plate (772), a substrate (774), and a liquid layer (776) inserted between the cover plate (772) and the substrate (774). A biological sample may be placed on the inner surface of the cover plate (772) (above the liquid layer (776)) and / or on the inner surface of the substrate (774) (below the liquid layer (776)).

[0063] The imaging assembly (700) of this embodiment includes a LGM (710) with two light sources (712, 714) arranged inside. The light sources (712, 714) may include laser diodes, diode-pumped solid-state lasers, or other light sources known in the art that emit laser beams of different wavelengths (e.g., red or green light). The light beams emitted from the light sources (712, 714) are directed through one or more beam-shaping lenses (716). In some implementations, one or more photo-shaping lenses may be used to shape the light beams emitted from each or both light sources. The LGM (710) may use one or more Powell lenses to diffuse and / or shape the laser beams from single-mode or nearly single-mode laser sources. Other beam-shaping optical systems, such as active beam expanders, attenuators, a single relay lens, a cylindrical lens, an actuated mirror, a diffracting element, and scattering components, may be used to control uniformity and increase tolerances. The laser beams may intersect at the back focus of the objective lens to provide better tolerances on the surface of the flow cell (770).

[0064] The LGM(710) in this embodiment further includes mirrors (718, 720). A light beam generated by a light source (712) is reflected by mirror (718) and directed into the EOM(740) through a single interface port via an aperture or semi-reflective surface of mirror (720). Similarly, a light beam generated by a light source (714) is reflected by mirror (720) and directed into the EOM(740) through a single interface port. In some embodiments, an additional set of articulated mirrors may be incorporated adjacent to the mirrors (718, 720) to provide an additional adjustment surface. Both light beams may be combined using a dichroic mirror (720). Each of the mirrors (718, 720) may be configured to articulate using manual or automatic control to align the light beams from the light sources (712, 714). In this embodiment, the light beam also passes through a shutter element (722).

[0065] The EOM(740) includes an objective lens assembly(756) and a z-stage(758) for moving the objective lens assembly(756) longitudinally closer to or further away from the flow cell(770). The LGM(710) is configured to produce uniform line illumination through the objective lens assembly(756). The z-stage(758) may then move the objective lens assembly(756) to focus the light beam onto one of the inner surfaces of the flow cell(770) (for example, onto a biological sample). In some implementations, the objective lens assembly(756) may be configured to focus the light beam beyond the flow cell(770) to a focal point, for example, to increase the linewidth of the light beam on the surface of the flow cell(770).

[0066] The EOM (740) in this embodiment also includes a semi-reflective mirror (754) for directing light through the objective lens assembly (756) while allowing light returned from the flow cell (774) to pass through. The EOM (740) further includes an imaging lens (744) and a corrector lens (748). The corrector lens (748) may be articulated longitudinally by the z-stage (746) either in a direction toward or away from the objective lens assembly (756) to ensure accurate imaging (e.g., to correct spherical aberration caused by moving the objective lens assembly (756) and / or from imaging through a thicker substrate). Light transmitted through the corrector lens (748) and the imaging lens (744) passes through a filter element (742) and enters the camera system (730). The camera system (730) includes one or more photosensors (732) for detecting light emitted from a biological sample in response to the incident light beam.

[0067] The EOM(740) in this embodiment further includes a semi-reflective mirror (752) that reflects the focus-tracking light beam emitted from the Focus Tracking Module (FTM)(760) to a flow cell (774), and then reflects the light returned from the flow cell (774) back to the FTM(760). The FTM(760) may include a focus-tracking light sensor that detects the characteristics of the returned focus-tracking light beam and generates a feedback signal for optimizing the focus of the objective lens assembly (756) on the flow cell (774).

[0068] The direction, size, and / or polarization of the laser beam may be adjusted by using lenses, mirrors, and / or polarizers. Optical lenses (e.g., cylindrical, spherical, or aspherical) may be used to actively adjust the illumination focus on the bifacial surface of the flow cell (770) target. The LGM(710) may also include multiple units, each designed for a specific / different wavelength and polarization. Stacking multiple units may be used to increase the laser power and wavelength options. Two or more laser wavelengths may be combined with dichroic and polarizers.

[0069] As merely an example, other components of the focus tracking module (560) and / or imaging assembly (522) may be constructed and operable in accordance with at least some of the teachings of U.S. Patent No. 10,416,428, issued September 17, 2019, titled "Systems and Methods for Improved Focus Tracking Using a Light Source Configuration" (the entire disclosure of which is incorporated herein by reference), U.S. Patent Application No. 63 / 300,531, filed January 18, 2022, titled "Dynamic Detilt Focus Tracking" (the entire disclosure of which is incorporated herein by reference), and U.S. Patent Application Publication No. 63 / 410,961, filed September 28, 2022, titled "Spot Error Handling for Focus Tracking" (the entire disclosure of which is incorporated herein by reference). As merely a further example, components of an imaging assembly (522) may be configured and operable in accordance with at least some of the teachings of U.S. Patent No. 10,774,371, entitled “Laser Line Illuminator for High Throughput Sequencing,” issued on 15 September 2020 (the entire disclosure of which is incorporated herein by reference), and U.S. Patent No. 9,958,465, entitled “Detection Apparatus having a Microfluorometer, a Fluidic System, and a Flow Cell Latch Clamp Module,” issued on 1 May 2018 (the entire disclosure of which is incorporated herein by reference).

[0070] VI. Examples of Multispectral Imaging Apparatus A. Multispectral imaging device with color separation assembly It may be desirable to provide imaging in two or more spectral channels during the sequencing process. In some embodiments, the emission assembly (550, 652) or line generation module (LGM) (602, 710) may be configured to simultaneously deliver multispectral light (i.e., light of multiple wavelengths) to two different locations on the flow cell (128, 368, 400, 450, 510), for example, via two or more excitation sources that generate excitation beams at different wavelengths. In some such scenarios, the corresponding emission captured from the reaction site on the flow cell (128, 368, 400, 450, 510) may also be at different wavelengths. Other scenarios may exist in which a fluorescent label at the reaction site on the flow cell (128, 368, 400, 450, 510) emits multispectral light in response to excitation from one or more illumination sources such as the emission assembly (550, 652) or line generation module (LGM) (602, 710).

[0071] When a fluorescent label at a reaction site on a flow cell (128, 368, 400, 450, 510) emits multispectral light in response to excitation from one or more illumination sources such as an emission assembly (550, 652) or a line generating module (LGM) (602, 710), it may be desirable to provide an arrangement of optical features that provides spatial separation of different wavelengths in the multispectral emitted light, while still maintaining a fixed optical path during sample scanning of the reaction site on the flow cell (128, 368, 400, 450, 510), regardless of whether such multispectral light is emitted from one spatial position on the flow cell (128, 368, 400, 450, 510) or from multiple spatial positions on the flow cell (128, 368, 400, 450, 510). In some implementations, multiple light emission path control is facilitated by the use of multiple imaging sensors within the camera system (540, 730), where the imaging sensors are displaced from one another.

[0072] In scenarios where different colors of a multispectral emission beam are spatially separated, it may be even more desirable to minimize the number of imaging lenses (620, 744) so ​​that a single imaging lens (620, 744) can be used despite the spatial separation of colors from the multispectral emission beam. Such minimization of the number of imaging lenses (620, 744) in an imaging system (116, 310) or imaging assembly (522, 700) may minimize manufacturing costs and reduce risks associated with different magnifications and distortions within each spectral channel, which may arise due to differences in manufacturing tolerances between two different optical components. As will be described in more detail below, an imaging system (116, 310) or imaging assembly (522, 700) can achieve multispectral imaging (e.g., providing four or more spectral channels) using a single imaging lens (620, 744) by including one or more correcting elements intended to induce astigmatism, without requiring a moving optical system.

[0073] Wavelength separation of a multispectral beam can be achieved using a color separation assembly (800) as shown in Figure 11. In this embodiment, the color separation assembly (800) is used to separate the wavelengths of a multispectral beam emitted from a reaction site on a flow cell (128, 368, 400, 450, 510), but the color separation assembly (800) may alternatively be used to separate the wavelengths of a multispectral excitation beam or in any other suitable situation. For example, the color separation assembly (800) may be placed in the optical paths of two excitation beams from a single fiber, and such excitation beams are used to illuminate two separate locations on a flow cell (128, 368, 400, 450, 510). It should also be understood that two or more color separation assemblies (800) may be positioned in the excitation pathway prior to the flow cells (128, 368, 400, 450, 510), and / or two or more color separation assemblies (800) may be positioned in the emission pathway from the flow cells (128, 368, 400, 450, 510).

[0074] The color separation assembly (800) in this embodiment includes two angled reflectors (802, 804) that form a right-angle reflector. The color separation assembly (800) may be used in infinity space (e.g., the collimated space between the objective lens assemblies (542, 606, 666, 756) and the imaging lenses (620, 744)), convergence space (e.g., the imaging space between the imaging lenses (620, 744) and one or more imaging sensors of the camera system (540, 730)), or elsewhere. To correct a multispectral radiation beam, the reflector (802) is in the form of a dichroic structure configured to induce spatial separation of the radiation beam, separating it into a first reflected wavelength beam from the first surface (802A) and a second reflected wavelength beam from the second surface (802B), and generating two distinct beam paths (806, 808) for each radiation wavelength beam, one for each surface, based on the gap between the first surface (802A) and the second surface (802B). A smaller gap may result in smaller induced spatial separation, and a larger gap may result in larger induced spatial separation.

[0075] In this embodiment, a transparent optical compensator (810) is introduced into the beam path (806) between the reflectors (802, 804) to compensate for the optical path length difference imposed by the dichroic reflector (802). In some embodiments, the optical path length and material of the optical compensator (810) are determined based on the desired wavelength of the emission reflected by the first surface (802A), the amount of optical path length delay induced by the size of the gap between the surfaces (802A, 802B), and the wavelength of the emission reflected by the second surface (802B). In some variations, the compensator (810) comprises an electro-optic compensator, and the amount of optical compensation is controlled by a signal from a controller (114, 308, 520). As just a further example, the compensator (810) may comprise a clock compensator.

[0076] For the compensating plate (810), two spatially separated emission beam paths (806, 808) are incident on the same optical path, depending on the implementation, to the imaging lens (620, 744) or to the exit reflector (812) coupled to spaced-apart image sensors (e.g., of a camera system (540, 730)). In a variant where the color separation assembly (800) is positioned in the convergence space between the imaging lens (620, 744) and the two image sensors, each of those image sensors may be configured to capture different respective emission wavelengths (e.g., positioned at an offset location, having a wavelength bandpass filter, or using some other configuration). Some variations of the color separation assembly (800) may omit one or both reflectors (804, 812). Some other variations of the color separation assembly (800) may include an additional reflector (not shown).

[0077] In this embodiment, the reflector (802) is shown as dichroic, but in some modifications, the reflector (804) may also be dichroic. Furthermore, in some modifications, both reflectors (802, 804) may be dichroic reflectors. In some modifications, the reflectors (802) and / or (804) may be composed of multiple separate components or as a single assembly. Furthermore, to ensure that the two emitted beams have the same optical path length, the compensator plate (810) may alternatively be positioned between the reflector (804) and the exit reflector (812). In some modifications, apertures may be introduced into one or both of the beam paths (806, 808) to prevent undesirable beam divergence and to ensure that the objective lens assemblies (542, 606, 666, 756) and imaging lenses (620, 744) form a relay lens configuration with sufficiently high resolution. For example, one or more apertures can be introduced to ensure that the beam path (806, 808) properly aligns with the entrance aperture of the imaging lens (620, 744).

[0078] Figure 12 shows one embodiment of an arrangement (900) in which a color separation assembly (930) is positioned in collimated / infinity space between an objective lens assembly (920) and an imaging lens (940). The configuration (900) may be incorporated into any of the various imaging systems (116, 310) or imaging assemblies (522, 700) described herein, or into any other suitable imaging context. The objective lens assembly (920) and the imaging lens (940) are schematically shown in Figure 12, respectively. It should be understood that the objective lens assembly (920) may include any suitable number of lens elements and / or other optical features, and the imaging lens (940) may include any suitable number of lens elements and / or other optical features. The objective lens assembly (920) is positioned on the flow cell (910), which may be configured and operable as in any of the modifications of the flow cells (128, 368, 400, 450, 510) described herein.

[0079] In this embodiment, a fluorescent label at a reaction site on the flow cell (910) emits light in response to excitation from one or more excitation sources (not shown), and the multispectral emission beam (960) passes through the objective lens assembly (920) to reach the color separation assembly (930). The objective lens (920) in this embodiment is configured to substantially collimate the multispectral emission beam (960), thereby positioning the color separation assembly (930) in the collimated / infinity space of the multispectral emission beam (960). In this embodiment, the multispectral emission beam (960) is generated from only one spatial location on the flow cell (910). In some other modifications, the multispectral emission beam (960) is generated from two or more different spatial locations on the flow cell (910). In some such modifications, two or more excitation sources are used to simultaneously illuminate each different spatial location on the flow cell (910). Such different excitation sources may emit excitation light at each different wavelength. Therefore, it should be understood that there may be configurations in which there are two or more multispectral emission beams (960) generated from two or more different spatial positions on the flow cell (910), just as the flow cell (910) may have two or more multispectral emission points simultaneously.

[0080] The color separation assembly (930) in this embodiment includes a filtering reflector (932) and a reflector (934) positioned behind the filtering reflector (932). The filtering reflector (932) is configured to reflect light from a multispectral emission beam (960) within a first spectral range, while allowing light from a multispectral emission beam (960) within a second spectral range to pass through and reach the reflector (934). In Figure 12, the reflected light within the first spectral range is represented by beam (962), while the reflected light within the second spectral range is represented by beam (964). Although the color separation assembly (930) is described herein as being formed from two distinct components, including a filtering reflector element (932) and a reflector element (934), some modifications may provide a color separation assembly (930) formed from a single component or element (e.g., a wedge-shaped optical component), so that not all forms of the color separation assembly (930) are necessarily formed from two distinct components. In this embodiment, the reflector element (934) comprises a simple mirror that reflects light in a second spectral range, such as represented by the beam (964). In some other modifications, the reflector element (934) comprises a dichroic element, thereby configuring the reflector element (934) to transmit light outside the second spectral range while reflecting light within the second spectral range. Therefore, it should be understood that terms such as “reflector element” and “reflective element” may include a simple mirror, a dichroic element, or other elements that can reflect light.

[0081] In some variations, the filtering reflector (932) is in the form of a dichroic element. For example, the filtering reflector (932) may be configured to reflect light of one color (e.g., blue) while allowing light of another color (e.g., green) to pass through and reach the reflector (934). In some other variations, the filtering reflector (932) is in the form of a multiband beam splitter. For example, the filtering reflector (932) may be configured to reflect two colors (e.g., blue and red) while allowing two other colors (e.g., green and orange) to pass through and reach the reflector (934). The colors referenced above are merely illustrative examples and are not intended to be limiting. In a scenario where the first spectral range (reflected by filtering reflector (932)) contains two or more colors and the second spectral range (reflected by reflector (934)) contains two or more colors, the beams (962, 964) may undergo further color separation, as described below with reference to Figures 13-15.

[0082] In this embodiment, the filtering reflective element (932) and the reflective element (934) are not parallel to each other. Therefore, the filtering reflective element (932) is tilted at a first angle about the y-axis, while the reflective element (934) is tilted at a second angle about the y-axis. (It should be understood that the x-axis and z-axis shown in Figure 12 do not necessarily correspond to the same axis, as shown in other drawings.) As a mere example, the filtering reflective element (932) and the reflective element (934) may be angularly offset from each other by about 1 degree. Alternatively, the filtering reflective element (932) and the reflective element (934) may have any other suitable angular offset. In either case, the angular offset between the filtering reflective element (932) and the reflective element (934) provides a corresponding angular offset between the first spectral range beam (962) and the second spectral range beam (964). This distinguishes the color separation assembly (930) in Figure 12 from the color separation assembly (800) in Figure 11.

[0083] The spatial distance between the reflective surface of the filtering reflector (932) and the reflective surface of the reflector (934) already provides a spatial offset between the first spectral range beam (962) and the second spectral range beam (964) (similar to the spatial offset provided in the arrangement of Figure 11, as described above), and an additional angular offset between the first spectral range beam (962) and the second spectral range beam (964) can effectively further enhance the spatial offset between the first spectral range beam (962) and the second spectral range beam (964), particularly as a function of the distance "downstream" of the color separation assembly (930). Therefore, depending on the type of additional optical components located "downstream" of the color separation assembly (930), if these additional optical components are located "downstream" of the color separation assembly (930), the angular offset between the filtering reflector (932) and the reflector (934) may be adjusted to provide a preferred degree of spatial separation between the beams (962, 964).

[0084] In this embodiment, the imaging lens (940) is positioned "downstream" of the color separation assembly (930), thereby allowing both beams (962, 964) to pass through the imaging lens (940) and enter the converged imaging space. The images formed at the focal point of the imaging lens (940) are separated laterally from each other by the angular and spatial separation between the beams (962, 964), as described above. The camera (950) is positioned within the converged imaging space generated by the imaging lens (940), thereby capturing these two images. As a mere example, the camera (950) may comprise a TDI camera. The camera (950) includes a first imaging sensor (952) and a second imaging sensor (954). In this embodiment, the imaging sensors (952, 954) are positioned adjacent to each other. The imaging sensor (952) is positioned to receive a first spectral range beam (962), while the imaging sensor (954) is positioned to receive a second spectral range beam (964). Thus, the configuration (900) enables a single camera (950) to simultaneously capture an image from a single multispectral beam (960) emitted from a single flow cell (910) using only one single objective lens assembly (920) and only one single imaging lens (940) with two separate imaging sensors (952, 954) in two distinct color channels.

[0085] B. Multispectral imaging device with one astigmatism correction element As described above, there may be several scenarios in which a first spectral range (e.g., reflected by filtering reflector (932)) contains two or more colors and a second spectral range (e.g., reflected by reflector (934)) contains two or more colors. In some such scenarios, a multispectral emission beam (such as multispectral beam (960)) may contain meaningful optical data within three or more spectral channels (e.g., within three or more colors), and it may be desirable to ultimately separate the multispectral emission beam into three or more spectral ranges for capture via three or more respective imaging sensors. Figures 13–15 show an embodiment of configuration (1000) in which a multispectral emission beam (such as multispectral beam (960)) can be further separated into four spectral ranges for capture via four respective imaging sensors (1052, 1054, 1062, 1064).

[0086] In configuration (1000), a first multispectral beam (1070) is transmitted from the color separation assembly (1010) along a first spatial orientation, while a second multispectral beam (1080) is transmitted from the color separation assembly (1010) along a second spatial orientation. As a mere example, the color separation assembly (1010) in configuration (1000) may be configured and operable in the same way as the color separation assembly (930) in configuration (900). Similarly, the first multispectral beam (1070) may be angularly and spatially offset from the second multispectral beam (1070), similar to the case where the first beam (962) is angularly and spatially offset from the second beam (964). It should also be understood that configuration (1000) may include a flow cell (910) and an objective lens assembly (920), respectively, such that the color separation assembly (1010) separates the multispectral emission beam from the flow cell into a first multispectral beam (1070) and a second multispectral beam (1070).

[0087] The beams (1070, 1080) pass through the imaging lens (1020) and finally reach a dichroic element (1030) positioned in the converged imaging space between the imaging lens (1020) and the first camera (1050). In this embodiment, the dichroic element (1030) is tilted at a first tilt angle about the y-axis. (It should be understood that the x, y, and z axes shown in Figures 13 to 15 do not necessarily correspond to the same axes as shown in other drawings.) The dichroic element (1030) is configured to reflect light from the first multispectral beam (1070) within a first spectral range, while allowing light from the first multispectral beam (1070) within a second spectral range to pass through the dichroic element (1030). Similarly, the dichroic element (1030) is configured to reflect light from a second multispectral beam (1080) in a third spectral range, while allowing light from a second multispectral beam (1080) in a fourth spectral range to pass through the dichroic element (1030). In Figures 13 to 15, light in the first spectral range is represented by beam (1072), light in the second spectral range is represented by beam (1074), light in the third spectral range is represented by beam (1082), and light in the fourth spectral range is represented by beam (1084). For the sake of illustration, the first spectral range may include red, the second spectral range may include blue, the third spectral range may include green, and the fourth spectral range may include orange. The colors referred herein are merely illustrative examples and are not intended to be limiting.

[0088] Due to the spatial separation between beams (1070, 1080) in the convergent imaging space, beams (1070, 1080) are incident on the dichroic element (1030) in two distinct regions. Thus, beams (1074, 1084) are reflected from the dichroic element (1030) in two distinct regions, and beams (1072, 1082) pass through the dichroic element in two distinct regions. A second camera (1060) is positioned to capture beams (1074, 1084). In particular, the images formed at the focal point of the imaging lens (1020) are separated laterally from each other due to the angular and spatial separation between beams (1074, 1084) as described above. The second camera (1060) is positioned within the convergent imaging space formed by the imaging lens (1020), thereby capturing these two images. As a mere example, the second camera (1060) may include a TDI camera. The second camera (1060) includes a first imaging sensor (1062) and a second imaging sensor (1064). In this embodiment, the imaging sensors (1062, 1064) are positioned adjacent to each other. The imaging sensor (1062) is positioned to receive a first spectral range beam (1074), while the imaging sensor (1064) is positioned to receive a second spectral range beam (1064).

[0089] As described above, the beam (1072, 1082) passes through the dichroic element (1030) in this embodiment. The passage of the beam (1072, 1082) through the dichroic element (1030) may induce astigmatism in the beam (1072, 1082). To correct this astigmatism, a corrector element (1040) is positioned in the path of the beam (1072, 1082) between the dichroic element (1030) and the first camera (1050). As just one example, the corrector element (1040) may comprise a flat plate or have any other preferred configuration. The corrector element (1040) is configured to offset the astigmatism induced in the beam (1072, 1082) by the dichroic element (1030), or to induce astigmatism in the beam (1072, 1082) that effectively offsets it. The compensating element (1040) is tilted at a second tilt angle about the z-axis. (As mentioned above, the x, y, and z axes shown in Figures 13 to 15 do not necessarily correspond to the same axes as those shown in other drawings.) Therefore, in this embodiment, the compensating element (1040) and the dichroic element (1030) are tilted about their respective mutually orthogonal axes. In some variations, the first tilt angle and the second tilt angle are equal. The compensating element (1040) may be configured with a tilt angle, thickness, and material selected to compensate for the aberrations of the dichroic element (1030).

[0090] The astigmatism-corrected beams (1072, 1082) are captured by the first camera (1050). In particular, the images formed at the focal point of the imaging lens (1020) are separated laterally from each other by the angular and spatial separation between the beams (1072, 1082) as described above. The first camera (1050) is positioned within the convergent imaging space formed by the imaging lens (1020), thereby capturing these two images using astigmatism correction provided by the correction element (1040). As a mere example, the first camera (1050) may comprise a TDI camera. The first camera (1050) includes a first imaging sensor (1052) and a second imaging sensor (1054). In this embodiment, the imaging sensors (1052, 1054) are positioned adjacent to each other. As described above, due to spatial separation between the beams (1070, 1080) in the convergent imaging space, the beams (1070, 1080) are incident on the dichroic element (1030) in two different regions, thereby allowing the beams (1072, 1082) to pass through the dichroic element (1030) in two different regions. The imaging sensor (1052) is positioned to receive the third spectral range beam (1072), while the imaging sensor (1054) is positioned to receive the fourth spectral range beam (1082).

[0091] Taking the above into consideration, arrangement (1000) allows two cameras (1050, 1060), in combination with a color separation assembly (1010), to simultaneously capture an image in four separate color channels using four respective imaging sensors (1052, 1054, 1062, 1064) from a single multispectral beam emitted from a single flow cell, using only one objective lens assembly and one imaging lens (1020).

[0092] C. Multispectral imaging device with two astigmatism correction elements Figures 16-17 show another embodiment of the array in which two cameras (1150, 1160), in combination with a color separation assembly (1110), enable simultaneous capture of an image in four distinct color channels using four separate imaging sensors (1152, 1154, 1162, 1164) from a single multispectral beam emitted from a single flow cell, using only one objective lens assembly and one imaging lens (1120). In configuration (1100), a first multispectral beam (1170) is transmitted from the color separation assembly (1110) along a first spatial orientation, while a second multispectral beam (1180) is transmitted from the color separation assembly (1110) along a second spatial orientation. As a mere example, the color separation assembly (1110) in configuration (1100) may be configured and operable in the same way as the color separation assembly (930) in configuration (900). Similarly, the first multispectral beam (1170) may be angularly and spatially offset from the second multispectral beam (1180) so as the first beam (962) is angularly and spatially offset from the second beam (964). It should also be understood that the configuration (1100) may include a flow cell (910) and an objective lens assembly (920), respectively, and a similar flow cell and objective lens assembly so as the color separation assembly (1110) separates the multispectral emission beam from the flow cell into a first multispectral beam (1170) and a second multispectral beam (1170).

[0093] The beams (1170, 1180) pass through the imaging lens (1120) and finally reach a first corrector element (1190) positioned in the convergent imaging space between the imaging lens (1120) and the dichroic element (1130). In this embodiment, the first corrector element (1190) is tilted at a first tilt angle about the z-axis. (It should be understood that the x, y, and z axes shown in Figures 16-17 do not necessarily correspond to the same axes as shown in other drawings.) As a mere example, the first corrector element (1140) may comprise a flat plate or have any other preferred configuration. The first corrector element (1140) is configured to induce astigmatism in the beams (1170, 1180). The first corrector element (1140) may consist of a tilt angle, thickness, and material selected to correct the aberration of the dichroic element (1130), as described below.

[0094] The dichroic element (1130) is also positioned in the convergent imaging space between the first correction element (1190) and the first camera (1150). In this embodiment, the dichroic element (1130) is tilted at a second tilt angle about the y-axis. (As described above, the x, y, and z axes shown in Figures 16-17 do not necessarily correspond to the same axes as those shown in other drawings.) The dichroic element (1130) is configured to reflect light from the first multispectral beam (1170) within a first spectral range, while allowing light from the first multispectral beam (1170) within a second spectral range to pass through the dichroic element (1130). Similarly, the dichroic element (1130) is configured to reflect light from a second multispectral beam (1180) in a third spectral range, while allowing light from a second multispectral beam (1180) in a fourth spectral range to pass through the dichroic element (1130). In Figures 16-17, light in the first spectral range is represented by beam (1172), light in the second spectral range is represented by beam (1174), light in the third spectral range is represented by beam (1182), and light in the fourth spectral range is represented by beam (1184). For the sake of illustration, the first spectral range may include red, the second spectral range may include blue, the third spectral range may include green, and the fourth spectral range may include orange. The colors referred herein are merely illustrative examples and are not intended to be limiting.

[0095] The beam (1174, 1184) reflected from the dichroic element (1130) passes through a second corrector element (1192) before reaching the second camera (1160). As a mere example, the second corrector element (1192) may comprise a flat plate or have any other preferred configuration. The second corrector element (1192) is configured to induce astigmatism in the beam (1174, 1184). In particular, the astigmatism induced in the beam (1174, 1184) by the second corrector element (1192) offsets or effectively offsets the astigmatism induced in the beam (1174, 1184) by the first corrector element (1190). In this embodiment, the second corrector element (1192) is tilted at a third tilt angle about the zy-axis. (As mentioned above, the x, y, and z axes shown in Figures 16-17 do not necessarily correspond to the same axes as those shown in other drawings.) Therefore, in this embodiment, the first corrector element (1190) and the second corrector element (1192) are tilted around mutually orthogonal axes. In some variations, the first tilt angle and the third tilt angle are equal. The second corrector element (1192) may consist of a tilt angle, thickness, and material selected to correct the aberration of the first corrector element (1190).

[0096] The astigmatism-corrected beams (1174, 1184) are captured by a second camera (1160). In particular, the images formed at the focal point of the imaging lens (1120) are separated laterally from each other by the angular and spatial separation between the beams (1174, 1184) as described above. The second camera (1160) is positioned in the converged image space generated by the imaging lens (1120) and thereby captures these two images. As a mere example, the second camera (1160) may comprise a TDI camera. The second camera (1160) includes a first imaging sensor (1162) and a second imaging sensor (1164). In this embodiment, the imaging sensors (1162, 1164) are positioned adjacent to each other. As described above, due to spatial separation between the beams (1170, 1180) in the convergent imaging space, the beams (1170, 1180) are incident on the dichroic element (1130) in two different regions, and thereby the beams (1072, 1082) are reflected from the dichroic element (1130) in two different regions. The imaging sensor (1162) is positioned to receive the first spectral range beam (1174), while the imaging sensor (1164) is positioned to receive the second spectral range beam (1164).

[0097] As described above, the beams (1172, 1182) pass through the dichroic element (1130) in this embodiment. The passage of the beams (1172, 1182) through the dichroic element (1130) may induce astigmatism in the beams (1172, 1182). This astigmatism induced by the dichroic element (1130) can effectively offset or offset the astigmatism induced in the beams (1170, 1180) by the first correction element (1190). As described above, the dichroic element (1130) is tilted at a second tilt angle about the y-axis. (Also, as mentioned above, the x, y, and z axes shown in Figures 13 to 15 do not necessarily correspond to the same axes as those shown in other drawings.) Therefore, in this embodiment, the first compensating element (1190) and the dichroic element (1130) are tilted around mutually orthogonal axes. In some variations, the first tilt angle (of the first compensating element (1190)) and the second tilt angle (of the dichroic element (1130)) are equal. Furthermore, the second tilt angle (of the dichroic element (1130)) and the third tilt angle (of the second compensating element (1192)) may be equal, thereby allowing the dichroic element (1130) and the second compensating element (1192) to be parallel to each other.

[0098] The astigmatism-corrected beams (1172, 1182) are captured by the first camera (1150). In particular, the images formed at the focal point of the imaging lens (1120) are separated laterally from each other by the angular and spatial separation between the beams (1172, 1182) as described above. The first camera (1150) is positioned in the converged image space generated by the imaging lens (1120) and thereby captures these two images. As a mere example, the first camera (1150) may comprise a TDI camera. The first camera (1150) includes a first imaging sensor (1152) and a second imaging sensor (1154). In this embodiment, the imaging sensors (1152, 1154) are positioned adjacent to each other. As described above, due to spatial separation between beams (1170, 1180) in the convergent imaging space, beams (1170, 1180) are incident on the dichroic element (1130) in two different regions, and as a result, beams (1172, 1182) pass through the dichroic element (1130) in two different regions. The imaging sensor (1152) is positioned to receive the third spectral range beam (1172), while the imaging sensor (1154) is positioned to receive the fourth spectral range beam (1182).

[0099] Taking the above into consideration, arrangement (1100) allows two cameras (1150, 1160), in combination with a color separation assembly (1110), to simultaneously capture an image in four separate color channels using four respective imaging sensors (1152, 1154, 1162, 1164) from a single multispectral beam emitted from a single flow cell, using only one objective lens assembly and one imaging lens (1120).

[0100] D. Multispectral imaging device with non-adjacent imaging sensors In the above configuration (900, 1000, 1100), each camera (950, 1050, 1060, 1150, 1160) includes its respective pair of adjacent imaging sensors (952, 954, 1052, 1054, 1152, 1154, 1162, 1164). In some scenarios, it may be desirable to modify the arrangement (900, 1000, 1100) to allow two spectrally distinct but spatially adjacent beams (962, 964, 1072, 1074, 1082, 1084, 1172, 1174, 1182, 1184) to be captured by non-adjacent imaging sensors. Figure 18 shows an embodiment of an arrangement (1200) that can be used to redirect two spectrally distinct but spatially adjacent beams (1240, 1250) to their respective non-adjacent imaging sensors (1220, 1230). In this embodiment, the first beam (1240) is in a first spectral range (e.g., blue), while the second beam (1250) is in a second spectral range (e.g., red). The beams (1240, 1250) may be transmitted from a color separation assembly (800, 930, 1010, 1110) as described herein, or from any other suitable light source.

[0101] The beams (1240, 1250) are adjacent and parallel to each other until they reach the deflection element (1210). In some other variations, the beams (1240, 1250) are not parallel to each other. The deflection element (1210) has a first reflecting surface (1212) and a second reflecting surface (1214). In this embodiment, the surfaces (1210, 1212) define a right angle to each other, but the surfaces (1210, 1212) may alternatively define any other preferred angle to each other. Surface (1212) is positioned to receive the first beam (1240) and reflect it toward the first image sensor (1220). Surface (1214) is positioned to receive the second beam (1250) and reflect it toward the second image sensor (1230). In this embodiment, the image sensors (1220, 1230) are oriented to face each other. In other variations, the image sensors (1220, 1230) have some other spatial relationship and orientation to each other, and the configuration of the deflection element (1210) is modified to accommodate such alternative spatial relationship / orientation. As just one example, the deflection element (1210) may comprise a knife-edge right-angle prism mirror. Alternatively, the deflection element (1210) may take any other preferred form.

[0102] In the context of configuration (900), the imaging sensors (952, 954) may be rearranged to be positioned similarly to the imaging sensors (1220, 1230), and the deflection element (1210) may be positioned to redirect beam (962) towards the imaging sensor (952) and beam (964) towards the imaging sensor (954). In the context of configuration (1000), the imaging sensors (1052, 1054) may be rearranged to be positioned similarly to the imaging sensors (1220, 1230), and the deflection element (1210) may be positioned to redirect beam (1072) towards the imaging sensor (1052) and beam (1082) towards the imaging sensor (1054). Similarly, the imaging sensors (1062, 1064) may be rearranged to be positioned similarly to the imaging sensors (1220, 1230), and the deflection element (1210) may be positioned to redirect beam (1074) towards imaging sensor (1062) and beam (1084) towards imaging sensor (1064). In the context of configuration (1100), the imaging sensors (1152, 1154) may be rearranged to be positioned similarly to the imaging sensors (1220, 1230), and the deflection element (1210) may be positioned to deflect beam (1172) towards imaging sensor (1152) and beam (1182) towards imaging sensor (1154). Similarly, the imaging sensors (1162, 1164) may be rearranged to be positioned similarly to the imaging sensors (1220, 1230), and the deflection element (1210) may be positioned to redirect the beam (1174) towards the imaging sensor (1162) and the beam (1184) towards the imaging sensor (1164).

[0103] Any of the above configurations (900, 1000, 1100) may be used in a scenario in which the excitation source irradiates the reaction site at a single position on the flow cell (128, 368, 400, 450, 510, 910), or in a scenario in which the reaction site is irradiated (simultaneously) at two or more different positions on the flow cell (128, 368, 400, 450, 510, 910). Furthermore, any of the above configurations (900, 1000, 1100) may be used in a scenario in which the excitation source irradiates the reaction site with two different wavelengths of light (simultaneously) at two or more different positions on the flow cell (128, 368, 400, 450, 510, 910). In the example above, the arrangement (900, 1000, 1100) separates emission from two object points (e.g., on the flow cell (128, 368, 400, 450, 510, 910)) into four image points (e.g., on the imaging sensor (952, 954, 1052, 1054, 1152, 1154, 1162, 1164)). Some variations may separate emission from more than two object points. Similarly, some variations may separate emission into two, three, or more than four image points.

[0104] In some variations, where the excitation source irradiates the reaction site (simultaneously) with light of two different wavelengths at two or more different locations on the flow cell (128, 368, 400, 450, 510, 910), the emitted beams from those two or more different locations will already have spatial separation (and in some cases, spectral separation as well). In such scenarios, it may not be necessary to include the spatial and angular separation capabilities of the color separation assembly (800, 930, 1010, 1110) as described above. Instead, dichroic elements (1030, 1130) may be positioned in collimated space to provide further separation within the two or more multispectral emission beams. In some such cases, a single dichroic element (1030, 1130) may be used for all of the two or more multispectral emission beams. In some other cases, each of the two or more multispectral emission beams may have its own dichroic element (1030, 1130). Alternatively, color separation assemblies (800, 930, 1010, 1110) may be positioned in the path of each multispectral emission beam to direct emission beams of a specific wavelength range to the corresponding imaging sensors (952, 954, 1052, 1054, 1152, 1154, 1162, 1164).

[0105] In yet another variation, the dichroic elements (1030, 1130) in configuration (1000, 1100) may be replaced with a dual-bandpass optical filter. The filtering reflector (932) in configuration (900) may also be replaced with a dual-bandpass optical filter. If the collimated / infinite space filter (e.g., filtering reflector (932)) is long-pass, then the converged / imaging space filter (e.g., dichroic elements (1030, 1130)) may have a single passband containing two intermediate wavelength sets, or a dual-passband filter may be used.

[0106] It should be understood from the foregoing that multispectral light emitted from a reaction site on a flow cell (128, 368, 400, 450, 510, 910) (or from any other type of irradiated sample) can be separated into any number of spectral channels by spatial separation of emission points (e.g., by irradiating two or more different points on the sample), by separation of the emitted beam into two or more spectral channels in collimated / infinite space, and / or by separation of the emitted beam into two or more spectral channels in converged imaging space. Furthermore, the teachings herein may also apply to situations where more than two fields of view are being imaged simultaneously, and where more than two, three, or four spectral channels are being imaged simultaneously. It should also be understood that the color separation stage does not necessarily have spectral symmetry. For example, a color separation assembly may divide three colors into a first channel having two colors and a second channel having only one color.

[0107] VII. Examples of Combinations The following embodiments relate to various non-exclusive methods to which the teachings herein may be combined or applied. The following embodiments are not intended to limit the scope of any claims that may be presented at any time in this application or any subsequent application. No waiver is intended. The following embodiments are provided for illustrative purposes only. It is intended that the various teachings herein may be constructed and applied in numerous other ways. Furthermore, it is intended that some variations may omit certain features mentioned in the following embodiments. Accordingly, none of the embodiments or features mentioned below should be considered important unless they are expressly indicated thereafter by the inventors or their successors. If any claim containing additional features beyond those mentioned below is presented in this application or any subsequent application relating to this application, those additional features should not be assumed to have been added for any reason relating to patentability.

[0108] Example 1 The apparatus comprises a sample stage region configured to provide an object surface, an optical assembly comprising an objective element providing a field of view, the object surface being within the field of view, an imaging lens configured to receive light transmitted through the objective element, and a first dichroic element, the optical assembly configured to transmit light from at least a first color channel, a second color channel, a third color channel, and a fourth color channel from the imaging lens toward the first dichroic element, the first dichroic element transmitting light from the first color channel and the third color channel, while the second An apparatus comprising: an optical assembly including a first dichroic element configured to reflect light from two color channels and a fourth color channel, the first dichroic element being further configured to induce a first astigmatism in the first and third color channels; a corrector element configured to induce a second astigmatism in the first and third color channels, the second astigmatism being configured to offset the first astigmatism; and a camera assembly configured to receive light from a first color channel, a second color channel, a third color channel, and a fourth color channel.

[0109] Example 2 The apparatus according to Example 1, wherein the first and second color channels are angularly offset from the third and fourth color channels on the incident surface of the first dichroic element.

[0110] Example 3 The apparatus according to Embodiment 2, wherein the optical assembly further comprises a color-molding assembly, the color-molding assembly configured to provide an angular offset of the light from the first and second color channels from the light from the third and fourth color channels.

[0111] Example 4 The apparatus according to Example 3, wherein the color-molded assembly includes a second dichroic element and a reflective element, the reflective element being angularly offset from the second dichroic element.

[0112] Example 5 The apparatus according to Embodiment 4, wherein a second dichroic element is configured to reflect light from the first and second color channels, and the second dichroic element is further configured to provide transmission of light from the third and fourth color channels to a reflecting element, and the reflecting element is configured to reflect light from the third and fourth color channels.

[0113] Example 6 The apparatus according to any one of Examples 3 to 5, wherein the color-molded assembly is positioned in the optical path between the objective element and the imaging lens.

[0114] Example 7 The apparatus according to any one of Examples 1 to 6, further comprising an excitation assembly, the excitation assembly being operable to emit light toward two different regions in the surface of an object, wherein the first and second color channels are associated with the first of the two different regions, and the third and fourth color channels are associated with the second of the two different regions.

[0115] Example 8 The apparatus according to any one of Examples 1 to 7, wherein the first dichroic element is inserted between the imaging lens and the camera assembly.

[0116] Example 9 The apparatus according to any one of Examples 1 to 8, wherein the first dichroic element comprises a first plate inclined at a first angle about a first axis.

[0117] Example 10 The apparatus according to Embodiment 9, wherein the correction element comprises a second plate inclined at a second angle about a second axis.

[0118] Example 11 The apparatus according to Example 10, wherein the second axis is perpendicular to the first axis.

[0119] Example 12 The apparatus according to any one of Examples 10 to 11, wherein the second angle is equal to the first angle.

[0120] Example 13 The camera assembly comprises a first image sensor configured to receive light from a first color channel, a second image sensor configured to receive light from a second color channel, a third image sensor configured to receive light from a third color channel, and a fourth image sensor configured to receive light from a fourth color channel, as described in any one of Examples 1 to 12.

[0121] Example 14 The apparatus according to Embodiment 13, wherein the first imaging sensor is positioned adjacent to the third imaging sensor, and the second imaging sensor is positioned adjacent to the fourth imaging sensor.

[0122] Example 15 The apparatus according to Embodiment 14, wherein the camera assembly comprises a first time-delay integral (TDI) camera and a second TDI camera.

[0123] Example 16 The apparatus according to Example 15, wherein the first TDI camera includes a first imaging sensor and a third imaging sensor, and the second TDI camera includes a second imaging sensor and a fourth imaging sensor.

[0124] Example 17 The apparatus according to any one of Examples 15 to 16, wherein the first TDI camera is oriented along a first axis, and the second TDI camera is oriented along a second axis, the second axis being perpendicular to the first axis.

[0125] Example 18 The apparatus according to Embodiment 13, further comprising: a first reflective member configured to reflect light from a first color channel toward a first image sensor, and further configured to reflect light from a third color channel toward a third image sensor; and a second reflective member configured to reflect light from a second color channel toward a second image sensor, and further configured to reflect light from a first color channel toward a first image sensor.

[0126] Example 19 The apparatus according to Embodiment 18, wherein the first reflective member comprises a first right-angle prism mirror, and the second reflective member comprises a second right-angle prism mirror.

[0127] Example 20 An apparatus comprising: an objective element that provides a field of view; an imaging lens configured to receive light transmitted through the objective element, and further configured to transmit light including spatially offset spectral channels through a converging imaging space; a dichroic element in the converging imaging space, configured to reflect light from at least one optical channel in the converging imaging space, while transmitting light from at least one other optical channel in the converging imaging space, and further configured to induce astigmatism in the light transmitted through the dichroic element; and a corrector element configured to induce a second astigmatism in the light transmitted through the dichroic element, wherein the second astigmatism offsets the first astigmatism.

[0128] Example 21 A method comprising transmitting light from an imaging lens through a dichroic element, wherein the transmitted light comprises a first color channel, a second color channel, a third color channel, and a fourth color channel, the first and second color channels being spatially offset from the third and fourth color channels at the incident surface on the dichroic element, and transmitting the light of the first and third color channels through a dichroic element, the dichroic element inducing a first astigmatism in the first and third color channels, and the second and fourth color channels A method comprising: reflecting light, wherein the light of the second and fourth color channels is reflected by a dichroic element; transmitting light from the dichroic element through a corrector element, wherein the corrector element induces a second astigmatism in the first and third color channels, the second astigmatism offsets the first astigmatism; receiving the light of the first and third color channels from the corrector element in a camera assembly; and receiving the light of the second and fourth color channels from the dichroic element in a camera assembly.

[0129] VIII. Others While the embodiments described herein are provided in the context of a system (100) that may be used in a nucleotide sequencing process, the teachings herein may also be readily applicable in other contexts, including systems that carry out other processes (i.e., procedures other than nucleotide sequencing). Therefore, the teachings herein are not necessarily limited to systems used to carry out a nucleotide sequencing process.

[0130] It should be understood that the subject matter described herein is not limited in its application to the configuration details and arrangement of components described herein or shown in the drawings herein. The subject matter described herein is capable of other implementations and can be carried out or performed in a variety of ways. Furthermore, it should be understood that the expressions and terms used herein are for illustrative purposes only and should not be considered limiting. When used herein, an element or process described in the singular and followed by the words "a" or "an" should be understood not to exclude multiple such elements or processes unless such exclusion is explicitly stated. Furthermore, references to "one example" are not intended to be construed as excluding the existence of additional embodiments that also incorporate the described features. The use of "including," "comprising," or "having" and their variations herein means that the items listed thereafter and their equivalents, as well as additional items, are included.

[0131] When used in patent claims, the term “set” should be understood as one or more things grouped together. Similarly, when used in patent claims, “based on” should be understood as indicating that one thing is at least partially determined by the thing that is designated as “based on.” When something needs to be exclusively determined by another, then that thing is referred to as “exclusively based on” the thing on which it is determined.

[0132] Unless otherwise specified or limited, the terms “mounted,” “connected,” “supported,” and “coupled,” as well as their variations, are used broadly and encompass both direct and indirect mounting, connection, support, and coupling. Furthermore, “connected” and “coupled” are not limited to physical or mechanical connection or coupling. Also, please understand that the expressions and terms used herein with respect to the orientation of devices or elements (e.g., “above,” “below,” “front,” “rear,” “distal,” “proximal”) are used solely to simplify the description of one or more embodiments described herein and do not in themselves indicate or imply that the devices or elements mentioned must have a particular orientation. Terms such as “outer” and “inner” are used herein for illustrative purposes only and are not intended to indicate or imply relative importance or significance.

[0133] It should be understood that the above description is illustrative and not restrictive. For example, the embodiments (and / or embodiments thereof) described above may be used in combination with each other. Furthermore, many modifications may be made to adapt specific circumstances or materials to the teachings of the subject matter described herein, without departing from the scope thereof. The dimensions, material types, and coatings described herein are intended to define parameters of the disclosed subject matter, but they are not restrictive and are rather illustrative. Many further examples will be apparent to those skilled in the art upon consideration of the above description. Thus, the scope of the disclosed subject matter should be determined by reference to the appended claims, along with the entire scope of equivalents to which such claims are granted. In the appended claims, the terms “including” and “in which” are used as plain English equivalents to “comprising” and “wherein,” respectively. Furthermore, in the following claims, terms such as “first,” “second,” and “third” are used merely as designations and are not intended to impose numerical requirements on those objects. Moreover, the following limitations on the claims are not written in means-plus-function form, and such limitations on the claims are not intended to be interpreted under paragraph 112(f) of the U.S. Patent Act unless, and until, the phrase “means for” is explicitly used and followed by a description of the function without further structure is explicitly used.

[0134] The following claims enumerate specific examples of the subject matter disclosed and are considered to be part of the above disclosure. These examples may be combined with each other.

[0135] [Implementation Method] (1) Apparatus, the apparatus, A sample stage region configured to provide an object surface, An optical assembly, The optical assembly is an objective element that provides a field of view, wherein the object plane is within the field of view, An imaging lens, wherein the imaging lens is configured to receive light that has passed through the objective element, A first dichroic element, wherein the optical assembly is configured to transmit light from at least a first color channel, a second color channel, a third color channel, and a fourth color channel from the imaging lens toward the first dichroic element, the first dichroic element is configured to transmit light from the first color channel and the third color channel, while reflecting light from the second color channel and the fourth color channel, and the first dichroic element is further configured to induce a first astigmatism in the first color channel and the third color channel, An optical assembly comprising: a correction element configured to induce a second astigmatism in the first and third color channels, wherein the second astigmatism is configured to offset the first astigmatism; A device comprising: a camera assembly configured to receive light from the first color channel, the second color channel, the third color channel, and the fourth color channel. (2) The apparatus according to Embodiment 1, wherein the first color channel and the second color channel are angularly offset from the third color channel and the fourth color channel on the incident surface of the first dichroic element. (3) The apparatus according to Embodiment 2, wherein the optical assembly further comprises a color-molded assembly, the color-molded assembly configured to provide the angular offset of the light from the first and second color channels from the light from the third and fourth color channels. (4) The apparatus according to Embodiment 3, wherein the color-molded assembly includes a second dichroic element and a reflective element, the reflective element being angularly offset from the second dichroic element. (5) The apparatus according to Embodiment 4, wherein the second dichroic element is configured to reflect light from the first and second color channels, and the second dichroic element is further configured to allow transmission of light from the third and fourth color channels to the reflecting element, and the reflecting element is configured to reflect light from the third and fourth color channels.

[0136] (6) The apparatus according to any one of embodiments 3 to 5, wherein the color-molded assembly is positioned in the optical path between the objective lens element and the imaging lens. (7) The apparatus according to any one of embodiments 1 to 6, further comprising an excitation assembly which is operable to emit light toward two different regions in the surface of the object, wherein the first and second color channels are associated with the first of the two different regions, and the third and fourth color channels are associated with the second of the two different regions. (8) The apparatus according to any one of embodiments 1 to 7, wherein the first dichroic element is inserted between the imaging lens and the camera assembly. (9) The apparatus according to any one of embodiments 1 to 8, wherein the first dichroic element comprises a first plate inclined at a first angle about a first axis. (10) The apparatus according to Embodiment 9, wherein the correction element comprises a second plate inclined at a second angle about a second axis.

[0137] (11) The apparatus according to embodiment 10, wherein the second axis is perpendicular to the first axis. (12) The apparatus according to either embodiment 10 or 11, wherein the second angle is equal to the first angle. (13) The camera assembly is A first imaging sensor configured to receive light from the first color channel, A second imaging sensor configured to receive light from the second color channel, A third imaging sensor configured to receive light from the third color channel, The apparatus according to any one of embodiments 1 to 12, comprising: a fourth imaging sensor configured to receive light from the fourth color channel. (14) The apparatus according to Embodiment 13, wherein the first imaging sensor is positioned adjacent to the third imaging sensor, and the second imaging sensor is positioned adjacent to the fourth imaging sensor. (15) The apparatus according to embodiment 14, wherein the camera assembly comprises a first time-delay integral (TDI) camera and a second TDI camera.

[0138] (16) The apparatus according to Embodiment 15, wherein the first TDI camera includes a first imaging sensor and a third imaging sensor, and the second TDI camera includes a second imaging sensor and a fourth imaging sensor. (17) The apparatus according to any one of embodiments 15 to 16, wherein the first TDI camera is oriented along a first axis, and the second TDI camera is oriented along a second axis, the second axis being perpendicular to the first axis. (18) The camera assembly is A first reflective member, wherein the first reflective member is configured to reflect light from the first color channel toward the first image sensor, and the first reflective member is further configured to reflect light from the third color channel toward the third image sensor, The apparatus according to embodiment 13, further comprising: a second reflective member, the second reflective member configured to reflect light from the second color channel toward the second image sensor, and the second reflective member further configured to reflect light from the first color channel toward the first image sensor. (19) The apparatus according to embodiment 18, wherein the first reflective member comprises a first right-angle prism mirror, and the second reflective member comprises a second right-angle prism mirror. (20) Apparatus, the apparatus, An objective element that provides a field of view, An imaging lens, wherein the imaging lens is configured to receive light transmitted through the objective element, and the imaging lens is further configured to transmit light including spatially offset spectral channels through a focusing imaging space, A dichroic element in the convergent imaging space, wherein the dichroic element is configured to reflect light from at least one optical channel in the convergent imaging space and transmit light from at least one other optical channel in the convergent imaging space, and the dichroic element is further configured to induce astigmatism in the light transmitted through the dichroic element, An apparatus comprising: a correction element, the correction element being configured to induce a second astigmatism in the light transmitted through the dichroic element, and the second astigmatism being configured to offset the first astigmatism.

[0139] (21) A method, The method involves transmitting light from an imaging lens through a dichroic element, wherein the transmitted light includes a first color channel, a second color channel, a third color channel, and a fourth color channel, and the first and second color channels are spatially offset from the third and fourth color channels at the incident surface on the dichroic element. The light of the first color channel and the third color channel is transmitted through the dichroic element, wherein the dichroic element transmits light in a way that induces a first astigmatism in the first color channel and the third color channel. Reflecting the light from the second color channel and the fourth color channel, wherein the light from the second color channel and the fourth color channel is reflected by the dichroic element, The light of the first color channel and the third color channel is transmitted from the dichroic element through a correction element, wherein the correction element induces a second astigmatism in the first color channel and the third color channel, and the second astigmatism offsets the first astigmatism. The camera assembly receives light from the first color channel and the third color channel from the correction element, A method comprising receiving light from the second color channel and the fourth color channel from the dichroic element in the camera assembly.

Claims

1. Apparatus, the apparatus, A sample stage region configured to provide an object surface, An optical assembly, The optical assembly is an objective element that provides a field of view, wherein the object plane is within the field of view, An imaging lens, wherein the imaging lens is configured to receive light that has passed through the objective element, A first dichroic element, wherein the optical assembly is configured to transmit light from at least a first color channel, a second color channel, a third color channel, and a fourth color channel from the imaging lens toward the first dichroic element, the first dichroic element is configured to transmit light from the first color channel and the third color channel, while reflecting light from the second color channel and the fourth color channel, and the first dichroic element is further configured to induce a first astigmatism in the first color channel and the third color channel, An optical assembly comprising: a correction element configured to induce a second astigmatism in the first and third color channels, wherein the second astigmatism is configured to offset the first astigmatism; A device comprising: a camera assembly configured to receive light from the first color channel, the second color channel, the third color channel, and the fourth color channel.

2. The apparatus according to claim 1, wherein the first color channel and the second color channel are angularly offset from the third color channel and the fourth color channel on the incident surface of the first dichroic element.

3. The apparatus according to claim 2, wherein the optical assembly further comprises a color-molded assembly, the color-molded assembly configured to provide the angular offset of the light from the first and second color channels from the light from the third and fourth color channels.

4. The apparatus according to claim 3, wherein the color-molded assembly includes a second dichroic element and a reflective element, the reflective element being angularly offset with respect to the second dichroic element.

5. The apparatus according to claim 4, wherein the second dichroic element is configured to reflect light from the first and second color channels, and the second dichroic element is further configured to allow transmission of light from the third and fourth color channels to the reflecting element, and the reflecting element is configured to reflect light from the third and fourth color channels.

6. The apparatus according to claim 3, wherein the color-molded assembly is positioned in the optical path between the objective lens element and the imaging lens.

7. The apparatus according to claim 1, further comprising an excitation assembly, the excitation assembly being operable to emit light toward two different regions in the surface of the object, the first and second color channels being associated with a first region of the two different regions, and the third and fourth color channels being associated with a second region of the two different regions.

8. The apparatus according to claim 1, wherein the first dichroic element is inserted between the imaging lens and the camera assembly.

9. The apparatus according to claim 1, wherein the first dichroic element comprises a first plate inclined at a first angle about a first axis.

10. The apparatus according to claim 9, wherein the correction element comprises a second plate inclined at a second angle about a second axis.

11. The apparatus according to claim 10, wherein the second axis is perpendicular to the first axis.

12. The apparatus according to claim 10, wherein the second angle is equal to the first angle.

13. The camera assembly is A first imaging sensor configured to receive light from the first color channel, A second imaging sensor configured to receive light from the second color channel, A third imaging sensor configured to receive light from the third color channel, The apparatus according to claim 1, further comprising: a fourth imaging sensor configured to receive light from the fourth color channel.

14. The camera assembly is A first reflective member, wherein the first reflective member is configured to reflect light from the first color channel toward the first image sensor, and the first reflective member is further configured to reflect light from the third color channel toward the third image sensor, The apparatus according to claim 13, further comprising: a second reflective member, the second reflective member configured to reflect light from the second color channel toward the second image sensor, and the second reflective member further configured to reflect light from the first color channel toward the first image sensor.

15. It is a method, The method involves transmitting light from an imaging lens through a dichroic element, wherein the transmitted light includes a first color channel, a second color channel, a third color channel, and a fourth color channel, and the first and second color channels are spatially offset from the third and fourth color channels at the incident surface on the dichroic element. The light of the first color channel and the third color channel is transmitted through the dichroic element, wherein the dichroic element transmits light in a way that induces a first astigmatism in the first color channel and the third color channel. The reflection of light from the second color channel and the fourth color channel, wherein the light from the second color channel and the fourth color channel is reflected by the dichroic element. The light of the first color channel and the third color channel is transmitted from the dichroic element through a correction element, wherein the correction element induces a second astigmatism in the first color channel and the third color channel, and the second astigmatism offsets the first astigmatism. The camera assembly receives light from the first color channel and the third color channel from the correction element, A method comprising receiving light from the second color channel and the fourth color channel from the dichroic element in the camera assembly.

Images