RU2805201C1 - Nozzle tube system for reagent mixing - Google Patents

Abstract

FIELD: mixing reagents.

SUBSTANCE: system comprises a flow cell channel configured to be fluidly coupled to the flow cell to retain analytes in the analytical system, a fluid system containing a plurality of flow channels for reagents, an additional flow channel, a mixing channel and a selector valve and a nozzle tube, fluidly coupled to an additional flow channel. Each of the plurality of reagent flow channels has a first end configured to be fluidly coupled to its reagent receiver of the plurality of reagent receivers, and a second end fluidly coupled to a selector valve. The additional flow channel has a first end configured to be in fluid coupling with the target receiver and a second end in fluid coupling with the selector valve. The mixing channel has a first end in fluid coupling with a selector valve. The fluid system is configured to suck reagents from a plurality of reagent receivers into the mixing channel by means of a selector valve, eject the sucked-in reagents from the mixing channel into the target receiver via the selector valve and an additional flow channel, and supply reagents from the target receiver to the flow channel of the flow cell. The nozzle tube has an elongated body having a central cavity extending between the proximal end of the elongated body and the distal end of the elongated body, and a nozzle insert located at the distal end of the elongated body. The fluid system is configured to both suck the mixed reagents from the target receiver and eject the mixed reagents back into the target receiver through the nozzle insert.

EFFECT: possibility of efficient mixing of reagents and matrix of samples in an automatic or semi-automatic manner.

15 cl, 11 dwg

Description

Cross reference to related applications

This application claims benefit from US Patent Application No. 15/841098, filed December 13, 2017, which claims benefit from US Patent Application No. 62/442765, filed January 5, 2017, as well as UK Patent Application No. 1704760.6, filed March 24, 2017 g., which also claims priority to US Patent Application No. 62/442,765, all of which prior applications are incorporated herein by reference in their entirety.

State of the art

In this field of technology, tools have been developed and continue to appear for sequencing the molecules under study, in particular, DNA, RNA and other biological samples. Prior to sequencing operations, samples of the molecules of interest are prepared to form a library or matrix, which will then be mixed with reagents and ultimately introduced into a flow cell, where individual molecules will be attached to sites and amplified to increase detection power. The sequencing operation involves repeating a cycle of steps to bind molecules at specified sites, label the associated components, image the components at specified sites, and process the resulting image data.

In such sequencing systems, fluid systems (or subsystems) provide the flow of substances (eg, reagents) under the control of some control system, such as a stored program computer and suitable interfaces.

Disclosure of the invention

Various features of one or more embodiments of the subject matter disclosed in the present invention are illustrated in the accompanying drawings and set forth in the description that follows. Other features, aspects and advantages of the present invention will become apparent upon examination of the specification, drawings and claims.

In some embodiments, a system may be provided comprising a flow channel configured to be fluidly coupled to a flow cell for retaining analytes in an analytical system; a fluid system for sucking reagents from the reagent receptacles, mixing the reagents, discharging the mixed reagents into a target receptacle, and supplying the mixed reagents from the target receptacle into a flow channel; and a nozzle tube configured to be in fluid communication with the fluid system, the nozzle tube having an elongated body having a central cavity extending between its ends, and a nozzle liner located at a distal end of the central cavity, the nozzle tube being configured to both suction mixed reagents from the receiver, and to eject the mixed reagents back to the target receiver through the nozzle insert.

In some embodiments of the system, the nozzle and cavity may be sized to promote vortex formation to provide mixing at the target receptacle while removing reagents from the nozzle tube through the nozzle liner and into the target receptacle.

In some embodiments of the system, the cavity may have a nominal internal diameter of about 0.5 mm, and the nozzle liner may be a tubular liner having a nominal internal diameter of about 0.25 mm.

In some embodiments of the system, the distal end of the nozzle tube may be wedge-shaped with edges intersecting at an apex that is offset from the central axis of the nozzle tube.

In some embodiments of the system, the nozzle insert may include a distal end having a shape corresponding to the wedge-shaped shape of the distal end of the nozzle tube.

In some embodiments of the system, the wedge shape may include four edges intersecting at a specified vertex.

In some embodiments of the system, the nozzle tube may extend a nominal distance of 2 mm from the bottom surface of the target receiver.

In some embodiments of the system, the proposed system may include a plurality of other tubes for suctioning the respective reagents, wherein said other tubes do not have nozzle liners.

In some embodiments of the system, the tube nozzle may be configured to accelerate the mixed reagents to a flow rate of at least about 1600 mm/s at a flow rate of at least about 5000 μL/min.

In some embodiments, a system may be provided that includes a flow cell for retaining analytes in the analytical system; a fluid system for sucking in reagents, mixing the reagents, discharging the mixed reagents into a target receiver, and supplying the mixed reagents from the target receiver into a flow cell; a nozzle tube configured to be in fluid communication with a fluid system, the nozzle tube having an elongated body having a central cavity extending between its ends and a nozzle located at a distal end of the elongated body, the nozzle reducing the nominal internal diameter of the central cavity; and a control circuit operably connected to the fluid system, wherein the control circuit is configured to control the fluid system so that the fluid system performs the following actions: sucking a certain set of reagents in turn, ejecting reagents from the reagent set into a target receiver through a nozzle, sucking the set reagents from the target receiver through the mixing nozzle, and expelling the set of mixed reagents back to the target receiver through the nozzle.

In some embodiments of the system, the nozzle may have a liner inserted into a central cavity at the distal end of the nozzle tube.

In some embodiments of the system, the target receiver may contain the analyte to be sequenced.

In some embodiments of the system, the central cavity may have a nominal internal diameter of 0.5 mm and the nozzle may have a nominal internal diameter of 0.25 mm.

In some embodiments of the system, the distal end of the nozzle tube may be wedge-shaped with edges intersecting at an apex that is offset from the central axis of the nozzle tube.

In some embodiments of the system, the nozzle may have a distal end that is shaped to match the wedge-shaped shape of the distal end of the nozzle tube.

In some embodiments, a method may be provided including the steps of: a) operating a pump to draw, in turn, a plurality of reagents from a corresponding plurality of reagent receivers; b) driving a pump to eject the reagents into a target receptacle through a nozzle tube in fluid communication with the pump, the nozzle tube having an elongated body having a central cavity extending between its ends and a nozzle located at a distal end of the elongated body, wherein the nozzle reduces the nominal internal diameter of the central cavity; c) actuating a pump to draw reagents from the target receiver and through the nozzle tube to further mix the reagents; and d) operating a pump to expel the reagents from the nozzle tube back to the target receiver.

In some embodiments of the method, the nozzle and cavity may be sized to promote vortex formation to provide mixing at the target receptacle as reagents are removed from the nozzle tube through the nozzle into the target receptacle.

In some embodiments of the method, the central cavity may have a nominal internal diameter of 0.5 mm, and the nozzle may include a liner inserted into the central cavity and having a nominal internal diameter of 0.25 mm.

In some embodiments of the method, the distal end of the nozzle tube may be wedge-shaped with edges intersecting at an apex that is offset relative to the central axis of the nozzle tube.

In some embodiments of the method, the reagents may include at least three reagents with different relative densities.

In some embodiments of the method, the proposed method may further include performing one or more repetitions of steps (b) and (c) before performing step (d).

Various features of one or more embodiments of the subject matter disclosed in the present invention are illustrated in the accompanying drawings and set forth in the description that follows. Other features, aspects and advantages of the present invention will become apparent upon examination of the specification, drawings and claims. It should be noted that relative dimensions in the following drawings may not be to scale.

Brief description of drawings

These and other features, aspects and advantages of the present invention will become more apparent upon reading the following detailed description in conjunction with the accompanying drawings, in which like reference numerals refer to like parts throughout the drawings, the drawings showing the following.

In fig. 1 is a general diagram of an exemplary sequencing system in which the technologies disclosed herein can be applied.

In fig. 2 is a general diagram of an exemplary fluid system provided in the sequencing system of FIG. 1.

In fig. 3 is a general diagram of an exemplary processing and control system provided in the sequencing system of FIG. 1.

In fig. Figure 4 shows an axonometric view of an example of a reagent manifold with selector valves.

In fig. 5 is a top view showing an example of the manifold and valve assembly of FIG. 4.

In fig. 6A is a schematic diagram of an exemplary reagent and sample matrix suction and mixing assembly; and in fig. Figure 6B shows how the reagents and sample matrix will be layered on top of each other before mixing.

In fig. 7 schematically illustrates an example of how the reagents to be mixed are individually drawn into the mixing volume.

In fig. 8 is a schematic cross-sectional view of an example target receiving vessel for mixed reagents and a sample matrix, also showing a nozzle tube ejecting the mixed reagents into the receiver.

In fig. 9A - 9D show an exemplary nozzle tube that may be used when mixing reagents.

In fig. Figure 10 shows a graph of example cycles during aspiration and mixing of reagents and sample matrix.

In fig. 11 is a block diagram illustrating an example logic circuit for aspirating and mixing reagents and a sample matrix.

Carrying out the invention

In fig. 1 shows an embodiment of a sequencing system 10 for processing molecular samples that can be sequenced to determine their components, the order of the components, and the overall structure of the sample. The system includes an instrument 12 that receives and processes a biological sample. Sample source 14 provides sample 16, which in many cases will contain a tissue sample. The sample source may be, for example, an individual or entity such as a human, animal, microorganism, plant, or other donor (including environmental samples), or any other entity that contains organic molecules of interest whose sequence needs to be determined. The system can be used with samples other than those taken from organisms, including synthesized molecules. In many cases, the molecules will contain DNA, RNA or other molecules having base pairs, the sequence of which may specify genes and gene variants having specific functions of greatest interest.

Sample 16 is introduced into sample preparation/library system 18. This system can isolate, digest, and otherwise prepare a sample for analysis. The resulting library contains molecules of interest with lengths that contribute to the sequencing operation. Next, the resulting library is fed to instrument 12, where the sequencing operation is performed. In practice, the library, sometimes referred to as a template, is coupled with reagents in an automated or semi-automated process and then introduced into the flow cell prior to sequencing.

In the embodiment illustrated in FIG. 1, the instrument contains a flow cell or array 20 into which a library of samples is placed. The flow cell has one or more fluid channels to enable chemical sequencing, including attachment of library molecules and amplification at sites or regions that can be detected during the sequencing operation. For example, flow cell/array 20 may contain sequencing arrays immobilized on one or more surfaces at specified locations or areas. The "flow cell" may contain a structured array, such as a micro-lattice, nano-lattice, etc. In practice, the spaces or areas may be arranged in a regular, repeating pattern, a complex non-repeating pattern, or a random pattern on one or more surfaces of the substrate. To enable chemical sequencing, various substances can also be introduced into the flow cell, including various reagents, buffers and other reaction media used for reactions, washing, etc. Substances flow through the flow cell and may contact molecules of interest at specific sites.

In the instrument, flow cell 20 is mounted on a movable table 22, which, in this embodiment, is configured to move in one or more directions, indicated by numeral 24. Flow cell 20 may, for example, be provided as a removable and replaceable cartridge that may interact with openings on the movable table 22 or other components of the system to allow reagents and other fluids to be introduced into or out of the flow cell 20. The table is coupled to an optical detection system 26 that can direct radiation or light 28 into the flow cell during sequencing . The optical detection system can use various techniques, such as fluorescence microscopy techniques, to detect analytes present in areas of the flow cell. As a non-limiting example, optical detection system 26 may employ confocal line scanning to generate progressive pixel image data that can be analyzed to locate individual sites in the flow cell and determine the type of nucleotide that has recently attached or bound to each site. It is also possible to use other suitable imaging technologies, for example, technologies that scan one or more emission points along the sample, or technologies that employ a step-and-shoot imaging technique. The detection optical system 26 and the stage 22 may cooperate with each other to hold the flow cell and detection system in a static relationship while obtaining an image of the area, or, as noted, the flow cell may be scanned in any suitable mode (for example, during a spot scan , line scanning, step-and-shoot scanning).

Although various technologies can be used to obtain images, or more generally to detect molecules at sites, the present invention may employ confocal optical imaging technology at wavelengths that excite fluorescent tags. Tags excited according to their absorption spectrum return fluorescent signals according to their emission spectrum. The optical detection system 26 is configured to intercept such signals, process the pixel image data at a resolution that allows signal-emitting areas to be analyzed, and process and store the resulting image data (or data derived from it).

During a sequencing operation, cycling operations or processes are performed automatically or semi-automatically where reactions are accelerated, for example with single nucleotides or oligonucleotides, followed by washing, imaging and unblocking in preparation for the next cycle. A sample library prepared for sequencing and immobilized in a flow cell may undergo a number of such cycles before all useful information is extracted from the library. The optical detection system can generate image data from scans of the flow cell (and portions thereof) during each cycle of the sequencing operation using detection electronics (eg, imaging cameras or imaging circuits or chips). Further, the resulting image data can be analyzed to determine the locations of individual regions in the image data, and to analyze and obtain characteristics of the molecules present at these regions, for example, taking into account the specific color or wavelength of light (the characteristic emission spectrum of a particular fluorescent label) detected for a specific location, as indicated by a group or cluster of pixels in the image data at a specified location. In the case of DNA or RNA sequencing, for example, four common nucleotides can be represented as a distinguishable spectrum of fluorescent emission (wavelengths or range of wavelengths of light). Each emission spectrum can then be assigned a value corresponding to that nucleotide. Based on this analysis and by tracking the cyclic values determined for each region, the individual nucleotides and their order can be determined for each region. These sequences can then be further processed to collect longer segments, including genes, chromosomes, etc. As used herein, the terms “automatic” and “semi-automatic” mean that operations are performed through system programming or system configuration with little or no human interaction once the operations have been initiated, or after processes involving such operations have been initiated.

In the illustrated embodiment, reagents 30 are drawn or drawn into the flow cell through a valve system 32. The valve system may access reagents from receivers or vessels in which they are stored, such as through pipettes or sippers (not shown). Fig. 1). The valve system 32 may allow the selection of reagents based on a given sequence of operations. The valve system may further receive commands to direct reagents through flow channels 34 to flow cell 20. Outlet or effluent flow channels 36 direct used reagents from the flow cell. In the illustrated embodiment, pump 38 is configured to move reagents through the system. The pump may also perform other useful functions, such as measuring reagents or other fluids through the system, intake air or other fluids, etc. An additional valve system 40 downstream of the pump 38 allows the used reagent to be properly directed to the waste collection vessels or receptacles 42.

The instrument further comprises various electronic circuits that assist in controlling the operation of various system components, monitoring their performance through feedback from sensors, collecting image data, and at least partially processing the image data. In the embodiment illustrated in FIG. 1, control/monitoring system 44 includes a control system 46 and a data acquisition and analysis system 48. Both systems include one or more processors (e.g., digital processing circuits such as microprocessors, multi-core processors, field programmable gate arrays, or any other suitable processing circuits) and associative memory circuits 50 (e.g., semiconductor memories, dynamic memories, internal and/or external storage devices, etc.) that can store machine-executable instructions for controlling, for example, one or more computers, processors, or other similar logical devices to provide specific functionality. Specialized or general-purpose computers may at least partially constitute a control system and a data acquisition and analysis system. The control system may comprise, for example, circuitry designed to (eg, programmed to) process commands for inkjet automation, optical instruments, table control, and any other useful functions of the tool. The data acquisition and analysis system 48 interfaces with the optical detection system to control the movement of the optical detection system or the specified table, or both, the emission of light for cyclic detection, the reception and processing of reflected signals, etc. The instrument may also contain various interfaces, indicated by reference number 52, such as an operator interface that allows control and monitoring of the instrument, transfer of samples, initiation of automated or semi-automatic sequencing operations, generation of reports, etc. Finally, in the embodiment shown in FIG. 1, external networks or systems 54 may be connected to or interact with the instrument, such as for analysis, control, monitoring, maintenance, or other operations.

It should be noted that although in FIG. 1 shows a single flow cell and fluid channels and a single optical detection system; some instruments may have multiple flow cells and fluid channels. For example, in one embodiment of the present invention, two such arrangements are provided to improve the sequencing process and increase productivity. In practice, any number of flow cells and flow channels may be provided. They may use the same or different reagent receivers, waste collection receivers, control systems, image analysis systems, etc. Where provided, a plurality of fluid systems may be controlled individually or may be controlled in concert. It should be understood that the expression "fluidly coupled" may be used herein to describe connections between two or more components that provide fluid communication between such components, in principle, just as the expression "electrically coupled" may be used to descriptions of the electrical connection between two or more components. The expression "fluidically interposed" may be used, for example, to describe a particular order of components. For example, if component B is located in a fluid between components A and C, then fluid flowing from component A to component C will flow through component B before reaching component C.

In fig. 2 shows an example of a fluid system provided in the sequencing system of FIG. 1. In the embodiment illustrated in this drawing, flow cell 20 includes a group of paths or lanes 56A and 56B, which can be grouped in pairs to receive liquid substances (eg, reagents, buffers, reaction media) during sequencing operations. Tracks 56A are connected to a common line 58 (the first common line), and tracks 56B are connected to a second common line 60. A bypass line 62 is also provided to allow fluids to bypass the flow cell without entering it. As mentioned above, an array of vessels or receptacles 64 provide the ability to store reagents and other fluids that may be used during a sequencing operation. Reagent selector valve 66 is mechanically coupled to a motor or actuator (not shown) to allow selection of one or more reagents for introduction into the flow cell. Next, the selected reagents are sent to the common line selector valve 68, which similarly contains a motor (not shown). The common line selector valve may be commanded to select one or more common lines 58 and 60, or both lines, to allow reagents 64 to flow into lanes 56A and/or 56B in a controlled manner, or bypass line 62 to allow one or more reagents to flow. through the specified bypass line. It should be noted that the bypass line can also be used to perform other useful operations, such as allowing all reagents (and liquids) to be charged into the reagent selector valve (or common line selector valve) without passing air through the flow cell, allowing flushing (e.g., automatic or semi-automatic flushing) of reagent channels and tubing independent of the flow cell, and allowing diagnostic functions (pressure and flow tests) to be performed on the system.

The used reagents exit the flow cell through lines connected between the flow cell and the pump 38. In the illustrated embodiment, the pump is a syringe pump having a pair of syringes 70 controlled and driven by an actuator 72 to draw in reagents and other fluids and to release of reagents and fluids during various testing, verification and sequencing cycles. The pump assembly may contain various other parts and components, including a valve system, instrumentation, actuators, etc. (not shown). In the illustrated embodiment, pressure sensors 74A and 74B measure pressures on the pump inlet lines, and pressure sensor 74C is configured to measure pressures at the outlet of the syringe pump.

The fluids used in the system flow from the pump to the selector valve 76 for used reagents. This valve provides the ability to select one of a variety of flow channels for used reagents and other fluids. In the illustrated embodiment, the first flow path leads to a first waste reagent receptacle 78, and the second flow path leads through the flow meter 80 to a second waste reagent receptacle 82. Depending on the reagents used, it is advisable to provide for the collection of reagents, or specific reagents, into separate waste collection vessels, with the waste reagent selector valve 76 allowing for such control.

It should be noted that the valve system within the pump assembly may be capable of drawing, through the pump, various fluids, including reagents, solvents, cleaners, air, etc., and injecting or circulating them through one or more common lines, bypass line and flow cell. Additionally, as noted above, in one embodiment of the present invention there are two parallel embodiments of the fluid system shown in FIG. 2, which are under common management. Each of these fluid systems can be part of a single sequencing instrument, and can perform functions in parallel, including sequencing operations, in different flow cells and with different sample libraries.

The fluid systems may be operated under the control of a control system 46 that implements prescribed protocols for testing, verification, sequencing, etc. The prescribed protocols will be created in advance and will contain a series of events or operations to carry out actions such as sucking in reagents, sucking in air, sucking in other fluids, releasing such reagents, air and fluids, etc. The protocols will provide the ability to coordinate such fluid operations with other instrument operations, such as the development of reactions in the flow cell, imaging of the flow cell and its sections, etc. In the illustrated embodiment, the control system 46 uses one or more valve interfaces 84 that are configured to issue command signals to the valves, as well as a pump interface 86 that is configured to control the operation of the pump actuator. Various input/output circuits 88 may also be provided for receiving feedback signals and processing such feedback signals, for example, from pressure sensors 74A - 74C and flow meter 80.

In fig. 3 shows specific functional components of the control/monitoring system 44. As shown in the drawing, memory circuit 50 stores prescribed paths that are executed during testing, commissioning, troubleshooting, maintenance, and sequencing operations. Many such protocols and routes can be implemented and stored in the memory circuit, and they can be updated or changed from time to time. As shown in FIG. 3, they may include a fluid control protocol 90 for controlling various valves, pumps, or any other fluid actuators, and for receiving and processing feedback signals from fluid sensors, such as valves, and pressure sensors and flow meters. Table control protocol 92 allows the flow cell to move at will, for example during image acquisition. Optical system control protocol 94 provides the ability to issue commands to imaging components to illuminate portions of the flow cell and receive reflected signals for processing. Imaging protocol 96 provides the ability to at least partially process image data to extract useful sequencing information. Other protocols and routes may be provided in the same or different memory circuits, as indicated by numeral 98. In practice, the memory circuit may be implemented as one or more storage devices, such as volatile and non-volatile storage devices. This storage device may be provided within the instrument, while some storage devices may be external.

One or more processors 100 access the stored protocols and execute them in the instrument. As noted above, the data processing circuitry may be part of dedicated computers, general purpose computers, or any suitable hardware, firmware, or software platform. The processors and operation of the tool can be controlled by a human operator through the operator interface 101 . The operator interface can provide the ability to perform testing, commissioning, troubleshooting and maintenance, as well as reporting any problems that may occur with the tool. The operator interface may also provide the ability to run and monitor sequencing operations.

In fig. 4 shows a valve assembly designed to remove reagents and fluids from the receivers and supply them to the flow cell. Valve assembly 102 includes a manifold structure 104 in which grooves are formed to define flow paths for reagents and other fluids. As can be seen in FIG. 4, valves 66 and 68 are actuated and controlled by motors 106 and 108. One or more motor interfaces or connections 110 supply power and optionally provide signals to or from the motors. As noted above, the motors (and thus the valves) are controlled by the control circuitry during testing, commissioning and maintenance, and during sequencing operations.

The reagent and fluid paths within the manifold are connected to tubes 112 which, during operation, remove reagents and other fluids from respective receptacles (not shown). Inlet channels for reagents and fluids, generally designated 114 in FIG. 4 may be produced by injection molding, etching, or any other suitable process to allow the movement of reactants and fluids from the tubes into the valves when the pump described above is commanded to suck in the reactants and fluids. At least one of the tubes is configured as a nozzle tube 116 that facilitates mixing of reagents during sequencing operations (eg, prior to reactions and imaging). In addition, in FIG. 4 also shows a mixing volume configured as a channel 118 into which reagents and fluids can be introduced and transferred for mixing. In some embodiments, the mixing volume may be part or all of the bypass line 62. For example, reagents may be drawn into the bypass line 62 in a desired sequence, but so that the reagents do not traverse the entire length of the bypass line (which may cause they will be sent to the waste collection vessel). Once the bypass line (or portion thereof serving as a mixing volume) has been loaded with the desired sequence of reagents, the end of the bypass line through which the reagents were introduced can be switched, using a valve, to make a fluid connection to the flow channel leading , for example, into the target receptacle, so that the entire set of reagents loaded into the bypass line can be further forced back out of the bypass line into the target receptacle. In other embodiments, the mixing volume may, for example, be a target receptacle, such as a target receptacle from which premixed fluids are supplied, or a separate target receptacle, such as a receptacle that is completely empty before being supplied with selected reagents.

In fig. 5 is a top view of valve assembly 102. Here again, valves 66 and 68 are visible in the manifold and connected to the reagent and fluid flow passages. The reagent selector valve 66 receives reagents from the tubes and directs the suction fluids to the common line selector valve 68 . Mixing port 118 is connected to a common reagent mixing line selector valve, as disclosed below. Also, in FIG. 5 shows openings 120 provided in the manifold to allow connection of the manifold to the tubes. One of the openings 120 (designated 122) will be connected to the nozzle tube to allow reagents to be injected into the target receptacle and to remove reagents from the target receptacle for mixing. The target receiver, for example, may be a container, tube, or other vessel designed to contain reagents. The target receiver may, for example, be used as a temporary working volume into which reagents and/or other materials can be transferred to prepare them for delivery, for example by mixing, to the flow cell. Thus, reagents and other fluids, after being prepared in the target receiver, can be transferred from the target receiver to the flow cells.

One embodiment of mixing channel 118 and reagent flow channels according to the present invention is shown in FIG. 6A. As noted above, the mixing channel 118 is connected to a common line selector valve 68, which in turn is connected to the output of the reagent selector valve 66. The mixing channel 118 is also connected to the pump 38 to allow the suction and ejection of reagents and fluids, as discussed below. In the embodiment illustrated in FIG. 6A, reagent receptacles or vessels 124, 126, and 128 contain reagents 130, 132, and 134, respectively. The additional or target receiver 136 contains, in this example, a pre-prepared matrix or library 138 of samples. For the mixing operation, reagents 130, 132 and 134 are premixed and further associated with the matrix 138. To provide such premixing, the reagents are drawn alternately into the mixing channel 118 through the respective flow channels indicated in FIG. 6A by reference number 140. An additional flow channel 142 provides the ability to introduce reagents into the target receiver 136 along with the matrix. In the illustrated embodiment, the mixing channel 118 defines a serpentine internal volume having loops 144 that allow the required amounts of reagents to be drawn and mixed in a relatively compact manifold area.

In one embodiment of the present invention, the reagents 130, 132 and 134 have different fluid properties that create a mixing problem. For example, the densities of the reagents differ, and there may be significant differences between the viscosities and surface tensions at the interface with the reagent oil. In one embodiment of the present invention, for example, viscosities vary from about 1.5 cP to 50 cP, for example, equal to 2.4 cP, at a temperature of 25 ° C, and surface tensions at the interface with the oil vary from about 5. 0 to approximately 19.2 dynes/cm. The matrix, by comparison, may have a different density and lower viscosity (eg, about 1 cP at 25°C) and a different surface tension at the interface with the oil (eg, about 9.8 dynes/cm). In fig. 6B illustrates the layering of reagents and matrix in target receiver 136 when they are not mixed. In the illustrated embodiment, the matrix represents approximately 30% of the total volume, reagent 130 represents approximately 22%, reagent 132 represents approximately 42%, and reagent 134 represents approximately 6%. In this context, the term “about” means that the values given are not exact and that the actual value may differ from these given values to an extent that does not materially change the operation in question.

To automatically mix reagents and matrix, the fluid system and its control enable the reagents to be selectively sucked one at a time into the mixing channel, injected into a target receptacle, and cyclically withdrawn and reinjected for mixing. In fig. 7 shows the technology proposed in the present invention for the absorption of reagents. As shown in the drawing, reagents 130, 132 and 134 are sucked in alternately by the control of valve 66. By means of a common line selector valve directing the reagents into the mixing channel, several sets of volumes of each reagent are drawn in, designated by reference numbers 146, 148, 150, 152 and 154. To enable reduction of pressure surges during mixing, the pump may also draw in some air before drawing in the reagents. This volume of air provides a cushion that limits positive and negative pressure surges during mixing. In the embodiment illustrated in FIG. 7, the intake air will be located at the top left of the reagent kits. In addition, it is possible to absorb a liquid buffer, which will facilitate filling, washing and unloading of reagents. After suction, as shown in FIG. 7, the valve system can be controlled to allow the pump to inject reagents into the matrix 138, which will be preloaded into the matrix receiver 136 disclosed above.

In another technology in which three or more reagents may be selected for mixing in a target receptacle, at least two reagents selected for mixing may be repeatedly introduced alternately into the mixing channel, with at least one other reagent selected for mixing remain in reserve until the reagents, repeatedly introduced alternately into the mixing channel, are completely supplied to the mixing channel. Next, the reagent in reserve can be added all at once to the mixing channel. For example, if reagents A and B are to be repeatedly introduced alternately into a mixing channel, followed by the introduction of a reserve reagent C, then the reagents in the mixing channel will generally be mixed in the form ABAWAWAWAWS, as opposed to the sequence ABCAWSAWSAWSABC (which will be obtained , for example, using technology similar to that discussed with reference to Fig. 7). This technology is believed to be advantageous because it prevents or reduces the likelihood of some reagents from forming undesirable reaction by-products. For example, a reserve reagent may react in a particular way with one of the other reagents individually, but may react differently with two or more of the remaining reagents in combination. The latter reaction may be desirable and may occur after the reagents have been thoroughly mixed, while the former reaction may occur during premixing when the reagents may still be relatively layered and only able to mix with the immediately adjacent adjacent reagent. In another example, the stored reagent may react with the material that forms the mixing channel structure to form an undesirable by-product. Since repeated, alternating introduction of reagents into the mixing channel may require several minutes, such as 5 minutes, 10 minutes, 15 minutes or more, depending on the number and quantity of each reagent required, delaying the introduction of potentially problematic reagents until until the remaining reagents are introduced one at a time into the mixing channel can significantly reduce the amount of time that the reserve reagent is in contact with other reagents and with the mixing channel structure, which in turn reduces the likelihood of unwanted reaction by-products forming. Obviously, in such embodiments, the reserve reagent may not benefit from premixing as does other reagents, but reducing the likelihood of unwanted reaction byproducts may be more important than not premixing the reserve reagent. In particular, if the reserve reagent is a liquid with a lower viscosity, then the lack of premixing with respect to the reserve reagent may, in fact, have little effect.

Using a mixing volume in the form of a channel, for example, a volume that is much longer than it is wide (for example, its length is at least 10, 100, 150 to 170, 160, 200, or 500 times its width ), can provide the ability to maintain the relatively layered structure of periodically supplied reagents relative to each other inside the channel by reducing the area of intersurface contact interface between each layer of reagents (reagents are liquids and, in this regard, it is likely that over time they will be in some degree to diffuse into each other across that boundary, so the interface/contact area concepts mentioned herein should be considered theoretical in nature; however, reducing such theoretical areas will result in a slower rate of diffusion). Moreover, for reagents that are somewhat incapable of mixing with each other, a mixing volume that is, for example, spherical in shape or that has a larger width/length ratio may cause different doses of reagents to be supplied to the mixing volume. mixing volumes will be able to float freely within the mixing volume and potentially recombine with previously administered doses of the same reagent, causing them to lose the layering that can be achieved in the channel-like mixing volume. For example, a mixing channel with a diameter of approximately 2.25 mm or a width of approximately 360 mm of its length can provide preferential layering in the feed reagents during the premixing process. After the required quantities of multiple sets of reagents are loaded into the mixing volume, the contents of the mixing volume can be fed to the target receiver (some of the fluids in the mixing volume may be lost in the dead space of the fluid system; the total volume of reagents supplied to the mixing volume , can be calibrated to account for such loss). Once supplied to the target receptacle, the supplied premixed reagents may be repeatedly sucked from and released back into the target receptacle to facilitate further mixing. In some embodiments, premixed (or subsequently premixed) reagents may be drawn from the target receptacle and pushed back into the mixing volume before being released back into the target receptacle. As a result, in such embodiments, the premixed reagents are allowed to move in and out of the mixing volume multiple times during the suction/ejection mixing operation.

The use of a mixing channel with a nozzle tube was found to promote vortex formation in the target receiver and provide excellent mixing of the reagents and sample matrix, despite significant differences in the properties of the reagent fluids. In addition, such structures and technologies enable automatic mixing with little or no human interaction. An example of a nozzle tube for use in these technologies is shown in FIG. 8 and 9A – 9C. As shown in FIG. 8, the nozzle tube has an elongated body with a central cavity (lumen) extending along its length and a tip 156 at its distal end. A nozzle is provided at said tip to reduce the internal diameter of the tube at that location and thereby increase the speed of fluids drawn in and expelled through the tube. In the illustrated embodiment, the nozzle is configured as a liner 158 secured to the distal end or tip of the tube. Other structures, such as machined caps, molded elements, upset areas, etc., may also form said nozzle.

In the illustrated embodiment, the tube has a nominal outside diameter 160 of about 0.125 inches (3.175 mm) and a nominal inside diameter 162 of 0.02 inches ± 0.001 inches (0.508 mm). The nozzle, on the other hand, has a nominal internal diameter 164 of 0.01 inch ± 0.001 inch (0.254 mm, although in some embodiments the nozzle internal diameter may range from 0.2 to 0.28 mm). Obviously, other sizes and dimensions can be used to provide the required mixing. Additionally, in the illustrated embodiment, the nozzle tube 116 is located at a height 166 above the bottom of the receiver 138 of approximately 2 mm. As reactants are injected into the receiver, as indicated by numeral 168, swirl within the receiver is increased by increasing the velocity of the reactants through the nozzle, which promotes the mixing process in the receiver, as indicated by arrow 170 in FIG. 8. Provision is then made for the mixed reagents to rise in the receiver as indicated by item number 172.

In fig. 9A shows the distal end of the nozzle tube in more detail. As can be seen in the drawing, the nominal inner diameter 162 of the tube is reduced by the nozzle liner 158, in this case to approximately half the inner diameter of the tube (the nozzle liner, in this example, is tubular in shape). The distal end shape of the present invention is best illustrated in FIG. 9B, 9C and 9D. As shown in these drawings, the nozzle tube has a multifaceted bottom edge containing four faces 174, which gives the tip of the nozzle tube a wedge-shaped appearance. The tube has a centerline 176 and the faces intersect at a vertex 178 that is offset or eccentric with respect to the centerline 176. This distal end geometry reduces or eliminates dragging or scraping of the receiver when lowering the tube into the receiver, or when lifting the receiver around the tube. It may be noted, however, that in the illustrated embodiment, the liner has a bottom contour that matches the contour of the tip (eg, one or more angled edges). In other words, the liner has a shape consistent with the polyhedral or wedge-shaped shape of the distal end of the nozzle tube. In addition, it may be noted that in one embodiment of the present invention, the tube and nozzle are made of an engineering plastic, for example, polyetheretherketone (PEEK). Such materials can provide chemical resistance to reagents and any solvents used in the process.

In fig. 10 is a graphical representation of exemplary cycles for suction, mixing and ejection of reagents and sample matrix, and FIG. 11 is a block diagram illustrating an example logic circuit for aspirating and mixing reagents and a sample matrix. In fig. 10, the cycle of suction, mixing and ejection of reagents is indicated by numeral 180, with the pressure generated by the pump indicated by axis 182 and the cycle time indicated by axis 184. Negative pressures indicate suction of one or more reagents, and positive pressures indicate ejection. The process can be considered to include a "transfer" sequence 186 followed by a "mix" sequence 196, as will be discussed below.

Based on the block diagram of Fig. 11, control logic 204 may begin by sucking air at step 206 to remove existing liquid from flow channels through which previous reactant mixtures may have been directed. For example, any remaining liquid present in the flow path 142 connecting the reagent selector valve 66 to the target receiver 136 may be drawn in the presence of air (i.e., thereby replacing the liquid with air), so that any new reagent mixture that is subsequently introduced into the target receiver through flow channel 142, does not mix with the remaining liquid. Next, a "transfer" sequence is performed, which begins with a "priming" sequence, as indicated by numeral 208 in FIG. 11. This "priming" sequence is indicated by a series of negative pressures or suction events, collectively indicated by reference number 188 in FIG. 10. Overall, these events enable the initial introduction of reagents into the system. As demonstrated in more detail in FIG. 11, a buffer may be sucked in, as indicated by numeral 210. This buffer may contain a liquid selected to be non-reactive or relatively inert with respect to said reactants and may be used as an incompressible working fluid passing through at least at least partially between the pump and the reagents, to enable more accurate measurement of the reagents in the mixing volume in subsequent steps, if required. Further, it is possible for the first reagent to be drawn in during the priming event, which is indicated by numeral 212 in FIG. 11, followed by the aspiration of any number of other reagents, until the last reagent is aspirated at step 214. In one embodiment of the present invention, for example, three such reagents are aspirated during a "priming" sequence.

According to the logic diagram illustrated in FIG. 11, the reagents to be mixed are then drawn in during a "transfer" sequence 218. The "transfer" sequence continues with the aspiration of the first reagent, indicated by item number 220, followed by the aspiration, in turn, of each of the additional reagents until the last reagent is absorbed, indicated by the item number 222. As mentioned above, in one of embodiments of the present invention, three reagents are absorbed during this sequence. As noted above, in one embodiment of the present invention, a number of sets of reagents are absorbed in relatively small quantities to create a sequence of reagents, thereby facilitating premixing. Thus, at logic step 214, it can be determined whether all reagent kits have been absorbed, and if not, it returns to step 220 to continue suctioning additional kits. It may also be noted that in the embodiment under consideration, all kits contain all the reagents selected for mixing, however this is not always the case. In addition, reagents may be absorbed in different volumes or quantities in different kits. After all reagents have been absorbed, control can move on and move beyond the “transfer” sequence. The "transfer" sequence is illustrated in terms of negative pressure events, collectively designated numeral 190 in FIG. 10.

As shown in FIG. 11, and as will be clear from the individual negative (and positive) pressure events of FIG. 10, each successive intake (or ejection) of reagents or premixed reagents involves controlling one or more of the valves disclosed above as well as a pump. That is, to suction individual reagents, the reagent selector valve will be switched to direct negative pressure into the tube for the corresponding receiver of the selected reagent. The pump will be controlled in a similar fashion to draw in a reagent (or air or buffer or matrix) and send out the sucked fluids according to a prescribed protocol. This mixing protocol will be predefined and stored in the memory circuit disclosed above, and executed automatically or semi-automatically based on the sequencing operation also defined in the memory circuit. These protocols are executed by means of a processing and control circuit which, through suitable interface circuitry, controls the operation of the valves and pump.

After all reagents have been sucked in, the sucked fluids can be discharged to the target receptacle, as indicated by numeral 226 in FIG. 11. As noted above, in the embodiment under consideration, this occurs through the nozzle tube, where mixing begins due to an increase in the speed of movement of the reagents through the nozzle and the resulting vortex in the target receiver. Such a release into the target receiver is indicated by positive pressure event 192 in FIG. 10. In specific embodiments, suction may be additionally provided as indicated by numeral 228 in FIG. 11. After this, the ejection of the sucked reagents into the target receiver can be ensured. This sequence may be followed by air suction, indicated by numeral 230 in FIG. 11, and negative pressure event 194 in FIG. 10 (for example, to remove as much liquid as possible from the bypass line, mixing channel, matrix channel and tube). It may also be noted that in some embodiments, the nozzle tube or receiver, or both, may move relative to each other (eg, vertically) during suction and ejection to further facilitate mixing of layered samples and reagents.

After suction and partial premixing in the mixing volume or channel through the operations disclosed above, mixing is carried out by repeatedly moving the reagents in the channel, and between the channel and the target receiver through the nozzle tube. To do this, a number of mixing cycles are performed during the "mixing" sequence 234. In this sequence, the associated reagents and matrix are aspirated at step 236 and ejected back to the target receptacle at step 238. The logic may repeatedly determine whether all of the required mixing cycles have been completed at step 240 and continues until all such cycles have been completed. will be completed. In the graph shown in FIG. 10, these cycles are collectively designated numeral 198. As can be seen in the figure, each of them includes a relatively short negative pressure event followed by a relatively short positive pressure event. These events effectively cause the bound reagents and matrix to be drawn into the mixing volume or channel through the nozzle tube, and the gradually mixed reagents and matrix to be returned to the target receiver through the nozzle. Since this process can displace any required volume, in the present embodiment, approximately 2000 μL is aspirated from and ejected into the target receptacle in each mixing cycle, although in other embodiments it is possible to pump approximately 500 μL or 1500 μL, depending on the size of the flow chambers used. At the end of the mixing process, the mixed reagents and template can be returned to the target receiver to resume the sequencing operation.

It should be noted that in one embodiment of the present invention, the nozzle tube effectively increases the speed of movement of the reagents (and mixed reagents) during their mixing during suction and ejection. This increase in speed results in increased kinetic energy, which promotes mixing. For example, in one embodiment of the present invention, the nozzle accelerates the mixture to about at least 1600 mm/s at a flow rate of at least about 5000 μL/min.

It should be understood that the use, if available, of ordinal indicators, for example, (a), (b), (c), ..., etc. in this specification and claims does not convey any particular order or sequence unless such order or sequence is expressly indicated. For example, if there are three steps identified as (i), (ii) and (iii), then it should be understood that these steps can be performed in any order (or even simultaneously, unless contraindicated) unless otherwise noted. For example, if step (ii) involves working with an element created in step (i), then step (ii) can be viewed as occurring at some point after step (i). By analogy, if step (i) involves working with an element that is created in step (ii), then the opposite is true.

It should also be understood that the use of the preposition "for", for example, "a valve for switching between two flow channels", can be replaced by an expression such as "intended for", for example, "a valve intended for switching between two flow channels", etc. d.

It should be understood that terms such as “about”, “approximately”, “substantially”, “nominal”, etc., when applied to quantities or similar quantifiable properties, include values within ±10 % of given values unless otherwise stated.

It should be understood that in addition to the claimed objects listed in this description, additional embodiments also fall within the scope of protection of the present invention.

Implementation option No. 1: A system containing: a flow cell for retaining analytes in the analytical system; a fluid system for sucking in reagents, mixing the reagents, and discharging the mixed reagents into a target receptacle; and a nozzle tube configured to be in fluid communication with the fluid system, the nozzle tube having an elongated body having a central cavity extending between its ends and a nozzle liner located at a distal end through which the nozzle tube draws reagents from the receiver and releases the mixed reagents back to the target receiver.

Embodiment No. 2: The system of Embodiment No. 1, wherein the nozzle and cavity are sized to promote vortex formation to provide mixing in said cavity as reagents are drawn into the tube through the nozzle liner.

Embodiment No. 3: The system of Embodiment No. 1, wherein the cavity has a nominal inside diameter of about 0.5 mm and the nozzle liner has a nominal inside diameter of about 0.25 mm.

Embodiment No. 4: The system according to Embodiment No. 1, wherein the distal end of the nozzle tube is wedge-shaped with the apex offset from the central axis of the nozzle tube.

Embodiment No. 5: The system according to Embodiment No. 4, wherein the nozzle insert includes a distal end having a shape corresponding to the wedge-shaped shape of the distal end of the nozzle tube.

Embodiment No. 6: The system according to Embodiment No. 4, wherein the wedge shape has four edges intersecting at a specified vertex.

Embodiment No. 7: The system according to Embodiment No. 1, wherein the nozzle tube has a length extending a nominal distance of 2 mm from the bottom surface of the receiver.

Embodiment No. 8: The system of Embodiment No. 1 having a plurality of other tubes for suctioning respective reagents, said other tubes not having nozzle liners.

Embodiment No. 9: The system of Embodiment No. 1, wherein the tube nozzle accelerates the mixed reagents to about at least 1600 mm/s at a flow rate of at least about 5000 μL/min.

Implementation option No. 10: A system containing: a flow cell for retaining analytes in the analytical system; a plurality of reagents located in appropriate receivers; a fluid system for sucking in reagents, mixing the reagents, and discharging the mixed reagents into a target receptacle; a nozzle tube configured to be in fluid communication with the fluid system; wherein the nozzle tube has an elongated body having a central cavity extending between its ends and a nozzle at the distal end; and a control circuit operatively coupled to the fluid system for controlling the fluid system to suck the plurality of reagents in turn, discharge the plurality of reagents into the target receiver through the nozzle, suck the plurality of reagents from the target receiver through the mixing nozzle, and discharge the mixed reagents back to the target receiver through the nozzle.

Embodiment No. 11: The system of Embodiment No. 10, wherein the nozzle has a liner at the distal end of the nozzle tube.

Embodiment No. 12: The system of Embodiment No. 10, wherein the target receiver contains the analyte to be sequenced.

Embodiment No. 13: The system of Embodiment No. 10, wherein the cavity has a nominal internal diameter of 0.5 mm and the nozzle has a nominal internal diameter of 0.25 mm.

Embodiment No. 14: The system according to Embodiment No. 10, wherein the distal end of the nozzle tube is wedge-shaped with the apex offset from the central axis of the nozzle tube.

Embodiment No. 15: The system of Embodiment No. 14, wherein the nozzle has a distal end whose shape corresponds to the wedge-shaped shape of the distal end of the nozzle tube.

Embodiment No. 16: A method comprising the steps of: driving a pump to draw a plurality of reagents from a target receptacle containing an analyte to be analyzed; sucking a plurality of reagents through a nozzle tube in fluid communication with a pump to mix the plurality of reagents, the nozzle tube having an elongated body having a central cavity extending between its ends and a nozzle at a distal end, and driving the pump to eject the mixed reagents to the target receiver.

Embodiment No. 17: The method of Embodiment No. 16, wherein the nozzle and cavity are sized to promote vortex formation to provide mixing in the cavity as reagents are drawn into the tube through the nozzle.

Embodiment No. 18: The method of Embodiment No. 16, wherein the cavity has a nominal internal diameter of 0.5 mm and the nozzle includes a liner having a nominal internal diameter of 0.25 mm.

Embodiment No. 19: The method according to Embodiment No. 16, wherein the distal end of the nozzle tube is wedge-shaped with an apex that is offset from the central axis of the nozzle tube.

Embodiment No. 20: The method according to Embodiment No. 16, wherein the reagents include at least three reagents with different relative densities.

It should be understood that all combinations of the above concepts (provided that such concepts are not mutually incompatible) are considered to be part of the subject matter of the present invention as disclosed herein. In particular, all combinations of claimed subject matter appearing at the end of this specification are considered to be part of the claimed subject matter of the present invention as disclosed in this specification. It should also be understood that terms used explicitly in this specification, which may also appear in any specification incorporated herein by reference, are intended to have a meaning consistent with the specific concepts disclosed herein.

Claims (34)

1. A system for mixing reagents, containing: a flow cell flow channel configured to be fluidly coupled to the flow cell for retaining analytes in the analytical system; a fluid system comprising a plurality of reagent flow channels, an additional flow channel, a mixing channel and a selector valve, wherein each of the plurality of reagent flow channels has a first end fluidly coupled to its reagent receiver of the plurality of reagent receivers, and a second end fluidly coupled to the selector valve, the additional flow channel has a first end configured to be fluidly connected to the target receiver, and a second end fluidly connected to the selector valve, the mixing channel has a first end in fluid communication with the selector valve, and the fluid system is configured to suck reagents from the plurality of reagent receivers into the mixing channel through a selector valve, eject the sucked reagents from the mixing channel into a target receiver through the selector valve and the additional flow channel, and supply reagents from the target receiver to the flow channel of the flow cell; And a nozzle tube fluidly connected to an additional flow channel, the nozzle tube having an elongated body having a central cavity extending between a proximal end of the elongated body and a distal end of the elongated body, and a nozzle liner located at the distal end of the elongated body, wherein the fluid system is configured to both suck mixed reagents from the target receptacle and eject the mixed reagents back into the target receptacle through the nozzle liner. 2. The system according to claim 1, characterized in that the nozzle liner and the elongated body are sized to promote vortex formation, providing mixing in the target receiver, when removing reagents from the nozzle tube through the nozzle liner into the target receiver. 3. The system of claim 1, wherein the central cavity has a nominal internal diameter of about 0.5 mm and tapers to about 0.25 mm at the nozzle liner. 4. The system according to claim 1, characterized in that the distal end of the nozzle tube has a wedge-shaped shape with edges intersecting at the apex, which is offset relative to the central axis of the nozzle tube. 5. The system according to claim 4, characterized in that the nozzle liner includes a distal end having a shape corresponding to the wedge-shaped shape of the distal end of the nozzle tube. 6. The system according to claim 4, characterized in that the wedge-shaped shape provides for the presence of four faces intersecting at the apex. 7. The system of claim 1, wherein the nozzle tube is configured to extend a nominal distance of 2 mm from the bottom surface of the target receiver when the nozzle tube is in fluid communication with the target receiver. 8. The system according to claim 1, characterized in that it contains a plurality of other tubes configured to suck appropriate reagents from a plurality of reagent receivers, said other tubes not having nozzle liners. 9. The system of claim 1, wherein the nozzle tube is sized to accelerate fluid flowing through the nozzle tube at a flow rate of at least about 5000 μL/min to a flow rate of at least about 1600 mm/s . 10. System for mixing reagents, containing: a flow cell to retain analytes in the analytical system; a fluid system comprising a plurality of reagent flow channels, an additional flow channel, a mixing channel and a selector valve, wherein each of the plurality of reagent flow channels has a first end fluidly coupled to its reagent receiver of the plurality of reagent receivers, and a second end fluidly coupled to the selector valve, the additional flow path has a first end configured to be fluidly coupled to the target receiver and a second end fluidly coupled to the selector valve, and the mixing channel has a first end in fluid communication with the selector valve; a nozzle tube fluidly connected to an additional flow channel, wherein the nozzle tube has an elongated body having a proximal end and a distal end, and a nozzle located at the distal end of the elongated body, wherein the nozzle tube has a central cavity extending from the proximal end of the elongated body to nozzles, wherein the nozzle reduces the nominal internal diameter of the central cavity; And a control circuit operatively coupled to the fluid system, the control circuit comprising one or more processors and one or more memories storing machine-executable instructions that, when executed by the one or more processors, cause the fluid system to execute: sucking a set of reagents alternately from a plurality of reagent receivers into the mixing channel by means of a selector valve, ejecting reagents in a specified set of reagents from the mixing channel into the target receiver by means of a selector valve and an additional flow channel through the nozzle, sucking a set of reagents from the target receiver through the mixing nozzle, and ejecting a set of mixed reagents back into the target receiver through the nozzle. 11. The system according to claim 10, characterized in that the nozzle includes a liner inserted into the distal end of the elongated body. 12. The system of claim 10, further comprising a target receiver, wherein the target receiver contains the analyte to be sequenced. 13. The system of claim 10, wherein the central cavity has a nominal internal diameter of 0.5 mm and tapers to about 0.25 mm at the nozzle. 14. The system according to claim 10, characterized in that the distal end of the nozzle tube has a wedge-shaped shape with edges intersecting at the apex, which is offset relative to the central axis of the nozzle tube. 15. The system according to claim 14, characterized in that the nozzle has a distal end, the shape of which corresponds to the wedge-shaped shape of the distal end of the nozzle tube.

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