Sequencing system comprising a base unit and a detachable cartridge

Abstract

Implementations include systems for sequencing a biological sample. The systems can include a reusable subsystem and a removable subsystem. The reusable subsystem can actuate and operate the removable subsystem to automate sequencing. A base unit of the reusable subsystem can form a fluidic connection between an integrated reagent cartridge and an integrated sensor cartridge of the removable subsystem. The integrated reagent cartridge can be configured to hold reagents and the integrated sensor cartridge can be configured with a biosensor for sequencing the biological sample.

Description

[0001] Related applications

[0002] This application claims priority and benefit on the filing date of U.S. Provisional Patent Application Serial No. 63 / 107,712, filed October 30, 2020, entitled “Sequencing Systems Including a BaseUnit and Removable Cartridge,” the entire contents of which are hereby incorporated by reference. Technical Field

[0003] Equipment and methods for nucleic acid sequencing, and more specifically, reagent kits and sensor cassettes used for sequencing. Background Technology

[0004] With advancements in nucleic acid sequencing technology, efforts have been made to reduce the complexity and cost of sequencers. Many of these technologies utilize microfluidics, which handles the behavior, precise control, and manipulation of fluids that may be geometrically confined to a very small (typically sub-millimeter) scale, with capillary penetration at this scale controlling mass transfer.

[0005] Sequencing is the process of determining the sequence of nucleic acids, or (for example, the nucleotide sequence in DNA). DNA sequencing includes methods or techniques for determining the sequence of four base nucleotides: adenine, guanine, cytosine, and thymine. Knowledge of DNA sequences has become essential for basic biological research and many applied fields, such as medical diagnostics, biotechnology, forensic biology, virology, and biosystematics. Comparing healthy and mutated DNA sequences can diagnose various diseases (including various cancers), characterize antibody libraries, and guide patient treatment. Rapid DNA sequencing methods enable faster and more personalized medical care and allow for the identification and classification of a wider range of organisms. Summary of the Invention

[0006] In this patent, we describe systems and methods for sequencing DNA and other nucleic acids. These systems and methods may include reusable basic units and removable cartridges. The removable cartridges may include integrated reagent kits (IRCs) and integrated sensor cartridges (ISCs).

[0007] There are many nucleic acid (e.g., DNA) sequencing methods, such as massively parallel sequencing. See, for example, Kumar, K., 2019, “Next-Generation Sequencing and Emerging Technologies,” Semin ThrombHemost 45(07):661-673. Traditional sequencing systems often encounter many challenges. For example, many traditional sequencing systems are not portable and are expensive due to their size. Many traditional sequencing systems also require external light sources, lasers, cameras, and platforms to accurately read and sequence DNA samples. The embodiments disclosed herein include sensor cassettes that allow for easy reconfiguration of the sequencing system. For example, the sensor cassettes may have larger or smaller sensor regions, different microfluidic channel configurations, or other properties that may be beneficial depending on a specific workflow. Furthermore, the kits and sensor cassettes are stored separately from the base unit. When the end user requires reagents and sensors, the kits and sensor cassettes can be coupled to the base unit to deliver reagents and sequenced DNA or other nucleic acid samples on demand. In some exemplary implementations, the system allows for a variety of sequencing applications with different read lengths (30bp to 700bp) and read throughputs (4M to 500M) within a relatively short turnaround time (3 hours to 48 hours).

[0008] The disclosed implementation offers several advantages over more traditional solutions. For example, the sequencing system provides appropriate sequencing reagent storage for both external (e.g., light-protected, frozen, airtight) and onboard (e.g., light-protected, suitable temperature, oxygen-permeable, light-protected) conditions to ensure optimal reagent chemical reactivity and proper reagent handling, thereby preventing inter-run contamination and corrosion of the base unit. As another example, the sequencing system provides the ability to accept and capture sequencing-significant nucleic acids in liquid sample form. Furthermore, the sequencing system provides appropriate reagent release functionality during base unit operation. Therefore, all reagents are optional during the sequencing reaction and are available for use. As another example, the sequencing system provides appropriate reagent delivery functionality, allowing all reagents to be programmed and delivered to the sequencing reaction site with volumetric accuracy on time, without the risk of cross-contamination of sensitive reagents. As another example, the sequencing system provides appropriate sequencing temperatures that allow for optimal sequencing reaction kinetics. In addition, the sequencing system provides detection functionality for sequencing reaction events and converts the signals into digital data formats. For example, sequencing systems provide automated sequencing base detection and related bioinformatics functions, including but not limited to decoding raw sequencing signal data into nucleic acid base sequences, base detection quality control, read alignment and genome / transcription assembly, as well as feature detection and quantification.

[0009] This summary provides a simplified description of different embodiments of the invention, which will be described in further detail below. This summary is not intended to limit the scope of the claimed subject matter. Other features, details, uses, and advantages of the claimed subject matter will become apparent from the following detailed description.

[0010] In one example, the system may include: (a) a detachable integrated sensor cassette (ISC) having: (i) a fluid control network including: a sample reservoir for receiving biological samples; a reaction chamber having at least one biosensor and an opaque surface spaced apart from the at least one biosensor; multiple reagent receiving ports; and multiple fluid channels for guiding biological samples and reagents to the reaction chamber; (ii) a reagent selection valve including: multiple valve ports; an output channel fluidly connected to the reaction chamber; and a bridging channel configured to fluidly couple one of the multiple valve ports to the output channel; (iii) a biosensor assembly including: at least one biosensor having a detector array for detecting bioanalytes on or near its functionalized surface; a substrate having electrical I / O pads providing connection to an electrical connector assembly of at least one biosensor and a base unit; and multiple electrical connections connecting at least one biosensor to I / O pads of the substrate; (b) a detachable assembly An integrated reagent kit (IRC) comprising: (i) a housing including: a plurality of fluid connectors located on a bottom surface configured to fluidly couple to a reagent receiving port of a removable integrated sensor cartridge (ISC); (ii) a plurality of reagent reservoirs disposed within the housing; and (iii) a waste container disposed within the housing; and (c) a base unit comprising: (i) a pump assembly fluidly connected to the removable integrated sensor cartridge (ISC) and the removable integrated reagent kit (IRC); (ii) a valve actuator for engaging a reagent selection valve of the removable integrated sensor cartridge (ISC); and (iii) an electrical connector assembly for controlling and receiving data from a biosensor assembly of the removable integrated sensor cartridge (ISC); wherein the removable integrated sensor cartridge (ISC), the removable integrated reagent kit (IRC), and the base unit are operatively coupled to each other via a system interface using at least one of fluid coupling, electrical coupling, or thermal coupling to jointly define a system interface.

[0011] In this example, the reaction chamber may have multiple reaction sites.

[0012] In this example, the functionalized surface of the biosensor can have multiple activity sensing regions.

[0013] In this example, the biosensor can be a CMOS image sensor with a functionalized surface.

[0014] In this example, the opaque surface of the reaction chamber can be the surface of a second biosensor.

[0015] In this example, the base unit may have a cooling unit that is fluidly connected to the detachable integrated kit for active cooling of the reagents.

[0016] In this example, the base unit may have a TEC unit that is coupled to a detachable integrated sensor box (ISC) to control the temperature of the reaction chamber.

[0017] In this example, the waste container can be a separate component of the pump assembly that is directly connected to the base unit.

[0018] In this example, the outer casing may have at least one opening to receive at least one reagent for the reaction.

[0019] In this example, the detachable integrated sensor box (ISC) may include a disposable pump.

[0020] In another example, the system includes: (a) a removable integrated sensor cassette (ISC) having: (i) a fluid control network including: a sample reservoir for receiving biological samples; a reaction chamber having at least one biosensor and an opaque surface spaced apart from the at least one biosensor; multiple reagent receiving ports; and multiple fluid channels guiding biological samples and reagents to the reaction chamber; (ii) a reagent selection valve including: multiple valve ports; an output channel fluidly connected to the reaction chamber; and a bridging channel configured to fluidly couple one of the multiple valve ports to the output channel; (iii) a biosensor assembly including: at least one biosensor having a detector array for detecting bioanalytes on or near its functionalized surface; a substrate having electrical I / O pads providing connection to an electrical connector assembly of at least one biosensor and a basic unit; and multiple electrical connections connecting the at least one biosensor to I / O pads of the substrate; and (b) a removable integrated reagent kit (IRC) having: (i) a housing containing Includes: (i) a plurality of fluid connectors located on the bottom surface configured to fluidly couple to a reagent receiving port of a removable integrated sensor cartridge (ISC); (ii) a plurality of reagent reservoirs disposed within a cartridge housing; (iii) a disposable pump; (iv) a plurality of flow control valves; (v) a waste container disposed within the cartridge housing; (c) a base unit having: (i) a pump actuator for engaging the disposable pump of the removable integrated reagent kit (IRC); (ii) a valve actuator for engaging the flow control valve of the removable integrated reagent kit (IRC); (iii) another valve actuator for engaging the reagent selection valve of the removable integrated sensor cartridge (ISC); and (iv) an electrical connector assembly for controlling and receiving data from a biosensor assembly of the removable integrated sensor cartridge (ISC); wherein the removable integrated sensor cartridge (ISC), the removable integrated reagent kit (IRC), and the base unit are operatively coupled to each other via a system interface using at least one of fluid coupling, electrical coupling, or thermal coupling to jointly define a system interface.

[0021] In this example, multiple flow control valves may be part of a detachable integrated sensor box (ISC).

[0022] In this example, the waste container can be a separate part that is directly connected to the pump assembly of the detachable integrated reagent kit (IRC).

[0023] In another example, a sequencing system includes: (a) a removable integrated reagent kit (IRC) including one or more reservoirs for containing one or more sequencing reagents, the IRC further including one or more connectors in fluid communication with the one or more reservoirs; (b) a removable integrated sensor cartridge (ISC) including: (i) one or more reagent receiving ports, which are positioned in fluid connection to the one or more connectors of the IRC when the IRC is engaged with the ISC; (ii) at least one biosample input; (iii) a reaction chamber including at least one sensor, the sensor including a functionalized surface and a detector array configured to detect a bioanalyte on or near the functionalized surface; (iv) a flow control network configured to selectively fluidly connect the reagent receiving port and the biosample input to the reaction chamber; and (c) a base unit configured to removably receive the IRC and the ISC, the base unit being configured to control the sequencing reaction in the reaction chamber and to receive sequencing data from the sensor when the IRC and the ISC are loaded into the base unit.

[0024] In this example, the flow control network further includes a multi-position valve that selectively connects the reagent receiving port and the biological sample input port to the reaction chamber, and the basic unit may include a valve actuator configured to actuate the multi-position valve.

[0025] In this example, the multi-position valve includes multiple valve ports fluidly connected to the reagent receiving port and the biological sample input port, and fluidly connected to the output channel of the reaction chamber; and a repositionable bridging channel configured to fluidly couple one of the multiple valve ports to the output channel.

[0026] In this example, the biological sample input may include a sample reservoir on the ISC, which is configured to receive the biological sample.

[0027] In this example, the reaction chamber may include an opaque surface spaced apart from the at least one sensor.

[0028] In this example, the opaque surface of the reaction chamber may be a second biosensor.

[0029] In this example, the sensor further includes a substrate comprising electrical contacts electrically coupled to the detector array, wherein the base unit further includes an electrical connector assembly configured to be electrically connected to the electrical contacts to receive sequencing data from the sensor.

[0030] In this example, the IRC may have a housing, the reservoir is located inside the housing, and the IRC may also include a waste container located inside the housing, the waste container being configured to receive used sequencing reagents.

[0031] In this example, the system may alternatively include a separate waste container configured to receive used sequencing reagents.

[0032] In this example, the base unit may include a pump assembly configured to be fluidly connected to the ISC and the IRC.

[0033] In this example, the IRC may alternatively include a disposable pump, wherein the base unit includes a pump actuator configured to engage the disposable pump.

[0034] In this example, the reaction chamber may include multiple reaction sites.

[0035] In this example, the functionalized surface of the sensor may include multiple active sensing regions.

[0036] In this example, the sensor may be a CMOS image sensor adjacent to the functionalized surface.

[0037] In this example, the base unit may further include a cooling unit fluidly connected to the IRC, the cooling unit being configured to actively cool the reagent.

[0038] In this example, the base unit may include a temperature control unit coupled to the ISC, the temperature control unit being configured to control the temperature of the reaction chamber.

[0039] In this example, the IRC may also include at least one opening configured to receive at least one sequencing reagent.

[0040] In this example, the base unit may also include one or more actuators configured to open the one or more stores of the IRC.

[0041] In this example, the IRC and ISC can be configured to be compressed together when loaded into the base unit.

[0042] In this example, the base unit can be configured to compress the IRC and ISC together.

[0043] In another example, a sequencing system may include: (a) a removable integrated reagent kit (IRC) configured to contain one or more sequencing reagents; (b) a removable integrated sensor cartridge (ISC) including a reaction chamber and at least one valve, the reaction chamber including at least one sensor electrically connected to a plurality of electrical contacts; and (c) a base unit, the system being configured to removably mount the IRC and ISC in the base unit, the base unit including a valve actuator and an electrical connector assembly, wherein the base unit is configured such that when the IRC and ISC are mounted in the base unit, the IRC is fluidly connected to the ISC, the valve actuator is operatively engaged with the at least one valve of the ISC, and the electrical connector assembly is electrically coupled to the plurality of electrical contacts of the ISC.

[0044] In this example, the at least one valve of the ISC may be a multi-position valve configured to selectively connect at least one of a plurality of reagent fluid channels and a biological sample fluid channel to the reaction chamber.

[0045] In this example, at least one of the IRC and ISC may be a plurality of additional flow control valves, and the base unit further includes at least one second valve actuator, and the base unit is configured such that, after the IRC and ISC are installed in the base unit, the at least one second valve actuator is operatively engaged with the plurality of additional flow control valves.

[0046] In this example, at least one of the IRC and ISC may also include a disposable pump, wherein the base unit includes a pump actuator, and wherein the base unit is configured such that the pump actuator is operatively engaged with the disposable pump after the IRC and ISC are mounted in the base unit.

[0047] In another example, an integrated sensor cassette (ISC) is configured for detachable mounting in a base unit of a sequencing system and includes: (a) one or more reagent receiving ports; (b) at least one biosample input; (c) a reaction chamber including at least one sensor comprising a functionalized surface and a detector array configured to detect a bioanalyte on or near the functionalized surface, the sensor including a substrate including electrical contacts electrically coupled to the detector array, the electrical contacts being configured to be electrically connected to the base unit when the ISC is mounted in the base unit; and (d) a flow control network configured to selectively fluidly connect the reagent receiving ports and the biosample input to the reaction chamber, the flow control network including a multi-position valve selectively connecting the reagent receiving ports and the biosample input to the reaction chamber, the multi-position valve being configured to be actuated by the base unit when the ISC is mounted in the base unit.

[0048] In another example, a sequencing method uses a sequencing system having a basic unit, a removable integrated reagent kit (IRC) including at least one sequencing reagent, and a removable integrated sensor cassette (ISC) including a reaction chamber, the method comprising the steps of: (a) mounting the IRC and ISC in the basic unit; (b) engaging at least one valve actuator of the basic unit with at least one valve of at least one of the IRC and ISC; and (c) electrically connecting an electrical connector assembly of the basic unit to an electrical contact of the ISC.

[0049] In this example, at least one valve may be a multi-position valve that selectively connects one or more fluid passages of a plurality of channels to the reaction chamber, wherein the base unit is configured to control the position of the multi-position valve using at least one valve actuator.

[0050] In this example, at least one of the IRC and ISC may include a disposable pump, wherein the base unit includes a pump actuator, and wherein the method includes engaging the pump actuator with the disposable pump.

[0051] In another example, the integrated flow control cassette includes: (a) a removable integrated reagent kit (IRC) comprising: (i) a housing having a plurality of fluid connectors on its bottom surface configured to fluidly couple to a reagent receiving port of a removable integrated sensor cassette (ISC); (ii) a plurality of reagent reservoirs disposed within the housing; (iii) a disposable pump; (iv) a plurality of flow control valves; and (v) a waste container disposed within the housing; (b) a removable integrated sensor cassette (ISC) having at least one opaque surface, the removable integrated sensor cassette (ISC) comprising: (i) a flow control network; (ii) a reaction chamber having at least one biosensor; and (iii) a biosensor assembly comprising: at least one biosensor with a detector array for detecting bioanalytes on or near its functionalized surface; a substrate having electrical I / O pads providing connections to an electrical connector assembly of at least one biosensor and a base unit; and a plurality of electrical connections connecting at least one biosensor to an I / O pad of the substrate.

[0052] In another example, the sequencing system includes: (a) a removable integrated cartridge having: (i) a reagent storage portion configured to contain one or more sequencing reagents; (ii) a flow control and sensing portion including a reaction chamber and at least one valve, the reaction chamber including at least one sensor electrically connected to a plurality of electrical contacts; and (b) a base unit configured to removably mount an IRC and an ISC in the base unit, the base unit including a valve actuator and an electrical connector assembly, and the base unit being configured such that when the integrated cartridge is mounted in the base unit, the IRC is fluidly connected to the ISC, the valve actuator is operatively engaged to at least one valve of the ISC, and the electrical connector assembly is electrically coupled to a plurality of electrical contacts of the ISC.

[0053] In another example, the sequencing system includes: (a) a removable integrated reagent kit (IRC) including at least one valve, the IRC being configured to contain one or more sequencing reagents; (b) a removable integrated sensor cassette (ISC) with a reaction chamber including at least one sensor electrically connected to a plurality of electrical contacts; and (c) a base unit configured to removably mount the IRC and ISC within the base unit, the base unit including a valve actuator and an electrical connector assembly, and the base unit being configured such that when the IRC and ISC are mounted in the base unit, the IRC is fluidly connected to the ISC, the valve actuator is operatively engaged to at least one valve of the IRC, and the electrical connector assembly is electrically coupled to a plurality of electrical contacts of the ISC. Attached Figure Description

[0054] It should be understood that, for the sake of simplicity and clarity, the elements shown in the figures are not necessarily drawn to scale. For example, for clarity, the dimensions of some elements are enlarged relative to each other. Furthermore, where deemed appropriate, reference numerals are repeated between figures to indicate corresponding elements.

[0055] Figure 1 An example of the basic unit of a sequencing system is shown.

[0056] Figure 2A-2C An example of an integrated kit for a sequencing system is shown.

[0057] Figures 3A-3C An example of an integrated sensor box for a sequencing system is shown.

[0058] Figures 4A-4B Examples of integrated kits and integrated sensor boxes located within a base unit are shown.

[0059] Figures 5A-5B An example of a basic unit that engages with an integrated sensor box is shown.

[0060] Figures 6A-6B An example of the basic unit of the integrated kit is illustrated. Detailed Implementation

[0061] In the following detailed description, reference is made to the accompanying drawings, which form a part of the invention, and specific embodiments in which the invention may be implemented are illustrated by way of illustration. The terms “height,” “top,” “bottom,” etc., are used with reference to the orientation in the described drawings. Because components of embodiments of the invention can be positioned in many different orientations, these terms are used for illustrative purposes and not for limitation.

[0062] As used herein, a “sequencing event” refers to the emission of a light signal (e.g., a fluorescent or luminescent signal) generated by the sequencing process. An exemplary sequencing process is a cycle of synthetic sequencing. In this approach, nucleotides are incorporated as primer extension products (e.g., using reversible termination nucleotides). In this approach, nucleotides can be labeled, for example, with fluorescent dyes or luminescent signal sources (e.g., luciferase or luciferase substrates). Luminescent signals include chemiluminescence and bioluminescence. Nucleotides can be directly labeled with fluorescent dyes or luminescent signal sources, or they can be bound to antibodies, aptamers, or other reagents labeled with signal-generating motifs. During sequencing, a defined light signal is generated at each site in the array by, for example, irradiating the fluorescent dye with an excitation wavelength, and the signal and corresponding position are recorded.

[0063] Figure 1An example of a base unit 100 of a sequencing system is shown. In some embodiments, the base unit 100 is a reusable subsystem of the sequencing system that can actuate and operate one or more removable subsystems (e.g., an integrated reagent kit (IRC) and an integrated sensor cassette (ISC)). Fluid coupling, electrical coupling, and / or thermal coupling can be established through interfaces of the base unit 100, the IRC, and the ISC. For example, a pump assembly of the base unit 100 can fluidly couple the base unit 100 to the IRC and / or the ISC. The base unit 100 may additionally include one or more valve actuators to engage components of the IRC and the ISC. For example, a first valve actuator may engage a flow control valve of the IRC, and a second valve actuator may engage a reagent selection valve of the ISC. In one example, the base unit 100 may include a loading region 104 with a gate. A removable subsystem can be inserted into the base unit 100 with the gate open prior to sequencing.

[0064] In some implementations, the base unit 100 includes modules for performing sequencing-related operations. The controller module of the base unit 100 may include a user interface 102 for selecting sequencing workflows and otherwise providing information input and / or output. The user interface 102 may be a touchscreen or other interface capable of receiving selections and / or displaying information. The controller module of the base unit 100 may communicate with additional modules of the base unit 100 during sequencing. For example, additional modules may include a loading module, one or more compression modules, one or more thermal control modules, a reagent selection module, a reagent dispensing module, a sensor readout module, and a data storage and processing module. The loading module may control the entry and exit of the removable subsystem. One or more compression modules may join the IRC, ISC, and / or components of the base unit 100 together. In one example, one or more compression modules may control: piercing the IRC of the removable subsystem, compressing the IRC and ISC to form a closed fluid pipeline, and pressing the thermoelectric cooler (TEC) and socket to the pad grid array (LGA) of the ISC. One or more thermal control modules may provide temperature regulation for the IRC and ISC. For example, one or more thermal control modules can provide temperature control for the IRC via non-contact air cooling and provide temperature ramping for the ISC reaction chamber. The thermal control modules can also dynamically adjust the TEC temperature based on data read from the ISC by sensors. For example, the thermal control module may include a cooling unit fluidly connected to the IRC to actively cool reagents. A reagent selection module can provide actuation to rotate the ISC's rotary valve to a desired position. A reagent dispensing module can control the supply of reagents to the ISC. For example, the reagent dispensing module can provide negative pressure to pull and meter sequencing reagents using a reagent selection valve and a motor-driven pump assembly. An electrical connector assembly, including a sensor readout module and a data storage and processing module, can control and receive data from the biosensor components of the ISC. The sensor readout module can provide a connection to the ISC's LGA to read analog signals of sequencing events and the temperature inside the reaction chamber. The sensor readout module can additionally convert analog signals into a digital format for data storage in the data storage and processing module.

[0065] Figure 2A-2C An example of an integrated reagent kit (IRC) 220 for a sequencing system is shown. The IRC 220 can be used as a sequencing reagent rack (optionally including a waste container for used reagents) before selecting a sequencing workflow. In some examples, the IRC 220 can hold between five and thirty different sequencing-related reagents, with volumes ranging from one to two hundred milliliters and a total volume up to six hundred milliliters. The dimensions of the IRC 220 can range from forty to one hundred and sixty millimeters in each orientation or other dimensions.

[0066] In some implementations, the IRC 220 may include a housing having a top cover 212 and a bottom cover 214. The top cover 212 may interface with the base unit, for example with... Figure 1 The basic unit 100 is docked. The bottom cover 214 can dock with the ISC, an example of which is shown in... Figures 3A-3B As further described herein, one or more reagent containers for receiving or storing reagents may be housed within the housing. The IRC 220 may additionally include multiple flow control valves to control the fluid flow between the reagent reservoir and the microfluidic channel of the ISC.

[0067] exist Figure 2A-2B In the specific example shown, the top cover 212 includes one or more inlet ports 202, one or more reagent ports 204, a fluid connection 206, one or more cantilever puncture devices 208, and one or more air ports 210. Inlet ports 202 allow one or more actuators of the base unit to push a sealed reagent reservoir within the IRC 220 down to an open state for reagent dispensing. Reagent ports 204 can receive reagents aspirated into the IRC 220 by a user, allowing for customized reagent modifications and / or additions. Fluid connections 206 can connect a pump line from the base unit to the IRC 220. Cantilever puncture devices 208 can be actuated by the base unit such that they puncture a covering (e.g., foil) on the reagent reservoir within the housing for ventilation. Opening the reagent reservoir via actuation of the base unit allows for reagent release. Air ports 210 can provide a path for the base unit to supply air into the IRC 220. For example, temperature-controlled air can be supplied from the thermal control module of the base unit to the IRC 220 through air port 210, thereby providing a suitable temperature environment for onboard reagents when the IRC 220 is operating in the base unit.

[0068] In the example shown, the bottom cover 214 of the IRC 220 includes a pump seal 216 and one or more reagent seals 218. The number of reagent seals 218 may equal the number of reagent reservoirs within the IRC 220. The pump seal 216 provides a seal between the pump lines in the IRC 220 and the pump lines in the ISC. The reagent seals 218 form a seal between the reagent reservoirs in the housing and the reagent receiving port of the ISC. The reagent seals 218 can function as a fluid connector between the IRC 220 and the ISC, allowing the IRC 220 to supply sequencing reagents to the sequencing reaction sites of the ISC. The pump seal 216 and reagent seals 218 may be rubber-based gaskets or other suitable materials.

[0069] In some implementations, the IRC 220 may additionally include a waste container within the casing. This waste container can receive the fluid after it has been used for the sequencing reaction in the ISC. Alternatively, the waste container may be external to the IRC 220 and may be directly connected to the pump assembly of the base unit. A disposable pump may also be located within the IRC 220 to fluidly connect the base unit and the ISC. Alternatively, the disposable pump may be a component of the ISC.

[0070] Figures 3A-3C An example of an integrated sensor cassette (ISC) 330 for a sequencing system is shown. The ISC 330 can be used with a basic unit (e.g., Figure 1 The basic unit 100 in the middle) and IRC (e.g. Figure 2A-2B The ISC330 may include a flow control network, a reagent selection valve 306, and a biosensor assembly 308. The ISC330 may also include multiple flow control valves located on either side of the biosensor assembly 308 to control fluid flow between components of the sequencing system.

[0071] refer to Figures 3A-3B In some implementations, the flow control network of the ISC 330 includes a sample reservoir 304, a reaction chamber 310, one or more reagent receiving ports 302, and one or more fluid channels 316. The sample reservoir 304 can receive biological samples. Biological samples are biological materials (blood, urine, tissue, cell cultures, saliva, etc.) from living or dead organisms (e.g., humans, animals, etc.). Biological samples can be processed and purified DNA from biological materials. In one example, the sample reservoir 304 can be a dome-shaped feature capable of receiving liquid volumes between ten and two hundred microliters. The dome-shaped feature minimizes the dead volume of the sample. The base unit can couple the reagent receiving ports 302 to the fluid connectors of the IRC to form a fluid connection for each reagent. The ISC 330 can have a number of reagent receiving ports 302 equal to the number of reagent reservoirs and fluid connectors in the IRC. Fluid channel 316 connects sample reservoir 304 and reagent receiving port 302 to reaction chamber 310, which may include one or more sequencing reaction sites. Although Figure 3A ISC 330 shows one example of fluid channel 316, but other examples may include different numbers or arrangements of fluid channels.

[0072] The reagent selection valve 306 may include a valve port, an output channel, and a bridging channel. The valve port provides a fluid connection between the reagent receiving port 302 and the reagent selection valve 306. The output channel fluidly connects the reagent selection valve 306 to the reaction chamber 310 via a main line 318. The bridging channel fluidly couples one of the multiple valve ports to the output channel, allowing reagents from the reagent receiving port 302 to be transferred to the reaction chamber 310. Depending on the sequencing workflow selected in the base unit, the bridging channel can be rotated to connect a specific valve port to the output channel. Rotation of the bridging channel can be controlled by the base unit.

[0073] refer to Figure 3C In some embodiments, reaction chamber 310 includes an opaque surface and at least one biosensor, which may be identical to biosensor assembly 308 and spaced apart from the opaque surface. The opaque surface may be a plastic material, and in some examples, it may also be the biosensor surface. For example, the biosensor may form the bottom surface of reaction chamber 310, and the opaque surface may be a coverslip covering reaction chamber 310. In another example, both the bottom and top surfaces of reaction chamber 310 may be biosensors, and the opaque surface may be an outer coating or component. The biosensor may be a silicon-based complementary metal-oxide-semiconductor (CMOS) sensor with a functionalized surface. In one example, the functionalized surface includes one or more activity sensing regions. The width and / or length of reaction chamber 310 may be between three and seventy millimeters, and the height between fifty and two hundred and fifty micrometers. The dimensions of reaction chamber 310 can be adjusted according to different sequencing applications. For example, a user can select an ISC with a reaction chamber 310 and / or fluidics, which has a specific size and other configurations for different ISCs with different sizes or configurations. The surface of the CMOS sensor can be exposed in the reaction chamber 310 to provide binding sites for DNA in biological samples for sequencing. The opacity of the opaque surface can be achieved by integrating light-shielding features (e.g., carbon dyes in plastic, or altering surface roughness) or by attaching an additional light-shielding cap to the surface. Additionally, the reaction chamber 310 may include an inlet 312 and an outlet 314. The inlet 312 can be connected to a main line for receiving sequencing reagents. Figure 3A (318 in the text). Outlet 314 can be connected to a waste line, which serves as a fluid connection to an external pump source or waste container.

[0074] In some implementations, the biosensor assembly 308 may include a biosensor to detect sequencing events. When a pixel of the biosensor detects light (e.g., bioluminescence, cold light, or chemiluminescence generated by a sequencing event), a voltage spike or some other electrical event will occur in the pixels connected to the LGA. The LGA includes a substrate having an array of electrical I / O pads (e.g., wires and contacts) surrounding the reaction chamber 310, the electrical I / O pad array being communicatively coupled to an input in the base unit, allowing the base unit to determine which pixels have detected a sequencing event. Analog signals detected by the biosensor can be transmitted via the electrical I / O pad array to a sensor readout module in the base unit. The temperature of the reaction chamber 310 can also be monitored, for example, by a thermistor, which can be transmitted via the electrical I / O pad array and read out by the base unit. This provides real-time temperature monitoring and feedback to the thermal control module of the base unit. In one example, the central portion of the electrical I / O pad array may be a thermally conductive material (e.g., copper), allowing the TEC module of the thermal control module to engage with the detector array and efficiently transfer heat to the biosensor.

[0075] Figures 4A-4B An example of loading an integrated reagent kit (IRC) and an integrated sensor cartridge (ISC) into a base unit 400 is shown. The bottom cover 414 of the IRC can be coupled to the integrated sensor cartridge (ISC) (below the IRC) via a loading module 450 of the base unit 400. The loading module 450 may include one or more alignment pins to properly align the ISC and IRC. Once loaded, the top cover 412 of the IRC can engage with additional modules of the base unit 400. For example, a compression module can control the puncture of the IRC and compress the IRC and ISC to form a closed fluidic pipeline. In an example where the IRC does not include a waste reservoir, a waste container 440 may also be positioned within the base unit 400 during sequencing. The waste container 440 may be disposed within the housing of the IRC or as a separate component that docks with the pump assembly of the base unit 400 (e.g., Figures 4A-4B (As shown).

[0076] Figures 5A-5BAn example of a base unit 500 coupled to an integrated sensor cassette (ISC) 530 is shown. The base unit 500 may include a valve actuator 532 coupled to a motor and a loading module 550. The loading module 550 may include one or more alignment pins to ensure proper alignment of the ISC 530 within the base unit 500. During sequencing, the valve actuator 532 may be coupled to a reagent selection valve 506. The reagent selection module of the base unit 500 may control the actuation force of the motor to control the rotation of the reagent selection valve 506 to a specific position. This specific position may correspond to a fluid connection from the reagent reservoir of the integrated reagent kit (IRC) to a reaction chamber located below the biosensor assembly 508. The specific position may be determined by the base unit 500 based on sequencing settings selected at a controller module of the base unit 500. The reaction chamber may receive a biological sample from a sample reservoir 504 and reagents from the reagent reservoir associated with the specific position. The biosensor assembly 508 can detect bioanalytes during interactions between biological samples and reagents and transmit signals indicating the bioanalytes to the sensor readout module of the base unit 500.

[0077] refer to Figure 5B The base unit 500 may additionally include a thermal control module 524 and a sensor readout module 526. The sensor readout module 526 can determine the analog signal from sequencing and the temperature inside the reaction chamber. The sensor readout module 526 can convert the analog signal into a digital format and transmit the digital signal containing sequencing information and temperature to the data storage and processing module of the base unit 500.

[0078] In some implementations, the thermal control module 524 can receive commands from other modules of the base unit 500 to dynamically control the temperature of the reaction chamber. For example, when a sequencing workflow is selected in the controller module, the thermal control module 524 can provide a heating feature to the reaction chamber to set it to a suitable temperature. Additionally, during sequencing, the thermal control module 524 can receive commands from the data storage and processing module to adjust the temperature of the reaction chamber. This command can be determined based on the temperature read by the sensor readout module 526 being outside a predetermined temperature range. The thermal control module 524 can dynamically adjust the TEC temperature target based on the command.

[0079] Figures 6A-6B An example of a base unit coupled with an Integrated Reagent Kit (IRC) 620 is shown. The IRC 620 can be loaded into the base unit on a loading module 650. After loading, the IRC can be positioned within the base unit's compression module 660. In non-compression mode, as... Figure 6AAs shown, the height of the compression module 660 can be greater than the height of the IRC 620. During sequencing, the compression module 660 can be compressed to pierce the IRC 620 and form a fluidic tube between the base unit, the IRC 620, and the integrated sensor cassette (ISC). Figure 6B The compressed module 660 is displayed.

[0080] In some implementations, the various components of the different embodiments described herein can be manufactured using injection molding. Such a process can result in low-cost components and make the kit cost-effective as a single-use consumable. Furthermore, because the IRC and ISC are separate from and isolated from the base unit, they can be stored separately under suitable conditions, resulting in improved functionality for both the reagents and the sensor in terms of sequencing accuracy and lifetime.

[0081] The system described herein, although typically constructed in the context of biological samples, can be used for the determination of non-biological analytes. In one approach, the system is used for any large-scale parallel determination, where optical signals identify the characteristics of the analyte.

[0082] It should be understood that the examples and embodiments described herein are for illustrative purposes only, and various modifications or changes made thereto will inspire those skilled in the art and will be included within the spirit and scope of this application and the appended claims. All publications, patents, and patent applications cited herein are incorporated herein by reference in their entirety for all purposes.

[0083] It should be understood that the above description is intended to be illustrative rather than restrictive. Many embodiments will be apparent to those skilled in the art upon reading the above description. Therefore, the scope of the invention should not be determined by reference to the above description, but rather by reference to the full scope of the appended claims and their equivalents.

[0084] While the foregoing disclosure illustrates illustrative aspects of this disclosure, it should be noted that various changes and modifications may be made herein without departing from the scope of this disclosure as defined by the appended claims. Furthermore, although elements of this disclosure may be described or claimed in the singular, plural forms are contemplated unless a limitation on the singular is expressly stated.

Claims

1. A sequencing system comprising: (a) A removable integrated reagent kit (IRC) comprising a housing surrounding one or more reservoirs for containing one or more sequencing reagents, the IRC further comprising one or more connectors in fluid communication with the one or more reservoirs, the one or more connectors comprising a plurality of reagent seals on the housing and a pump seal on the housing; (b) A detachable integrated sensor housing (ISC), said ISC comprising: (i) Multiple reagent receiving ports, wherein when the IRC is engaged with the ISC, the reagent receiving ports are positioned to be fluidly connected to the reagent seal of the IRC; (ii) At least one biological sample input terminal; (iii) A reaction chamber comprising at least one sensor, the sensor comprising a functionalized surface and a detector array configured to detect a bioanalyte on or near the functionalized surface; (iv) A flow control network configured to selectively fluidly connect the reagent receiving port and the biological sample input port to the reaction chamber; and (c) A base unit configured to detachably receive the IRC and the ISC, wherein the IRC and the ISC are configured to be compressed together when loaded into the base unit, the base unit being configured to control the sequencing reaction in the reaction chamber and to receive sequencing data from the sensor when the IRC and the ISC are loaded into the base unit.

2. The sequencing system according to claim 1, wherein, The flow control network also includes a multi-position valve that selectively connects the reagent receiving port and the biological sample input port to the reaction chamber, wherein the basic unit also includes a valve actuator configured to actuate the multi-position valve.

3. The sequencing system according to claim 2, wherein, The multi-position valve includes multiple valve ports fluidly connected to the reagent receiving port and the biological sample input port, and fluidly connected to the output channel of the reaction chamber; And a repositionable bridging channel configured to fluidly couple one of the plurality of valve ports to the output channel.

4. The sequencing system according to claim 1, wherein, The biological sample input terminal includes a sample reservoir on the ISC, which is configured to receive the biological sample.

5. The sequencing system according to claim 1, wherein, The reaction chamber also includes an opaque surface spaced apart from the at least one sensor.

6. The sequencing system of claim 5, wherein the opaque surface of the reaction chamber comprises a second biosensor.

7. The sequencing system of claim 1, wherein the sensor further includes a substrate including electrical contacts electrically coupled to the detector array, wherein the base unit further includes an electrical connector assembly configured to be electrically connected to the electrical contacts to receive sequencing data from the sensor.

8. The sequencing system according to claim 1, wherein, The IRC also includes a waste container located within the housing, the waste container being configured to receive used sequencing reagents.

9. The sequencing system according to claim 1, wherein, The system also includes a separate waste container configured to receive used sequencing reagents.

10. The sequencing system according to claim 1, wherein, The basic unit also includes a pump assembly configured to be fluidly connected to the ISC and the IRC.

11. The sequencing system according to claim 1, wherein, The IRC also includes a disposable pump, wherein the base unit further includes a pump actuator configured to engage the disposable pump.

12. The sequencing system according to claim 1, wherein, The reaction chamber includes multiple reaction sites.

13. The sequencing system according to claim 1, wherein, The functionalized surface of the sensor includes multiple active sensing regions.

14. The sequencing system of claim 13, wherein the sensor comprises a CMOS image sensor adjacent to the functionalized surface.

15. The sequencing system according to claim 1, wherein, The basic unit also includes a cooling unit fluidly connected to the IRC, the cooling unit being configured to actively cool the reagent.

16. The sequencing system according to claim 1, wherein, The base unit also includes a temperature control unit coupled to the ISC, the temperature control unit being configured to control the temperature of the reaction chamber.

17. The sequencing system according to claim 1, wherein, The IRC also includes at least one opening configured to receive at least one sequencing reagent.

18. The sequencing system according to claim 1, wherein, The basic unit also includes one or more actuators configured to open the one or more stores of the IRC.

19. The sequencing system according to claim 1, wherein, The base unit is configured to compress the IRC and ISC together.

20. A sequencing system comprising: (a) A detachable integrated reagent kit (IRC) configured to contain one or more sequencing reagents; (b) A detachable integrated sensor box (ISC) comprising a reaction chamber and at least one valve, the reaction chamber comprising at least one sensor electrically connected to a plurality of electrical contacts; as well as (c) A base unit, wherein the system is configured to detachably mount the IRC and ISC in the base unit, the base unit including a valve actuator and an electrical connector assembly. The base unit is configured such that when the IRC and ISC are mounted in the base unit, the base unit compresses the IRC and ISC together, fluidly connecting the IRC to the ISC, the valve actuator is operatively engaged with at least one valve of the ISC, and the electrical connector assembly is electrically coupled to the plurality of electrical contacts of the ISC.

21. The sequencing system of claim 20, wherein the at least one valve of the ISC comprises a multi-position valve configured to selectively connect at least one of a plurality of reagent fluid channels and a biological sample fluid channel to the reaction chamber.

22. The sequencing system of claim 21, wherein at least one of the IRC and ISC further comprises a plurality of additional flow control valves, wherein the base unit further comprises at least one second valve actuator, and wherein the base unit is configured such that, after the IRC and ISC are mounted in the base unit, the at least one second valve actuator is operatively engaged with the plurality of additional flow control valves.

23. The sequencing system of claim 22, wherein at least one of the IRC and ISC further comprises a disposable pump, wherein the base unit further comprises a pump actuator, and wherein the base unit is configured such that the pump actuator is operatively engaged with the disposable pump after the IRC and ISC are mounted in the base unit.

24. An integrated sensor cassette (ISC) configured for detachable mounting in a base unit of a sequencing system, said ISC comprising: (a) One or more reagent receiving ports, the reagent receiving ports being configured to be in fluid communication with a reagent reservoir via a reagent seal; (b) At least one biological sample input terminal; (c) A reaction chamber comprising at least one sensor, the sensor comprising a functionalized surface and a detector array configured to detect a bioanalyte on or near the functionalized surface, the sensor further comprising a substrate comprising electrical contacts electrically coupled to the detector array, the electrical contacts being configured to be electrically connected to the base unit when the ISC is mounted in the base unit; as well as (d) A flow control network configured to selectively connect the reagent receiving port and the biological sample input to the reaction chamber, the flow control network including a multi-position valve that selectively connects the reagent receiving port and the biological sample input to the reaction chamber, the multi-position valve being configured to be driven by the base unit when the ISC is installed in the base unit.

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