Patent Grant
10730046
It has been demonstrated that cancerous cells are more resistant to fluid shear stress than normal (non-cancerous) cells. (See, Barnes J. M., Nauseef, J. T., Henry, M. D. (2012) Resistance to Fluid Shear Stress Is a Conserved Biophysical Property of Malignant Cells. PLoS ONE 7(12): e50973. doi:10.1371/journal.pone.0050973.) In particular, the repeated exposure of fluid samples comprising both cancerous and normal cells to a fluid shear stress has been found to impart a selective resistance to the fluid shear stress to the cancerous cells and to selectively kill the non-cancerous cells, thereby providing a fluid sample enriched in the cancerous cells. (Id.) The concentration and isolation of fluid shear stress-resistant cancerous cells allows for further characterization of the cancerous cells, ultimately leading to improved clinical diagnostic tests for prognostic and therapeutic applications.
Fluid handling systems for applying a plurality of pulses of fluid shear stress to a fluid sample are provided. Related methods for the fluid handling systems are also provided.
In a first aspect, a first embodiment of a fluid handling system for applying a plurality of pulses of fluid shear stress to a fluid sample is provided comprising a first sample chamber; a second sample chamber; a plurality of conduits mounted between and in fluid communication with the first sample chamber and the second sample chamber, each conduit having an inner diameter of less than about 1000 μm; and a force delivery system mounted to the first sample chamber and configured to apply a force sufficient to push the fluid sample at a substantially constant flow rate from the first sample chamber through each of the conduits to the second sample chamber. The dimensions of each conduit may be substantially the same. The fluid handling system may further comprise a control system operably coupled to the force delivery system to repeatedly apply the force, each application of force having a selected magnitude and a selected duration time.
In the first embodiment, the conduits may be arranged in series and are each separated by an additional sample chamber.
In some such embodiments, the force delivery system may be a gas delivery system configured to deliver gas to pressurize the sample chambers to a selected pressure, the gas delivery system comprising a plurality of gas valves, each gas valve in fluid communication with an associated sample chamber.
In some such embodiments, the fluid handling system may further comprise a syringe stack comprising a plurality of stackable syringe assemblies, each stackable syringe assembly in fluid communication with an adjacent stackable syringe assembly, each stackable syringe assembly comprising: a syringe body defining a sample chamber; a gas inlet port in fluid communication with the sample chamber; and a conduit through which the fluid sample may pass from the sample chamber into an adjacent sample chamber of the adjacent stackable syringe assembly; wherein the syringe stack comprises the first sample chamber, the second sample chamber, the plurality of conduits and the additional sample chambers, and wherein each gas valve is in fluid communication with an associated stackable syringe assembly via an associated gas inlet port.
Alternatively, in the first embodiment, the conduits may be arranged substantially parallel to one another.
In some such embodiments, the force delivery system may be a linear drive assembly configured to translate a first surface in a first direction towards the fluid sample in the first sample chamber at a selected speed over a selected distance and to translate a second surface in an opposing, second direction towards the fluid sample at the selected speed over the selected distance in the second sample chamber.
In some such embodiments, the fluid handling system may further comprise a moveable sample receptacle assembly comprising a first syringe body defining the first sample chamber, a second syringe body defining the second sample chamber and the plurality of substantially parallel conduits mounted between the first syringe body and the second syringe body; a first fixed piston mounted in a first bore of the first syringe body; and a second fixed piston mounted in a second bore of the second syringe body; wherein the linear drive assembly is configured to translate the moveable sample receptacle assembly back and forth along the longitudinal axis of the sample receptacle assembly.
In a second embodiment, a fluid handling system for applying a plurality of pulses of fluid shear stress to a fluid sample comprises a first sample chamber; a second sample chamber; a plurality of conduits mounted between and in fluid communication with the first sample chamber and the second sample chamber, wherein the conduits are arranged in series and are each separated by an additional sample chamber; and a gas delivery system mounted to the first sample chamber, the gas delivery system configured to deliver gas to pressurize the sample chambers to a selected pressure, the gas delivery system comprising a plurality of gas valves, each gas valve in fluid communication with an associated sample chamber. Each conduit may have an inner diameter of less than about 1000 μm. The dimensions of each conduit may be substantially the same.
In the second embodiment, the fluid handling system may further comprise: a syringe stack comprising a plurality of stackable syringe assemblies, each stackable syringe assembly in fluid communication with an adjacent stackable syringe assembly, each stackable syringe assembly comprising a syringe body defining a sample chamber; a gas inlet port in fluid communication with the sample chamber; and a conduit through which the fluid sample may pass from the sample chamber into an adjacent sample chamber of the adjacent stackable syringe assembly; wherein the syringe stack comprises the first sample chamber, the second sample chamber, the plurality of conduits and the additional sample chambers, and wherein each gas valve is in fluid communication with an associated stackable syringe assembly via an associated gas inlet port.
In some such embodiments, the syringe body may further comprise a bottom end portion configured such that the bottom end portion is insertable into a top opening of an adjacent syringe body of the adjacent stackable syringe assembly to form a pressure-tight seal.
In some such embodiments, the stackable syringe assembly may further comprise an arm defining a bore through which fluid may pass, the arm mounted to the syringe body and extending from the gas inlet port, the arm configured to mount to a gas line coupler and to a receptacle configured to collect a portion of the fluid sample. The receptacle may be a pipette tip insertably mounted in the bore.
In some such embodiments, the syringe body may comprise a partition assembly mounted in the sample chamber and configured to reduce foaming of the fluid sample as it passes into the sample chamber en route to the conduit. The partition assembly may comprise a central portion mounted to a side wall of the syringe body at a bottom end of the central portion, the central portion extending upwardly such that a top end of the central portion is positioned within a substantially central location within the sample chamber, and first and second lateral portions mounted to opposite sides of the top end of the central portion, wherein the central portion and first and second lateral portions define gaps through which the fluid sample may pass.
In some such embodiments, the gas delivery system may comprise a gas valve stack, the gas valve stack comprising a plurality of gas valve assemblies, the plurality of gas valve assemblies comprising the plurality of gas valves. The plurality of gas valve assemblies may comprise three-way solenoid valves.
In some such embodiments, the fluid handling system may further comprise a support assembly mounted to the syringe stack and configured to position the syringe stack vertically.
In the second embodiment, the fluid handling system may further comprise a control system operably coupled to the gas delivery system and configured to sequentially deliver gas to each sample chamber at the selected pressure for a selected duration time to sequentially pass the fluid sample through each conduit. The control system may be configured to perform operations comprising (a) pressurizing the first sample chamber with gas to the selected pressure; (b) maintaining the pressurization at the selected pressure until an indicator indicates the substantially complete delivery of the fluid sample through a first conduit in the plurality of conduits into an adjacent sample chamber; (c) venting the first sample chamber for a selected hold time; and (d) repeating steps (a)-(c) such that the fluid sample passes through each conduit in the plurality of conduits.
A method of using the fluid handling system according to the second embodiment may comprise (a) pressurizing the first sample chamber comprising the fluid sample with gas to the selected pressure; (b) maintaining the pressurization at the selected pressure until an indicator indicates the substantially complete delivery of the fluid sample through a first conduit in the plurality of conduits into an adjacent sample chamber; (c) venting the first sample chamber for a selected hold time; and (d) repeating steps (a)-(c) such that the fluid sample passes through each conduit in the plurality of conduits. The method may further comprise withdrawing a portion of the fluid sample from the adjacent sample chamber during the selected hold time.
In another aspect, a stackable syringe assembly for use in a fluid handling system is provided comprising a syringe body defining a sample chamber, the syringe body comprising a bottom end portion configured such that the bottom end portion is insertable into a top opening of an adjacent syringe body of an adjacent stackable syringe assembly to form a pressure-tight seal; a gas inlet port in fluid communication with the sample chamber; and a conduit through which the fluid sample may pass from the sample chamber into an adjacent sample chamber of the adjacent stackable syringe assembly.
In a third embodiment, a fluid handling system for applying a plurality of pulses of fluid shear stress to a fluid sample is provided comprising: a first sample chamber; a second sample chamber; a plurality of substantially parallel conduits mounted between and in fluid communication with the first sample chamber and the second sample chamber; and a linear drive assembly mounted to the first sample chamber and configured to translate a first surface in a first direction towards the fluid sample in the first sample chamber at a selected speed over a selected distance and to translate a second surface in an opposing, second direction towards the fluid sample in the second sample chamber at the selected speed over the selected distance.
In the second embodiment, the fluid handling system may further comprise: a moveable sample receptacle assembly comprising a first syringe body defining the first sample chamber, a second syringe body defining the second sample chamber and the plurality of substantially parallel conduits mounted between the first syringe body and the second syringe body; a first fixed piston mounted in a first bore of the first syringe body; and a second fixed piston mounted in a second bore of the second syringe body; wherein the linear drive assembly is configured to translate the moveable sample receptacle assembly back and forth along the longitudinal axis of the sample receptacle assembly. The conduits may each have an inner diameter of less than about 1000 μm. The dimensions of each conduit may be substantially the same.
In some such embodiments, the plurality of substantially parallel conduits may be embedded in a conduit holding block and arranged in an array.
In some such embodiments, the first fixed piston and the second fixed piston may each be provided by a syringe plunger.
In some such embodiments, the first fixed piston and the second fixed piston may each be adjustably mounted to the linear drive assembly via a first piston anchor assembly and a second piston anchor assembly, respectively.
In some such embodiments, the fluid handling system may further comprise a control system operably coupled to the linear drive assembly and configured to translate the sample receptacle assembly back and forth over the selected distance between an initial position and a selected position at the selected speed to repeatedly pass the fluid sample through the plurality of substantially parallel conduits. The control system may configured to perform operations comprising: (a) translating the sample receptacle assembly from the initial position to the selected position in the first direction along the longitudinal axis at the selected speed to transfer the fluid sample from the first sample chamber to the second sample chamber through the plurality of substantially parallel conduits; (b) holding the sample receptacle assembly at the selected position for a selected hold time; (c) translating the sample receptacle assembly from the selected position to the initial position in the opposing direction along the longitudinal axis at the selected speed to transfer the fluid sample from the second sample chamber to the first sample chamber through the plurality of substantially parallel conduits; (d) holding the sample receptacle assembly at the initial position for the selected hold time; and (e) repeating steps (a)-(d) such that the fluid sample passes through the plurality of substantially parallel conduits a selected number of times.
A method of using the fluid handling system according to the third embodiment may comprise (a) translating the sample receptacle assembly comprising the fluid sample in the first sample chamber from an initial position to a selected position in the first direction along the longitudinal axis at the selected speed to transfer the fluid sample from the first sample chamber to the second sample chamber through the plurality of substantially parallel conduits; (b) holding the sample receptacle assembly at the selected position for a selected hold time; (c) translating the sample receptacle assembly from the selected position to the initial position in the opposing direction along the longitudinal axis at the selected speed to transfer the fluid sample from the second sample chamber to the first sample chamber through the plurality of substantially parallel conduits; and (d) holding the sample receptacle assembly at the initial position for the selected hold time. The method may further comprise repeating steps (a)-(b) or steps (a)-(d) such that the fluid sample passes through the plurality of substantially parallel conduits a selected number of times. The method may further comprise withdrawing a portion of the fluid sample from one of the first or second sample chambers during one of the selected hold times.
Other principal features and advantages of the invention will become apparent to those skilled in the art upon review of the following drawings, the detailed description, and the appended claims.
Exemplary embodiments of the invention will hereafter be described with reference to the accompanying drawings, wherein like numerals denote like elements.
As used herein, the term “mount” includes join, unite, connect, couple, associate, insert, hang, hold, affix, attach, fasten, bind, paste, secure, bolt, screw, rivet, solder, weld, glue, form over, form in, layer, mold, rest on, rest against, abut, and other like terms. The phrases “mounted on”, “mounted to”, and equivalent phrases indicate any interior or exterior portion of the element referenced. These phrases also encompass direct mounting (in which the referenced elements are in direct contact) and indirect mounting (in which the referenced elements are not in direct contact, but are connected through an intermediate element). Elements referenced or shown as mounted to each other herein may further be integrally formed together, for example, using a molding or thermoforming process as understood by a person of skill in the art. As a result, elements described herein as being mounted to each other need not be discrete structural elements. The elements may be mounted permanently, removably, or releasably unless specified otherwise.
Use of directional terms, such as top, bottom, right, left, front, back, etc. are merely intended to facilitate reference to various surfaces that form components of the devices referenced herein and are not intended to be limiting in any manner.
Fluid handling systems for applying a plurality of pulses of fluid shear stress to a fluid sample are provided. By “pulse,” it is meant that the fluid sample is exposed to a selected magnitude of fluid shear stress for a selected duration of time. As described in U.S. Pat. Pub. No. 20140038231, which is hereby incorporated by reference in its entirety, fluid sample being passed through a conduit experiences a range of magnitudes of fluid shear stress from zero to a maximum value, with the magnitude depending upon its position relative to the longitudinal axis of the conduit. The “selected magnitude of fluid shear stress” may refer to the shear stress calculated at the wall of the conduit through which the fluid sample is passed, calculated using Poiseuille's equation, τ=4Qη/πr3, wherein τ is shear stress; Q is flow rate; η is the viscosity of the medium; and r is the radius of the conduit. A pulse program may be characterized by the number of pulses, the magnitude of fluid shear stress for each pulse, the duration of time for each pulse and the hold time between pulses. The fluid handling systems may be used to apply a variety of different pulse programs, including those disclosed in U.S. Pat. Pub. No. 20140038231.
Similarly, the fluid samples may include the preparations disclosed in U.S. Pat. Pub. No. 20140038231. Thus, the fluid samples may include cancerous cells, e.g., an in vitro preparation of cancerous cells or a blood sample from a patient. The repeated exposure of such fluid samples to fluid shear stress via the disclosed fluid handling systems imparts an increased resistance to fluid shear stress to the cancerous cells and provides a population of fluid shear stress-resistant cancerous cells in the fluid samples. Since normal cells (non-cancerous cells) do not experience an increase in their resistance to fluid shear stress or experience a lower increase than the cancerous cells, the normal cells may be selectively killed by the application of fluid shear stress provided by the disclosed fluid handling systems. Thus, the fluid handling systems may be used to carry out the methods disclosed in U.S. Pat. Pub. No. 20140038231, including methods for purifying/concentrating fluid samples comprising cancerous cells. Such methods are useful as part of clinical diagnostic tests for prognostic applications to assess the likely health outcome for a patient having, or at risk of developing, cancer or metastases and therapeutic applications to assess the effect of a treatment program for such a patient.
The fluid handling systems are also applicable to the field of cytologic pathology in which cancer cells may be admixed with a variety of non-cancer cells. This includes, but is not limited to, fine needle aspirates and fluid specimens including urine, pleural effusion, peritoneal fluid, and cerebrospinal fluid. The fluid handling systems may be used to enrich the relative abundance of cancer to non-cancer cells in the sample preparation, prior to a variety of subsequent analytic procedures. This facilitates standard cytolologic workup as well as molecular analyses including genetic analysis and immunohistochemical staining.
As shown schematically in
As discussed above, passage of the fluid sample through the conduits results in exposure of the fluid sample to a range of magnitudes of fluid shear stress from zero to a maximum value, in which the magnitude of the fluid shear stress depends upon the flow rate achieved by the application of the force (as well as parameters of the conduit and the fluid sample itself). Thus, the magnitude of the force may be selected to provide a selected flow rate and thus, a selected magnitude of fluid shear stress. The selected magnitude of fluid shear stress may vary over a wide range, including high and/or supra-physiologic levels. In some embodiments, the selected magnitude of fluid shear stress is in the range of from about 300 to about 6500 dyn/cm2. In some embodiments, the selected magnitude of fluid shear stress is at least 500 dyn/cm2, at least 1000 dyn/cm2, at least 3500 dyn/cm2, or at least 6000 dyn/cm2. Other selected magnitudes of fluid shear stress may be used, including those disclosed in U.S. Pat. Pub. No. 20140038231.
As shown in
In some embodiments of the fluid handling system 2200, the force delivery assembly 2216 may be a gas delivery system, e.g., the force to push the fluid sample through the conduits 2212 is generated by exposing the fluid sample to a pressurized gas. In some embodiments of the fluid handling system 2200, the force delivery assembly 2216 may be a linear drive assembly, e.g., the force to push the fluid sample through the conduits 2212 is generated by the mechanical translation of a surface against the fluid sample contained in the sample chambers 2204, 2208. These embodiments are described in separate sections immediately below. Other types of force delivery assemblies may be used, including those based on the use of centripetal force, gravitational force and shear force.
With reference to
In the exemplary embodiment, an uppermost stackable syringe assembly 202 may include a syringe body 210. A bottom wall (not shown) of the syringe body 210 and side walls extending from the bottom wall define a uppermost sample chamber (not shown) which is accessible via a top opening 214 at a top end of the syringe body 210. Each stackable syringe assembly may be similarly configured. A bottommost stackable syringe assembly 206 may also include a syringe body 218. A bottom end portion 222 extends from a bottom wall (not shown) of the syringe body 218. A bottommost sample chamber (not shown) of the bottommost stackable syringe assembly 206 is also accessible via a bottom opening 226 in the bottom end portion 218. Each stackable syringe assembly may be similarly configured.
Each stackable syringe assembly in the syringe stack 104 may include a bottom end portion configured as shown in
The syringe stack 104 may be sealed via a cap configured to insert into the top opening 214 of the uppermost stackable syringe assembly 202 and to form another pressure-tight seal. The cap may include an o-ring on an end thereof to facilitate formation of the pressure-tight seal.
The stackable syringe assembly 106 may include the syringe body 302. A bottom wall 303 of the syringe body 302 and side walls 304a-d extending from the bottom wall 303 define a sample chamber 306. A bottom end portion 318 extends from the bottom wall 303. The sample chamber 306 is accessible via a top opening 310 at a top end and via a bottom opening 314 in the bottom end portion 318. A bore 307 defined in the bottom wall 303 connects the sample chamber 306 to the hollow interior of the bottom end portion 318 and the bottom opening 314. As described above with respect to the bottommost stackable syringe assembly 206 (with reference to
The stackable syringe assembly 106 may include a mounting member 326 mounted to the syringe body 302. The mounting member 326 may be configured to mount to an element of the support assembly 112 (e.g., a syringe stack support rod 738, with reference to
The stackable syringe assembly 106 may include an arm 334 extending from a gas inlet port 338 defined the side wall 304b of the syringe body 302. The walls of the arm 334 may define a bore 342 leading to the gas inlet port 338 at one end of the bore 342 and to a gas line coupler 346 at an opposing end of the bore 342. In this exemplary embodiment, the arm 334 is oriented substantially at a 45° angle with respect to the longitudinal axis of the syringe body 302, although other orientations may be used, e.g., 90° (see arm 2008 in
The dimensions of the syringe body 302 may depend upon the desired volume of the fluid sample to be held in the sample chamber 306. For example, the dimensions of the syringe body 302 may be that which is sufficient to hold about 10 mL, about 5 mL, etc. of the fluid sample.
The stackable syringe assembly 106 may include the gas line coupler 346 mounted to a top end of the arm 334. The gas line coupler 346 may be configured to connect a gas line mounted to the gas line coupler 346 at one end and an associated gas valve assembly 502 (with reference to
As shown in
The stackable syringe assembly 106 may include a conduit 352 in fluid communication with the sample chamber 306. As shown in the exemplary embodiment, the conduit 352 may be provided via a needle 350 which may be mounted on a threaded projection 320 which surrounds the bore 307 and extends into the hollow interior of the bottom end portion 318. The conduit 352 may be mounted and positioned such that it receives the fluid sample passing from the sample chamber 306 via the bore 307. Commercially available needles, e.g., hypodermic needles, may be used for the needle 350. The dimensions of the conduit 352 may be selected to provide a selected magnitude and selected duration time for the pulses of fluid shear stress to be applied (for a given pressure of gas applied to the fluid sample). The conduit may be micron-sized, e.g., the inner diameter of the conduit may be less than about 1000 μm, less than about 500 μm, less than about 200 μm, less than about 150 μm, etc. The conduits may have an inner diameter of about 150 μm and a length of about 1.27 cm. The conduits of the stackable syringe assemblies may be substantially uniform such that the dimensions of each conduit are substantially the same as the dimensions of another conduit in the plurality of stackable syringe assemblies. A variety of materials may be used for the conduits, e.g., stainless steel.
For clarity,
These figures also show that the syringe body 302 may include a partition assembly 358 mounted to the side walls 304a, c, d of the syringe body 302 which extends into the sample chamber 306. The partition assembly 358 may be configured to reduce foaming of the fluid sample as it passes into the sample chamber 306 en route to the conduit 352. Various configurations of the partition assembly 358 may be used. In the exemplary embodiment, the partition assembly 358 may include a central portion 362 mounted to the side wall 304d of the syringe body 302 at a bottom end 364 of the central portion 362. The central portion 362 may extend upwardly such that a top end 366 of the central portion 362 is positioned at a substantially central location within the sample chamber 306. The partition assembly 358 may include first and second lateral portions 370a, b mounted to opposite sides of the top end 366 of the central portion 362. The central portion 362 and first and second lateral portions 370a, b define multiple gaps 374a-c through which the fluid sample may pass into the bore 307 and subsequently, into the conduit 352, with a reduced amount of foaming.
With reference to
With reference to
With reference to
As described above, the fluid handling system 100 may include the gas delivery system in fluid communication with the syringe stack 104. The gas delivery system may include the gas valve stack 108 which is configured to deliver gas to each sample chamber of each stackable syringe assembly. Various configurations of the gas valve stack 108 may be used. A perspective view of the exemplary gas valve stack 108 is shown in
Various configurations of the gas valve assemblies may be used. A perspective view of the exemplary gas valve assembly 502 is shown in
With reference back to
The gas valves of the gas valve assemblies may be three-way gas valves such that when the gas valve is open, the gas will continue to flow to a sample chamber of an associated stackable syringe assembly in the syringe stack 104 while the sample chamber is isolated from atmosphere. When the gas valve is closed, gas no longer flows to the sample chamber in the associated stackable syringe assembly in the syringe stack 104 and the sample chamber is vented to atmosphere. The gas valve stack 108 may be sealed via a cap 548 configured to insert into a top opening of a block 552 of an uppermost gas valve assembly 556 and to form another pressure-tight seal.
As described above, the fluid handling system 100 may include the support assembly 112 mounted to various components of the fluid handling system 100 and configured to support and to position various components of the fluid handling system 100 in certain orientations, e.g., vertically (with reference to
The support assembly 112 may include a collar 734 mounted to the base plate 702 and positioned between the first support post 706 and the second support post 710, but offset from a line connecting the first support post 706 and the second support post 710. The collar may mount to (e.g., via a fastener 742) the syringe stack support rod 738 which is inserted into a bore defined in the collar 734. As described above, the syringe stack support rod 738 may also be inserted into the bores of the tubular mounting members of the stackable syringe assemblies (as shown in
The support assembly 112 may include the holder 122 (with reference to
The components of the fluid handling system 100 may be formed from a variety of materials having sufficient strength and rigidity for the described application. With respect to the syringe bodies of the stackable syringe assemblies, materials suitable for stereolithographic fabrication or injection molding may be used, e.g., various plastics.
With reference to
The control system 800 may include an input interface 806, an output interface 808, a communication interface 810, a computer-readable medium 812, a processor 814, and a control application 816. The control system 800 may include fewer or additional components as compared to those shown in
Input interface 806 provides an interface for receiving information from the user (e.g., a selected hold time between pulses of fluid shear stress) for processing by control system 800. Although not shown, input interface 806 may further provide an interface for receiving information from gas delivery system 804 (e.g., a flowmeter output signal) for processing by control system 800. Input interface 806 may interface with various input technologies including, but not limited to, a display 818, a keyboard 820, a mouse 822, a touch screen, a track ball, a keypad, etc. to allow the user to enter information into control system 800 or to make selections presented in a user interface displayed on display 818. Display 818 may be a thin film transistor display, a light emitting diode display, a liquid crystal display, or any of a variety of different displays known to those skilled in the art. control system 800 may have one or more input interfaces that use the same or a different input interface technology.
Output interface 808 provides an interface for outputting information for review by a user of fluid handling system 802. Such information may include the open/close status of each of the gas valves of each of the gas valve assemblies as a function of time as shown in
Communication interface 810 provides an interface for receiving and transmitting data between devices using various protocols, transmission technologies, and media as known to those skilled in the art. Communication interface 810 may support communication using various transmission media that may be wired or wireless. Exemplary wireless communication devices include antennas that receive and transmit electromagnetic radiation at various frequencies. Control system 800 may have one or more communication interfaces that use the same or a different communication interface technology. Data and messages may be transferred between any input or output device and controller 800 using communication interface 810. Thus, communication interface 810 provides an alternative (or additional) interface to either or both of input interface 806 and output interface 808.
Control system 800 may be linked to one or more interfaced devices. For example, control system 800 may interface with another fluid handling system, an external computing device, an external system for analyzing certain characteristics of collected processed fluid samples. If connected, control system 800 and the one or more interfaced devices may be connected directly or through a network. The network may be any type of wired and/or wireless public or private network including a cellular network, a local area network, a wide area network such as the Internet, etc. Control system 800 may send and receive information to/from one or more of the interfaced devices. For example, control system 800 may send results obtained for the fluid sample for storage on one or more of the interfaced devices. As another example, control system 800 may receive software updates from one or more of the interfaced devices and/or receive commands from one or more of the interfaced devices. The commands may control operation of one or more components of fluid handling system 802 including control system 800. The one or more interfaced devices may include a computing device of any form factor such as a personal digital assistant, a desktop computer, a laptop computer, an integrated messaging device, a cellular telephone, a smart phone, a pager, etc. without limitation.
Computer-readable medium 812 is an electronic holding place or storage for information so that the information can be accessed by processor 814 as known to those skilled in the art. Computer-readable medium 812 can include, but is not limited to, any type of random access memory (RAM), any type of read only memory (ROM), any type of flash memory, etc. such as magnetic storage devices (e.g., hard disk, floppy disk, magnetic strips, . . . ), optical disks (e.g., CD, DVD, . . . ), smart cards, flash memory devices, etc. Control system 800 may have one or more computer-readable media that use the same or a different memory media technology. Control system 800 also may have one or more drives that support the loading of a memory media such as a CD or DVD.
Processor 814 executes instructions as known to those skilled in the art. The instructions may be carried out by a special purpose computer, logic circuits, or hardware circuits. Thus, processor 814 may be implemented in hardware, firmware, or any combination of these methods and/or in combination with software. The term “execution” is the process of running an application or the carrying out of the operation called for by an instruction. The instructions may be written using one or more programming language, scripting language, assembly language, etc. Processor 814 executes an instruction, meaning that it performs/controls the operations called for by that instruction. Processor 814 operably couples with input interface 806, with computer-readable medium 812, with communication interface 810, and with output interface 808 to receive, to send, and to process information. Processor 814 may retrieve a set of instructions from a permanent memory device and copy the instructions in an executable form to a temporary memory device that is generally some form of RAM. Control system 800 may include a plurality of processors that use the same or a different processing technology.
Control application 816 performs operations associated with controlling the operation of fluid handling system 802 and/or performs operations associated with processing output signals or input signals received by various components of fluid handling system 802. Some or all of the operations described herein may be controlled by instructions embodied in control application 816. The operations may be implemented using hardware, firmware, software, or any combination of these methods. With reference to the example embodiment of
With reference to
The following exemplary operations may be carried out when using any of the disclosed fluid handling systems based on gas pressure to apply a plurality of pulses of fluid shear stress to a fluid sample. At least some of these operations may be performed by the control system 800 (including the control application 816) or the control system 928. To begin, the fluid sample may be loaded into an uppermost stackable syringe assembly of a syringe stack via a top opening in the uppermost stackable syringe assembly. The top opening may then be sealed via a cap. The fluid sample will be contained in an uppermost sample chamber of the uppermost stackable syringe assembly.
In a first operation, pressurize the uppermost sample chamber of the uppermost stackable syringe assembly to a selected pressure while each sample chamber of each other stackable syringe assembly in the syringe stack are each vented to atmosphere. This may be accomplished by opening a gas valve of a gas valve assembly associated with the uppermost stackable syringe assembly and closing each gas valve of each other gas valve assembly. The pressurization provides a force which pushes the fluid sample from the uppermost sample chamber through a conduit of the uppermost stackable syringe assembly into a second sample chamber of a second stackable syringe assembly (i.e., the stackable syringe assembly immediately below the uppermost stackable syringe assembly), thereby exposing the fluid sample to a first pulse of fluid shear stress. The selected pressure may be that which provides a selected flow rate of the fluid sample through the conduit (for a conduit of a particular inner diameter). The selected flow rate provides a selected duration time for the first pulse of fluid shear stress (for a conduit of a particular length). Similarly, the selected flow rate provides a selected magnitude of fluid shear stress for the first pulse of fluid shear stress (flow rate and fluid shear stress are related via Poiseuille's equation as described above). An exemplary selected pressure may be about 300 psig to provide a selected flow rate of about 250 μL/sec. Other selected pressures may be that sufficient to provide a selected flow rate in the range of about 25 μL/sec to about 250 μL/sec. A gas regulator (or another similar device) in fluid communication with a gas source may be used to provide for variable control of the pressure and thus, variable flow rates.
In a second operation, maintain pressurization until a first indicator (e.g., a first flowmeter output signal) indicates a jump in gas flow corresponding to the complete delivery of the fluid sample through the conduit of the uppermost stackable syringe assembly. The fluid sample will now be in the second sample chamber of the second stackable syringe assembly.
In a third operation, vent the uppermost sample chamber of the uppermost stackable syringe assembly for a selected hold time, e.g., a few minutes. The hold time may be eliminated such that the hold time is effectively zero. This may be accomplished by closing the gas valve of the gas valve assembly associated with the uppermost stackable syringe assembly.
In a fourth operation, pressurize the uppermost sample chamber of the uppermost stackable syringe assembly and the second sample chamber of the second stackable syringe assembly to the selected pressure while each sample chamber of each other stackable syringe assembly in the syringe stack are each vented to atmosphere. The pressurization provides a force which pushes the fluid sample from the second sample chamber through a conduit of the second stackable syringe assembly into a third sample chamber of a third stackable syringe assembly (i.e., the stackable syringe assembly immediately below the second stackable syringe assembly), thereby exposing the fluid sample to a second pulse of fluid shear stress.
In a fifth operation, maintain pressurization until a second indicator (e.g., a second flowmeter output signal) indicates a jump in gas flow corresponding to the complete delivery of the fluid sample through the conduit of the second stackable syringe assembly. The fluid sample will now be in the third sample chamber of the third stackable syringe assembly.
In a sixth operation, vent sample chambers of the uppermost and the second stackable syringe assemblies for the selected hold time (or a different selected hold time).
In subsequent operations, repeat the pressurizing, maintaining pressurization and venting operations until the fluid sample has passed through each stackable syringe assembly.
In a final operation, vent each sample chamber of each stackable syringe assembly to atmosphere.
After the application of one or more pulses of fluid shear stress, the fluid sample may be referred to as “a processed fluid sample” which may be collected and analyzed via a variety of techniques, e.g., techniques for determining the concentration of viable cells in the processed fluid sample, including those described in U.S. Pat. Pub. No. 20140038231. As shown in
However, processed fluid sample which has been exposed to a smaller number of pulses of fluid shear stress may also be collected and similarly analyzed. As described above with reference to the stackable syringe assembly 106 shown in
Alternatively, as described above with reference to the stackable syringe assembly 2000 shown in
With reference to
Various configurations of the sample receptacle assembly 1112 may be used which are capable of holding a selected volume of fluid sample (e.g., 5 mL, 10 mL, etc.).
The walls of the first syringe body 1220a may define a first bore 1228a which is accessible via a first opening 1232a at a first end 1236a and a second opening 1240a at a second, opposing end 1244a. Similarly, the walls of the second syringe body 1220b define a second bore 1228b which is accessible via a third opening 1232b at a third end 1236b and a fourth opening 1240b at a fourth, opposing end 1244b. The first syringe body 1220a may be mounted to the second syringe body 1220b by shaping the first end 1236a of the first syringe body 1220a and the fourth end 1244b of the second syringe body 1220b such that the first end 1236a of the first syringe body 1220a may be inserted into the second syringe body 1220b at the fourth opening 1240b, i.e., press fitted into the second syringe body 1220b. For example, the outside diameter of the first syringe body 1220a may be reduced at the first end 1236a to form an inner tubular projection 1248 extending from the first end 1236a of the first syringe body 1220a. The inside diameter of the second syringe body 1220b may be increased by a substantially similar amount at the fourth end 1244b to form a recess into which the inner tubular projection 1248 may be inserted. The first bore 1228a and the second bore 1228b may have substantially similar diameters, thereby defining a substantially continuous bore in the sample receptacle assembly 1112. A variety of materials may be used for the first syringe body 1220a and the second syringe body 1220b, e.g., stainless steel or a plastic, e.g., Radel® by Solvay Specialty Polymers.
The plurality of substantially parallel conduits may be embedded in the conduit holding block 1252 having a first face 1256 and second face (not shown) via the plurality of substantially parallel channels 1224 formed therein. The plurality of substantially parallel conduits may be arranged in an array within the conduit holding block 1252. The dimensions of each conduit and the total number of conduits (e.g., 0.028 inches outer diameter, 0.006 inches inner diameter, 1.27 inches length) may be selected to provide a selected magnitude and selected duration time for the pulses of fluid shear stress to be applied (for a given mechanical pressure applied to the fluid sample). The conduits may be micron-sized, e.g., the inner diameter of the conduit may be less than about 1000 um, less than about 500 μm, less than about 200 μm, less than about 150 μm, etc. The conduits may have a wall thickness (e.g., 0.011 inches) sufficient to facilitate insertion into the channels 1224 of the conduit holding block 1252 while maintaining a substantially straight lumen. The conduits may be substantially uniform such that the dimensions of each conduit are substantially the same as the dimensions of another conduit in the plurality of conduits.
A variety of materials may be used for the conduits, e.g., stainless steel, plastic or glass. Commercially available conduits having such dimensions and made from such a material may be used (e.g., Hypo tubes by Micro Group). The number of conduits (e.g., 10, 20, 30, etc.) may be selected to provide a sufficient capacity for the selected volume of fluid sample to be passed through the conduits. Use of a relatively large number of conduits may be useful to ensure the continued operation of the fluid handling system 1100 even if one or a few conduits is blocked or otherwise fails. The surfaces of the conduits exposed to the fluid sample may be made substantially smooth (e.g., polishing, deburring, etc.) to facilitate the flow of the fluid sample through the conduits. A variety of materials may be used for the conduit holding block 1252, e.g., plastics including Delrin® available from DuPont. The conduits may be press fit into the channels 1224 of the conduit holding block 1252 and, optionally, adhered with an adhesive suitable for the materials used (e.g., Loctite® available from Henkel). The conduit holding block 1252 may be mounted to the first and second syringe bodies 1220a,b by inserting the conduit holding block 1252 into the first bore 1228a of the first syringe body 1220a at the first opening 1232a and inserting the inner tubular projection 1248 at the first end 1236a of the first syringe body 1220a into the second syringe body 1220b at the fourth opening 1240b. The dimensions of the conduit holding block 1252, e.g., the outer diameter, may be selected to provide a sufficiently close fit within the first bore 1228a of the first syringe body 1220a to provide a seal against the passage of the fluid sample around the outer surface of the conduit holding block 1252.
Another embodiment of a conduit holding block 2100 is shown in
Channel 2112a of the plurality of substantially parallel channels 2112 is labeled (half of channel 2112a is indicated with a dotted line) in
Various alternative embodiments may be used. For example, in some embodiments, the first funnel region 2116a does not have the cylindrical section 2122a at all, i.e., only has the conical section 2124a. In some embodiments, channels of the plurality of substantially parallel channels 2112 may each have a single funnel region (rather than two), extending from the first face 2108 of the conduit holding block 2100 towards an elongated intermediate region which extends towards the second face 2110 of the conduit holding block. The single funnel region may have various shapes (e.g., a conical shape) and various dimensions (e.g., a length of about 3 mm). The elongated intermediate region may have various dimensions (e.g., a length of about 19.05 mm).
The syringe assembly 1104 may include a first piston (not shown) mounted in the first bore 1228a of the first syringe body 1220a and a second piston (not shown) mounted in the second bore 1228b of the second syringe body 1220b. A variety of pistons may be used for the first and second pistons. For example, as shown in
With reference to
As shown in
As described further below, the sample receptacle assembly 1112 may translate back and forth along the longitudinal axis 1128 (with reference to
Since it is the relative motion between the sample receptacle assembly 1112 and the first and second pistons that is relevant, it is to be understood that the operational states shown in
With reference to
With reference to
With reference to
The linear drive assembly 1108 may include a bearing rail 1530 mounted to the base plate 1522. The linear drive assembly 1108 may include a device support assembly 1534 which may include a carriage base 1538 mounted to the bearing rail 1530. The carriage base 1538 may include side projections 1542a, b which extend from the sides of the carriage base 1538 through gaps defined between the top plate 1528 and the side walls 1510a, b, respectively, of the housing 1502. The side projections 1542a, b include horizontal support bars 1546a, b which extend upwardly from top surfaces of each side projection 1542a, b and run substantially parallel to the top plate 1518. A bore 1550 defined in the carriage base 1538 forms a lead screw interface with a lead screw (not shown) mounted between the end walls and extending substantially parallel to an axis parallel to the longitudinal axis 1128 of the syringe assembly 1104 (with reference to
The linear drive assembly 1108 may be configured to move a device, e.g., the sample receptacle assembly 1112 in other directions, in addition to linear translation along the longitudinal axis 1128 (with reference to
As shown in
Unless otherwise described, the components of the fluid handling system 1100 may be formed from a variety of materials having sufficient strength and rigidity for the described application.
With reference to
The control system 1600 may include an input interface 1606, an output interface 1608, a communication interface 1610, a computer-readable medium 1612, a processor 1614, and a control application 1616. The control system 1600 may include fewer or additional components as compared to those shown in
Input interface 1606 provides an interface for receiving information from the user for processing by control system 1600. Although not shown, input interface 1606 may further provide an interface for receiving information from the linear drive assembly 1604 for processing by control system 1600. Input interface 1606 may interface with various input technologies including, but not limited to, a display 1618, a keyboard 1620, a mouse 1622, a touch screen, a track ball, a keypad, etc. to allow the user to enter information into control system 1600 or to make selections presented in a user interface displayed on display 1618. Display 1618 may be a thin film transistor display, a light emitting diode display, a liquid crystal display, or any of a variety of different displays known to those skilled in the art. Control system 1600 may have one or more input interfaces that use the same or a different input interface technology.
Output interface 1608 provides an interface for outputting information for review by a user of fluid handling system 1602. Such information may include an output signal from a pressure sensor mounted to the fluid handling system 1602 or a voltage signal from an actuator of the linear drive assembly 1604. Monitoring such signals during the operation of the fluid handling system 1602 provides a diagnostic on the fluid sample transfer conditions which may inform the user of abnormal conditions, e.g., an undesired flow rate or conduit plugging. Output interface 1608 may further provide an interface for outputting information to the linear drive assembly 1604. Control system 1600 may have one or more output interfaces that use the same or a different interface technology.
Communication interface 1610 provides an interface for receiving and transmitting data between devices using various protocols, transmission technologies, and media as known to those skilled in the art. Communication interface 1610 may support communication using various transmission media that may be wired or wireless. Exemplary wireless communication devices include antennas that receive and transmit electromagnetic radiation at various frequencies. Control system 1600 may have one or more communication interfaces that use the same or a different communication interface technology. Data and messages may be transferred between any input or output device and controller 1600 using communication interface 1610. Thus, communication interface 1610 provides an alternative (or additional) interface to either or both of input interface 1606 and output interface 1608.
Control system 1600 may be linked to one or more interfaced devices. For example, control system 1600 may interface with another fluid handling system, an external computing device, an external system for analyzing certain characteristics of collected processed fluid samples. If connected, control system 1600 and the one or more interfaced devices may be connected directly or through a network. The network may be any type of wired and/or wireless public or private network including a cellular network, a local area network, a wide area network such as the Internet, etc. Control system 1600 may send and receive information to/from one or more of the interfaced devices. For example, control system 1600 may send results obtained for the fluid sample for storage on one or more of the interfaced devices. As another example, control system 1600 may receive software updates from one or more of the interfaced devices and/or receive commands from one or more of the interfaced devices. The commands may control operation of one or more components of fluid handling system 1602 including control system 1600. The one or more interfaced devices may include a computing device of any form factor such as a personal digital assistant, a desktop computer, a laptop computer, an integrated messaging device, a cellular telephone, a smart phone, a pager, etc. without limitation.
Computer-readable medium 1612 is an electronic holding place or storage for information so that the information can be accessed by processor 1614 as known to those skilled in the art. Computer-readable medium 1612 can include, but is not limited to, any type of random access memory (RAM), any type of read only memory (ROM), any type of flash memory, etc. such as magnetic storage devices (e.g., hard disk, floppy disk, magnetic strips, . . . ), optical disks (e.g., CD, DVD, . . . ), smart cards, flash memory devices, etc. Control system 1600 may have one or more computer-readable media that use the same or a different memory media technology. Control system 1600 also may have one or more drives that support the loading of a memory media such as a CD or DVD.
Processor 1614 executes instructions as known to those skilled in the art. The instructions may be carried out by a special purpose computer, logic circuits, or hardware circuits. Thus, processor 1614 may be implemented in hardware, firmware, or any combination of these methods and/or in combination with software. The term “execution” is the process of running an application or the carrying out of the operation called for by an instruction. The instructions may be written using one or more programming language, scripting language, assembly language, etc. Processor 1614 executes an instruction, meaning that it performs/controls the operations called for by that instruction. Processor 1614 operably couples with input interface 1606, with computer-readable medium 1612, with communication interface 1610, and with output interface 1608 to receive, to send, and to process information. Processor 1614 may retrieve a set of instructions from a permanent memory device and copy the instructions in an executable form to a temporary memory device that is generally some form of RAM. Control system 1600 may include a plurality of processors that use the same or a different processing technology.
Control application 1616 performs operations associated with controlling the operation of fluid handling system 1602 and/or performs operations associated with processing output signals or input signals received by various components of fluid handling system 1602. Some or all of the operations described herein may be controlled by instructions embodied in control application 1616. The operations may be implemented using hardware, firmware, software, or any combination of these methods. With reference to the example embodiment of
In an exemplary embodiment, the fluid handling system 1602 may be configured as shown in
The following exemplary operations may be carried out when using any of the disclosed fluid handling systems based on mechanical pressure to apply a plurality of pulses of fluid shear stress to a fluid sample. At least some of these operations may be performed by the control system 1600 (including the control application 1616). The operations will be described with respect to the fluid handling system 1100 described in
In a first operation, translate the sample receptacle assembly 1112 to an initial position. An exemplary initial position is shown in
In a second operation, translate the sample receptacle assembly 1112 from the initial position to the selected position in a first direction (e.g., −z) along the longitudinal axis 1128 (with reference to
The second operation is illustrated in
In a third operation, hold the sample receptacle assembly 1112 at the selected position for a selected hold time, e.g., 10 seconds, 30 seconds, etc. The hold time may be eliminated such that the hold time is effectively zero.
In a fourth operation, translate the sample receptacle assembly 1112 from the selected position back to the initial position in an opposing direction (e.g., +z) along the longitudinal axis 1128 at the selected speed (or a different selected speed) (with reference to
In a fifth operation, hold the sample receptacle assembly 1112 at the initial position for the selected hold time (or a different selected hold time).
In subsequent operations, repeat the translating and holding operations until the fluid sample has passed through the plurality of substantially parallel conduits a selected total number of times (e.g., 5, 10, 15, etc.).
The exemplary operations above refer to translating the sample receptacle assembly 1112 relative to the first and second pistons. However, as described above, such operations may alternatively involve translating the first and second pistons relative to the sample receptacle assembly 1112.
After the application of one or more plurality of pulses of fluid shear stress, the fluid sample may be referred to as “a processed fluid sample” which may be collected and analyzed via a variety of techniques, e.g., techniques for determining the concentration of viable cells in the processed fluid sample, including those described in U.S. Pat. Pub. No. 20140038231. As shown in
It is to be understood that the use of the phrases “syringe body” and “bore” and the like in this disclosure is not limited to structures having circularly-shaped cross-sections, although such structures may be used.
A fluid handling system similar to those described in the section “Fluid Handling System Based on Gas Pressure” was used to apply pluralities of pulses of fluid shear stress to fluid samples which included cancerous cells, e.g., PC-3 cells. The fluid samples were prepared according to methods as described in U.S. Pat. Pub. No. 20140038231. The fluid samples were loaded onto the fluid handling system and processed using the operations described in “Fluid Handling System Based on Gas Pressure” to provide processed fluid samples.
The word “exemplary” is used herein to mean serving as an example, instance, or illustration. Any aspect or design described herein as “exemplary” is not necessarily to be construed as preferred or advantageous over other aspects or designs. Further, for the purposes of this disclosure and unless otherwise specified, “a” or “an” means “one or more”.
The foregoing description of exemplary embodiments of the invention has been presented for purposes of illustration and of description. It is not intended to be exhaustive or to limit the invention to the precise form disclosed, and modifications and variations are possible in light of the above teachings or may be acquired from practice of the invention. The embodiments were chosen and described in order to explain the principles of the invention and as practical applications of the invention to enable one skilled in the art to utilize the invention in various embodiments and with various modifications as suited to the particular use contemplated. It is intended that the scope of the invention be defined by the claims appended hereto and their equivalents.
The present application claims priority to U.S. Provisional Patent Application No. 62/073,142 that was filed Oct. 31, 2014, the entire contents of which are hereby incorporated by reference.
| Filing Document | Filing Date | Country | Kind |
|---|---|---|---|
| PCT/US2015/058255 | 10/30/2015 | WO | 00 |
| Publishing Document | Publishing Date | Country | Kind |
|---|---|---|---|
| WO2016/070007 | 5/6/2016 | WO | A |
| Number | Name | Date | Kind |
|---|---|---|---|
| 20010052460 | Chien | Dec 2001 | A1 |
| 20020006359 | Mathies | Jan 2002 | A1 |
| 20020036018 | McNeely | Mar 2002 | A1 |
| 20030041652 | Spaid | Mar 2003 | A1 |
| 20030221996 | Svoronos et al. | Dec 2003 | A1 |
| 20040142408 | Kirk et al. | Jul 2004 | A1 |
| 20040182788 | Dorian | Sep 2004 | A1 |
| 20040213699 | Berndtsson | Oct 2004 | A1 |
| 20070269355 | Malmqvist | Nov 2007 | A1 |
| 20080269076 | Ermakov | Oct 2008 | A1 |
| 20080314454 | Delattre | Dec 2008 | A1 |
| 20090298067 | Irimia et al. | Dec 2009 | A1 |
| 20100041128 | Banes et al. | Feb 2010 | A1 |
| 20130089869 | Blobe et al. | Apr 2013 | A1 |
| 20130236879 | Berry et al. | Sep 2013 | A1 |
| 20140087412 | Fouras et al. | Mar 2014 | A1 |
| 20140093891 | Kern | Apr 2014 | A1 |
| 20140274739 | Rinker et al. | Sep 2014 | A1 |
| 20170153217 | Johnston | Jun 2017 | A1 |
| Number | Date | Country |
|---|---|---|
| 2012129360 | Sep 2012 | WO |
| 2013086509 | Jun 2013 | WO |
| 2014070776 | May 2014 | WO |
| Entry |
|---|
| Barnes, et al., “Resistance to fluid shear stress is a conserved biophysical property of malignant cells.”, PLoS One 7 (12), 1-12 (2012). |
| Chen, et al., “Cardiac-like flow generator for long-term imaging of endothelial cell responses to circulatory pulsatile flow at microscale”, Lab on a Chip 13(15), 2999-3007 (2013). |
| Lu, et al., “Microfluidic Shear Devices for Quantitative Analysis of Cell Adhesion”, Anal Chem 76(18), 5257-5264 (2004). |
| Nagrath, et al., “Isolation of rare circulating tumour cells in cancer patients by microchip technology”, Nature 450 (7173), 1235-1239 (2007). |
| Patent Cooperation Treaty, International Searching Authority, Search Report and Written Opinion for PCT/US2015/058255, 9 pages, dated Feb. 15, 2016. |
| Number | Date | Country | |
|---|---|---|---|
| 20170333896 A1 | Nov 2017 | US |
| Number | Date | Country | |
|---|---|---|---|
| 62073142 | Oct 2014 | US |
