Disposable system for analysis of haemostatic function

Abstract

To provide devices, systems, and methods for assessing hemostasis in a subject by preparation and analysis of a test sample from an elephant.SOLUTION: In some embodiments, the disposable system includes a multi-channel or multi-chamber test cartridge device configured to operate with a test system for evaluation of hemostasis in a subject by in vitro evaluation of a test sample from the subject. In some embodiments, the disposable system is configured to interrogate a test sample to evaluate the clot hardness, strength, or other mechanical properties of the test sample to evaluate the function of various physiological processes during clotting and / or dissolution of the resulting clot.SELECTED DRAWING: Figure 17

Description

Detailed Description of the Invention

[0001] Government Licensing Rights This invention was made with government support under grant R44HL103030 awarded by the National Heart Lung and Blood Institute. The government has certain rights in this invention. [Technical field]

[0002] CROSS-REFERENCE TO RELATED APPLICATIONS This application is filed under the title "Disposable System for Analysis of This application claims priority to and benefit of U.S. Provisional Application No. 62 / 488,045, filed April 20, 2017, entitled "Hemostatic Function," which is incorporated herein by reference in its entirety.

[0003] The present application relates to devices, systems, and methods for assessing hemostasis in a subject by preparation and analysis of a test sample from the subject. [Background technology]

[0004] Hemostasis, the physiological control of bleeding, is a complex process incorporating vascular structures, platelets, coagulation factors, fibrinolytic proteins, and various activators and inhibitors. Disruption of hemostasis plays a central role in the development of myocardial infarction, stroke, pulmonary embolism, deep vein thrombosis, and excessive bleeding. Consequently, in vitro diagnostics (IVDs) are critically needed to quantify hemostatic function / dysfunction and direct appropriate treatment.

[0005] The process of coagulation is highly dependent, among other things, on the temperature it occurs at. Under normal conditions, coagulation occurs at body temperature, which is optimal for proper enzymatic action of the clotting factors in the cascade.

[0006] The preparation of the blood to be tested is also important, as the way in which the blood sample is prepared prior to its evaluation can affect, for example, the action of vasculature components, platelets and other cellular components, coagulation factors, fibrinolytic components, and any inhibitors or activators of hemostasis. [Summary of the Invention]

[0007] Devices, systems, and methods for the evaluation of hemostasis are provided. For example, a disposable system for the analysis of hemostatic function is provided. In some embodiments, the disposable system includes a multi-channel or multi-chamber test cartridge device configured to operate with a test system for the evaluation of hemostasis in a subject by in vitro evaluation of a test sample from the subject. In some embodiments, the disposable system is configured to interrogate the test sample to evaluate clot firmness, strength, or other mechanical properties of the test sample to evaluate the function of various physiological processes during the clotting and / or lysis of the resulting clot. The sample can comprise, in whole or in part, whole blood, plasma, platelet-rich plasma, or platelet-poor plasma. Additionally, the sample can include one or more reagents (such as anticoagulants or antiplatelet agents that may be present in the blood when collected), or one or more pharmacological treatments (such as in the case of heparin or low molecular weight heparin), or other inert components (such as polystyrene beads) that are added to the test sample before the cartridge device is used. The disposable system is robust (e.g., can be performed in a non-laboratory environment), rapid (e.g., taking only minutes to perform), easy to use, and provides unambiguous results. The present invention facilitates point-of-care assessment of hemostasis of a test sample (e.g., characterized for functional components of hemostasis) and facilitates accurate identification of hemostasis defects. The illustrated devices automate one or more pre-measurement steps that minimize sample manipulation steps required by the user, thereby improving test reproducibility and / or test quality. In some embodiments, a disposable system includes multiple test circuits, each having a pathway defined by channels and chambers configured to prepare a test sample of blood for evaluation by the measurement device. In each test circuit, a portion of the test sample is introduced to a reagent or combination of reagents specific to that test circuit.

[0008] In some embodiments, the disposable system is configured to condition each test sample before, during, and / or after mixing with the reagent(s) to optimize the proper action of the applicable blood constituents and chemicals being evaluated (e.g., vasculature constituents, platelets or other cellular constituents, coagulation factors, fibrinolytic constituents, and any other inhibitors or activators of hemostatic function, etc.).

[0009] In one aspect, a device (eg, a cartridge) for assessment of hemostasis is disclosed. The device includes a housing, an input port integrally formed with the housing and structurally configured to establish fluid communication and eject the contents of a sample holding tube, and a first chamber in fluid communication with the input port, configured to receive a sample contained in the sample holding tube and to condition the received sample to a desired temperature (e.g., a predefined temperature range) before allowing the received sample to contact one or more reagents located in one or more fluid circuits downstream of the first chamber, each of the one or more fluid circuits comprising: i) a second chamber in fluid communication with the first chamber that meters the sample in the first chamber into an aliquot, wherein the metered sample is introduced into a reagent or combination of reagents (e.g., in the form of lyophilized reagent beads) located in a corresponding fluid circuit (e.g., a reagent pocket) to form a mixed sample; and ii) a test chamber in fluid communication with the second chamber, structurally configured for interrogation by a measurement system configured to determine properties (e.g., mechanical or viscoelastic properties) of the mixed sample.

[0010] In some embodiments, at least one of the one or more fluid circuits comprises one or more pockets (e.g., each configured to contain lyophilized reagent beads containing a reagent or combination of reagents).

[0011] In some embodiments, at least one of the one or more fluid circuits comprises one or more liquid holding pockets (e.g., each configured to contain an analytical sample in liquid form containing a reagent or combination of reagents).

[0012] In some embodiments, at least one of the one or more fluidic circuits comprises one or more lyophilized reagents located on one or more surfaces thereof (e.g., lyophilized onto each of the surfaces; lyophilized as a film disposed on or attached to one or more of the surfaces).

[0013] In some embodiments, at least one of the one or more fluid circuits includes one or more reagents fabricated onto its surface (e.g., dried onto the surface, spray coated onto the surface, baked onto the surface).

[0014] In some embodiments, the input port is communicatively coupled to a pressure port, and pressure applied to the pressure port causes the contents of the sample holding tube to be ejected through the input port into the first chamber.

[0015] In some embodiments, the input port comprises a needle assembly.

[0016] In some embodiments, the needle assembly includes an input port and a second port, the second port configured to release a liquid or gas into the sample holding tube to facilitate ejection of contents therein, hi some embodiments, the input port is located within (e.g., concentrically located with) the second port configured to release a liquid or gas into the sample holding tube to cause ejection of contents therein.

[0017] In some embodiments, the input port comprises a luer lock configured to connect to a sample holding tube, the sample holding tube being a syringe.

[0018] In some embodiments, the input port is communicatively coupled to a first pressure port, and pressure applied to the first pressure port causes the contents of the sample holding tube to be ejected through the input port into the first chamber.

[0019] In some embodiments, the first chamber is configured to mate with a corresponding thermal conditioning system (e.g., a heating / cooling system) of the measurement system to condition the received sample at or near a desired temperature.

[0020] In some embodiments, the shape and / or materials of the first chamber are optimized to facilitate thermal regulation (eg, heating and / or cooling) of the sample to or near a desired temperature.

[0021] In some embodiments, the first chamber is configured to mate with a corresponding thermal regulation surface of a subsystem component of the measurement system to condition the received sample at or near a desired temperature. In some embodiments, the channel portion of the one or more fluid circuits is configured to mate with a corresponding heating / cooling system of the measurement system to condition the received sample at or near a desired temperature.

[0022] In some embodiments, the one or more fluidic circuit channel portions are configured to mate with a corresponding thermal regulation system of the measurement system to regulate a received sample to a desired temperature. In some embodiments, the first chamber and / or the one or more fluidic circuit channel portions are in physical proximity (e.g., in physical contact or near physical contact) with a sensor configured to measure the temperature of a sample received in the first chamber.

[0023] In some embodiments, the sensor is selected from the group consisting of a thermistor, a thermocouple, and an optical sensor (eg, an IR sensor).

[0024] In some embodiments, the device includes a first pressure port in fluid communication with the first chamber, the first pressure port configured to receive a negative or differential pressure (e.g., to fill the first chamber), and a filter positioned within the first pressure port in at least one of the fluid circuits (e.g., such that the filter is clogged by sample received in the first chamber when the first chamber is full). In some embodiments, the filter is configured to allow air to move through the first pressure port but prevent fluid from moving therethrough.

[0025] In some embodiments, the device includes a first pressure port configured to receive a negative or differential pressure to fill the first chamber, and a first fluid pathway extending from the first pressure port to the first chamber, wherein the filter is positioned within the first pressure port. do.

[0026] In some embodiments, for each of the one or more fluid circuits, fluid communication between the first chamber and the second chamber is through a second fluid path that originates from a side (e.g., a side wall, a bottom wall, etc.) of the first chamber (e.g., such that bubbles present in the received sample are trapped away from the second chamber).

[0027] In some embodiments, each of the one or more fluid circuits includes a third fluid path in fluid communication with the second chamber, the third fluid path leading to a second pressure port configured to receive a negative or differential pressure to fill the second chamber.

[0028] In some embodiments, the second pressure port has a second filter therein, the second filter being configured to clog when the second chamber is filled.

[0029] In some embodiments, the device includes one or more fluid paths in fluid communication with the second pressure port for all of the one or more fluid circuits, the one or more fluid paths configured to provide a negative pressure to the second pressure port for all of the one or more fluid circuits.

[0030] In some embodiments, for each of the one or more fluid circuits, the second chamber is in fluid communication with an ejection port, the ejection port being configured to be closed while the sample is being metered into aliquots in the second chamber and further configured to be open to atmospheric pressure after the sample is being metered into aliquots in the second chamber.

[0031] In some embodiments, each of the one or more fluid circuits includes a third set of fluid paths in fluid communication between a respective second chamber (e.g., a metering chamber) and the test chamber, and a portion of the third set of fluid paths is arranged as a serpentine-shaped conduit or channel.

[0032] In some embodiments, each of the one or more fluid circuits further comprises a serpentine reservoir between the test chamber and the second chamber.

[0033] In some embodiments, the metered sample is selectively directed through one or more portions of the fluid circuit to promote mixing of the metered sample with a reagent or combination of reagents.

[0034] In some embodiments, the metered sample is selectively and multiplicatively directed between a first location in the fluid circuit (e.g., a second chamber) and a second location in the fluid circuit (e.g., a location within a serpentine reservoir) for each of one or more fluid circuits.

[0035] In some embodiments, each of the one or more fluid circuits further comprises a third pressure port in fluid communication with the second chamber and the test chamber, the third pressure port configured to receive a negative or differential pressure (e.g., to draw an aliquot from the second chamber into the test chamber), and the third pressure port further configured to receive alternating pressure, e.g., to alternately draw an aliquot from the second chamber along the serpentine reservoir and push the aliquot through the serpentine reservoir into the second chamber.

[0036] In some embodiments, the serpentine reservoir includes an optical detection zone to facilitate optical detection of a metered sample within the serpentine reservoir or the location of a sample within the serpentine reservoir.

[0037] In some embodiments, each of the one or more fluid circuits includes a test chamber and a second chamber. The mixing path further comprises a mixing channel between the mixing channel and the bar, the mixing channel comprising one or more ferromagnetic beads or rods therein.

[0038] In some embodiments, at least one of the one or more fluid circuits comprises one or more quality test portals.

[0039] In some embodiments, the one or more quality test portals are configured to be optically sensed and the quality test ports are transparent.

[0040] In some embodiments, the one or more quality test portals are configured to be electrically sensed, and the quality test port comprises one or more sensing electrodes.

[0041] In some embodiments, one or more quality test ports are configured to be sampled for a characteristic of the metered sample (eg, for pressure, presence of flow, flow rate, temperature).

[0042] In some embodiments, for each of one or more fluid circuits, the test chamber includes a mechanism for coupling energy into the test chamber to perform measurements, such as in the case of a lens configured to direct ultrasonic pulses into the test chamber.

[0043] In another aspect, a device for assessing hemostasis is disclosed, the device comprising: a housing; an input port integrally formed with the housing, the input port establishing fluid communication with a sample-holding tube and structurally capable of evacuating the contents of the sample-holding tube; a first chamber in fluid communication with the input port, the first chamber receiving a sample contained in a vacuum tube, whereby the sample temperature is adjusted to a desired temperature before the sample contacts one or more reagents; one or more second chambers in fluid communication with the first chamber, the one or more second chambers configured to meter the sample in the first chamber into one or more aliquots; one or more reagent pockets, each filled with one or more lyophilized reagent beads, the one or more reagent pockets in fluid communication with each of the aliquot chambers and allowing the sample present in each aliquot to be mixed with said one or more reagent beads; and one or more test chambers in fluid communication with the aliquot chambers and structurally capable of being interrogated to determine viscoelastic properties of such sample after it has been mixed with one or more reagents.

[0044] In some embodiments, a reagent or combination of reagents located in one or more fluid circuits includes, or in combination with, an intrinsic pathway activator (e.g., kaolin, celite, glass, ellagic acid, micronized silica, Hageman factor, etc.).

[0045] In some embodiments, a reagent or combination of reagents located in one or more fluid circuits includes, or in combination with, an extrinsic pathway activator (e.g., tissue factor, recombinant tissue factor, thromboplastin, etc.).

[0046] In some embodiments, a reagent or combination of reagents located in one or more fluid circuits includes a coagulation activator (e.g., thrombin, factor Xa, reptilase, ecarin, Russell's viper venom, or other snake venom, etc.), or a combination thereof.

[0047] In some embodiments, a reagent or combination of reagents located in one or more fluid circuits is a platelet activator or platelet inhibitor (e.g., a GPIIb / IIIa inhibitor (e.g., abciximab, eptifibatide, tirofiban, roxifiban, orbofiban), cytochalasin D, blebbistatin, PAR1 inhibitors, PAR4 inhibitors, glycoprotein IB inhibitors, TRAP, ADP, arachidonic acid, ADP inhibitors, nonsteroidal anti-inflammatory drugs, platelet-activating factor, ristocetin, epinephrine, etc.), or combinations thereof.

[0048] In some embodiments, a reagent or combination of reagents located in one or more fluid circuits comprises a fibrinolytic function activator or inhibitor (e.g., tPA, uKA, streptokinase, TAFIa, plasmin / plasminogen, aprotinin, epsilon-aminocaproic acid, tranexamic acid, plasminogen activator inhibitor 1 (PAI1), α2-antiplasmin (α2-AP), or plasmin-antiplasmin complex, carboxypeptidase inhibitor), or combination thereof.

[0049] In some embodiments, a reagent or combination of reagents located in one or more of the fluid circuits comprises or in combination with an FXIIIa inhibitor.

[0050] In some embodiments, a reagent or combination of reagents located in one or more of the fluid circuits comprises thrombomodulin or a combination thereof.

[0051] In some embodiments, a reagent or combination of reagents located in one or more of the fluid circuits comprises low molecular weight heparin or a combination thereof.

[0052] In some embodiments, a reagent or combination of reagents located in one or more fluid circuits comprises hexadimethrine bromide (polybrene) or a combination thereof.

[0053] In some embodiments, a reagent or combination of reagents located in one or more of the fluid circuits includes heparin or a combination thereof.

[0054] In some embodiments, a reagent or combination of reagents located in one or more of the fluid circuits comprises a corn trypsin inhibitor or a combination thereof.

[0055] In some embodiments, a reagent or combination of reagents located in one or more of the fluid circuits includes adenosine or a combination thereof.

[0056] In some embodiments, a reagent or combination of reagents located in one or more of the fluid circuits comprises GPRP (Gly-Pro-Arg-Pro) or a combination thereof.

[0057] In some embodiments, a reagent or combination of reagents located in one or more of the fluid circuits includes calcium or a combination thereof.

[0058] In some embodiments, a reagent or combination of reagents located in one or more of the fluid circuits comprises fibronectin or a combination thereof.

[0059] In some embodiments, a reagent or combination of reagents located in one or more of the fluid circuits comprises collagen or a combination thereof.

[0060] In some embodiments, a reagent or combination of reagents located in one or more of the fluid circuits comprises or in combination with an immunodetection reagent.

[0061] In some embodiments, a reagent or combination of reagents located in one or more of the fluid circuits comprises heparinase I or a combination thereof.

[0062] In some embodiments, a reagent or combination of reagents located in one or more fluid circuits comprises endothelial cells or activated endothelial cells.

[0063] In some embodiments, the measurement system is selected from the group consisting of a sonorheometry-based system, a thromboelastography-based system, a thromboelastometry-based system, an optical-based system, a fluorescence-based system, a colorimetric-based system, an aggregometry-based system, a resonance-based system, and an electrical impedance-based system.

[0064] In another aspect, a method is disclosed for mixing a sample with one or more reagents in a device (e.g., cartridge) and testing the mixed sample for assessment of hemostasis. The method includes receiving a plurality of metered samples from a plurality of metering chambers that have received test fluid from sample-holding tubes (e.g., via a mechanical linkage connecting the device to the sample-holding tubes or via an opening into which the sample from the sample-holding tubes is disposed), and flowing each of the aliquots alternately and multiplicatively until the aliquot is mixed with the reagent or combination of reagents to form a mixed aliquot, wherein at least one aliquot: i) flows a small portion of the aliquot in a first direction from the metering chamber, through one or more reagent pockets having one or more reagents (e.g., lyophilized reagent beads) therein, and along a serpentine path in communication with the metering chamber; and ii) alternately and periodically flowing the mixed aliquot from the detection zone through at least a portion of the serpentine path toward a metering chamber until the mixed aliquot reaches a detection zone located within or after the serpentine path, and ii) from the detection zone in a second direction opposite to the first direction, through at least a portion of the serpentine path toward a metering chamber until a trigger event; and driving the mixed aliquot into a test chamber in fluid communication with the metering chamber, the test chamber structurally configured for interrogation by a measurement system configured to determine a property (e.g., a mechanical property or a viscoelastic property) of the mixed aliquot, the interrogation of the test chamber being performed with the mixed aliquot located therein.

[0065] In some embodiments, the method includes receiving a fluid in a first chamber configured to substantially adjust the temperature of the test sample toward body temperature or other desired temperature, and a metered sample received in a metering chamber is received from the first chamber.

[0066] In some embodiments, the test fluid moves to the first chamber in response to an applied pressure applied by or generated from the measurement system.

[0067] In some embodiments, the method includes adjusting the test fluid in the first chamber to or substantially near a desired temperature, and the test fluid is mixed with one or more reagents after exiting the first chamber.

[0068] In some embodiments, the method includes isolating (e.g., blocking via a valve) the test fluid in the metering chamber to prevent the test fluid from contacting the one or more reagents during filling of the metering chamber.

[0069] In some embodiments, a second pressurized positive or negative pressure is applied by or generated from the measurement system at a second port in communication with the tortuous path (e.g., applied at a second pressure port in communication therewith) to move at least one aliquot in a second direction.

[0070] In some embodiments, the first applied positive or negative pressure is inversely applied by or generated from the measurement system to move at least one aliquot in a second direction.

[0071] In some embodiments, the act of receiving the mixed aliquot into the test chamber includes a third Further comprising receiving a negative pressure through the pressure port, the third pressure port further being in fluid communication with the test chamber.

[0072] In some embodiments, the test chamber is downstream of the serpentine path and the third pressure port is downstream of the test chamber.

[0073] These and other features and advantages of the present invention will become readily apparent to those skilled in the art in view of the following detailed description and accompanying drawings which describe both preferred and alternative embodiments of the invention. [Brief explanation of the drawings]

[0074] The accompanying drawings, which are incorporated in and constitute a part of this specification, illustrate embodiments and, together with the description, serve to explain the principles of the method and system. [Figure 1] 1 illustrates a perspective view of an exemplary biological sample input device of a cartridge for use in a disposable system, according to an exemplary embodiment. [Figure 2] 2 illustrates a side cross-sectional view of the exemplary biological sample input device of FIG. 1 with a casing, according to an exemplary embodiment. [Figure 3] 3 illustrates a side cross-sectional view of the exemplary biological sample input device of FIG. 2 with a sample-holding tube mounted thereon, according to an exemplary embodiment. [Figure 4] 4 illustrates a detailed view of the exemplary biological sample input device of FIG. 3, according to an exemplary embodiment. [Figure 5A] 1 illustrates the biological fluid pathways of four test circuits (e.g., hemostasis test circuits) located on the sample preparation surface, according to an exemplary embodiment. [Figure 5B] 5B shows the biological fluid pathways of four test circuits (e.g., hemostasis test circuits) located on the sample preparation surface according to an exemplary embodiment, and further shows the cartridge body of FIG. 5A further connected to a sample holding tube according to an exemplary embodiment. [Figure 6A] 5A, 5B, and 8 with corresponding labels for heating chamber filling. [Figure 6B] 5A, 5B, and 8 with corresponding labels for heating chamber filling. [Figure 6C]5A, 5B, and 8 with corresponding labels for sample chamber filling. [Figure 6D] 5A, 5B, and 8 with corresponding labels for sample chamber filling. [Figure 6E] 5A, 5B, and 8 with labels corresponding to sample mixing and test chamber filling. [Figure 6F] 5A, 5B, and 8 with labels corresponding to sample mixing and test chamber filling. [Figure 7] 5B illustrates the backside of the cartridge of FIG. 5A according to an exemplary embodiment, including interconnecting surfaces that abut the sample preparation surfaces, which collectively form a biological fluid pathway for the test circuit. [Figure 8] 8 illustrates a portion of a biological fluid pathway on the interconnection surface of FIG. 7, according to an exemplary embodiment. [Figure 9] 1 illustrates an exemplary test chamber section for use with an exemplary cartridge, according to an exemplary embodiment. [Figure 10] 1 illustrates a cross-sectional view of an exemplary test chamber within an exemplary test chamber section, according to an exemplary embodiment. [Figure 11] 11 illustrates a detailed cross-sectional view of the exemplary test chamber of FIG. 10 according to an exemplary embodiment. [Figure 12] 1 illustrates a cross-sectional view of a disposable system operably coupled to a measurement system, according to an exemplary embodiment. [Figure 13] 1 illustrates an exemplary shear modulus versus time curve according to an exemplary embodiment. [Figure 14] An example of shear modulus curves obtained with an activator of coagulation and with and without a fibrinolytic inhibitor is shown. A differential comparison of these curves can provide information about the fibrinolytic activity of the sample. [Figure 15] 1 illustrates potential embodiments of difference criteria that can be measured from shear modulus curves obtained with an activator of coagulation and with and without a fibrinolysis inhibitor. [Figure 16]10 shows a photograph of the exemplary cartridge of FIGS. 1, 2, 3, 4, 5A, 5B, 7, 8, and 9 for use in a disposable system, according to an exemplary embodiment. [Figure 17] 10 illustrates a front view of the exemplary cartridge of FIGS. 1, 2, 3, 4, 5A, 5B, 7, 8, and 9 according to an exemplary embodiment. [Figure 18] 10 illustrates a front view of the exemplary cartridge of FIGS. 1, 2, 3, 4, 5A, 5B, 7, 8, and 9 according to an exemplary embodiment. DETAILED DESCRIPTION OF THE INVENTION

[0075] The present invention will now be described in more detail below with reference to specific embodiments thereof. Indeed, the invention may be embodied in many different forms and should not be construed as limited to the embodiments set forth herein; rather, these embodiments are provided so that this disclosure will satisfy applicable legal requirements.

[0076] As used in this specification and the appended claims, the singular forms "a," "an," and "the" include plural referents unless the context clearly dictates otherwise.

[0077] As used herein, the term "comprises" and variations thereof are used synonymously with the term "includes" and variations thereof and are open, non-limiting terms.

[0078] As used throughout, "subject" refers to an individual. A subject may be a vertebrate, more specifically a mammal (e.g., a human, horse, pig, rabbit, dog, sheep, goat, non-human primate, cow, cat, guinea pig, or rodent), fish, bird, or reptile or amphibian. The term does not denote a particular age or sex.

[0079] The devices described herein include single-use cartridge devices configured to facilitate in vitro assessment of one or more hemostatic functions. Hemostatic function refers to the functional role of various blood components, such as clotting factors, fibrinogen, platelets, fibrinolytic factors, and vasculature components. In some embodiments, the cartridge device and associated measurement system are configured to assess hemostatic function by measuring a change in at least one mechanical property of a sample being tested when the sample is exposed to one or more reagents. In some embodiments, the cartridge device and its test chamber are configured to facilitate measurement of viscoelastic properties, for example, based on probing using ultrasonic pulses or ultrasonic energy. However, other probing systems may be used with cartridge devices having the features described herein. Examples of other probing systems include, but are not limited to, systems using cup / pin technology (such as in thromboelastography and thromboelastometry), an oscillating piston for measuring changes in mechanical impedance, optical sensing, fluorescence sensing, colorimetric sensing, aggregometry, resonance sensing, or electrical impedance sensing, among others.

[0080] A wide range of reagents may be utilized in the cartridge device, including intrinsic pathway activators (including, but not limited to, kaolin, Hageman factor, celite, glass, ellagic acid, micronized silica, etc.), extrinsic pathway activators (including, but not limited to, tissue factor, recombinant tissue factor, thromboplastin, etc.), other coagulation activators (including, but not limited to, thrombin, factor Xa, reptilase, ecarin, Russell's viper venom, or other snake venom), platelet activators, or platelet inhibitors (including, but not limited to, GPIIb / IIIa inhibitors (abciximab, eptifibatide, tirofiban, roxifiban, orbofiban, etc.), cytochalasin D, blebbistatin, PAR1 inhibitors, PAR4 inhibitors, glycoprotein IB inhibitors, TRAP, ADP, arachidonic acid, ADP inhibitors, nonsteroidal anti-inflammatory drugs, etc.), fibrinolytic function activators or fibrinolytic function inhibitors (including, but not limited to, tPA, uKA, streptokinase, TAFIa, plasmin / plasminogen, aprotinin, epsilon-aminocaproic acid, tranexamic acid, plasminogen activator inhibitor 1 (PAI1) ), α2-antiplasmin (α2-AP), or plasmin-antiplasmin complex, carboxypeptidase inhibitors, etc.), as well as others (FXIIIa inhibitors, hexadimethrine bromide (polybrene), heparinases (e.g., heparinase I), ristocetin, heparin, low molecular weight heparin, corn trypsin inhibitor, adenosine, GPRP, calcium, fibronectin, collagen, epinephrine, immunodetection reagents, direct thrombin inhibitors, factor Xa inhibitors, reagents aimed at reversing or eliminating the effects of new oral anticoagulants (such as direct thrombin inhibitors and factor Xa inhibitors), thrombomodulin, etc. Additional non-functional reagents may also be used to preserve the functionality of other reagents (buffers and stabilizers for lyophilization or desiccation, dyes, etc.).

[0081] In some embodiments, the reagents are disposed and stored within chambers (e.g., pockets located within the fluidic circuit) of the cartridge device, although in alternative embodiments, the reagents may be disposed and stored within various chambers or fluidic channels within the fluidic circuit of the cartridge device. A fluidic circuit generally refers to one or more fluid pathways established between a sample preparation and one or more test chambers where the sample is ultimately measured.

[0082] In some embodiments, the reagents are placed and stored in the cartridge device in liquid form, or can be lyophilized in spheres (such as in the case of Lyopheres™ from BioLyph LLC), lyophilized in films, lyophilized onto plastic surfaces, dried onto plastic surfaces, or spray coated to improve shelf-life stability. Those skilled in the art will recognize that these reagents are not all inclusive and that other reagents or combinations of reagents that are inhibitors or activators of one or more hemostatic functions can be used in this cartridge.

[0083] The cartridge device disclosed herein is one component of a measurement system (e.g., a hemostasis measurement system). The measurement system (also referred to as an instrument) includes at least an interface element that couples the cartridge device with a measurement element configured to measure viscoelastic or mechanical properties of a sample processed within the cartridge device. The measured viscoelastic or mechanical properties are output as results to a user interface. An exemplary user interface is described in commonly assigned U.S. Publication No. 2011 / 0252352 (Viola et al.), which is incorporated herein by reference in its entirety.

[0084] In some embodiments, the interface element includes one or more heating and / or cooling elements.

[0085] In some embodiments, the interface element includes a fluid manifold that facilitates connection to one or more pump elements and one or more valves.

[0086] In some embodiments, the interface element includes one or more sensors configured to, for example, perform hemostasis measurements. In some embodiments, the one or more sensors include ultrasound sensors. In other embodiments, the one or more sensors include sensors for thromboelastography, thromboelastometry (e.g., thromboelastography-based systems or These include other investigative devices that measure changes in mechanical impedance, changes in perturbations as observed via a thromboelastometry-based system), or optical-based systems (e.g., having an optical sensor), fluorescence, colorimetric-based systems, aggregometry-based systems (e.g., having an optical sensor, acoustic sensor, or electrode that measures agglutination using a test sample), resonance-based systems (e.g., having an optical, acoustic, or mechanical position sensor that measures the sample when it is at or near resonance), electrical impedance-based systems (e.g., having electrodes configured to measure electrical impedance), or combinations thereof.

[0087] In some embodiments, the interface element includes a mechanical clamp configured to position the cartridge device in a desired orientation relative to components of the measurement system (such as one or more sensors, a fluid manifold, heating and / or cooling elements). When the interface element interfaces with the components of the measurement system, the cartridge device, in some embodiments, is driven through a controlled series of operations orchestrated by the measurement system to prepare the test sample for measurement. In some embodiments, the preparation operations include sample aspiration from a sample container (also referred to as a sample holding tube), sample heating and / or cooling, sample metering, sample mixing with reagents, and sample measurement. Each step for various embodiments is described below. After the measurement is completed, the results are output at the instrument user interface.

[0088] In some embodiments, the cartridge device and its internal components are the only components in direct contact with the sample being analyzed.

[0089] In some embodiments, the cartridge includes computer-readable information (e.g., an RFID tag, a computer-readable medium such as a flash IC, a QR code, a barcode, etc.) and / or human-readable information (e.g., a label) that can be optically or communicatively interrogated.

[0090] The various embodiments described below do not utilize any variable valve elements in the cartridge design, but instead rely on a fluid manifold and one or more valves located within the instrument. Fluid moves through the various cartridge components via pressure differentials, gravity, material properties (such as hydrophobic or hydrophilic), and / or capillary forces.

[0091] In these embodiments, the cartridge is configured to couple with an instrument via one or more connection ports aligned via alignment slots. The connection ports include one or more pressure ports and one or more release ports. However, in alternative embodiments, actuated valves (such as in the case of elastomeric valves) may be included within the cartridge design to control fluid flow. These valves, in some embodiments, are actuated by corresponding hardware and software components of the measurement system.

[0092] The surface properties and texture of cartridge surfaces in direct contact with the sample can be optimized to promote sample attachment and / or sample flow. In some embodiments, the interior surfaces of the test chamber and / or other interior surfaces of the fluidic circuitry within the cartridge device are plasma-treated to optimize the surface energy and texture for attachment of specific plasma proteins. In other embodiments, the interior surfaces of the test chamber and / or other interior surfaces of the fluidic circuitry are treated with surface roughness texturing, material coating (such as in the case of gold plating), biomaterial coating (e.g., in the case of fibronectin or collagen coating), raw material selection (e.g., the use of specific plastics or other materials for plating that do not require additional processing), etc. Such treatments can be used independently or in conjunction with plasma treatment. Similarly, cartridge materials can be selected or engineered to achieve a desired hydrophobic or hydrophilic property. These properties can be altered by plasma treatment or by surface coating.

[0093] As described in more detail below, the cartridge and associated measurement system may utilize one or more sensors of one or more types (e.g., optical, pressure, ultrasonic, etc.) as part of the automated operation of the cartridge. Additionally, the output of such one or more sensors may be further utilized to perform quality control checks. These checks may be performed before, during, or after cartridge testing to ensure the functionality of one or more of the subsystems (e.g., ultrasound or other interrogation system, fluidics, fluid level, clamping, cartridge positioning / orientation system, or temperature control), ensure that the cartridge is functional, ensure that accurate sample preparation is performed before or has already been performed for the measurement, and may be used to pass or fail the test result or even to abort the test before the measurement begins.

[0094] In the following discussion, it is noted that a fluidic circuit includes a channel having fluidic components connecting one or more chambers together. A fluidic circuit is also referred to as a test channel among multiple channels that can be individually and controllably processed within a single cartridge device.

[0095] Cartridge Input Section 1, 2, 3, and 4 are schematic diagrams of an exemplary biological sample input section of a cartridge 100 for assessing hemostasis. Specifically, FIG. 1 shows a perspective view of an exemplary biological sample input device of a cartridge for use in a disposable system, according to an exemplary embodiment. FIG. 2 shows a side view of the exemplary biological sample input device of FIG. 1 with a casing, according to an exemplary embodiment. FIG. 3 shows a side cross-sectional view of the exemplary biological sample input device of FIG. 2 with a sample-holding tube mounted thereon, according to an exemplary embodiment. FIG. 4 shows a detailed view of the exemplary biological sample input device of FIG. 3, according to an exemplary embodiment. In an alternative embodiment, the input section of the cartridge comprises a well into which a fluid sample can be placed, for example, by a pipette or tube.

[0096] In some embodiments, and as shown in FIG. 1, cartridge 100 has double mating tabs 28a, 28b for coupling cartridge 100 to sample vessel guide 1 (shown in FIG. 2). As shown in FIG. 2, sample vessel guide 1 aligns sample vessel 2 with sample input port 3 of cartridge 100 when mated with cartridge 100. Cartridge 100 also includes alignment tab 29 configured to slide into alignment groove 30 of sample vessel guide 1 to further stabilize the coupling of sample vessel guide 1 to cartridge 100. Sample vessel guide 1 can further provide hard stop 5 (shown in FIG. 3) for holding sample vessel 2 at the proper height to establish fluid communication with cartridge 100.

[0097] In various embodiments, sample container 2 is a vacuum tube (also referred to herein as sample holding tube 2), such as a BD Vacutainer™ tube, and sample input port 3 includes one or more needles required for sample transfer 3a and discharge 4 (see FIG. 1). While shown as concentric in the drawings, the needles can be configured to be concentric, parallel, or integral. In some embodiments, and as shown in FIG. 1, sample transfer needle 3a includes inlets (3b and 3d) and outlet 3c that terminates in sample inlet chamber 26 of cartridge 100. Sample inlet chamber 26 is in fluid communication with inlet pathway 8 leading to holding / heating chamber 6 (see FIG. 5A). In some embodiments, and as shown in FIG. 1, discharge needle 4 terminates within sample container 2 when attached and spaced apart from inlet 3d to prevent bubbles from forming. Including an outlet 4a configured to minimize being drawn into inlets 3b and 3d. The dispensing needle 4 also has an inlet 4b that terminates at the dispensing inlet chamber 27 of the cartridge. The dispensing inlet chamber 27 is in fluid communication with a dispensing path 9 that terminates, in some embodiments, at a filter chamber 9a (shown in FIG. 5A) that houses a filter. Alternative sample containers 2 such as syringes that require a luer lock connection on the cartridge 100 may be utilized. Also, as described above, in other embodiments, the input section of the cartridge comprises wells, for example, where a fluid sample can be placed by a pipette or tube.

[0098] Dispensing path FIGS. 5A, 5B, 7, and 8 are schematic diagrams of the biological fluid path of an exemplary cartridge 100 according to one embodiment. Specifically, FIGS. 5A and 5B each show the biological fluid path of four test circuits (corresponding to test chambers 16a, 16b, 16c, and 16d shown in FIG. 5A), also referred to herein as a hemostasis test circuit, located on a sample preparation surface according to an exemplary embodiment. Although shown with four test circuits, additional circuits or fewer circuits may be included, for example, including two, three, five, six, seven, eight, etc. FIG. 5B further shows the cartridge body of FIG. 5A further connected to a sample holding tube 2 according to an exemplary embodiment. FIG. 7 shows the back side of the cartridge of FIG. 5A according to an exemplary embodiment and includes an interconnect surface in contact with the sample preparation surface that collectively forms the biological fluid path for the test circuits. FIG. 8 shows a portion of the biological fluid path on the interconnect surface of FIG. 7 according to an exemplary embodiment.

[0099] As described above, biological fluid pathways are formed on and across multiple planes defined within cartridge 100. A first plane of the fluid pathways of cartridge 100 is shown in FIGS. 5A and 5B. The first plane of the fluid pathways of cartridge 100 may alternatively be referred to as the front surface of cartridge 100. FIGS. 7 and 8 each show a second plane of the fluid pathways of cartridge 100, with FIG. 8 showing the second plane of the fluid pathways separated from the remainder of the structure of cartridge 100 for ease of understanding. The second plane of the fluid pathways of cartridge 100 may alternatively be referred to as the rear surface of cartridge 100. The fluid pathways between the first and second planes are connected by fluidic vias that traverse the various planes of cartridge 100.

[0100] As discussed above in connection with FIG. 1 , in some embodiments, the discharge inlet chamber 27 is in fluid communication with the discharge path 9. The discharge path 9 may terminate in a filter chamber 9 a (shown in FIG. 5A ) that may house a filter therein. The filter chamber 9 a in a first plane of the fluid path of the cartridge 100 is in fluid communication with a discharge port 22 i (shown in FIG. 8 ) in a second plane of the fluid path of the cartridge 100. As discussed in more detail below, the cartridge 100 interfaces with a measurement system (also referred to herein as an instrument) via the discharge port 22 i to provide air pressure to the discharge path 9.

[0101] Heating Chamber Path As discussed above in connection with FIG. 1, in some embodiments, the sample transfer needle outlet 3c terminates in the sample inlet chamber 26 of the cartridge 100. The sample inlet chamber 26 is in fluid communication with the inlet pathway 8. The sample inlet pathway 8 provides a fluid communication path between the sample inlet chamber 26 and the holding / heating chamber 6 (also referred to herein as the heating chamber 6 or "first chamber") (shown in FIG. 5A). As used herein, the labels "first," "second," and "third" are provided merely as labels and are not intended to imply any ordering. The heating chamber 6 is configured to mate with a corresponding thermal regulation (e.g., heating / cooling) system of the measurement system to heat or cool the sample to or near a predefined temperature.

[0102] The heating chamber 6 provided herein allows fluid to be metered or aliquoted into its respective test This promotes uniform conditioning of the test fluid before heating, thus reducing test sample variability that may affect subsequent measurements and analyses. The shape of heating chamber 6 may be optimized for heating / cooling transfer, as is the case herein when thin cross-sections with thin walls are used. The materials of cartridge 100 may also be optimized to promote heating / cooling. In some embodiments, the sample heating / cooling conditioning step may also be performed within one or more chambers / channels of the cartridge design and is not limited to occurring solely within heating chamber 6. In some embodiments, stirring, rotating, or vibrating elements (not shown) may be placed in heating chamber 6 that may be controlled by the measurement system to promote uniform temperature heating or cooling. In other embodiments, the test fluid in heating chamber 6 may be vibrated by the measurement system, which vibrates cartridge 100 to promote uniform temperature conditioning of the test fluid.

[0103] In some embodiments, temperature measurements are performed on the test sample in cartridge 100. To measure the temperature, a sensor may be incorporated into the measurement system or cartridge 100. In some embodiments, a thermistor or thermocouple may be placed in physical contact with cartridge 100 or the biological sample (such as blood). In other embodiments, an IR thermometer is directed toward cartridge 100 or the biological sample. In either case, cartridge 100 may incorporate a small well through which the incoming blood passes rather than being in direct contact with the blood. In some embodiments, the temperature of the test sample may be assessed at or near heating chamber 6. In other embodiments, the temperature of the test sample may be assessed while the test sample flows through the channel as it is directed toward test chamber 16.

[0104] 5A, 5B, and 8, the sample inlet pathway 8 terminates at a first corner 6a of the heating chamber 6, as shown in the upper left corner of the heating chamber 6 in FIGS. 5A and 5B. In some embodiments, chambers along the fluid pathway are generally filled from the top to prevent blood from backflowing into the inlet. A fill outlet channel 10a extends from a second corner 6b of the heating chamber 6 opposite the first corner 6a.

[0105] The fill outlet channel 10a extends to the filter chamber 10, which contains the filter. The filter chamber 10 (e.g., as shown in Figures 5A and 5B) in the first plane of the cartridge 100's fluid path is in fluid communication with the heating chamber fill channel 10b (see Figure 8) shown in the second plane of the cartridge 100's fluid path. The fill conduit 10b is in fluid communication with a pressure port 22a (also see Figure 8) that facilitates filling of the heating chamber 6. The conduit 10b is part of a network of conduits used to integrate the cartridge 100's pressure ports (e.g., 22a as discussed, and 22b-22i, also discussed later) into one or more areas where a measurement system can be coupled with its pressure control interface. Such a configuration reduces the complexity of the measurement system to control the movement of fluid within the cartridge 100. In practice, the conduits that handle the control of the movement of the fluid sample in the first plane of the cartridge 100 are primarily located in the cartridge 100's second plane. 6A and 6B show front and rear perspective views of FIGS. 5A, 5B, and 8 with additional labeling corresponding to the descriptions in this section.

[0106] Heating chamber filling In operation, the instrument's fluid pump draws sample through input port 3 (see FIGS. 1-2) of cartridge 100, via connection port 22 (see FIG. 7) (also referred to herein as a pressure port), and into heating chamber 6 (see FIG. 5A or 5B). For example, the instrument's fluid pump may communicate with pressure port 22a (see FIG. 8) and apply a differential pressure (e.g., positive or negative) thereto. This creates increased pressure along fill conduit 10b, within heating chamber 6, and along inlet path 8, drawing the sample into heating chamber 6. At the same time, the inner discharge needle 4 is connected to the separation channel 9, which receives air pressure from the instrument via discharge port 22i (see Figure 8), to neutralize the pressure in the sample container 2 when the sample is aspirated into the heating chamber 6 of the cartridge 100. During filling of the heating chamber 6, all other ports (e.g., 22b-22i) are closed, for example, by the measurement system.

[0107] When the heating chamber 6 fills, the filter in the filter chamber 10 becomes clogged, creating a pressure spike that is detected by the instrument, causing the instrument to turn off the fluid pump. The instrument may also close the discharge port 22i or otherwise interrupt the supply of air pressure through the discharge port 22i upon detecting the pressure spike. Alternative fill detection techniques, i.e., an optical sensor placed at the desired fill level, a volume control placed at the desired fill level, a pressure change (negative and / or positive pressure) over a period of time, an ultrasonic detector, etc., may also be used. For example, the sample remains in the heating chamber until it reaches a desired temperature, which may be at or near normal and typical subject body temperature (e.g., approximately 37°C for a healthy human). Alternatively, other desired temperatures may be recognized. The geometry of the heating chamber 6 and the channel leading to the sample metering chamber 11 (described below) is configured to trap any bubbles that may be present in the fluid sample away from the rest of the fluid circuit. The shape of inlet pathway 8 includes anti-wicking feature 8a (see Figures 5A and 5B) configured to reduce the formation of bubbles within heating chamber 6 and prevent wicking into and out of sample container 2. To this end, additional unprocessed test sample (e.g., unheated blood) cannot be wicked into the heating chamber after the first extracted test sample has been heated and / or cooled, for example, when a processed test sample is drawn into metering chamber 11 (also referred to herein as sample chamber 11 and the "second" chamber). Figures 6A and 6B also show labels corresponding to the description in this section.

[0108] Sample aliquot (metering) chamber pathway 5A, 5B, and 8, a first side 6c (see FIG. 5A) of the heating chamber 6 extends between a first corner 6a and a second corner 6b.

[0109] One or more of the outlet ports 6e-6h (see FIG. 5A) are positioned along the length of the second side 6d of the heating chamber. Each of the one or more outlet ports 6e-6h may be positioned in a different one of the valleys along the second side 6d of the heating chamber, which valleys direct the sample into a conduit leading to each respective test channel. In some embodiments, a test channel collectively refers to the associated fluid path structure and test chamber used to perform a measurement of a given aliquot sample. In some embodiments, and as shown in FIG. 5B, each of the one or more outlet ports 6e-6h in the first plane of the fluid path of the cartridge 100 is in fluid communication with a first end of one or more corresponding channels 20a-20d (see FIG. 8) in the second plane of the fluid path of the cartridge 100, collectively referred to as channel 20 (see FIG. 5B). The second end of each of one or more channels 20a-20d (see FIG. 8) in the second plane of the fluid pathway of cartridge 100 is similarly in fluid communication with the first end of one or more corresponding channels 11a-11d (see FIG. 5B) in the first plane of the fluid pathway of cartridge 100. The second end of each of one or more channels 11a-11d terminates in one or more corresponding sample chambers 11 (shown with an "o" symbol inside and overlapping ("x4") in FIG. 5B). In the example shown in FIGS. 5A, 5B, 7, and 8, there are four sample chambers 11. More or fewer sample chambers 11 and corresponding fluid communication paths with heating chamber 6 may be present on cartridge 100 in some configurations.

[0110] The sample chamber 11 is fed by one or more channels 20 that start at the bottom of the heating chamber 6. This geometry avoids bubbles being drawn into the sample chamber 11 when they reach the upper part of the heating chamber 6.

[0111] Each sample chamber 11 has a corresponding fill channel 11e that is in fluid communication with a corresponding filter chamber 12 (shown with a "+" symbol and overlapping ("x4") in FIG. 5B) that has a filter therein. The filter chambers 12 in the first plane of the fluid path of the cartridge 100 are in fluid communication with channel 12a (see FIG. 8) in the second plane of the fluid path of the cartridge 100. Channel 12a is in fluid communication with pressure port 22g (see FIG. 8).

[0112] In some configurations, when two or more sample chambers 11 are implemented on cartridge 100, channel 12a (see FIG. 8) is in fluid communication with all of the sample chambers 11 (see FIG. 5B) via corresponding fill channel 11e and filter chamber 12. Channel 12a therefore acts as a manifold for applying negative pressure to all of the sample chambers 11 through a single pressure port 22g. Thus, separate pressure ports are beneficially not required for filling each of the sample chambers 11. Figures 6C and 6D show front and back perspective views of Figures 5A, 5B, and 8, with additional labels corresponding to the descriptions in this section.

[0113] Heating chamber discharge path 5A, 5B, and 8, heating chamber 6 includes discharge channels 31a, 31b, and 31c along a first side 6c of heating chamber 6 for discharging heating chamber 6 when sample chamber 11 is filled. Discharge channel 31 (not shown) includes a fluid path through conduit elements 31a-31d. Channels 31a-31b terminate at one end along first side 6c at heating chamber 6 and at the other end at filter chamber 31c having a filter therein. Channels 31a-b thus provide a fluid path between heating chamber 6 and filter chamber 31c. Filter chamber 31c (see FIG. 5A) in a first plane of cartridge 100's fluid path is in fluid communication with channel 31d (see FIG. 8) in a second plane of cartridge 100's fluid path, and channel 31d is in fluid communication with discharge port 22c (see FIG. 8). As discussed in more detail below, cartridge 100 interfaces with the instrument via vent port 22c for providing air pressure for the instrument to vent heating chamber 6. Figures 6C and 6D also show labels corresponding to the descriptions in this section.

[0114] Sample chamber discharge path 5A, 5B, and 8, each sample chamber 11 includes an outlet path 18 that terminates at a first end of the corresponding sample chamber 11. The second ends of the outlet paths 18 in the first plane of the cartridge 100's fluid path are in fluid communication with an outlet manifold 18 in the second plane of the cartridge 100's fluid path. When two or more sample chambers 11 are present, all of the sample chamber outlet paths 18 (see FIG. 5B) are in fluid communication with an outlet manifold 18a (see FIG. 8). The outlet manifold 18a (see FIG. 8) in the second plane of the cartridge 100's fluid path is further in fluid communication with a first end of a channel 18b (see FIG. 5B) in the first plane of the fluid path. The second end of the channel 18b (see FIG. 5B) is in fluid communication with a filter chamber 18c (see FIG. 5B) having a filter therein. Filter chamber 18c (see FIG. 5B) in a first plane of the fluid pathway of cartridge 100 is in fluid communication with discharge port 22e (see FIG. 8). Figures 6C and 6D also show labels corresponding to the descriptions in this section.

[0115] Sample aliquot (weighing) chamber filling In operation, once the sample is at or near the desired temperature, the sample is aliquoted (or metered) into one or more separate sample chambers 11 (see FIG. 5B). Unless otherwise noted, with reference to FIG. 5B, in various embodiments, the sample chambers are pumped via the instrument's pumps. The heating chamber 6 is filled by applying negative pressure to pressure port 22g (see Figure 8) while simultaneously venting the heating chamber 6 through vent port 22c (see Figure 8). Each filter chamber 12 has an internal filter that clogs when the corresponding sample chamber 11 is filled, triggering a pressure sensor in the instrument to turn off the pump as the heating chamber 6 is filled. As mentioned above, alternative fill detection techniques can be used. In various embodiments, all sample chambers 11 are controlled by a single valve and fluid path in the instrument via pressure port 22g (see Figure 8). The cutoff pressure is not activated until the filters of all sample chambers are clogged within the corresponding filter chambers 12. The sample chambers 11 are used to separate the sample into independent functional channels, aliquot the sample into known volumes, and sort the sample for mixing with reagents. While the sample chambers 11 are being filled, pressure ports 22b, 22d, 22f, and 22h (see FIG. 8) and discharge port 22e (see FIG. 8) are closed by the instrument to prevent fluid from leaking past location 19 located below the sample chambers 11. Once the sample chambers 11 are filled, discharge port 22e (see FIG. 8) is opened to atmospheric pressure, which allows one or more sample chambers 11 to be fluidly isolated from each other and from the heating chamber 6. Discharge port 22e (see FIG. 8) remains open to atmospheric pressure during sample mixing, as discussed below. Figures 6C and 6D also show labels corresponding to the description in this section.

[0116] Mixing and Testing Pathways Unless otherwise noted, with reference to FIG. 5B, each of the sample chambers 11 is in fluid communication with one or more corresponding reagent pockets 14 (see FIG. 5A) configured to accommodate at least one lyophilized bead containing a reagent. As shown in FIGS. 5A and 5B, two reagent pockets 14 (shown as 14a and 14b in FIG. 5A for one of the test channels) are provided. In other embodiments, a single reagent pocket is used for each test channel. In yet other embodiments, three or more reagent pockets 14 are used for each test channel. The reagent pockets 14 are in fluid communication with the serpentine channels 13 (see FIG. 5A and shown with an overlapping symbol (“×4”)). Each of the serpentine channels 13 has a first end in fluid communication with the reagent pocket and a second end terminating in an optical detection zone 15. As discussed below, an instrument (i.e., a measurement system) may optically interrogate the optical detection zone 15 of the cartridge 100 to facilitate control of a pump that promotes mixing of the individual aliquots with the corresponding reagent(s). Each of the serpentine channels 13 (see FIG. 5A) is in fluid communication with a test chamber 16 (see FIG. 5B, having a duplicate symbol ("x4")), and the test chamber 16 is in fluid communication with a filter chamber 17 having a filter therein. The filter chamber 17 in the first plane of the fluid path of the cartridge 100 is in fluid communication with a first end of a corresponding one of the fluid channels 17a-17d (see FIG. 8) in the second plane of the fluid path of the cartridge 100. The second end of each of the pressure ports 22b, 22d, 22f, and 22h (see FIG. 8) is in fluid communication with a corresponding pressure port 22b, 22d, 22f, and 22h (see FIG. 8). Cartridge 100 interfaces with an instrument via pressure ports 22b, 22d, 22f, and 22h (see FIG. 8) to provide positive and negative pressures to facilitate sample mixing and testing, as described below. Figures 6E and 6F also show front and rear perspective views of Figures 5A, 5B, and 8, with additional labels corresponding to the descriptions in this section.

[0117] Sample Mixing Unless otherwise noted, with reference to FIG. 5B, each individual aliquot within sample chamber 11 is drawn into a separate reservoir or channel (serpentine channel path 13 (see FIG. 5A) in various embodiments) and brought into contact with a channel-specific reagent located in one (or both) of two reagent pockets 14 (see FIG. 5A). Specifically, the instrument's pump applies negative pressure to pressure ports 22b, 22d, 22f, and 22h (see FIG. 8) to pump the reagent into reagent pockets 14 (see FIG. 5A) and serpentine channel path 13 (see FIG. 5A). The sample is drawn through the serpentine channel 13 (see FIG. 5A) until the sample activates the instrument's optical sensor (the detection zone is the top of the serpentine channel near the optical detection zone 15 (see FIG. 5A)), which blocks the channel from the pump. Mixing in the serpentine channel, or in zones or regions therein, can be controlled by one or more independent valves and pathways that allow for individual channel control. Alternative sensor technologies may be utilized, such as pressure, pass-through optical sensors, ultrasonic detection, time, or volume control. Once the optical sensors on all channels are activated, the pump reverses, applying positive pressure to pressure ports 22b, 22d, 22f, and 22h (see FIG. 8). The positive pressure pushes the biological sample, such as blood, down serpentine path 13 (see FIG. 5A) for a specified time or until a second set of optical sensors on the instrument is captured (in an alternative embodiment). This process of pulling the sample up serpentine path 13 (see FIG. 5A) to the optical detection zone where it is detected by the optical sensors of the instrument and pushed back for a given time is repeated until complete sample mixing is achieved. Figures 6E and 6F also show labels corresponding to the descriptions in this section.

[0118] Other sensors (e.g., impedance sensors), pressure sensors, etc. may be used. Alternatively, additional sensors may be used to detect both ends of the optical detection zone. Alternating path geometries, obstacles to create turbulence, cycle times, and cycle speeds are all design alternatives that may be used with different test types to achieve optimal results. In an alternative embodiment, mixing may be accomplished with one or more ferromagnetic beads or rods placed within the cartridge and controlled by the instrument.

[0119] Test chamber filling Referring to Figures 5A, 5B, and 8, one or more test chambers 16 are filled after mixing is complete. Using one or more independent valves and pathways, each test chamber 16 is filled with sample via the application of a pressure perturbation (negative and / or positive pressure) at pressure ports 22b, 22d, 22f, and 22h. Specifically, the instrument pump applies negative pressure to pressure ports 22b, 22d, 22f, and 22h until all of the filters clog in one or more filter chambers, causing a pressure spike and forcing the instrument to turn off the pump, similar to filling the heating chamber 6. As mentioned above, alternative fill detection techniques can be used. The test chambers 16 have design features, such as ridges 24a, that prevent bubble formation within the test chambers 16 during filling. Once filled, the instrument begins viscoelastic testing of the sample. Figures 6E and 6F also show labels corresponding to the descriptions in this section.

[0120] In some embodiments, the cartridge device includes at least four independent fluidic circuits configured with different sets of reagents for performing measurements (and / or sample preparation) in parallel. Measurements are performed on each of the at least four channels of the cartridge. In some embodiments, the measurements include viscoelastic properties such as sample shear modulus. In other embodiments, the measurements include other properties such as sample viscosity, elastic modulus, or any other mechanical property, or a combination thereof.

[0121] Table 1 provides an exemplary set of reagents and measurement parameters for use in an exemplary cartridge device (e.g., device 100, among others). As shown in Table 1, channel number 1 of the exemplary cartridge device contains kaolin, an activator of the intrinsic pathway of coagulation. Channel number 2 is investigated to measure the clotting time of the test sample in the presence of kaolin and in the additional presence of heparinase I, a neutralizer of the anticoagulant heparin, as shown in Table 1. Channel number 3 is investigated to measure the overall clot firmness of the test sample in the presence of i) thromboplastin, an activator of the extrinsic pathway of coagulation, and ii) polybrene, a neutralizer of the anticoagulant heparin, as shown in Table 1. Channel number 4 is investigated to measure the clot firmness of the test sample either with the same reagents as channel number 3 or with the addition of abciximab (e.g., Clotinab® and / or ReoPro®), an inhibitor of platelet aggregation / contraction. As shown in Table 1, calcium is added to all reagent formulations when the assay is configured to run on citrated whole blood samples. [Table 1]

[0122] Table 2 provides an additional exemplary set of reagents and measurements for use with an exemplary cartridge device (e.g., device 100, among others). As shown in Table 2, channel number 2 contains an extrinsic pathway activator, accompanied by inhibition of fibrinolysis with tranexamic acid (TXA). In addition to the measurements already shown in Table 1, channel numbers 2, 3, and 4 are also interrogated to measure, for example, changes in clot firmness, which may be related to the fibrinolysis process. In some embodiments, other channels can contain reagents that inhibit fibrinolysis and can also be interrogated to measure changes in clot firmness. For example, channel number 4 can also contain TXA or other fibrinolysis inhibitors to measure clot firmness in the absence of fibrinolysis. [Table 2]

[0123] In some embodiments, clotting time and clot firmness are measured by analyzing shear modulus (thrombus firmness) versus time curves generated within each measurement channel of the cartridge. Figure 13 shows an exemplary shear modulus versus time curve, according to an exemplary embodiment. Clotting time can be determined by identifying when clot firmness meets or exceeds a threshold, when a first or higher derivative of such a measured property meets or exceeds a threshold, or by identifying the time point of maximum acceleration in the rate of clot firmness, or some combination of the above methods. Clot firmness can be estimated by clot firmness at a certain time after clotting time, or the maximum overall clot firmness measured within a certain time limit, or clot firmness at the time of maximum rate of change in clot firmness, or some combination of the above methods. Similar methods can also be applied to measure the effects of fibrinolysis (i.e., thrombolysis) and the corresponding reduction in clot firmness. In some embodiments, the change in clot firmness can be calculated as a percentage reduction in clot firmness over a certain time window, as a rate of change in clot firmness over time, as the area under or above the clot firmness versus time curve within a given time window, as the time required to achieve a given reduction in clot firmness, or a combination thereof. Curves similar to those just described and similar measurements can be generated by plotting Young's modulus, viscosity, or other viscoelastic properties of the sample being measured. [Table 3]

[0124] Those skilled in the art will recognize that clotting time and clot firmness can be estimated using many methods and criteria. Clotting time and clot firmness values ​​obtained from at least four channels / measurements can be combined to provide at least six parameters that can indicate the functional status of a patient's hemostatic system. The indices are summarized in Table 3. The correlation between results from different channels (clotting time, clot firmness, change in clot firmness, etc.) can be verified to be within expected ranges as an additional quality control check to verify instrument, cartridge, and sample function.

[0125] In other embodiments, other reagents may be used, such as a fibrinolytic index, an index corresponding to the functionality of antiplatelet therapy, an index corresponding to the functionality of anticoagulant therapy, or other hemostatic indexes. Output parameters may be obtained.

[0126] For example, one or more fibrinolytic indices can be formed using the change in clot firmness measured in any of the channels shown in Table 2, preferably channels 3 and 4. Alternatively, a fibrinolytic index can be formed by combining the difference in the change in clot firmness measured in channels 2 and 3 shown in Table 2. Such combinations can be in the form of ratios, differences, or combinations thereof. One advantage of using a combination of the change in clot firmness measured with and without an antifibrinolytic reagent is the ability to mitigate the interfering effect of non-fibrinolytic decreases in clot firmness values. In some embodiments, TXA or other fibrinolysis inhibitor reagents can be included in both channels 2 and 4 of the exemplary cartridge in Table 2. With such modifications, parameters for clot firmness, platelet contribution, and fibrinogen contribution can be derived without the influence of fibrinolysis by combining the clot firmness measurements obtained in channels 2 and 4.

[0127] As noted above, an exemplary user interface is described in commonly assigned U.S. Publication No. 2011 / 0252352 (Viola et al.), which is incorporated herein by reference in its entirety. The exemplary user interface may be used to display, among other parameters, the measured hemostatic index discussed in connection with Table 4. [Table 4]

[0128] As described above, in various embodiments, the test chamber 16 is shaped to facilitate ultrasonic testing of viscoelastic properties of a sample, although alternative shapes may be implemented to measure other test types. Such ultrasonic testing systems are described in commonly assigned U.S. Pat. No. 9,726,647 and U.S. Publication No. 2016 / 0139159, both of which are incorporated by reference in their entireties. The ultrasonic transducer of the measurement system interfaces with the test chamber 16 of the cartridge 100 via a flexible, deformable elastomer 21 secured to a test block 21d on the cartridge 100.

[0129] Exemplary elastomer materials optionally include Dynaflex D3202, Versaflex OM9-802CL, Maxelast 54740, RTP6035, and Versaflex CL2003X, among others. Now, unless otherwise noted, with reference to FIG. 9, test block 21d is aligned with test chamber 16 (see FIG. 5B) via alignment slots 23 and 24 in cartridge 100. Still referring to FIG. 9, elastomer 21 may be secured to test block 21d via flange 21a on elastomer 21. Flange 21a may have multiple alignment holes 21b that can receive corresponding alignment pegs (not shown) from test block 21d. Soft elastomers 21 may also each include a lens 21c that focuses ultrasonic energy into the sample in test chamber 16.

[0130] Figure 16 shows a photograph of the exemplary cartridge of Figures 1, 2, 3, 4, 5A, 5B, 7, 8, and 9 for use in a disposable system according to an exemplary embodiment. Figure 17 shows a front view of the exemplary cartridge of Figures 1, 2, 3, 4, 5A, 5B, 7, 8, and 9 according to an exemplary embodiment. Figure 18 shows a front view of the exemplary cartridge of Figures 1, 2, 3, 4, 5A, 5B, 7, 8, and 9 according to an exemplary embodiment.

[0131] As described in U.S. Pat. No. 9,272,280, incorporated herein by reference in its entirety, in various embodiments, the consumable cartridge includes a lens assembly that focuses ultrasonic energy into the sample, which can be used to generate flow and mixing. The lens assembly or sound-focusing assembly is designed using a soft material, such as a thermoplastic elastomer 134 (previously referred to as 21), in conjunction with a rigid substrate 132 (e.g., formed from test block 21d), such as polystyrene, as shown in Figures 10, 11, and 12. This combination provides dry ultrasonic coupling without the need for any fluid or gel couplants. Note that the same lens and ultrasonic driver used for hemostasis measurements can be used in this manner to provide mixing. Increased acoustic energy for mixing can be delivered, for example, by increasing pulse length, pulse amplitude, or pulse repetition frequency.

[0132] 10, a top cross-sectional view of test chamber 116 (previously referred to as test chamber 16) is shown. To seal each test chamber, e.g., test chamber 116, lens assembly 131 includes rigid substrate 132 and couplant 134 that may be positioned at the rear end of each test chamber.

[0133] 10 , each couplant 134 includes an elastomeric material. Optionally, the elastomeric material is a thermoplastic elastomer (TPE). Exemplary elastomeric materials optionally include Dynaflex D3202, Versaflex OM9-802CL, Maxelast 54740, RTP6035, and Versaflex CL2003X, among others. Optionally, the couplant is overmolded onto a rigid substrate. Optionally, the couplant is mechanically secured to the rigid substrate.

[0134] 10 , between each couplant 134 and the open space of each test chamber is a rigid substrate 132. The rigid substrate and couplant form an interface (e.g., a lens assembly) that focuses ultrasound waves transmitted by the ultrasound transducer into the open space of the chamber and onto any biological fluids and / or reagents within the chamber. The rigid substrate of the lens can include a material that allows sound to pass through and can act to focus ultrasound waves at a level within the space. Optionally, the rigid substrate includes styrene.

[0135] Referring now to FIG. 11, the lens assembly is mounted in test block 21d (element 21b in FIG. 11). The lens assembly may be glued or welded to surface 101 of test block 21d (shown as 132) to secure the lens in place in an orientation that allows for the desired focusing of sound. Alternatively, the lens assembly is optionally fabricated together with surface 101 of test block 21d. In this regard, rigid substrate 132 may be molded with surface 101 of test block 21d, and couplant 134 may be overmolded or mechanically secured to the rigid substrate. A wide range of materials may be used to construct the device. For example, plastic may be used for single-use disposable cartridges.

[0136] 11, each of the test chambers 116 can have a lens assembly positioned across the large opening of the open space of each chamber. In this way, each chamber can be separately interrogated with focused ultrasound.

[0137] 11 , when placed in the instrument, the couplant 134 can be placed in acoustic communication with the transducer for delivering ultrasound waves through the lens assembly and into the test chamber 116. Optionally, an intermediate layer of acoustically transparent material is positioned between the ultrasound transducer and the couplant. For example, an intermediate layer or block of Rexolite® or TPX® can be used. The intermediate layer can be pressed against the couplant and can be in acoustic contact with the transducer.

[0138] Referring further to FIG. 11 , sound generated by the transducer passes through the intermediate layer, couplant, and rigid substrate and is focused into the biological sample, such as blood, and reagents within the test chamber. A portion of the sound directed into the chamber contacts the distal inner surface 111 of the test chamber, defined by surface 126. Optionally, the surface is polystyrene. The distal inner surface has a known shape and is positioned a known distance from the ultrasound source. The distal inner surface 111 is used as a calibrated reflector to estimate the speed of sound and sound attenuation of the test chamber at baseline and during the clot formation and thrombolysis processes. These measurements can be used, for example, to estimate the subject's hematocrit level along with an index of hemostasis. The sound generated by the transducer can be focused into the biological sample within the test chamber using a parabolic mirror coupled to the biological sample via an elastomer.

[0139] Other exemplary cartridge devices and measurement systems, and methods thereof, are described in U.S. Patent No. 9,031,701, U.S. Provisional Application No. 61 / 443,084, U.S. Patent No. 9,272,280, U.S. Patent No. 9,410,971, U.S. Provisional Application No. 61 / 443,088, U.S. Publication No. 2011 / 0252352, Published PCT Publication No. WO2011 / 127436, U.S. Publication No. 2012 / 0294767, U.S. Patent No. 7,892,188, U.S. Patent No. 8,740,818, and U.S. Publication No. 2016 / 0274067, each of which is incorporated by reference herein in its entirety.

[0140] As noted, the cartridges and features described herein may be modified for use with other types of measurement systems, such as thromboelastography-based systems, thromboelastometry-based systems, optical-based systems, fluorescence-based systems, colorimetric-based systems, aggregometry-based systems, resonance-based systems, and electrical impedance-based systems, among others.

[0141] Many modifications and other embodiments of the inventions described herein will come to mind to one skilled in the art to which these inventions pertain having the benefit of the teachings presented in the foregoing description. Therefore, it is to be understood that the invention is not limited to the specific embodiments disclosed and that modifications and other embodiments are intended to be included within the scope of the appended claims.

[0142] Although specific terms are employed herein, they are intended to be generic and descriptive only and not for purposes of limitation. It is used in a clear sense.

[0143] When used in the claims, the terms "first," "second," and "third" are provided merely as labels and are not intended to imply any order.

Claims

1. The housing and an input port integrally formed with the housing, the input port being structurally configured to receive the contents of a sample holding tube; a first chamber in fluid communication with the input port, the first chamber configured to receive a sample contained in the sample holding tube and to condition the received sample to or near a desired temperature before allowing the received sample to contact one or more reagents located in one or more fluid circuits downstream of the first chamber; The device, wherein each of the one or more fluid circuits comprises: i) a second chamber in fluid communication with the first chamber that meters the sample in the first chamber into an aliquot, wherein the metered sample is introduced to a reagent or combination of reagents located in a corresponding fluid circuit to form a mixed sample; and ii) a test chamber in fluid communication with the second chamber, the test chamber being structurally configured for interrogation by a measurement system configured to determine at least one viscoelastic property of the mixed sample.

2. 10. The device of claim 1, wherein at least one of the one or more fluid circuits comprises one or more pockets configured to accommodate at least one lyophilized bead containing the reagent or combination of reagents.

3. 3. The device of claim 1, wherein at least one of the one or more fluid circuits comprises one or more liquid-retaining pockets for containing the reagent or combination of reagents.

4. 4. The apparatus of claim 1, wherein the input port is communicatively connected to a first pressure port, and when pressure is applied to the first pressure port, the pressure causes the contents of the sample holding tube to be expelled through the input port into the first chamber.

5. 5. The apparatus of claim 1, wherein the input port forms part of a needle assembly structurally configured to establish fluid communication and expel the contents of the sample holding tube.

6. 6. The device of claim 5, wherein the needle assembly comprises the input port and a second port, the second port configured to release a liquid or gas into the sample holding tube to facilitate evacuation of the contents therein.

7. The apparatus of any one of claims 1 to 6, wherein the input port is structurally configured to couple to a luer lock configured to connect to the sample holding tube.

8. 8. The apparatus of claim 1, wherein the first chamber is configured to mate with a corresponding thermal conditioning surface of a subsystem component of the measurement system to condition the received sample at or near the desired temperature.

9. 9. The apparatus of claim 1, wherein the first chamber comprises a shape and / or material optimized to facilitate heating and / or cooling of the sample to or near the desired temperature.

10. The channel portions of the one or more fluid circuits are connected to a corresponding thermal regulation system of the measurement system.

10. The apparatus of claim 1, configured to mate with a temperature control system to adjust the received sample to the desired temperature.

11. 11. The apparatus of claim 10, wherein the first chamber and / or the channel portion of the one or more fluid circuits are in physical proximity to a sensor of the measurement system configured to measure the temperature of the sample received in the first chamber.

12. 12. The apparatus of any one of claims 1 to 11, comprising a filter positioned in the first pressure port, the filter configured to allow air to move through the first pressure port but to prevent fluid from moving therethrough.

13. 13. The apparatus of claim 1, wherein each of the one or more fluid circuits comprises a third set of fluid paths in fluid communication between a respective second chamber and a test chamber, a portion of the third set of fluid paths being arranged as a serpentine-shaped conduit.

14. 14. The apparatus of claim 13, wherein each of the second chambers is connected to a second pressure port, and pressure applied to the second pressure ports causes the second chambers to fill.

15. 15. The apparatus of claim 14, wherein each of the second chambers is connected to a discharge port, the discharge port being configured to be closed while the sample is being metered into the aliquots in the second chambers, and the discharge port being further configured to be vented to atmospheric pressure after the sample is being metered into the aliquots in the second chambers.

16. 16. The device of any one of claims 1 to 15, wherein the serpentine-shaped conduit forms a serpentine reservoir between the test chamber and the second chamber, and a metered sample is directed through a portion of the serpentine-shaped conduit to promote mixing of the metered sample with the reagent or combination of reagents.

17. 17. The apparatus of claim 16, wherein the metered sample is selectively and multiplicatively directed for each of the one or more fluid circuits between a first position in the fluid circuit and a second position in the fluid circuit, the length of the first position and the second portion comprising at least a portion of the serpentine conduit.

18. 18. The apparatus of claim 17, wherein each of the test chambers is connected to a third pressure port, and pressure applied to the third pressure port causes the test sample to flow through a respective serpentine conduit toward the test chamber.

19. 20. The apparatus of claim 18, wherein pressure applied in a reverse direction to the third pressure port causes the test sample to flow through the respective serpentine conduit and away from the test chamber.

20. 20. The device of claim 19, wherein the serpentine reservoir comprises an optical detection area.

21. 21. The device of claim 1, wherein each of the one or more fluid circuits further comprises a mixing zone between the test chamber and the second chamber, the mixing zone comprising one or more ferromagnetic beads or rods therein.

22. At least one of the one or more fluid circuits includes one or more quality test portals, the one or more quality test portals configured to be optically or electrically sensed. The device according to any one of claims 1 to 21,

23. 23. The apparatus of claim 22, wherein at least one of the one or more quality test ports is configured to be sampled for a characteristic of the metered sample.

24. Apparatus according to any preceding claim, wherein the test chamber comprises a lens configured to direct ultrasonic pulses generated by the measurement system into the test chamber.

25. 25. The device of claim 1, further comprising, for each of the one or more fluid paths, a second chamber in fluid communication with the first chamber, the one or more reagent pockets, and the test chamber.

26. 26. The device of any one of claims 1-25, wherein at least one of the reagents or combinations of reagents located in the one or more fluid circuits is selected from the group consisting of an intrinsic pathway activator, an extrinsic pathway activator, and a coagulation activator.

27. 27. The device of any one of claims 1 to 26, wherein at least one of the reagents or combinations of reagents located in the one or more fluid circuits is selected from the group consisting of a platelet activator, a platelet inhibitor, and a fibrinolytic function inhibitor.

28. 28. The device of any one of claims 1-27, wherein at least one of the reagents or combinations of reagents located in the one or more fluid circuits is selected from the group consisting of a FXIIIa inhibitor, thrombomodulin, polybrene, heparin, corn trypsin inhibitor, adenosine, GPRP (Gly-Pro-Arg-Pro), calcium, fibronectin, collagen, an immunodetection reagent, and heparinase I, or a combination thereof.

29. 29. The apparatus of any one of claims 1 to 28, wherein the measurement system is selected from the group consisting of a sonorheometry-based system, a thromboelastography-based system, a thromboelastometry-based system, an optical-based system, a fluorescence-based system, a colorimetric-based system, an aggregometry-based system, a resonance-based system, and an electrical impedance-based system.

30. 30. The device of any one of claims 1-29, comprising at least four test channels, a first test channel containing an intrinsic pathway activator, a second test channel containing the intrinsic pathway activator and a heparin neutralizer, a third test channel containing an extrinsic pathway activator, and a fourth test channel containing the extrinsic pathway activator and a platelet inhibitor.

31. 30. The device of any one of claims 1-29, comprising at least four test channels, a first test channel containing an intrinsic pathway activator, a second test channel containing an extrinsic pathway activator and a fibrinolytic function inhibitor, a third test channel containing an extrinsic pathway activator, and a fourth test channel containing the extrinsic pathway activator and a platelet inhibitor.

32. 32. The device of claim 30 or 31, wherein the third channel and the fourth channel each contain hexadimethrine bromide (polybrene).

33. 32. The device of claim 31, wherein the fourth test channel further comprises a fibrinolytic function inhibitor.

34. 34. The device of any one of claims 1-33, comprising at least two test channels, a first test channel containing an intrinsic / extrinsic pathway activator and a second test channel containing the intrinsic / extrinsic pathway activator and heparin.

35. 1. A method of mixing a sample with one or more reagents in a device and testing the mixed sample for assessment of hemostasis, comprising: receiving a plurality of metered samples from a plurality of metering chambers that have received test fluid from the sample holding tubes; flowing at least one of the aliquots alternately and multiplicatively until the at least one aliquot is mixed with a reagent or combination of reagents to form a mixed aliquot, wherein the at least one aliquot i) flows from the metering chamber in a first direction through one or more reagent pockets having the one or more reagents therein and along a serpentine path in communication with the metering chamber until at least a portion of the at least one aliquot reaches a detection zone located within or after the serpentine path, and ii) alternately and cyclically flows from the detection zone in the second direction reserved in the first direction through at least a portion of the serpentine path toward the metering chamber; and driving the mixed aliquot into a test chamber in fluid communication with the metering chamber, the test chamber being structurally configured for interrogation by a measurement system configured to determine a characteristic of the mixed aliquot, and the interrogation of the test chamber being performed by the measurement system within which the mixed aliquot is located.

36. 36. The method of claim 35, further comprising receiving the test fluid in a first chamber configured to adjust the temperature of the test sample substantially toward body temperature, and wherein the metered sample received in the metering chamber is received from the first chamber.

37. 37. The method of claim 35 or 36, wherein the test fluid moves to the first chamber in response to a first applied positive or negative pressure applied by or generated from the measurement system.

38. 38. The method of any one of claims 35 to 37, further comprising conditioning the test fluid in the first chamber to or substantially near a desired temperature, wherein the test fluid is mixed with the one or more reagents after exiting the first chamber.

39. 39. The method of any one of claims 35 to 38, further comprising isolating the test fluid in the metering chamber to prevent the test fluid from contacting the one or more reagents during filling of the metering chamber.

40. 40. The method of any one of claims 35 to 39, wherein a second pressurized positive or negative pressure is applied by or generated from the measurement system at a second port in communication with the tortuous path to move the at least one aliquot in the second direction.

41. 41. The method of any one of claims 35 to 40, wherein the first applied positive or negative pressure is inversely applied by or generated from the measurement system to move the at least one aliquot in the second direction.

42. said receiving said mixed aliquot in said test chamber; receiving a negative or differential pressure through the third pressure port; The method of any one of claims 35 to 41, wherein the pressure port is further in fluid communication with the test chamber.

43. The method of any one of claims 35 to 42, wherein the test chamber is downstream of the tortuous path and the third pressure port is downstream of the test chamber.

44. Apparatus configured to perform any one of the methods according to claims 35 to 43.

Images