Disposable system for hemostatic function analysis

Abstract

In some embodiments, a disposable system includes a multi-channel or multi-chamber test cartridge device configured to operate with a test system for assessing hemostasis in a subject by assessing a test sample from the subject in vitro. In some embodiments, the disposable system is configured to interrogate a test sample to assess clot hardness, strength, or other mechanical properties of the test sample to assess the function of various physiological processes occurring during coagulation and / or dissolution of the resulting clot.

Description

[0001] This is a reissue application based on the parent Chinese divisional patent application with the filing date of April 20, 2018, patent application number 202211271508.6, entitled "Disposable system for hemostatic function analysis."

[0002] Government Rights This invention was made with government support under R44 HL103030 awarded by the National Heart, Lung, and Blood Institute. The government has certain rights in the invention.

[0003] Related Applications This application claims priority to and the benefit of U.S. Provisional Application No. 62 / 488,045, filed April 20, 2017, entitled "Disposable system for hemostatic function analysis," which is incorporated by reference herein in its entirety. TECHNICAL FIELD

[0004] The present application relates to devices, systems, and methods for assessing hemostasis in a subject by preparing and analyzing a test sample from the subject. BACKGROUND

[0005] Hemostasis (physiological control of bleeding) is a complex process involving the vasculature, platelets, coagulation factors, fibrinolytic proteins, and various activators and inhibitors.

[0006] Disruptions in hemostasis play a central role in the pathogenesis of myocardial infarction, stroke, pulmonary embolism, deep vein thrombosis, and excessive bleeding. Thus, there is an urgent need for in vitro diagnostics (IVD) to quantify hemostatic function / dysfunction and guide appropriate treatment.

[0007] Furthermore, the coagulation process is highly dependent on the temperature at which it occurs. Normally, coagulation occurs at body temperature, which is optimal for proper enzymatic action of the cascade of coagulation factors.

[0008] Preparation of the blood to be tested is of great importance, as the manner in which the blood sample is prepared prior to evaluation can affect, for example, the action of the vasculature components, platelets and other cellular components, coagulation factors, fibrinolytic components, and any inhibitors or activators of hemostasis. SUMMARY

[0009] The present disclosure provides devices, systems, and methods for assessing hemostasis. For example, disposable systems for hemostasis function analysis are provided. In some embodiments, a disposable system includes a multi-channel or multi-chamber test cartridge device configured to operate with a test system for assessing hemostasis in a subject by assessing a test sample from the subject in vitro. In some embodiments, the disposable system is configured to interrogate the test sample to assess clot firmness, strength, or other mechanical properties of the test sample to assess the function of various physiological processes that occur during clotting and / or dissolving the resulting clot. The sample can include, in whole or in part, whole blood, plasma, platelet-rich plasma, or platelet-poor plasma. In addition, the sample can include one or more reagents (such as an anticoagulant or antiplatelet drug that can be present in the blood at the time of collection), or one or more pharmacological treatment substances (such as in the case of heparin or low molecular weight heparin) or other inert components (such as polystyrene beads) added to the test sample prior to use of the cartridge device. The disposable system facilitates point-of-care assessment of hemostasis in a test sample that is robust (e.g., can be performed in a non-laboratory setting), fast (e.g., can be completed in just a few minutes), easy to use, and provides clear results (e.g., directly related to the functional components of hemostasis), and facilitates identification of accurate hemostasis defects. Exemplary devices automatically perform one or more pre-measurement steps, minimizing the number of sample manipulation steps required by the user, thereby improving test reproducibility and / or test quality. In some embodiments, the disposable system includes a plurality of test circuits, each test circuit having a path defined by channels and chambers configured to prepare a blood test sample for assessment by a 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.

[0010] In some embodiments, the disposable system is configured to condition the respective test sample prior to, during, and / or after mixing with the reagent to optimize the proper action of the available blood components and chemicals being assessed (e.g., vasculature components, platelet or other cellular components, clotting factors, fibrinolytic components, and any other inhibitors or activators of hemostasis function, etc.).

[0011] In an aspect, a device (e.g., a cartridge) for hemostasis assessment is disclosed. The device includes a housing; an input port integrally formed with the housing, the input port structurally configured to establish fluid communication and expel contents of a sample holding tube; and a first chamber in fluid communication with the input port. The first chamber is configured to receive a sample contained in the sample holding tube and to condition the received sample to a desired temperature (e.g., a predetermined temperature range) prior to allowing the received sample to contact one or more reagents in one or more fluidic circuits located downstream of the first chamber, wherein each of the one or more fluidic circuits includes 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 a combination of reagents (e.g., in the form of lyophilized reagent beads) located in a corresponding fluidic circuit (e.g., a reagent bag) to form a mixed sample, and ii) a test chamber in fluid communication with the second chamber, the test chamber structurally configured to be interrogated by a measurement system configured to determine a property (e.g., mechanical or viscoelastic) of the mixed sample.

[0012] In some embodiments, at least one of the one or more fluidic circuits includes one or more bags (e.g., each bag configured to hold lyophilized reagent beads, the lyophilized reagent beads including a reagent or a combination of reagents).

[0013] In some embodiments, at least one of the one or more fluidic circuits includes one or more reservoir bags (e.g., each bag configured to hold a liquid form of an assay, the assay including a reagent or a combination of reagents).

[0014] In some embodiments, at least one of the one or more fluidic circuits includes one or more lyophilized reagents disposed on one or more surfaces thereof (e.g., lyophilized on each surface; lyophilized into a thin film disposed on or adhered to the one or more surfaces).

[0015] In some embodiments, at least one of the one or more fluidic circuits includes one or more reagents treated on a surface thereof (e.g., dried on the surface; sprayed on the surface; baked on the surface).

[0016] In some embodiments, the input port is communicatively coupled to a pressure port, wherein a pressure applied to the pressure port causes the contents of the sample holding tube to be expelled through the input port to the first chamber.

[0017] In some embodiments, the input port includes a needle assembly.

[0018] In some embodiments, the needle assembly includes an input port and a second port, where the second port is configured to discharge liquid or gas into the sample holding tube so as to facilitate the expulsion of contents therein. In some embodiments, the input port is located (e.g., concentrically located) within the second port, which is configured to discharge liquid or gas into the sample holding tube, thereby expelling the contents of the sample holding tube.

[0019] In some embodiments, the input port includes a luer lock configured to connect to the sample holding tube, where the sample holding tube is a syringe.

[0020] In some embodiments, the input port is communicatively coupled to the first pressure port, where the pressure, when applied to the first pressure port, causes the contents of the sample holding tube to be expelled through the input port to the first chamber.

[0021] In some embodiments, the first chamber is configured to cooperate with a corresponding thermal regulation system (e.g., heating / cooling system) of the measurement system to regulate the received sample to or near a desired temperature.

[0022] In some embodiments, the shape and / or material of the first chamber is optimized to facilitate the thermal regulation (e.g., heating and / or cooling) of the sample to or near a desired temperature.

[0023] In some embodiments, the first chamber is configured to cooperate with a corresponding thermal regulation surface of a subsystem component of the measurement system to regulate the received sample to or near a desired temperature. In some embodiments, the channel portion of the one or more fluidic circuits is configured to cooperate with a corresponding heating / cooling system of the measurement system to regulate the received sample to or near a desired temperature.

[0024] In some embodiments, the channel portion of the one or more fluidic circuits is configured to cooperate with a corresponding thermal regulation system of the measurement system to regulate the received sample to a desired temperature. In some embodiments, the first chamber and / or the channel portion of the one or more fluidic circuits is in physical proximity (e.g., in physical contact or near contact) with a sensor configured to measure the temperature of the sample received in the first chamber.

[0025] In some embodiments, the sensor is selected from the group consisting of a thermistor, a thermocouple, and an optical sensor (e.g., an infrared sensor).

[0026] In some embodiments, the device includes a first pressure port in fluid communication with the first chamber, the first pressure port configured to accept a negative or differential pressure (e.g., to fill the first chamber); and a filter within the first pressure port in the at least one fluidic circuit (e.g., such that when the first chamber is filled, the filter is occluded by the sample received in the first chamber). In some embodiments, the filter is configured to allow air to move through the first pressure port, but prevent fluid from moving through.

[0027] In some embodiments, the device includes a first pressure port configured to accept a negative or differential pressure to fill the first chamber; and a first fluidic path extending from the first pressure port to the first chamber, wherein a filter is within the first pressure port.

[0028] In some embodiments, for each of the one or more fluidic circuits, fluidic communication between the first chamber and the second chamber is through a second fluidic path originating from a side (e.g., a sidewall, a bottom wall, etc.) of the first chamber (e.g., such that air bubbles present in a received sample are captured away from the second chamber).

[0029] In some embodiments, each of the one or more fluidic circuits includes a third fluidic path in fluidic communication with the second chamber, wherein the third fluidic path leads to a second pressure port configured to accept a negative or differential pressure to fill the second chamber.

[0030] In some embodiments, the second pressure port has a second filter therein, wherein the second filter is configured to occlude when the second chamber is filled.

[0031] In some embodiments, the device includes one or more fluidic paths for all of the one or more fluidic circuits in fluidic communication with the second pressure port, wherein the one or more fluidic paths are configured to provide a negative pressure to the second pressure port for all of the one or more fluidic circuits.

[0032] In some embodiments, for each of the one or more fluidic circuits, the second chamber is in fluidic communication with a vent port, wherein the vent port is configured to close when an aliquot of sample is metered into the second chamber, and further configured to open to atmospheric pressure after the aliquot of sample is metered into the second chamber.

[0033] In some embodiments, each of the one or more fluidic circuits includes a third set of fluidic paths in fluidic communication between a respective second chamber (e.g., a metering chamber) and a test chamber, wherein a portion of the third set of fluidic paths is arranged as a serpentine conduit or channel.

[0034] In some embodiments, each of the one or more fluidic circuits further includes a serpentine reservoir between the test chamber and the second chamber.

[0035] In some embodiments, the metered sample is optionally directed through a portion of one or more fluidic circuits to facilitate mixing of the metered sample and a reagent or combination of reagents.

[0036] In some embodiments, for each of the one or more fluidic circuits, the metered sample is alternatively and multiply directed between a first location in the fluidic circuit (e.g., a second chamber) and a second location in the fluidic circuit (e.g., a location in a serpentine reservoir).

[0037] In some embodiments, each of the one or more fluidic circuits further comprises a third pressure port in fluid communication with the second chamber and the test chamber, the third pressure port configured to accept a negative or differential pressure (e.g., for drawing an aliquot from the second chamber to the test chamber), wherein the third pressure port is further configured to alternatively accept an alternating pressure, e.g., for alternately drawing an aliquot from the second chamber along the serpentine reservoir and pushing the aliquot through the serpentine reservoir to the second chamber.

[0038] In some embodiments, the serpentine reservoir comprises an optical detection zone to facilitate optical detection of the metered sample in the serpentine reservoir or a location of the sample in the serpentine reservoir.

[0039] In some embodiments, each of the one or more fluidic circuits further comprises a mixing path between the test chamber and the second chamber, the mixing path comprising one or more ferromagnetic beads or rods therein.

[0040] In some embodiments, at least one of the one or more fluidic circuits comprises one or more mass test inlets.

[0041] In some embodiments, the one or more mass test inlets are configured to be optically sensed, wherein the mass test inlets are transparent.

[0042] In some embodiments, the one or more mass test inlets are configured to be electrically sensed, wherein the mass test inlets comprise one or more sensing electrodes.

[0043] In some embodiments, the one or more mass test inlets are configured to be sampled for a property of the metered sample (e.g., pressure, presence of flow, flow rate, temperature).

[0044] In some embodiments, for each of the one or more fluidic circuits, the test chamber comprises a mechanism to couple energy into the test chamber to perform a measurement, e.g., in the case of a lens configured to direct an ultrasonic pulse into the test chamber.

[0045] 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 being structurally capable of establishing fluid communication with a sample holding tube and expelling contents of the sample holding tube; a first chamber in fluid communication with the input port, the input port receiving a sample contained in a vacuum tube and thereby conditioning the sample to a desired temperature prior to the sample contacting one or more reagents; one or more second chambers in fluid communication with the first chamber, the one or more second chambers being configured to aliquot the sample in the first chamber into one or more aliquots; one or more reagent pockets, each reagent pocket being filled with one or more lyophilized reagent beads in fluid communication with each aliquot chamber and allowing the sample present in each aliquot to mix with the one or more reagent beads; and one or more test chambers in fluid communication with the aliquot chambers and being structurally capable of being interrogated to determine a sample viscoelasticity after the sample mixes with one or more reagents.

[0046] In some embodiments, the reagent or combination of reagents located in one or more fluidic circuits comprises an intrinsic pathway activator (e.g., kaolin, celite, glass, ellagic acid, micronized silicon dioxide, heparin, etc.) or combinations thereof.

[0047] In some embodiments, the reagent or combination of reagents located in one or more fluidic circuits comprises an extrinsic pathway activator (e.g., tissue factor, recombinant tissue factor, thromboplastin, etc.) or combinations thereof.

[0048] In some embodiments, the reagent or combination of reagents located in one or more fluidic circuits comprises a coagulation activator (e.g., thrombin, factor Xa, reptilase, ecarin, Russell's viper venom, or other snake venom, etc.) or combinations thereof.

[0049] In some embodiments, the reagent or combination of reagents located in one or more fluidic circuits comprises a platelet activator or platelet inhibitor (e.g., a GPIIb / IIIa inhibitor (e.g., abciximab, eptifibatide, tirofiban, roxifiban, orbofiban), cytochalasin D, blebbistatin, a PAR1 inhibitor, a PAR4 inhibitor, a glycoprotein IB inhibitor, TRAP, ADP, arachidonic acid, an ADP inhibitor, a non-steroidal anti-inflammatory drug, platelet-activating factor, ristocetin, epinephrine, etc.) or combinations thereof.

[0050] In some embodiments, the reagent or combination of reagents located in one or more fluid circuits comprises an activator or inhibitor of fibrinolysis (e.g., tPA, uKA, streptokinase, TAFIa, plasmin / plasminogen, aprotinin, epsilon-aminocaproic acid, tranexamic acid, plasminogen activator inhibitor 1 (PAI1), alpha 2-antiplasmin (a2-AP), or plasmin-antiplasmin complex, carboxypeptidase inhibitor) or combinations thereof.

[0051] In some embodiments, the reagent or combination of reagents located in one or more fluid circuits comprises an inhibitor of FXIIIa or combinations thereof.

[0052] In some embodiments, the reagent or combination of reagents located in one or more fluid circuits comprises thrombomodulin or combinations thereof.

[0053] In some embodiments, the reagent or combination of reagents located in one or more fluid circuits comprises low molecular weight heparin or combinations thereof.

[0054] In some embodiments, the reagent or combination of reagents located in one or more fluid circuits comprises hirudin (polybrene) or combinations thereof.

[0055] In some embodiments, the reagent or combination of reagents located in one or more fluid circuits comprises heparin or combinations thereof.

[0056] In some embodiments, the reagent or combination of reagents located in one or more fluid circuits comprises corn trypsin inhibitor or combinations thereof.

[0057] In some embodiments, the reagent or combination of reagents located in one or more fluid circuits comprises adenosine or combinations thereof.

[0058] In some embodiments, the reagent or combination of reagents located in one or more fluid circuits comprises GPRP (glycine-proline-arginine-proline) or combinations thereof.

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

[0060] In some embodiments, the reagent or combination of reagents located in one or more fluid circuits comprises fibronectin or combinations thereof.

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

[0062] In some embodiments, the reagent or combination of reagents located in one or more fluid circuits comprises an immunoassay reagent or combinations thereof.

[0063] In some embodiments, the reagent or combination of reagents located in one or more fluidic circuits comprises heparinase I or a combination therewith.

[0064] In some embodiments, the reagent or combination of reagents located in one or more fluidic circuits comprises endothelial cells or activated endothelial cells.

[0065] In some embodiments, the measurement system is selected from the group consisting of a system based on acoustic rheometry, a system based on thromboelastography, a system based on thromboelastometry, an optical based system, a fluorescence based system, a chromometry based system, an aggregation measurement based system, a resonance based system, and an electrical impedance based system.

[0066] In another aspect, a method of mixing a sample with one or more reagents and testing the mixed sample to assess hemostasis in a device (e.g., a cartridge) is disclosed. The method comprises: receiving a plurality of metered samples from a plurality of metering chambers, the metering chambers receiving a test fluid from a sample holding tube (e.g., via a mechanical coupler connecting the device to the sample holding tube or via an opening into which the sample holding tube places its sample); alternately and multiplicatively flowing each aliquot until the aliquot is mixed with a reagent or combination of reagents to form a mixed aliquot, wherein at least one aliquot is alternately and cyclically flowed i) from the metering chamber in a first direction through one or more reagent pockets having one or more reagents (e.g., lyophilized reagent beads) and along a serpentine path in communication with the metering chamber until at least a portion of the aliquot reaches a detection zone located in or after the serpentine path, and ii) from the detection zone in a second direction opposite the first direction through at least a portion of the serpentine path toward the metering chamber until a triggering event occurs; and driving the mixed aliquot in a test chamber in fluid communication with the metering chamber, wherein the test chamber is structurally configured to be interrogated by a measurement system configured to determine a property (e.g., mechanical or viscoelastic) of the mixed aliquot, and wherein the interrogation of the test chamber is performed with the mixed aliquot located therein.

[0067] In some embodiments, the method comprises receiving a fluid in a first chamber, the first chamber configured to substantially regulate a temperature of a test sample to a body temperature or other desired temperature, wherein the metered sample received in the metering chamber is received from the first chamber.

[0068] In some embodiments, the test fluid is moved into the first chamber in response to an applied pressure exerted or generated by the measurement system.

[0069] In some embodiments, the method includes adjusting the test fluid in the first chamber to or substantially near a desired temperature, where the test fluid mixes with the one or more reagents after exiting the first chamber.

[0070] In some embodiments, the method includes isolating (e.g., blocking via a valve) the test fluid in the metering chamber from contacting the one or more reagents to prevent the test fluid from contacting the one or more reagents during filling of the metering chamber.

[0071] In some embodiments, the second applied positive or negative pressure is applied or generated by the measurement system at a second port in communication with the serpentine path (e.g., at a second pressure port in communication) to move the at least one aliquot in a second direction.

[0072] In some embodiments, the first applied positive or negative pressure is applied or generated in reverse by the measurement system to move the at least one aliquot in a second direction.

[0073] In some embodiments, the operation of receiving the mixed aliquot in the test chamber further includes accepting a negative pressure via a third pressure port, where the third pressure port is further in fluid communication with the test chamber.

[0074] In some embodiments, the test chamber is downstream of the serpentine path, and the third pressure port is downstream of the test chamber.

[0075] These and other features and advantages of the present application will become more apparent from the following detailed description and accompanying drawings, which describe and illustrate preferred and alternate embodiments of the present application. BRIEF DESCRIPTION OF DRAWINGS

[0076] The accompanying drawings incorporated in and forming a part of the specification, illustrate embodiments of the present application and, together with the description, serve to explain the principles of the method and system.

[0077] Figure 1 A perspective view of an example biological sample input for a cartridge for a disposable system is shown in accordance with illustrative embodiments.

[0078] Figure 2 A side cross-sectional view of an example biological sample input having a housing is shown in accordance with illustrative embodiments. Figure 1

[0079] A side cross-sectional view of an example biological sample input having a housing is shown in accordance with illustrative embodiments. Figure 3 Figure 2 A side cross-sectional view of an example biological sample input having a housing is shown in accordance with illustrative embodiments.

[0080] Figure 4 A side cross-sectional view of an example biological sample input having a housing is shown in accordance with illustrative embodiments. Figure 3 ​a detailed view of an example biological sample input.

[0081] Figure 5A and Figure 5B Each illustrates a biological fluid path of four test loops (e.g., hemostasis test loops) on a sample preparation plane, according to an illustrative embodiment. Figure 5B Further illustrates a cartridge body of Figure 5A , further coupled to a sample holding tube, according to an illustrative embodiment.

[0082] Figure 6A and Figure 6B Illustrates a front perspective view and a rear perspective view of Figure 5A , Figure 5B and Figure 8 , with markers corresponding to heating chamber fill.

[0083] Figure 6C and Figure 6D Illustrates a front perspective view and a rear perspective view of Figure 5A , Figure 5B and Figure 8 , with markers corresponding to sample chamber fill.

[0084] Figure 6E and Figure 6F Illustrates a front perspective view and a rear perspective view of Figure 5A , Figure 5B and Figure 8 , with markers corresponding to sample mixing and test chamber fill.

[0085] Figure 7 Illustrates a back side of a cartridge of Figure 5A , according to an illustrative embodiment, and includes an interconnect plane that interfaces with a sample preparation plane, which together form a biological fluid path for test loops.

[0086] Figure 8 Illustrates a portion of a biological fluid path on an interconnect plane of Figure 7 , according to an illustrative embodiment.

[0087] Figure 9 Illustrates an example test chamber portion for use with an example cartridge, according to an illustrative embodiment.

[0088] Figure 10 Illustrates a cross-sectional view of an example test chamber in an example test chamber portion, according to an illustrative embodiment.

[0089] Figure 11 Illustrates a detailed cross-sectional view of an example test chamber of Figure 10 , according to an illustrative embodiment.

[0090] Figure 12A cross-sectional view of a disposable system operatively coupled to a measurement system, according to an illustrative embodiment, is shown.

[0091] Figure 13 An example shear modulus versus time curve is shown according to an illustrative embodiment.

[0092] Figure 14 Examples of shear modulus curves obtained with coagulation activators and with and without fibrinolysis inhibitors are shown. Comparisons of these curves can provide information about the fibrinolytic activity of the samples.

[0093] Figure 15 A possible embodiment of a difference measure is shown, which can be measured based on shear modulus curves obtained with and without a coagulation activator and fibrinolysis inhibitor.

[0094] Figure 16 The illustration shows a system used in a disposable system according to an illustrative embodiment. Figure 1 , Figure 2 , Figure 3 , Figure 4 , Figure 5A , Figure 5B , Figure 7 , Figure 8 and Figure 9 A photograph of an exemplary medicine cartridge.

[0095] Figure 17 An illustrative embodiment is shown. Figure 1 , Figure 2 , Figure 3 , Figure 4 , Figure 5A , Figure 5B , Figure 7 , Figure 8 and Figure 9 A front view of an exemplary cartridge.

[0096] Figure 18 An illustrative embodiment is shown. Figure 1 , Figure 2 , Figure 3 , Figure 4 , Figure 5A , Figure 5B , Figure 7 , Figure 8 and Figure 9 A front view of an exemplary cartridge. Detailed Implementation

[0097] The present application will now be described more fully hereinafter with reference to the accompanying drawings, in which embodiments of the application are shown. The present application may, however, 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. Like reference numerals refer to like elements throughout.

[0098] As used in the specification and the appended claims, the singular forms "a," "an" and "the" include plural referents unless the context clearly dictates otherwise.

[0099] As used herein, the term "comprises" and variations thereof are synonymous with the term "includes" and variations thereof and are open-ended, non-limiting terms.

[0100] As used throughout, "subject" refers to an individual. The subject can be a vertebrate, more specifically a mammal (e.g., a human, a horse, a pig, a rabbit, a dog, a sheep, a goat, a non-human primate, a cow, a cat, a guinea pig, or a rodent), a fish, a bird, or a reptile or amphibian. The term does not connote a particular age or sex.

[0101] The devices described herein include single-use cartridge devices configured to facilitate the assessment of one or more hemostatic functions in vitro. Hemostatic functions refer to the functional roles of various blood components, such as coagulation factors, fibrinogen, platelets, fibrinolytic factors, and components of the vasculature. In some embodiments, the cartridge devices and associated measurement systems are configured to assess hemostatic functions by measuring changes 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 devices and their test chambers are configured to facilitate the measurement of viscoelasticity, for example based on interrogation using ultrasound pulses or ultrasound energy. However, other interrogation systems can be used with cartridge devices having the features described herein. Examples of other interrogation systems include, for example, but are not limited to, systems employing cup / spike technology (as in the case of thromboelastography and thromboelastometry), vibrating pistons to measure changes in mechanical impedance, optical sensing, fluorescence sensing, colorimetric sensing, aggregation measurements, resonance sensing, or electrical impedance sensing, etc.

[0102] The cartridge device can use a variety of reagents, including intrinsic pathway activators (not limited to kaolin, Haegmann factor, diatomaceous earth, glass, ellagic acid, micronized silica, etc.), extrinsic pathway activators (not limited to tissue factor, recombinant tissue factor, thrombin, etc.), other coagulation activators (not limited to thrombin, factor Xa, hemostatic agents, snake vein enzyme, Russell venom or other snake venom, etc.), platelet activators or platelet inhibitors (not limited to GPIIb / IIIa inhibitors (such as abciximab, epifetabide, tirofiban, roxifiban, olpofibrateban), cytochalasin D, blisterstatin, PAR1 inhibitors, PAR4 inhibitors, glycoprotein IB inhibitors, TRAP, ADP, arachidonic acid, ADP inhibitors, nonsteroidal anti-inflammatory drugs, etc.), fibrinolytic function activators. Or fibrinolytic inhibitors (not limited to tPA, uKA, streptokinase, TAFIa, plasmin / plasminogen, aprotinin, ε-aminocaproic acid, tranexamic acid, plasminogen activator inhibitor 1 (PAI1), α2-anti-plasmin (α2-AP) or plasmin-anti-plasmin complex, carboxypeptidase inhibitors, etc.), as well as others (FXIIIa inhibitors, hymexazol (polyhexamethylenetetramine), heparinase (e.g., heparinase I), ristocetin, heparin, low molecular weight heparin, zeatinase inhibitors, adenosine, GPRP, calcium, fibronectin, collagen, adrenaline, immunoassay reagents, direct thrombin inhibitors, factor Xa inhibitors, reagents designed to reverse or eliminate the effects of new oral anticoagulants (such as direct thrombin inhibitors and factor Xa inhibitors), thrombomodulin, etc.). Additional non-functional reagents can also be used to preserve the functionality of other reagents (buffers and stabilizers for lyophilization or drying, dyes, etc.).

[0103] In some embodiments, reagents are placed and stored in a chamber of the cartridge device (e.g., a bag located in a fluid loop), but in alternative embodiments, reagents may be placed and stored in various chambers or fluid channels within the fluid loop of the cartridge device. A fluid loop generally refers to one or more fluid paths established between one or more test chambers for sample preparation and final measurement of the sample.

[0104] In some embodiments, the reagent is placed and stored in liquid form in a cartridge device, or it can be freeze-dried in spheres (such as Lyopheres manufactured by BioLyph LLC). TM In some cases, the reagents are freeze-dried in a film, freeze-dried on a plastic surface, dried on a plastic surface, or sprayed, etc., to improve stability during the shelf life. Those skilled in the art should recognize that these reagents are not exhaustive; other reagents or combinations of reagents, acting as inhibitors or activators of one or more hemostatic functions, may be used in this cartridge.

[0105] The cartridge device disclosed herein is a component of a measurement system (e.g., a hemostasis measurement system). The measurement system (also referred to as an instrument) includes at least one interface element coupled between the cartridge device and a measurement element configured to measure the viscoelasticity or mechanics of a sample being processed within the cartridge device. The measured viscoelasticity or mechanics is output as a result to a user interface. An example user interface is described in commonly assigned U.S. Publication No. 2011 / 0252352 to Viola et al., which is incorporated by reference herein in its entirety.

[0106] In some embodiments, the interface element includes one or more heating and / or cooling elements.

[0107] In some embodiments, the interface element includes a fluidic manifold that facilitates connection to one or more pump elements and one or more valves.

[0108] In some embodiments, the interface element includes one or more sensors configured to perform, for example, a hemostasis measurement. In some embodiments, the one or more sensors include an ultrasonic sensor. In other embodiments, the one or more sensors include other interrogation devices that are based on thromboelastography, thromboelastometry (e.g., a system based on thromboelastography or a system based on thromboelastometry), or that measure changes in mechanical impedance, changes in observed perturbations via an optical-based system (e.g., with optical sensors), fluorescence, a colorimetric-based system, a system based on aggregation measurements (e.g., with optical sensors, acoustic sensors, or electrodes that measure aggregation of a test sample), a system based on resonance (e.g., with optical, acoustic, or mechanical position sensors that measure a sample when it is at or near resonance), a system based on electrical impedance (e.g., with electrodes configured to measure electrical impedance), or combinations thereof.

[0109] In some embodiments, the interface element includes a mechanical clamp configured to position the cartridge device in a desired orientation with respect to components of the measurement system (one or more sensors, fluidic manifolds, heating and / or cooling elements, etc.). When the interface element is engaged with the components of the measurement system, in some embodiments, the cartridge device is driven via a series of controlled actions coordinated by the measurement system to prepare a test sample for measurement. In some embodiments, the preparation operations include aspirating a sample from a sample container (also referred to as a sample holding tube), heating and / or cooling the sample, performing a sample quantification, mixing the sample with reagents, and measuring the sample. Each of these steps is described below with reference to various embodiments. Upon completion of the measurement, the results are output in the instrument user interface.

[0110] In some embodiments, the cartridge device and its internal components are the only components that come into direct contact with the sample to be analyzed.

[0111] In some embodiments, the cartridge includes computer readable information (e.g., radio frequency identification tags, computer readable media such as flash memory integrated circuits, two-dimensional codes, bar codes, etc.) and / or human readable information (e.g., indicia) that can be optically or communicatively interrogated.

[0112] The various embodiments described below do not use any active valve elements in the cartridge design, but rather rely on fluidic manifolds and one or more valves placed in the instrument. Fluid moves through the various cartridge components via pressure differential and / or gravity and / or material properties (as in the case of hydrophobic or hydrophilic) and / or capillary forces.

[0113] In these embodiments, the cartridge is configured to couple with the instrument through one or more connection ports that are aligned via alignment slots. The connection ports include one or more pressure ports and one or more vent ports. However, in alternative embodiments, actuated valves (as in the case of elastomeric valves) can be included in the cartridge design to control fluid flow. In some embodiments, these valves are driven by corresponding hardware and software components in the measurement system.

[0114] The surface properties and texture of the cartridge surfaces that come into direct contact with the sample can be optimized to promote sample adhesion and / or sample flow. In some embodiments, the interior surfaces of the test chamber and / or other interior surfaces of the fluidic circuit within the cartridge device are plasma treated to optimize surface energy and texture for adhesion of particular plasma proteins. In other embodiments, the interior surfaces of the test chamber and / or other interior surfaces of the fluidic circuit are textured with surface roughness, material coated (as in the case of gold plating), biological material coated (e.g., in the case of fibronectin or collagen coating), raw material selection (e.g., using a particular plastic or other material for the plate that does not require additional treatment), etc. Such treatment can be performed independently or in combination with plasma treatment. Similarly, the cartridge material can be selected or manipulated to obtain a desired hydrophobicity or hydrophilicity. These properties can be altered through plasma treatment or surface coating.

[0115] As described in greater detail below, the cartridge and associated measurement system can 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. In addition, the output of such one or more sensors can be further used to perform quality control checks. These checks can be performed before, during, or after cartridge testing to ensure the functionality of one or more subsystems (e.g., ultrasonic or other interrogation system, fluidics, liquid level, clamping, cartridge positioning / orientation system, or temperature control), to ensure proper cartridge functionality, to ensure proper sample preparation before a measurement is taken or before a measurement is completed, and can also be used to accept or reject test results, or even abort a test before a measurement is started.

[0116] Note that in the following discussion, a fluidic circuit includes a channel with fluidic components connecting one or more chambers together. The fluidic circuit is also referred to as a test channel in a plurality of channels that can be individually and controllably processed in a single cartridge device.

[0117] Cartridge input section Figure 1 、 Figure 2 、 Figure 3 and Figure 4 is a schematic view of an example biological sample input portion of a cartridge 100 for assessing hemostasis. In particular, Figure 1 shows a perspective view of an example biological sample input of a cartridge for use in a disposable system, according to an illustrative embodiment. Figure 2 shows a side view of an example biological sample input of a cartridge having a housing, according to an illustrative embodiment. Figure 1 shows a side view of an example biological sample input of a cartridge having a housing, according to an illustrative embodiment. Figure 3 shows a side view of an example biological sample input of a cartridge having a housing, according to an illustrative embodiment. Figure 2 shows a side view of an example biological sample input of a cartridge having a housing, according to an illustrative embodiment. Figure 4 shows a detailed view of an example biological sample input of a cartridge, according to an illustrative embodiment. In alternative embodiments, the input portion of the cartridge includes a well into which a fluid sample can be placed, for example, by a pipette or test tube. Figure 3

[0118] In some embodiments, as shown in Figure 1 , the cartridge 100 has dual connection tabs 28a, 28b for coupling the cartridge 100 to the sample container guide 1, as shown in Figure 2 . As shown in Figure 2 , when mated with the cartridge 100, the sample container guide 1 aligns the sample container 2 to the sample input port 3 of the cartridge 100. The cartridge 100 also includes an alignment tab 29 configured to slide into an alignment groove 30 of the sample container guide 1 to further stabilize the coupling of the sample container guide 1 to the cartridge 100. The sample container guide 1 can also provide a hard stop 5, as shown in Figure 3 , to hold the sample container 2 at an appropriate height to establish fluid communication with the cartridge 100.

[0119] In various embodiments, the sample container 2 is a vacuum tube, such as a BD Vacutainer TM tube, and the sample input port 3 includes a sample transfer 3a and one or more needles required for venting 4 (see Figure 1 ). Although shown as concentric in the figures, the needles can be configured as concentric, side-by-side, or integrated. In some embodiments, as shown in Figure 1 ​As shown, the sample delivery needle 3a includes an inlet (3b and 3d) and an outlet 3c, with the outlet 3c terminating at the sample inlet chamber 26 of the cartridge 100. The sample inlet chamber 26 is in fluid communication with an inlet path 8 leading to the holding / heating chamber 6 (see [reference]). Figure 5A In some embodiments, such as Figure 1 As shown, the vent needle 4 includes an outlet 4a configured to terminate within the sample container 2 upon attachment and spaced apart from the inlet 3d to minimize air bubbles drawn into the inlets 3b and 3d. The vent needle 4 also has an inlet 4b terminating in the vent inlet chamber 27 of the cartridge. The vent inlet chamber 27 is in fluid communication with the vent passage 9, which in some embodiments terminates in a filter chamber 9a housing a filter (e.g., ...). Figure 5A (As shown in the diagram). An alternative sample container 2, such as a syringe, can be used, which requires a Luer lock connection on the cartridge 100. Furthermore, as mentioned above, in other embodiments, the inlet portion of the cartridge includes a well into which a fluid sample can be placed, for example, via a pipette or tube.

[0120] Exhaust path Figure 5A , Figure 5B , Figure 7 and Figure 8 This is a schematic diagram of the biofluid path of an exemplary cartridge 100 according to an embodiment. Specifically, Figure 5A and Figure 5B Each illustrates four test loops located on the sample preparation plane according to an illustrative embodiment (corresponding to...). Figure 5A The biofluid pathways of test chambers 16a, 16b, 16c, and 16d shown herein are also referred to herein as hemostasis test circuits. Although four test circuits are shown, more or fewer circuits may be included, such as two, three, five, six, seven, eight, etc. Figure 5B Further illustrations are shown according to illustrative embodiments. Figure 5A The cartridge body is further coupled to the sample holding tube 2. Figure 7 An illustrative embodiment is shown. Figure 5A The back of the cartridge includes an interconnecting plane that engages with the sample preparation plane, together forming a biofluidic path for the test circuit. Figure 8 The illustration shows an example of a method in which... Figure 7 The portion of the biofluid pathway on the interconnected plane.

[0121] As described above, the biofluid pathway is formed on and through a plurality of planes defined in the cartridge 100. Figure 5A and 5BA first fluid path plane of the cartridge 100 is shown. The first fluid path plane of the cartridge 100 can alternatively be referred to as the front plane of the cartridge 100. Figure 7 and Figure 8 Each shows a second fluid path plane of the cartridge 100, wherein for ease of understanding, Figure 8 The fluid path in the second plane is shown in isolation from the rest of the structure of the cartridge 100. The second fluid path plane of the cartridge 100 can alternatively be referred to as the back plane of the cartridge 100. The fluid path between the first and second planes is connected by fluid passageways that pass through different planes of the cartridge 100.

[0122] As discussed above in connection with Figure 1 In some embodiments, the vent inlet chamber 27 is in fluid communication with a vent path 9. The vent path 9 can terminate in a filter chamber 9a (as shown in Figure 5A The filter chamber 9a in the first fluid path plane of the cartridge 100 is in fluid communication with a vent port 22i (as shown in Figure 8 the cartridge 100 via the vent port 22i to provide atmospheric pressure to the vent path 9, as discussed in more detail below.

[0123] Heating chamber path As discussed above in connection with Figure 1 In some embodiments, the sample delivery needle outlet 3c terminates in a sample inlet chamber 26 of the cartridge 100. The sample inlet chamber 26 is in fluid communication with an inlet path 8. The sample inlet path 8 provides a fluid communication path between the sample inlet chamber 26 and a holding / heating chamber 6 (also referred to herein as the heating chamber 6 or “first chamber”), as shown in Figure 5A The designations “first,” “second,” and “third” as used herein are provided merely as designations and are not intended to imply an order. The heating chamber 6 is configured to cooperate with a corresponding thermal regulation (e.g., heating / cooling) system in the measurement system to heat or cool the sample to or near a predetermined temperature.

[0124] As provided herein, the heating chamber 6 helps to uniformly condition the test fluids prior to being metered or aliquoted into their respective tests, thereby reducing variability in the test samples that can affect subsequent measurements and analysis. The shape of the heating chamber 6 can be optimized for heating / cooling transfer, as is the case here with a thin cross-section having a thin wall. The material of the cartridge 100 can also be optimized to facilitate heating / cooling. In some embodiments, the sample heating / cooling conditioning phase can also be implemented in one or more chambers / channels of the cartridge design and is not limited to occurring only in the heating chamber 6. In some embodiments, a stirring, rotating, or oscillating element (not shown) can be placed in the heating chamber 6, which can be controlled by the measurement system to facilitate uniform temperature heating or cooling. In other embodiments, the test fluids in the heating chamber 6 can be vibrated by the measurement system vibrating the cartridge 100 to facilitate uniform temperature conditioning of the test fluids.

[0125] In some embodiments, temperature measurements are taken of the test samples in the cartridge 100. To measure temperature, a sensor can be incorporated in the measurement system or the cartridge 100. In some embodiments, a thermistor or thermocouple can be placed in physical contact with the cartridge 100 or the biological sample, such as blood. In other embodiments, an infrared thermometer is directed at the cartridge 100 or the biological sample. In either case, the cartridge 100 can include a small well for the blood to pass into, rather than being in direct contact with the blood. In some embodiments, the temperature of the test sample can be assessed at or near the heating chamber 6. In other embodiments, the temperature of the test sample can be assessed while the test sample is flowing through the channel as it is being directed toward the test chamber 16

[0126] Referring now to Figure 5A , Figure 5B and Figure 8 , the sample inlet path 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 Figure 5A and Figure 5B . In some embodiments, the chamber along the fluid path is typically filled from the top to prevent backflow of blood into the inlet. A fill outlet channel 10a extends from a second corner 6b of the heating chamber 6 opposite the first corner 6a.

[0127] The fill outlet channel 10a extends to a filter chamber 10 having a filter therein. The filter chamber 10 in the first fluid path plane of the cartridge 100 (e.g., as shown in Figure 5A and Figure 5B ) is in fluid communication with a heating chamber fill channel 10b shown in the second fluid path plane of the cartridge 100 (see Figure 8 ). The fill conduit 10b is in fluid communication with the pressure port 22a (see also Figure 8) in fluid communication, pressure port 22a facilitates filling of the heating chamber 6. Conduit 10b is part of a network of conduits that serve to integrate the pressure ports of the cartridge 100 (e.g., 22a, as discussed, and 22b-22i, to be discussed later) to one or more regions with which the measurement system can couple its pressure control interface. This configuration reduces the complexity of the measurement system that controls the movement of fluids within the cartridge 100. In fact, conduits that handle the control of movement of fluid samples in the first plane of the cartridge 100 are primarily placed in the second plane of the cartridge 100. Figure 6A and Figure 6B front and rear perspective views of Figure 5A , Figure 5B and Figure 8 are shown with additional markings corresponding to the portion of the description.

[0128] Heating chamber fill In operation, the fluid pump of the instrument draws a sample through the input port 3 (see Figure 7 ), also referred to herein as a pressure port, via the connection port 22 (see Figure 1 to Figure 2 ) and sucks it into the heating chamber 6 (see Figure 5A or Figure 5B ) of the cartridge 100. For example, the fluid pump of the instrument can be in communication with and apply a differential pressure (e.g., positive or negative pressure) to the pressure port 22a (see Figure 8 ). This, in turn, creates a pressure applied along the fill conduit 10b, within the heating chamber 6, and along the inlet path 8 to draw the sample into the heating chamber 6. At the same time, the internal vent needle 4 is connected to the isolated path 9 that receives atmospheric pressure from the instrument via the vent port 22i (see Figure 8 ) to counteract the pressure in the sample container 2 as the sample is drawn into the heating chamber 6 of the cartridge 100. During the filling of the heating chamber 6, all other ports (e.g., 22b-22i) are closed, e.g., by the measurement system.

[0129] When the heating chamber 6 is filled, the filter within the filter chamber 10 becomes clogged and creates a pressure spike that is detected by the instrument, causing the instrument to shut off the jet pump. The instrument can also shut off the vent port 22i or stop supplying atmospheric pressure via the vent port 22i upon detecting the pressure spike. Alternative fill detection techniques can also be used, i.e., optical sensors placed at the desired fill level, volume control, fixed time of pressure change (negative and / or positive), ultrasonic detectors placed at the desired fill level, etc. The sample remains in the heating chamber until a desired temperature is reached, which can be, for example, (or close to) normal and typical subject body temperature (e.g., about 37°C for a healthy human). In other cases, other desired temperatures can be needed. The shape of the heating chamber 6 and the passage to the sample metering chamber 11 (described below) are configured so that air bubbles that can be present in the fluid sample are captured away from the rest of the fluidic circuit. The shape of the inlet path 8 includes an anti-siphon feature 8a (see Figure 5A and Figure 5B ) and is configured to reduce the occurrence of air bubbles forming in the heating chamber 6 and to prevent siphoning to and from the sample container 2. To this end, after heating and / or cooling the first drawn test sample, e.g., when the processed test sample is pulled into the metering chamber 11 (also referred to herein as the sample chamber 11 and the “second” chamber), additional unprocessed test sample (e.g., unheated blood) is prevented from being siphoned into the heating chamber. Figure 6A and Figure 6B Also shown are indicia corresponding to this portion of the description.

[0130] Sample aliquot chamber path Referring to Figure 5A , Figure 5B and Figure 8 , a first side 6c (see Figure 5A ) of the heating chamber 6 extends between a first corner 6a and a second corner 6b.

[0131] One or more outlet ports 6e-6h (see Figure 5A ) are arranged along the length of the second side 6d of the heating chamber. Each of the one or more outlets 6e-6h can be arranged in a different one of the recesses along the second side 6d of the heating chamber, where the recesses direct the sample into a conduit leading to each respective test channel. In some embodiments, a test channel refers to the associated fluid path structure and test chamber collectively used to make a measurement on a given aliquot. In some embodiments, as shown in Figure 5B , each of the one or more outlets 6e-6h in the first fluid path plane of the cartridge 100 is in fluid communication with a first end of a respective one or more channels 20a-20d (see Figure 8 ) in the second fluid path plane of the cartridge 100, which are collectively referred to as channels 20 (seeFigure 5B ). The second end of each of the one or more channels 20a-20d (see Figure 8 ) in the second fluid path plane of the cartridge 100 is likewise in fluid communication with the first end of a corresponding one or more channels 11a-11d (see Figure 5B ) in the first fluid path plane of the cartridge 100. The second end of each of the one or more channels 11a-11d terminates in a respective one or more sample chambers 11 (shown repeatedly in the “o” symbol (“x4”) in Figure 5B . In the example shown in Figure 5A , Figure 5B , Figure 7 and Figure 8 , there are four sample chambers 11. In some configurations, there can be more or fewer sample chambers 11 on the cartridge 100 and corresponding fluid communication paths with the heating chamber 6.

[0132] The sample chambers 11 are fed by one or more channels 20 originating from the bottom of the heating chamber 6. This geometry avoids the aspiration of air bubbles into the sample chambers 11 when air bubbles rise to the upper portion of the heating chamber 6.

[0133] Each sample chamber 11 has a respective fill channel 11e that is in fluid communication with a respective filter chamber 12 (shown repeatedly in the “+” symbol (“x4”) in Figure 5B ) having a filter therein. The filter chambers 12 in the first fluid path plane of the cartridge 100 are in fluid communication with a channel 12a (see Figure 8 ) in the second fluid path plane of the cartridge 100. The channel 12a is in fluid communication with the pressure port 22g (see Figure 8 ).

[0134] In some configurations, when more than one sample chamber 11 is implemented on the cartridge 100, the channel 12a (see Figure 8 ) is in fluid communication with all of the sample chambers 11 (see Figure 5B ) via the respective fill channels 11e and filter chambers 12. Thus, the channel 12a acts as a manifold for applying negative pressure to all of the sample chambers 11 via a single pressure port 22g. Thus, it is advantageous that a separate pressure port is not needed to fill each sample chamber 11. Figure 6C and Figure 6D show front and rear perspective views of Figure 5A , Figure 5B and Figure 8 , with additional labeling corresponding to the description of this section.

[0135] Heating chamber exhaust path Reference is made to Figure 5A , Figure 5B and Figure 8Heating chamber 6 includes venting paths 31a, 31b, 31c along a first side 6c of the heating chamber for venting the heating chamber 6 as the sample chamber 11 is filled. The venting channel 31 (not shown) includes a fluid path through conduit elements 31a-31d. Channels 31a-31b terminate at one end of the heating chamber 6 along the first side 6c and at the other end in a filter chamber 31c having a filter therein. Thus, channels 31a-b provide a fluid path between the heating chamber 6 and the filter chamber 31c. The filter chamber 31c (see Figure 5A ) in the first fluid path plane of the cartridge 100 is in fluid communication with a channel 31d (see Figure 8 ) in the second fluid path plane of the cartridge 100, which in turn is in fluid communication with the vent port 22c (see Figure 8 ). As discussed in greater detail below, the cartridge 100 is coupled with the instrument via the vent port 22c of the instrument to provide atmospheric pressure to vent the heating chamber 6. Figure 6C and Figure 6D are also shown corresponding to this section of the description.

[0136] Sample chamber exhaust path Referring to Figure 5A , Figure 5B and Figure 8 , each sample chamber 11 includes a venting path 18 that terminates at a first end of the respective sample chamber 11. The second end of the venting path 18 in the first fluid path plane of the cartridge 100 is in fluid communication with a vent manifold 18a in the second fluid path plane of the cartridge 100. When there is more than one sample chamber 11, all of the venting paths 18 (see Figure 5B ) of the sample chambers are in fluid communication with the vent manifold 18a (see Figure 8 ). The vent manifold 18a (see Figure 8 ) in the second fluid path plane of the cartridge 100 is further in fluid communication with a first end of a channel 18b (see Figure 5B ) in the first fluid path plane. A second end of the channel 18b (see Figure 5B ) is in fluid communication with a filter chamber 18c (see Figure 5B ) having a filter therein. The filter chamber 18c (see Figure 5B ) in the first fluid path plane of the cartridge 100 is in fluid communication with the vent port 22e (see Figure 8 ). Figure 6C and Figure 6D are also shown corresponding to this section of the description.

[0137] Sample aliquot chamber fill During operation, once the sample is at or near the desired temperature, the sample is aliquoted (or metered) into one or more independent sample chambers 11 (see Figure 5B). Referring to Figure 5B , unless otherwise noted, in various embodiments, the sample chambers are filled by applying negative pressure at pressure port 22g (see Figure 8 ) via a pump in the instrument while the heating chamber 6 is vented through vent port 22c (see Figure 8 ). Each filter chamber 12 has a filter that will clog when the corresponding sample chamber 11 is filled, and will trigger the instrument’s pressure sensor to shut off the pump, similar to filling the heating chamber 6. Alternative fill detection techniques can be used, as described above. 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 triggered until the filters of all sample chambers are clogged in the corresponding filter chambers 12. The sample chambers 11 are used to separate the sample into independent functional channels, to aliquot a known volume of sample, and to position the sample to mix with reagents. When the sample chambers 11 are filled, pressure ports 22b, 22d, 22f, and 22h (see Figure 8 ) and vent port 22e (see Figure 8 ) are closed by the instrument to prevent fluid leakage through the location 19 disposed below the sample chambers 11. Once the sample chambers 11 have been filled, vent port 22e (see Figure 8 ) is opened to atmospheric pressure so that one or more sample chambers 11 can be fluidically isolated from each other and from the heating chamber 6. Vent port 22e (see Figure 8 ) remains open to atmosphere during sample mixing, as described below. Figure 6C and Figure 6D also show indicia corresponding to this section of the description.

[0138] Mixing and testing path Referring to Figure 5B , unless otherwise noted, each sample chamber 11 is in fluid communication with a corresponding one or more reagent pouches 14 (see Figure 5A ), which are configured to hold at least one lyophilized bead containing a reagent. As shown in Figure 5A and Figure 5B , two reagent pouches 14 are provided (shown as 14a and 14b for one test channel in Figure 5A ). In other embodiments, a single reagent pouch is used per test channel. In other embodiments, more than two reagent pouches 14 are used per test channel. The reagent pouches 14, in turn, are in fluid communication with the serpentine channel 13 (see Figure 5Aand shown with repeating notation ("x4")). Each serpentine channel 13 has a first end in fluid communication with a reagent bag and a second end terminating at an optical detection zone 15. As described below, the instrument (i.e., the measurement system) can optically interrogate the optical detection zone 15 of the cartridge 100 to facilitate control of a pump that facilitates mixing of each aliquot with a corresponding reagent. Each serpentine channel 13 (see Figure 5A ) is in fluid communication with a test chamber 16 (see Figure 5B , repeating notation "x4") which in turn is in fluid communication with a filter chamber 17 having a filter therein. The filter chamber 17 in the first fluid path plane of the cartridge 100 is in fluid communication with a first end of a corresponding one of fluid channels 17a-17d (see Figure 8 ) in the second fluid path plane of the cartridge 100. A second end of each of the fluid communication channels 17a-17d (see Figure 8 ) is in fluid communication with a corresponding pressure port 22b, 22d, 22f, and 22h (see Figure 8 ). The cartridge 100 is coupled with the instrument via the pressure ports 22b, 22d, 22f, and 22h (see Figure 8 ) to provide positive and negative pressure to facilitate mixing and testing of the sample as described below. Figure 6E and Figure 6F also show front and rear perspective views of Figure 5A , Figure 5B and Figure 8 with additional labeling corresponding to the description of this section.

[0139] Sample mixing Referring to Figure 5B , unless otherwise noted, each individual aliquot in the sample chamber 11 is drawn into a separate reservoir or channel (in various embodiments, a serpentine channel path 13 (see Figure 5A ) and contacted with channel-specific reagents located in one (or both) of the two reagent bags 14 (see Figure 5A ). Specifically, a pump in the instrument applies negative pressure to the pressure ports 22b, 22d, 22f, and 22h (see Figure 8 ) to draw the sample through the reagent bags 14 (see Figure 5A ) and the serpentine channel path 13 (see Figure 5A) Aspiration of sample. The reagents and sample remain separate from each other during filling of the sample chamber 11 to avoid the reagents floating on the blood and being trapped in the filter in the filter chamber 12, and to ensure that the sample chamber(s) 11 are fluidically isolated from the heating chamber 6. To this end, the exact time when the test sample comes into contact with the reagents can be measured, thereby facilitating accurate and precise measurement of the clotting time (e.g., from the time of mixing). Furthermore, in some embodiments, the reagents and test sample remain separate from each other until all channels are metered, to prevent unwanted siphoning of the test sample from untreated samples in other channels or fluidic paths. The sample is aspirated through the serpentine channel 13 (see Figure 5A ) until it triggers an optical sensor in the instrument (the top of the serpentine channel near the optical detection zone 15 (see Figure 5A )), which isolates the channel from the pump. Mixing in the serpentine channel or zone, or site, can be controlled by one or more independent valves and paths that allow individual channel control. Alternative sensor technologies can be used: pressure, by optical sensor, ultrasonic detection, time, volume control, etc. Once the optical sensor of all channels is triggered, the pump is reversed and positive pressure is applied to the pressure ports 22b, 22d, 22f, and 22h (see Figure 8 ). The positive pressure pushes the biological sample (e.g., blood) down the serpentine path 13 (see Figure 5A ) for a specified time, or until a second set of optical sensors in the instrument trips (in alternative embodiments). The process is repeated, pulling the sample up the serpentine path 13 (see Figure 5A ) to the optical detection zone (where the sample is detected by the optical sensors of the instrument and pushed back for a given time), until complete mixing of the sample is achieved. Figure 6E and Figure 6F Also shown are indicia corresponding to this portion of the description.

[0140] Other sensors can be used (e.g., impedance sensors), pressure sensors, etc. Alternatively, additional sensors can be used to detect both ends of the optical detection zone. Alternative path geometries, obstacles to create turbulence, number of cycles, and cycle speed are all design alternatives that can be used for different test types to achieve optimal results. In alternative embodiments, mixing can be achieved with one or more ferromagnetic beads or rods placed within the cartridge and controlled by the instrument.

[0141] Test chamber fill Referring to Figure 5A , Figure 5B and Figure 8After mixing is complete, one or more test chambers 16 are filled. Using one or more independent valves and pathways, each test chamber 16 is caused to be filled with sample via pressure perturbation (negative and / or positive pressure) applied at pressure ports 22b, 22d, 22f, and 22h. Specifically, the instrument pump will apply negative pressure to pressure ports 22b, 22d, 22f, and 22h until all filters are clogged in one or more filter chambers and a pressure spike is generated, causing the instrument to shut off the pump, similar to filling the heating chamber 6. Alternative fill detection techniques can be used, as described above. Test chambers 16 have design features, such as ridges 24a, that prevent air bubbles from forming in the test chambers 16 during filling. Once filled, the instrument begins viscoelastic testing of the sample. Figure 6E and Figure 6F Also shown are indicia corresponding to this portion of the description.

[0142] 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. The measurements are performed on each of the at least four channels of the cartridge. In some embodiments, the measurements include viscoelasticity, such as the shear modulus of the sample. In another embodiment, the measurements include other properties, such as the viscosity, elastic modulus, or any other mechanical property of the sample, or combinations thereof.

[0143] Table 1 provides a set of example reagents and measurement parameters used in an example reagent cartridge device (e.g., device 100, etc.). As shown in Table 1, Channel #1 of the example cartridge device is interrogated to measure the clotting time of a test sample in the presence of kaolin, which is an activator of the intrinsic coagulation pathway. As shown in Table 1, Channel #2 is interrogated to measure the clotting time of a test sample in the presence of kaolin and further in the presence of heparinase I, which is a neutralizer of the anticoagulant heparin. As shown in Table 1, Channel #3 is interrogated to measure the overall clot firmness of a test sample in the presence of i) thromboplastin, which is an activator of the extrinsic coagulation pathway, and ii) polybrene, which is a neutralizer of the anticoagulant heparin. As shown in Table 1, Channel #4 is interrogated to measure the clot firmness of a test sample with the same reagents as Channel #3, but with the addition of the platelet aggregation / contraction inhibitor, abciximab (e.g., Clotinab ® and / or ReoPro ® ), for example. As shown in Table 1, when the assay is configured to operate with a citrated whole blood sample, calcium is added to all reagent formulations.

[0144] Table 1. Reagents used in preferred embodiments

[0145] Table 2 provides another set of example reagents and measurements used in an example reagent cartridge device (e.g., device 100, etc.). As shown in Table 2, channel #2 includes an exogenous pathway activator that inhibits fibrinolysis by tranexamic acid (TXA). In addition to the measurements given previously in Table 1, channel #2, channel #3, and channel #4 are interrogated to measure clot firmness changes, e.g., which can be related to the fibrinolysis process. In some embodiments, other channels can include reagents that inhibit fibrinolysis, and can also be interrogated to measure clot firmness changes. For example, channel #4 can also include TXA or other fibrinolysis inhibitors in order to measure clot firmness in the absence of fibrinolysis.

[0146] Table 2. Reagents used in preferred embodiments

[0147] In some embodiments, clotting time and clot firmness are measured by analyzing the shear modulus (clot firmness) versus time curve generated within each measurement channel in the cartridge. Figure 13 Example shear modulus versus time curves are shown according to illustrative embodiments. Clotting time can be determined by identifying when the clot firmness reaches or exceeds a threshold value, or when the first or higher order derivative of such a property being measured reaches or exceeds a threshold value, or at the point of maximum acceleration of the clot firmness rate, or some combination of the above methods. Clot firmness can be estimated by the clot firmness at a fixed time after clot time, or the maximum overall clot firmness measured within some time limit, or the clot firmness at the point of maximum rate of change of clot firmness, or some combination of the above methods. Similar methods can also be used to measure the effect of fibrinolysis (i.e., clot dissolution) and the corresponding decrease in clot firmness. In some embodiments, clot firmness change can be calculated as the percentage drop in clot firmness across a fixed time window, the rate of change of clot firmness with time, the area under or over the clot firmness versus time curve within a predetermined time window, the time required to achieve a predetermined drop in clot firmness, or combinations thereof. By plotting Young's modulus, viscosity, or other viscoelastic properties of the sample being measured, similar curves and measurements can be formed as just described.

[0148] Table 3. Parameters reported by measurements of the preferred embodiments discussed in connection with Table 1.

[0149]

[0150] Those of ordinary skill in the art will recognize that a variety of methods and standards can be used to estimate clotting time and clot firmness. The clotting time and clot firmness values obtained from at least four channels / measurement values can be combined to provide at least six parameters to describe the functional status of a patient's hemostatic system. These indicators are summarized in Table 3. The relationship 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 indicator to verify instrument, cartridge, and sample functionality.

[0151] In other embodiments, other reagents can be used, and other hemostasis indicators or output parameters can be obtained, such as in the case of fibrinolysis indicators, indicators corresponding to anti-platelet therapy function, indicators corresponding to anti-coagulation therapy function, etc.

[0152] For example, a fibrinolysis index or indices can be formed using the change in clot firmness measured in any of the channels shown in Table 2, but preferably channels #3 and #4. Alternatively, a fibrinolysis index can be formed by the combination of the difference in change in clot firmness measured in channels #2 and #3 shown in Table 2. Such a combination can be in the form of a ratio, difference, or combination thereof. One benefit of utilizing a combination of change in clot firmness measured with and without an anti-fibrinolytic agent is the ability to mitigate the interfering effects of non-fibrinolysis induced clot firmness value reductions. In some embodiments, TXA or other fibrinolysis inhibitor reagent can be included in channels #2 and #4 of the example cartridges of Table 2. With such a modification, the parameters 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.

[0153] As noted above, an example user interface is described in commonly assigned U.S. Publication No. 2011 / 0252352 to Viola et al., which is incorporated herein by reference in its entirety. The example user interface can be used to display the measured hemostasis indices as discussed in connection with Table 4, as well as other parameters.

[0154] Table 4. Parameters reported by the measured values of the preferred embodiments discussed in connection with Table 2.

[0155]

[0156] As previously mentioned, in various embodiments, the test chamber 16 is shaped to facilitate ultrasonic testing of the viscoelastic properties of the sample, although alternative geometries can be implemented to facilitate other types of testing. Such ultrasonic testing systems have been described in commonly assigned U.S. Patent No. 9,726,647 and U.S. Publication No. 2016 / 0139159, both of which are incorporated herein by reference in their entirety. The ultrasonic transducers in the measurement system are connected to the test chamber 16 of the cartridge 100 via a flexible and deformable elastomer 21, and the elastomer 21 is secured to a test block 21d on the cartridge 100.

[0157] Example elastomer materials can optionally include Dynaflex D3202, Versaflex OM 9-802CL, Maxelast 54740, RTP 6035, Versaflex CL2003X, etc. Referring now to Figure 9 , unless otherwise noted, the test block 21d is aligned with the test chamber 16 via alignment slots 23 and 24 on the cartridge 100 (see Figure 5B . Still referring to Figure 9 , the elastomer 21 can be secured to the test block 21d via a flange 21a on the elastomer 21. The flange 21a can have a plurality of alignment holes 21b, which can receive corresponding alignment pins (not shown) from the test block 21d. The soft elastomer 21 can also each include a lens 21c that focuses ultrasonic energy within the sample at the test chamber 16.

[0158] Figure 16 FIGS. 1-4 show photographs of example cartridges according to illustrative embodiments, Figure 1 , Figure 2 , Figure 3 , Figure 4 , Figure 5A , Figure 5B , Figure 7 , Figure 8 , and Figure 9 according to illustrative embodiments. Figure 17 FIGS. 5A-5D show front views of example cartridges according to illustrative embodiments, Figure 1 , Figure 2 , Figure 3 , Figure 4 , Figure 5A , Figure 5B , Figure 7 , Figure 8 , and Figure 9 according to illustrative embodiments. Figure 18 FIGS. 6A-6D show front views of example cartridges according to illustrative embodiments, Figure 1 , Figure 2 , Figure 3 , Figure 4 , FIG. 5 A, Figure 5B , Figure 7 ,Figure 8 and Figure 9 front view of an exemplary cartridge.

[0159] As described in U.S. Patent No. 9,272,280, which is incorporated by reference in its entirety, in various embodiments, a cartridge for consumption contains a lens assembly that focuses ultrasonic energy within a sample that can be used to generate flow and mixing. As shown in Figure 10 , Figure 11 and Figure 12 , the lens assembly or sound focusing assembly is designed using a soft material such as thermoplastic elastomer 134 (previously referred to as 21) in combination with a rigid substrate 132 such as polystyrene (e.g., formed by test block 21d). This combination provides dry ultrasonic coupling that does not require the use of any fluid or gel couplant. Note that the same lens and ultrasonic driver used for hemostasis measurements can be used for this to provide mixing. Increasing the acoustic energy for mixing can be delivered by, for example, increasing the pulse length, pulse amplitude, or pulse repetition frequency.

[0160] Referring now to Figure 10 , a top cross-sectional view of test chamber 116 (previously referred to as test chamber 16) is shown. To seal each test chamber, such as test chamber 116, the lens assembly 131 includes a rigid substrate 132 and a couplant 134, which can be located at the back end of each test chamber.

[0161] Still referring to Figure 10 , each couplant 134 includes an elastomeric material. Optionally, the elastomeric material is a thermoplastic elastomer (TPE). Example elastomeric materials can include Dynaflex D3202, Versaflex OM 9-802CL, Maxelast 54740, RTP 6035, Versaflex CL2003X, etc. Optionally, the couplant is overmolded onto the rigid substrate. Optionally, the couplant is mechanically anchored to the rigid substrate.

[0162] Still referring to Figure 10 , there is a rigid substrate 132 between each couplant 134 and the open space of each test chamber. The rigid substrate and couplant form an interface that focuses the ultrasonic waves emitted by the ultrasonic transducer (e.g., lens assembly) into the open space of the chamber and onto any biological fluid and / or reagents in the chamber. The rigid substrate of the lens can include a material that allows sound to pass through and can focus the ultrasonic waves at a certain horizontal plane within the space. Optionally, the rigid substrate includes styrene.

[0163] Referring now to Figure 11 , the lens assembly can be glued or welded to the surface 101 of the test block 21d (in Figure 11A lens assembly (shown as element 132) can be used to secure the lens in place in an orientation that allows for the desired focusing of sound. Alternatively, the lens assembly can be optionally manufactured with the surface 101 of the test block 21d. In this regard, the rigid substrate 132 can be molded with the surface 101 of the test block 21d, and the couplant 134 can be overmolded or mechanically anchored on the rigid substrate. A variety of materials can be used to construct the device. For example, plastic can be used for single use, disposable cartridges.

[0164] Still referring to Figure 11 Each test chamber 116 can have a lens assembly positioned above the large opening of the open space of each chamber. In this way, each chamber can be individually interrogated by focused ultrasound.

[0165] Still referring to Figure 11 When placed in the instrument, the couplant 134 can be placed in acoustic communication with the transducer for providing 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.

[0166] Still referring to Figure 11 The sound produced by the transducer passes through the intermediate layer, the couplant, the rigid substrate, and is focused within the biological sample (e.g., blood) and the reagent in the test chamber. Some of the sound introduced into the chamber contacts the distal interior surface 111 of the test chamber defined by the surface 126. Optionally, the surface is polystyrene. The distal interior surface has a known geometry and is positioned at a known distance from the source of ultrasound. The distal interior surface 111 acts as a calibrated reflector that is used to estimate the speed of sound and sound attenuation in the test chamber at baseline and during clot formation and clot lysis. For example, these measurements can be used to estimate the hematocrit of the subject and the hemostatic index. The sound produced by the transducer can be focused within the biological sample in the test chamber using a parabolic mirror that is acoustically coupled to the biological sample using an elastomer.

[0167] 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. WO 2011 / 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 herein by reference in its entirety.

[0168] As noted above, the cartridges and features described herein can 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, aggregation measurement-based systems, resonance-based systems, and electrical impedance-based systems, among others.

[0169] Many modifications and other embodiments of the applications set forth herein will occur to those skilled in the art upon reading of the foregoing description. Thus, it is to be understood that the application is not to be limited to the particular embodiments disclosed herein and that modifications and other embodiments are intended to be included within the scope of the appended claims.

[0170] Although specific terms are employed herein, they are used in a generic and descriptive sense only and not for purposes of limitation.

[0171] As used in the claims, the terms "first," "second," and "third," and the like, merely denote different occurrences of analogous elements and do not imply a sequence or order unless specifically stated.

Claims

1. A method comprising: receiving a sample from a sample holding tube in a cartridge; metering the sample in a plurality of metering chambers to form a plurality of metered samples; mixing each of the plurality of metered samples with a respective set of one or more reagents, or with a reagent combination, to form a respective mixed sample, wherein a first metered sample of the plurality of metered samples is mixed with a first set of one or more reagents or with a reagent combination over a plurality of cycle periods by repeated flow in i) a first direction along a mixing path defined along (a) a first metering chamber, (b) by a first set of one or more reagent pockets having the first set of one or more reagents or reagent combination therein, and (c) along a first serpentine path in communication with the first metering chamber and the first set of one or more reagent pockets, until at least a portion of the first metered sample reaches a first detection zone located in or after the first serpentine path, and ii) a second direction opposite the first direction from the first detection zone through at least a portion of the first serpentine path toward the first metering chamber, wherein the first metering chamber is connected to a first test chamber by the first serpentine path and the first set of one or more reagent pockets, and wherein the first metered sample is repeatedly flowed along the first and second directions over the mixing path according to varying pressure applied at a first port by a measurement system, wherein the first port is in communication with the first serpentine path; and driving the first mixed sample into a first test chamber; interrogating the first test chamber by a measurement system to determine one or more viscoelastic properties of the first mixed sample, wherein the determined one or more viscoelastic properties of the first mixed sample are provided for assessment of hemostasis of the sample.

2. A method of mixing a sample with one or more reagents and testing the sample for assessment of hemostasis in a device, the method comprising: receiving a plurality of metered samples from a plurality of metering chambers, the metering chambers receiving a test fluid from a sample holding tube; mixing each of the plurality of metered samples with a respective set of one or more reagents, or with a reagent combination, to form a respective mixed sample, wherein mixing is performed by applying a first positive or negative pressure at a first port in communication with a first serpentine path from a measurement system to move the first metered sample in a first direction through the first serpentine path, and applying a second positive or negative pressure from a measurement system to move the first metered sample in a second direction through the first serpentine path, wherein the second positive or negative pressure is opposite the first positive or negative pressure; driving the first mixed sample into a first test chamber; and and interrogating the first test chamber with a measurement system configured to determine one or more viscoelastic properties of the first mixed sample, wherein the determined one or more viscoelastic properties of the first mixed sample are provided for assessment of hemostasis of the sample.

3. A method of mixing a sample with one or more reagents and testing the sample for assessment of hemostasis in a device, the method comprising: receiving a plurality of metered samples from a plurality of metering chambers that received a test fluid from a sample holding tube; mixing each of the plurality of metered samples with a respective set of one or more reagents, or with a combination of reagents, to form a respective mixed sample by varying pressure from a measurement system at a first port; driving the first mixed sample into a first test chamber by applying negative or differential pressure with a second port in fluid communication with the first test chamber; and interrogating the first test chamber with a measurement system configured to determine one or more viscoelastic properties of the first mixed sample, wherein the determined one or more viscoelastic properties of the first mixed sample are provided for assessment of hemostasis of the sample.

4. A method of mixing a sample with one or more reagents and testing the sample for assessment of hemostasis in a device, the method comprising: receiving a plurality of metered samples from a plurality of metering chambers that received a test fluid from a sample holding tube; mixing each of the plurality of metered samples with a respective set of one or more reagents, or with a combination of reagents, to form a respective mixed sample, wherein at least one or more reagents or combination of reagents is in the form of one or more lyophilized beads; driving the first mixed sample into a first test chamber; and interrogating the first test chamber with a measurement system configured to determine one or more viscoelastic properties of the first mixed sample, wherein the determined one or more viscoelastic properties of the first mixed sample are provided for assessment of hemostasis of the sample.

5. A method of mixing a sample with one or more reagents and testing the sample for assessment of hemostasis in a device, the method comprising: receiving a plurality of metered samples from a plurality of metering chambers that received a test fluid from a sample holding tube; mixing each of the plurality of metered samples with a respective set of one or more reagents, or with a combination of reagents, to form a respective mixed sample, wherein at least one of the reagents or combination of reagents is selected from the group consisting of an intrinsic pathway activator, an extrinsic pathway activator, and a coagulation activator; driving the first mixed sample into a first test chamber; and interrogating the first test chamber with a measurement system configured to determine one or more viscoelastic properties of the first mixed sample, wherein the determined one or more viscoelastic properties of the first mixed sample are provided for assessment of hemostasis of the sample.

6. A method of mixing a sample with one or more reagents and testing the sample for assessment of hemostasis in a device, the method comprising: receiving a plurality of metered samples from a plurality of metering chambers, the metering chambers having received test fluid from a sample holding tube; mixing each of the plurality of metered samples with a respective set of one or more reagents, or with a combination of reagents, to form a respective mixed sample, wherein at least one of the reagents or combination of reagents comprises an intrinsic pathway activator; driving the first mixed sample into a first test chamber; and inquiring the first test chamber by a measurement system configured to determine one or more viscoelastic properties of the first mixed sample, wherein the determined one or more viscoelastic properties of the first mixed sample are provided for assessment of hemostasis of the sample.

7. A method of mixing a sample with one or more reagents and testing the sample for assessment of hemostasis in a device, the method comprising: receiving a plurality of metered samples from a plurality of metering chambers, the metering chambers having received test fluid from a sample holding tube; mixing each of the plurality of metered samples with a respective set of one or more reagents, or with a combination of reagents, to form a respective mixed sample, wherein at least one of the reagents or combination of reagents comprises an extrinsic pathway activator; driving the first mixed sample into a first test chamber; and inquiring the first test chamber by a measurement system configured to determine one or more viscoelastic properties of the first mixed sample, wherein the determined one or more viscoelastic properties of the first mixed sample are provided for assessment of hemostasis of the sample.

8. A method of mixing a sample with one or more reagents and testing the sample for assessment of hemostasis in a device, the method comprising: receiving a plurality of metered samples from a plurality of metering chambers, the metering chambers having received test fluid from a sample holding tube; mixing each of the plurality of metered samples with a respective set of one or more reagents, or with a combination of reagents, to form a respective mixed sample, wherein at least one of the reagents or combination of reagents is selected from the group consisting of a platelet activator, a platelet inhibitor, and an agonist; driving the first mixed sample into a first test chamber; and inquiring the first test chamber by a measurement system configured to determine one or more viscoelastic properties of the first mixed sample, wherein the determined one or more viscoelastic properties of the first mixed sample are provided for assessment of hemostasis of the sample.

9. A method of mixing a sample with one or more reagents and testing the sample for assessment of hemostasis in a device, the method comprising: receiving a plurality of metered samples from a plurality of metering chambers, the metering chambers having received test fluid from a sample holding tube; mixing each of the plurality of metered samples with a respective set of one or more reagents, or a combination of reagents, to form a respective mixed sample, wherein the one or more reagents or combination of reagents comprises a coagulation activator selected from the group consisting of: kaolin, heparin, celite, glass, ellagic acid, micronized silica, tissue factor, recombinant tissue factor, thromboplastin, thrombin, factor Xa, reptilase, hirudin, and Russell's viper venom; driving the first mixed sample into a first test chamber; and interrogating the first test chamber by a measurement system configured to determine one or more viscoelastic properties of the first mixed sample, wherein the determined one or more viscoelastic properties of the first mixed sample are provided for assessment of hemostasis of the sample.

10. A method of mixing a sample with one or more reagents and testing the sample for assessment of hemostasis in a device, the method comprising: receiving a plurality of metered samples from a plurality of metering chambers, the metering chambers having received a test fluid from a sample holding tube; mixing each of the plurality of metered samples with a respective set of one or more reagents, or a combination of reagents, to form a respective mixed sample, wherein the one or more reagents or combination of reagents comprises a platelet activator or platelet inhibitor selected from the group consisting of: abciximab, eptifibatide, tirofiban, roxifiban, orbofiban, cytochalasin D, blesstatin, a PARl inhibitor, a PAR4 inhibitor, a glycoprotein IB inhibitor, a TRAP, ADP, arachidonic acid; driving the first mixed sample into a first test chamber; and interrogating the first test chamber by a measurement system configured to determine one or more viscoelastic properties of the first mixed sample, wherein the determined one or more viscoelastic properties of the first mixed sample are provided for assessment of hemostasis of the sample.

11. A method of mixing a sample with one or more reagents and testing the sample for assessment of hemostasis in a device, the method comprising: receiving a plurality of metered samples from a plurality of metering chambers, the metering chambers having received a test fluid from a sample holding tube; mixing each of the plurality of metered samples with a respective set of one or more reagents, or a combination of reagents, to form a respective mixed sample, wherein the one or more reagents or combination of reagents comprises a fibrinolytic agonist or antagonist selected from the group consisting of: tPA, uKA, streptokinase, TAFIa, plasmin / plasminogen, aprotinin, epsilon-aminocaproic acid, tranexamic acid, plasminogen activator inhibitor 1 (PAI1), alpha 2-antiplasmin (a2-AP), plasmin-antiplasmin complex, and carboxypeptidase inhibitor; driving the first mixed sample into a first test chamber; and interrogating the first test chamber by a measurement system configured to determine one or more viscoelastic properties of the first mixed sample, wherein the determined one or more viscoelastic properties of the first mixed sample are provided for assessment of hemostasis of the sample. interrogating the first test chamber with a measurement system configured to determine one or more viscoelastic properties of the first mixed sample, wherein the determined one or more viscoelastic properties of the first mixed sample are provided for assessment of hemostasis of the sample.

12. A method of mixing a sample with one or more reagents and testing the sample for assessment of hemostasis in a device, the method comprising: receiving a plurality of metered samples from a plurality of metering chambers that received a test fluid from a sample holding tube; mixing each of the plurality of metered samples with a respective set of one or more reagents, or with a combination of reagents, to form a respective mixed sample, wherein the one or more reagents or combination of reagents comprises a reagent selected from the group consisting of: FXIIIa inhibitor, hirudin (polybrene), heparinase, reductase, heparin, low molecular weight heparin, corn trypsin inhibitor, adenosine, GPRP, calcium, fibronectin, collagen, epinephrine, immunoassay reagents, direct thrombin inhibitors, factor Xa inhibitors, direct thrombin inhibitors, and thrombomodulin; driving the first mixed sample into a first test chamber; and interrogating the first test chamber with a measurement system configured to determine one or more viscoelastic properties of the first mixed sample, wherein the determined one or more viscoelastic properties of the first mixed sample are provided for assessment of hemostasis of the sample.

13. A method of mixing a sample with one or more reagents and testing the sample for assessment of hemostasis in a device, the method comprising: receiving a plurality of metered samples from a plurality of metering chambers that received a test fluid from a sample holding tube; mixing each of the plurality of metered samples with a respective set of one or more reagents, or with a combination of reagents, to form a respective mixed sample, wherein the first set of one or more reagents comprises an intrinsic pathway activator, wherein a second set of one or more reagents comprises an extrinsic pathway activator and a fibrinolysis inhibitor, wherein a third set of one or more reagents comprises an extrinsic pathway activator, and wherein a fourth set of one or more reagents comprises an extrinsic pathway activator and a platelet inhibitor; driving the first mixed sample into a first test chamber; and interrogating the first test chamber with a measurement system configured to determine one or more viscoelastic properties of the first mixed sample, wherein the determined one or more viscoelastic properties of the first mixed sample are provided for assessment of hemostasis of the sample.

14. A method of mixing a sample with one or more reagents and testing the sample for assessment of hemostasis in a device, the method comprising: receiving a plurality of metered samples from a plurality of metering chambers that received a test fluid from a sample holding tube; mixing each of the plurality of metered samples with a respective set of one or more reagents, or with a combination of reagents, to form respective mixed samples, wherein the set of one or more reagent pouches associated with the test chamber comprises a combination of kaolin, calcium, a buffer, and a stabilizer; driving the first mixed sample into a first test chamber; and inquiring the first test chamber by a measurement system configured to determine one or more viscoelastic properties of the first mixed sample, wherein the determined one or more viscoelastic properties of the first mixed sample are provided for assessment of hemostasis of the sample.

15. A method of mixing a sample with one or more reagents and testing the sample for assessment of hemostasis in a device, the method comprising: receiving a plurality of metered samples from a plurality of metering chambers, the metering chambers having received a test fluid from a sample holding tube; mixing each of the plurality of metered samples with a respective set of one or more reagents, or with a combination of reagents, to form respective mixed samples, wherein the set of one or more reagent pouches associated with the test chamber comprises a combination of kaolin, heparinase I, calcium, a buffer, and a stabilizer; or, a combination of thromboplastin, polybrene, calcium, a buffer, and a stabilizer; driving the first mixed sample into a first test chamber; and inquiring the first test chamber by a measurement system configured to determine one or more viscoelastic properties of the first mixed sample, wherein the determined one or more viscoelastic properties of the first mixed sample are provided for assessment of hemostasis of the sample.

16. A method of mixing a sample with one or more reagents and testing the sample for assessment of hemostasis in a device, the method comprising: receiving a plurality of metered samples from a plurality of metering chambers, the metering chambers having received a test fluid from a sample holding tube; mixing each of the plurality of metered samples with a respective set of one or more reagents, or with a combination of reagents, to form respective mixed samples, wherein the set of one or more reagent pouches associated with the test chamber comprises a combination of thromboplastin, polybrene, calcium, a buffer, and a stabilizer; driving the first mixed sample into a first test chamber; and inquiring the first test chamber by a measurement system configured to determine one or more viscoelastic properties of the first mixed sample, wherein the determined one or more viscoelastic properties of the first mixed sample are provided for assessment of hemostasis of the sample.

17. A method of mixing a sample with one or more reagents and testing the sample for assessment of hemostasis in a device, the method comprising: receiving a plurality of metered samples from a plurality of metering chambers, the metering chambers having received a test fluid from a sample holding tube; mixing each of the plurality of metered samples with a respective set of one or more reagents, or with a combination of reagents, to form respective mixed samples, wherein the set of one or more reagent pouches associated with the test chamber comprises a combination of thromboplastin, polybrene, at least one of acyloxime and cytochalasin D, calcium, a buffer, and a stabilizer; driving the first mixed sample into a first test chamber; and inquiring the first test chamber by a measurement system configured to determine one or more viscoelastic properties of the first mixed sample, wherein the determined one or more viscoelastic properties of the first mixed sample are provided for assessment of hemostasis of the sample. driving the first mixed sample into a first test chamber; and interrogating the first test chamber by a measurement system configured to determine one or more viscoelastic properties of the first mixed sample, wherein the determined one or more viscoelastic properties of the first mixed sample are provided for assessment of hemostasis of the sample.

18. A method of mixing a sample with one or more reagents and testing the sample for assessment of hemostasis in a device, the method comprising: receiving a plurality of metered samples from a plurality of metering chambers that received a test fluid from a sample holding tube; mixing each of the plurality of metered samples with a respective set of one or more reagents, or with a combination of reagents, to form a respective mixed sample, wherein the set of one or more reagent pouches associated with a test chamber comprises: thromboplastin, eponistin, abciximab, calcium, a buffer, and a stabilizer; driving the first mixed sample into a first test chamber; and interrogating the first test chamber by a measurement system configured to determine one or more viscoelastic properties of the first mixed sample, wherein the determined one or more viscoelastic properties of the first mixed sample are provided for assessment of hemostasis of the sample.

19. A method of mixing a sample with one or more reagents and testing the sample for assessment of hemostasis in a device, the method comprising: receiving a plurality of metered samples from a plurality of metering chambers that received a test fluid from a sample holding tube; mixing each of the plurality of metered samples with a respective set of one or more reagents, or with a combination of reagents, to form a respective mixed sample, wherein a first set of one or more reagents comprises kaolin, calcium, a buffer, and a stabilizer, a second set of one or more reagents comprises a combination of kaolin, heparinase I, calcium, a buffer, and a stabilizer; a third set of one or more reagents comprises thromboplastin, eponistin, calcium, a buffer, and a stabilizer, a fourth set of one or more reagents comprises thromboplastin, eponistin, abciximab, a buffer, and a stabilizer; driving the first mixed sample into a first test chamber; and interrogating the first test chamber by a measurement system configured to determine one or more viscoelastic properties of the first mixed sample, wherein the determined one or more viscoelastic properties of the first mixed sample are provided for assessment of hemostasis of the sample.

20. A method of mixing a sample with one or more reagents and testing the sample for assessment of hemostasis in a device, the method comprising: receiving a plurality of metered samples from a plurality of metering chambers that received a test fluid from a sample holding tube; ​ each of the plurality of metered samples is mixed with a respective set of one or more reagents, or a combination of reagents, to form a respective mixed sample, wherein a first set of one or more reagents comprises kaolin, calcium, a buffer, and a stabilizer, a second set of one or more reagents comprises a combination of thromboplastin, polybrene, calcium, a buffer, and a stabilizer; a third set of one or more reagents comprises thromboplastin, polybrene, calcium, a buffer, and a stabilizer, a fourth set of one or more reagents comprises thromboplastin, polybrene, acicimab, calcium, a buffer, and a stabilizer; driving the first mixed sample into a first test chamber; and interrogating the first test chamber by a measurement system configured to determine one or more viscoelastic properties of the first mixed sample, wherein the determined one or more viscoelastic properties of the first mixed sample are provided for assessment of hemostasis of the sample.

21. A system comprising: a device comprising: a housing; an input port integral with the housing, the input port structurally configured to receive contents of a sample holding tube; and a first chamber in fluid communication with the input port, the first chamber configured to receive a sample, after which the received sample is allowed to contact one or more reagents in one or more fluidic circuits downstream of the first chamber, wherein each of the one or more fluidic circuits comprises i) a second chamber in fluid communication with the first chamber, the second chamber metering the sample in the first chamber into aliquots, wherein the metered samples are introduced into reagents or combinations of reagents in the respective fluidic circuit to form respective mixed samples, and ii) a test chamber in fluid communication with the second chamber, the test chamber structurally configured to be interrogated by a measurement system, a measurement system configured to determine at least one viscoelastic property of the mixed sample.

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