Acoustic concentration of cells or particles in blood products

Abstract

A system for trapping cells or particles in a blood sample may include a device having a channel with a predefined geometry, the channel having an input and a single output, the channel configured to receive a sample including particles and a residual portion at the input; and configured to flow the sample through the channel to the single output, and a transducer creating acoustic excitation of the device at an excitation frequency relative to the predefined geometry, wherein, upon the transducer creating acoustic excitation at the excitation frequency, the channel retains the particles as the residual portion of the sample continues to flow through the channel to the single output.

Description

ACOUSTIC CONCENTRATION OF CELLS OR PARTICLES IN BLOOD PRODUCTSCROSS-REFERENCE TO RELATED APPLICATIONS[0001| This application claims the benefit of U.S. provisional application Serial No. 63 / 666,868 filed July 2, 2024, the disclosure of which is hereby incorporated in its entirety by reference herein.TECHNICAL FIELD

[0002] Aspects of the disclosure generally relate to acoustic concentration of cells or particles in blood products.BACKGROUND

[0003] Rapid detection of pathogens in blood may be challenging due to the abundance of a pathogen bein far lower than the abundance of endogenous blood cells. For example, in abacterial infection in blood, there may be as many as one billion blood cells for every bacterium in circulation. Rapid detection can be improved by devices and methods that can purify and / or enrich the pathogen while removing most of the normal blood cells. Furthermore, the concentration of the pathogen in a blood sample may be very low, such that it is below the limit of detection of a detector, unless the pathogen is first concentrated in solution before being presented to the detector. For these reasons, devices and methods are desired that can rapidly enrich and concentrate pathogens from blood and do so in a format that they can incorporated into automated detection systems.SUMMARY10004] A system for trapping cells or particles in a blood sample may include a device having a channel with a predefined geometry, the channel having an input and a single output, the channel configured to receive a sample including target particles or cells and a residual portion at the input; and configured to flow the sample through the channel to the single output, and a transducer creating acoustic excitation of the device at an excitation frequency that is determined relative tothe predefined geometry, the channel is configured to, upon the transducer creating acoustic excitation at the excitation frequency, retain the particles as the residual portion of the sample continues to flow through the channel to the single output.

[0005] A method for trapping cells or particles in a blood sample may include flowing a sample including particles and a residual portion through an input of a channel of an acoustic concentrator device; and applying acoustic excitation at an excitation frequency to the channel to retain the particles within the channel as the residual portion of the sample continues to flow through the channel.

[0006] A system for trapping cells or particles in a blood sample may include a device having a channel with a predefined geometry, the channel configured to receive a sample including particles and a residual portion and to flow the sample through the channel to an output, and wherein the channel is configured to receive acoustic excitation at an excitation frequency relative to the predefined geometry, and upon the acoustic excitation at the excitation frequency, retain the particles as the residual portion of the sample continues to flow through the channel to the output.BRIEF DESCRIPTION OF THE DRAWINGS

[0007] The embodiments of the present disclosure are pointed out with particularity in the appended claims. However, other features of the various embodiments will become more apparent and will be best understood by referring to the following detailed description in conjunction with the accompanying drawings herein.

[0008] FIG. 1 illustrates a cross-sectional top view of a microchannel component of a concentration device for a blood diagnostic system, having a channel configured to receive a sample at an input and concentrating particles into an output.

[0009] FIG. 2 illustrates a side view of channel component of the device of FIG. 1 .

[0010] FIG. 3 illustrates a sequential process of fluid flow through the concentration device with acoustic actuation during some of the steps.[0011 FIG. 4 illustrates an example block diagram for a system for operating at least a portion of the process 300 of FIG. 3 utilizing the device 100 of FIG. 1.

[0012] FIGs. 5A-C illustrates examples of numerical simulation of flow, pressure, and particle trajectories within the device of FIG. 1, during acoustic actuation.DETAILED DESCRIPTION[0013| As required, detailed embodiments of the present invention are disclosed herein; however, it is to be understood that the disclosed embodiments are merely exemplary of the invention that may be embodied in various and alternative forms. The figures are not necessarily to scale; some features may be exaggerated or minimized to show details of particular components. Therefore, specific structural and functional details disclosed herein are not to be interpreted as limiting, but merely as a representative basis for teaching one skilled in the art to variously employ the present invention.

[0014] As explained above, rapid detection of pathogens in blood may be challenging due to the abundance of a pathogen being far lower than the abundance of endogenous blood cells. For example, in a bacterial infection in blood, there may be as many as one billion blood cells for every bacterium in circulation. Rapid detection can be improved by devices and methods that can purify and / or enrich the pathogen while removing most of the normal blood cells. Furthermore, the concentration of the pathogen in a blood sample may be very low, such that it is below the limit of detection of a detector, unless the pathogen is first concentrated in solution before being presented to the detector. For these reasons, devices and methods are desired that can rapidly enrich and concentrate pathogens from blood and do so in a format that they can incorporated into automated detection systems.

[0015] Disclosed herein is a trap, or concentration device, constructed from an acoustically actuated microchannel. With suitable channel dimensions, actuator coupling, and actuator frequency, particles or cells can be retained in the microchannel by the acoustic forces even as fluid continues to flow through it. Hence if a suspension of cells flows through the device, a fraction of the cells will remain trapped in the device while the rest of the suspension exits. If the acoustic excitation is then stopped while fluid flow continues, the cells can be swept out of thedevice as a bolus with higher concentration than that of the original input suspension. This bolus may then be delivered to a downstream detection system or to further process steps.|0016| FIG. 1 illustrates a cross-sectional top view of a concentration device 100 for a blood cleansing system having a channel 102 configured to receive a sample at an input 104 and concentrating molecules into an output 106. FIG. 2 illustrates a side view of the device 100 of FIG. 1.

[0017] Referring to FIGs. 1 and 2, the channel 102 (or microchannel 102) may have dimensions configured to retain particles or cells within the microchannel 102 by applying acoustic forces. The sample comprises a predetermined concentration of non-target endogenous particles that are retained within the channel 102. The remaining residual portion of the fluid will flow through the output 106. The actuator frequency may facilitate the appropriate cell retention. If the acoustic excitation is then stopped while fluid flow continues, the cells may be swept out of the device 100 as a bolus, released into an output 106, with higher concentration than that of the original input suspension. The bolus may then be delivered to a downstream detection system or for further processing.

[0018] FIG. 3 illustrates a process 300 of fluid flow through the concentration device 100. At step 305, a sample flows through the channel 102 and target cells are trapped within the channel by acoustic forces created by a transducer 304 while fluid continues to flow through the channel 102. The transducer 304 may be an ultrasonic transducer configured to covert electrical energy into sound waves or acoustic energy. Such energy may then exert mechanical forces on objects acoustically coupled to the transducer 304, in this case, on particles within the channel 102. The pressure field in the fluid due to the acoustic wave may displace the particles within the channel 102. This force may cause some particles to separate from other particles or cause particles to travel along a certain direction that is different than their travel due to the fluid drag force.

[0019] At step 310, a second fluid may flow through the channel 102 to wash residual fluid or to prepare the trapped cells in a new fluid. Similar to step 305, acoustic power is applied to the particles via the transducer 304. The second fluid may be any fluid capable of facilitating acoustic wave propagation on the particles. The second fluid may have certain properties such as acousticimpedance, viscosity, density, and biocompatibility. Example fluids may include water, saline, glycerol, lysis buffer, etc.|0020| At step 315, the acoustic power may be turned off to release the cells into the flowing fluid and out of the channel 102 and into a collection vessel 306 or another processing step. In steps 305 and 310, waste may be collected at a waste vessel 308. The flowing fluid containing the released particles may be driven toward the outlet 102 by a second fluid, a gas, or an immiscible fluid.[00211 Referring back to FIGs. 1 and 2, and as explained, the acoustic forces trap particles via pressure nodes. Stable pressure nodes may arise when a mechanical resonance is established by oscillating the microchannel 102 at an appropriate frequency. The resonant frequency may depend on channel dimensions, wall dimensions, and channel materials. Because of this, a height H of the channel 102 may be approximately 0.2 to 2 mm. A width W of the channel 102 may be approximately 0.3 to 1 mm. In addition to the channel height and width, the wall thickness T may also affect node creation and be considered as part of the channel geometry. Walls enclosing the channel 102 may be approximately 1 to 3mm. Such example dimensions would facilitate the trap at approximately 10MHz, or at least between 5-15MHz when the fluid is a saline buffer or blood. Such a combination of frequency and geometry create numerous pressure nodes through the crosssection of the channel 102. This is best illustrated and described with respect to FIG. 5C. Further, the dimensions of the channel 102 and the excitation frequency may vary, but may also be interrelated. In one example, the channel height H may be two to twenty times that of the wavelength of sound in the fluid at the applied frequency. Further, while the height H is discussed herein, relative widths may also be accommodated for numerous node creation.|0022| The channel 102 may be made of a wide range of materials including metals, glass, silicon, and polymers, especially rigid thermoplastics such as polystyrene, acrylic, and cyclic olefin polymers. It is desirable for the channel 102 to be low cost for single use disposable applications where contamination from previous samples is unacceptable. Further, the channel 102 includes a single output to facilitate the trapping of endogenous blood cells or contaminants.

[0023] The acoustic excitation may be applied by permanently or reversibly coupling a piezoelectric actuator, such as the transducer 306, to one or more surfaces on the exterior of the microchannel 102. The piezoelectric may be lead zirconate titanate, aluminum nitride, or other materials. The coupling may be achieved with adhesive or with a liquid such as glycerol. The transducer 306 may be further mounted to a temperature control system to dissipate heat generated by the transducer 306.

[0024] The device 100 may be operated with a predetermined optimum concentration of blood cells or other residual cells or particles present in the input sample. To enhance trapping of the smaller target bacteria or particles. Larger particles (or other inhomogeneities in the suspension) can create local variations in the acoustic field that improve trapping of smaller particles. For example, trapping of bacteria is improved when a low concentration of red blood cells is present in the solution, even though the red blood cells are not the target of the concentrator. Reagents that produce aggregates of blood cells or platelets may also be intentionally added to create desirable clusters of non-target cells that improve the trapping of the target bacteria. The predetermined concentration of blood cells may be the result of upstream separation processes. The blood cells may be a fraction of those present in the original sample, and need not be added to the sample.

[0025] Further, such trapping within the channel 102 is done without the addition of ligands, binding agents, or antibodies that are complementary to the target cells. While the sample or blood may be diluted or adjusted for mechanical performance, for example with density medium or aggregation additives, no specific recognition molecules are required for the trapping by implementing a channel size relative to an excitation frequency.

[0026] The channel 102 may be incorporated into a microfluidic system that includes valves or switches and microchannel networks to perform several unit operations. This is described in more detail with respect to FIG. 4.

[0027] In another example, a fluidic switching network of valves and channels may be situated downstream of the trap so that waste flow is directed to one system outlet while the released bolus (i.e., eluate) is directed to a different system outlet.

[0028] Further, a fluidic switching network of valves and channels may be situated upstream of the trap to enable flow of washing or reagent solutions that flush out the original suspending fluid while the trap is activated and bathe the trapped cells in a new fluid. The cells may be released to flow out of the trap in this new fluid. The reagent fluid may contain dyes, antibodies, reporters, lysis agents, primers, etc. to aid in downstream detection or analysis.

[0029] In yet another example, the elution or release fluid bolus may be driven by a meniscus such as an air — fluid boundary or an oil — aqueous boundary to keep the concentrated targets within a small and well-defined fluid volume. In this example the air or oil may be provided from a source connected to the fluidic network.

[0030] In some examples, the trap may be operated repeatedly as steps of loading, washing, and eluting are performed on partial volumes of the sample and repeated until the entire sample is processed.

[0031] Additionally or alternatively, the trap may include optical or electrical devices or other means to measure the amount of trapping or analyze the cells while they are trapped. Further, the channel may be clamped or temporarily adhered to an acoustic transducer to enable user-friendly replacement of the channel while re-using the transducer. In some examples, the channel may be a single use component within a system of multi-use components, such as pumps, valves, sensors, temperature control components, optics, electronics, transducers, etc.

[0032] In some embodiments, the assay of the target cells may occur within the trap, omitting the elution step. For example the target cells may be detected by optical mechanisms while they are retained in the trap.[00331 FIG. 4 illustrates an example block diagram for a system 400 for operating at least a portion of the process 300 of FIG. 3 utilizing the device 100 of FIG. 1.

[0034] In this example and generally, the trap may be directly connected to receive flow from an upstream separation / purification step, including an acoustic separator that operates at different frequencies and has different channel dimensions than the frequencies and dimensions of the concentrator. The operating parameters of the upstream separation step may be tuned to maximizeperformance of the downstream concentration step. For example, a sensor system may measure the constituents of the output of the separator step and adjust the separator parameters to achieve a desired composition of the sample for delivery to the concentrator.

[0035] The system 400 may receive an input sample source 401 at a reservoir. The system 400 may include a separator 402, assay buffer source or reservoir 404, concentrator 406 (i.e., device 100), assay subsystem 408 and a waste collector or reservoir 410. The concentrator 406 may be incorporated into the system 400 where the input sample is first purified by a different component and subsequently concentrated and delivered to the downstream assay 408. Various valves 412a- d (collectively referred to herein as valves 412) are included between the components and switched to different open and closed states to accomplish the unit operations. For example, a first valve 412a may be arranged between the separator 402 and the concentrator 406. A second valve 412b may be arranged between the assay buffer source 404 and the concentrator 406. A third valve 412c may be arranged between the waste collector 410 and the output of the concentrator 406. A fourth valve 412d may be arranged between the concentrator 406 and the assay 408, downstream from the third valve 412c.

[0036] The first valve 412a may control the sample flow from the separator 402 to the concentrator 406. During step 310, the second valve 412b may control the flow of the wash fluid to the concentrator 406. The third valve 412c may control the flow to the waste collector 410 from the concentrator 406 during steps 305 and 310, while the fourth valve 412d may control the flow to the downstream assay 408 (or sample collector 306). The valves 412 may be controlled by a controller (not shown) according to a predefined or customizable algorithm. This may include the controller receiving data from a sensor system regarding the measure of constituents at the output of the separator step and providing instructions to the valves 412 to adjust the separator parameters to achieve a desired composition of the sample for delivery to the concentrator 406.[0037| Accordingly, the transducer 306 and channel 102 comprise a resonant system and are optimized together, such that channel dimensions, transducer dimensions, transducer material, coupling layer between channel 102 and transducer 306, and positioning of channel 102 on the transducer 306 are all variables subject to optimization for best results in terms of concentrator performance. The mounting condition of the transducer 306 to any further fixturing may alsoimpact performance, and may include air backing, thin metal backing, damped backing, and specific temperature control configurations. Additionally flow rates and applied acoustic power can be optimized and these settings may be different for different steps of initial trapping, washing, and elution of targets. For example flow rate for trapping may be lower than flow rate for elution. System controls or software may automatically adjust the parameters according to predetermined values or to closed-loop sensor readout.|0038| Thus, the disclosed device, system, and method may be used for capturing pathogens or other small bodies in blood, such as platelets, exosomes, viral particles, or nanoparticles. This is due at least in part to the high frequency, high energy, and small channel dimensions to trap these smaller particles. Prior devices, including surface acoustic wave device, have been costly to build and have low fluid velocity and low sample throughput. The disclosed device does not require any added ‘seed’ particles or synthetic particle additives.

[0039] While the device is described herein for processing bacteria in blood, the device may also be used for concentrating targets in other samples such as other biofluids or environmental samples. Similar to the use of blood cells to enhance trapping, existing particles such as dirt in environmental samples could be employed to aid trapping of smaller target species.

[0040] FIGs. 5A-C illustrate example simulations within the device 100. The simulation are shown in half of the symmetrical device having polystyrene walls. The simulations indicate particle trajectories do not escape, that the acoustic force exceeds a drag force.

[0041] FIG. 5A illustrates an example channel having a fluid velocity of 0.1-1 mL / min and ordinary continuous flow of liquid through the rectangular channel.[0042| FIG. 5B illustrates an example channel having an acoustic pressure field in fluid at 8.82 MHz. High pressure is represented by 502 and low pressure is represented by 504. Areas between the high and low pressure streams are pressure nodes 506. As explained above, through a combination of a channel height and excitation frequency, numerous nodes 506 may be created to occupy as much of the cross-sectional area of the channel 102 as possible.

[0043] FIG. 5C illustrates example simulated trajectories of 1 um particles.10044] Computing devices described herein generally include computer-executable instructions, where the instructions may be executable by one or more computing devices such as those listed above. Computer-executable instructions may be compiled or interpreted from computer programs created using a variety of programming languages and / or technologies, including, without limitation, and either alone or in combination, Java™, C, C++, C#, Visual Basic, Java Script, Perl, etc. In general, a processor (e.g., a microprocessor) receives instructions, e.g., from a memory, a computer-readable medium, etc., and executes these instructions, thereby performing one or more processes, including one or more of the processes described herein. Such instructions and other data may be stored and transmitted using a variety of computer-readable media.(0045] While exemplary embodiments are described above, it is not intended that these embodiments describe all possible forms of the invention. Rather, the words used in the specification are words of description rather than limitation, and it is understood that various changes may be made without departing from the spirit and scope of the invention. Additionally, the features of various implementing embodiments may be combined to form further embodiments of the invention.

Claims

WHAT IS CLAIMED IS:

1. A system for trapping cells or particles in a blood sample, comprising: a device having a channel with a predefined geometry, the channel having an input and a single output, the channel configured to receive a sample including particles and a residual portion at the input and to flow the sample through the channel from the input to the single output; and a transducer configured to generate an acoustic excitation of the device at an excitation frequency relative to the predefined geometry, wherein the channel is configured to, upon the transducer creating the acoustic excitation at the excitation frequency, retain the particles as the residual portion of the sample continues to flow through the channel to the single output.

2. The system of claim 1, wherein a dimension of the predefined geometry is between two and twelve times a wavelength of sound in the fluid at the excitation frequency.

3. The system of claim 1, wherein a dimension of the predefined geometry is at least one of a channel height, channel width, and wall thickness.

4. The system of claim 3, wherein the channel height is between 0.2 to 2 mm.

5. The system of claim 1, wherein the excitation frequency is 5-15MHz.

6. The system of claim 1, further comprising a plurality of valves arranged downstream from the channel output configured to control the flow of the residual portion at the output.

7. A method for trapping cells or particles in a blood sample, comprising: flowing a sample including particles and a residual portion through an input of a channel of an acoustic concentrator device; andapplying acoustic excitation at an excitation frequency to the channel to retain the particles within the channel as the residual portion of the sample continues to flow through the channel.

8. The method of claim 7, wherein the channel has a predefined geometry related to the excitation frequency.

9. The method of claim 8, wherein a dimension of the predefined geometry is between two and twelve times a wavelength of sound in the fluid at the acoustic excitation.

10. The method of claim 8, wherein a dimension of the predefined geometry includes at least one of a channel height, channel width, and wall thickness.

11. The method of claim 10, wherein the channel height is between 0.2 to 2 mm.

12. The method of claim 7, wherein the excitation frequency is approximately 5-15MHz.

13. The method of claim 7, wherein the residual portion flows to and is collected at a single output.

14. The method of claim 7, wherein the sample comprises a predetermined concentration of non-target endogenous particles that are retained within the channel.

15. The method of claim 13, further comprising controlling a plurality of valves arranged downstream from the channel output to control the flow at the output.

16. The method of claim 7, further comprising flowing a second fluid through the input of a channel of the acoustic concentrator device to wash residual fluid from the sample.

17. A system for trapping cells or particles in a blood sample, comprising: a device having a channel with a predefined geometry, the channel configured to receive a sample including particles and a residual portion and to flow the sample through the channel to an output; and wherein the channel is configured to receive acoustic excitation at an excitation frequency relative to the predefined geometry, and upon the acoustic excitation at the excitation frequency, retain the particles as the residual portion of the sample continues to flow through the channel to the output.

18. The system of claim 17, wherein a dimension of the predefined geometry is between two and twelve times a wavelength of sound in the fluid at the excitation frequency.

19. The system of claim 17, wherein a dimension of the predefined geometry is at least one of a channel height, channel width, and wall thickness.

20. The system of claim 19, wherein the channel height is between 0.2 to 2 mm and the excitation frequency is 5-15MHz.

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