On-plate lysis, internal controls, and volume expansion in fluidic cartridges

Abstract

A fluid cartridge chamber containing non-magnetic beads and magnetic elements. Movement imparted to the magnetic elements by a varying magnetic field imparts movement to the non-magnetic beads, which in turn lyse cells of a fluid sample within the chamber. An internal control is provided within the chamber to validate assay results and / or to validate effectiveness of cell lysis. The internal control article can be in a non-liquid form deposited on a portion of the chamber, the non-magnetic beads and / or the magnetic elements, and is dissolved when contacted by a fluid sample. The internal control can be provided in a pellet form that dissolves when contacted by a fluid sample or is broken when exposed to the lysing power of the non-magnetic beads. A fluid cartridge comprising fluidly connected pools, and one pool comprising an extension pool to which a chamber extender is attached to extend the volumetric capacity of the extension pool.

Description

Cross-reference to related applications

[0001] This application claims the benefit of U.S. Provisional Application No. 63 / 685,122, filed August 20, 2024, and U.S. Provisional Application No. 63 / 752,023, filed January 31, 2025, on the filing date of the application. Technical Field

[0002] This disclosure relates to systems and methods for performing mechanical lysis of a sample in a sample chamber of a test platform (such as a fluidic cartridge) by providing the sample to a lysis chamber comprising a magnetic element and a plurality of non-magnetic beads and agitating the magnetic element with a magnetic field, thereby contacting the agitated magnetic element and agitating the non-magnetic beads to lyse the cells contained in the sample. Additionally, this disclosure relates to a fluidic cartridge having an expansion well and a chamber expander, the chamber expander being hermetically sealed to the body of the cartridge to expand the volumetric capacity of the expansion well. This disclosure further relates to apparatus and methods for providing an internal control to a sample chamber prior to sample addition. The internal control may be provided in a dry (non-liquid) soluble form, or it may be contained in a capsule or pellet that disintegrates during mechanical lysis, thereby releasing the internal control into the sample chamber. Background Technology

[0003] Molecular assays performed in testing platforms (such as fluidic chambers) often require lysing cells contained in a sample to release nucleic acids from the cells. Lysis can be performed using, for example, chemical, acoustic, mechanical (physical destruction), and / or enzymatic methods. Cells can be lysed before the sample is introduced into the fluidic chamber, thus requiring additional sample handling and treatment before introduction if, for example, acoustic or mechanical lysis methods are used. Lysing cells at least partially on the fluidic chamber eliminates the need for such sample handling and treatment before introducing the sample into the fluidic chamber.

[0004] Additionally, fluid cartridges used to perform molecular assays or other tests comprise multiple pools or chambers interconnected by channels, which often have valves to control the flow through them. The volumetric capacity of each chamber is determined by the width and height of its internal space. Since such fluid cartridges are often made of molded plastic, limitations in molding techniques can restrict the variability of the metric volumetric capacity achievable within the cartridge. For example, limitations in molding techniques may make it impractical to mold a cartridge with multiple pools, one of which is significantly larger than the others. Therefore, if one pool requires a significantly larger volumetric capacity than the others, the only way to achieve this larger capacity might be to make that pool much wider than the others in the cartridge. This would increase the overall width of the cartridge, or, if the allowable width of the cartridge is limited, for example, by the size of the instrument in which the cartridge is handled, the other pools in the cartridge would need to be made smaller.

[0005] Accordingly, there is a need to increase the volumetric capacity of at least one chamber of the fluid cartridge.

[0006] In the case of performing molecular assays on a fluidic cartridge, it may be desirable for the reaction mixture to include an internal control. The internal control (such as, for example, a plasmid, nucleic acid transcript, or nucleic acid extracted from a whole organism (such as yeast)) will be exposed to the same assay conditions as the sample (such as lysis, sample purification, combination with amplification reagents and detection probes, thermal cycling, etc., in the case of a whole organism containing the internal control), such that if the amplification and detection protocols are performed correctly—that is, all steps of the molecular assay procedure have been properly performed with feasible reagents used in the assay—a signal indicating the presence of the internal control (i.e., a positive result for the internal control nucleic acid) can be expected. On the other hand, failure to detect a signal indicating the presence of the internal control (i.e., a negative result for the internal control nucleic acid), or detection of less internal control than expected, may indicate an error or malfunction in one or more of the following steps: sample preparation (e.g., lysis or analyte purification), material delivery, amplification, and / or detection, and / or reagent failure. Such errors or malfunctions can be system-based (e.g., a failure of the instrument or a module within the instrument) and / or material-based (e.g., one or more reagents have degraded or become unstable).

[0007] The internal control can be provided to the reaction mixture by simply dispensing a certain amount of reagent containing the internal control (“internal control reagent” or “ICR”) together with the sample into the sample chamber, or the internal control can be provided to the sample before dispensing into the sample chamber. However, these approaches introduce additional steps into the sample preparation process, which may reduce throughput and may lead to errors, spills, or contamination. Summary of the Invention

[0008] The following is a simplified overview to provide a basic understanding of some aspects described herein. This overview is not a comprehensive summary of the claimed subject matter. It is intended neither to identify key or essential elements of the claimed subject matter nor to depict its scope. Its sole purpose is to present some concepts in a simplified form as a prelude to the more detailed description that follows.

[0009] The implementation of this disclosure can be described in light of the following embodiments, the features of which can be combined in any reasonable manner.

[0010] Some implementation plans cover:

[0011] A1. A lysis capsule for performing a cell lysis procedure on a fluid sample, wherein the lysis capsule comprises: a hollow body having an open first end and an open second end; a first porous membrane attached to the body, the first porous membrane covering the open first end; a second porous membrane attached to the body, the second porous membrane covering the open second end, wherein the hollow body defines a lysis chamber between the first porous membrane and the second porous membrane; a plurality of non-magnetic beads disposed within the lysis chamber; and at least one magnetic element disposed within the lysis chamber, wherein the pore sizes of the first porous membrane and the second porous membrane are designed to retain the plurality of non-magnetic beads and the at least one magnetic element within the lysis chamber.

[0012] A2. The lysis capsule according to embodiment A1, wherein the first porous membrane comprises a mesh.

[0013] A3. The lysis capsule according to embodiment A1 or A2, wherein the first porous membrane is hydrophilic.

[0014] A4. The lysis capsule according to any one of embodiments A1 to A3, wherein the second porous membrane comprises a mesh.

[0015] A5. The rupture capsule according to any one of embodiments A1 to A3, wherein the second porous membrane is a filter matrix.

[0016] A6. The rupture capsule according to any one of embodiments A1 to A5, wherein the first porous membrane and the second porous membrane comprise different porosities.

[0017] A7. The rupture capsule according to embodiment A6, wherein the porosity or porosity range of the first porous membrane is greater than the porosity or porosity range of the second porous membrane.

[0018] A8. The rupture capsule according to embodiment A7, wherein the first porous membrane has a porosity of 70 µm to 500 µm and the second porous membrane has a porosity of 30 µm to 100 µm.

[0019] A9. The rupture capsule according to any one of embodiments A1 to A8, wherein the first porous membrane is attached to the top edge of the body, the top edge defining the open first end, and wherein the second porous membrane is attached to the bottom edge of the body, the bottom edge defining the open second end.

[0020] A10. The rupture capsule according to any one of embodiments A1 to A9, wherein each of the plurality of non-magnetic bead-like elements is composed of ceramic or glass.

[0021] A11. The rupture capsule according to any one of embodiments A1 to A10, wherein each of the plurality of non-magnetic bead-like elements has a spherical shape.

[0022] A12. The lysis capsule according to embodiment A11, wherein each of the plurality of nonmagnetic bead-like elements has a diameter of 100 µm to 2000 µm.

[0023] A13. The rupture capsule according to any one of embodiments A1 to A12, wherein the at least one magnetic element is coated or encapsulated with a non-magnetic material.

[0024] A14. The rupture capsule according to embodiment A13, wherein the non-magnetic material is metal or plastic.

[0025] A15. The rupture capsule according to any one of embodiments A1 to A14, wherein the at least one magnetic element occupies a larger volume than any one of the plurality of non-magnetic bead-like elements.

[0026] A16. The rupture capsule according to any one of embodiments A1 to A15, wherein the at least one magnetic element comprises a plurality of edges.

[0027] A17. The rupture capsule according to embodiment A16, wherein each of the plurality of edges is circular.

[0028] A18. The rupture capsule according to embodiment A16 or A17, wherein the at least one magnetic element has a cubic shape.

[0029] A19. The rupture capsule according to embodiment A18, wherein the width of each face of the cube is 2.0 mm to 4.3 mm.

[0030] A20. The pyrolysis capsule according to any one of embodiments A1 to A19, wherein the at least one magnetic element is composed of neodymium.

[0031] A21. The pyrolysis capsule according to embodiment A20, wherein the neodymium is N52 or N42 grade.

[0032] A22. The rupture capsule according to any one of embodiments A1 to A21, wherein each of the plurality of non-magnetic bead-like elements and the at least one magnetic element are inert.

[0033] A23. The lysis capsule according to any one of embodiments A1 to A22, wherein the plurality of non-magnetic bead-like elements occupy 50% to 75% of the volume of the lysis chamber.

[0034] A24. The pyrolysis capsule according to embodiment A23, wherein the at least one magnetic element occupies 4.5% to 11% of the volume of the pyrolysis chamber.

[0035] A25. The lysis sac-like device according to any one of embodiments A1 to A24, further comprising an internal control contained within the lysis chamber, wherein the internal control is provided to verify assay results and / or to verify the effectiveness of the cell lysis procedure.

[0036] A26. The lysis capsule according to embodiment A25, wherein at least a portion of the internal control is contained in an internal control reagent, the internal control reagent is disposed on at least one of the first porous membrane and the second porous membrane, and wherein the internal control reagent is adapted to dissolve upon contact with the fluid sample.

[0037] A27. The lysis capsule according to embodiment A25, wherein at least a portion of the internal control is contained in an internal control reagent, the internal control reagent is disposed on at least a portion of the plurality of nonmagnetic beads and / or the at least one magnetic element, and wherein the internal control reagent is adapted to dissolve upon contact with the fluid sample.

[0038] A28. The lysis capsule according to embodiment A25, wherein at least a portion of the internal control is contained in an internal control reagent disposed on the inner wall of the hollow body, and wherein the internal control reagent is adapted to dissolve upon contact with the fluid sample.

[0039] A29. The lysis capsule according to embodiment A25, wherein at least a portion of the internal control is embedded in or contained in internal control pellets contained within the lysis chamber, and wherein the internal control pellets are adapted to dissolve upon contact with the fluid sample and / or adapted to disintegrate upon agitation of the plurality of magnetic beads.

[0040] A30. The lysis capsule according to embodiment A29, wherein the internal control pellet comprises: a core comprising an excipient in which the internal control is embedded; and a coating surrounding the core and adapted to be broken by a mechanical lysis shear force applied by movement of the plurality of nonmagnetic beads within the lysis chamber, wherein the excipient is adapted to at least partially dissolve upon exposure to a fluid after the coating is broken.

[0041] A31. The lysis capsule according to embodiment A30, wherein the excipient comprises at least one of microcrystalline cellulose and hydroxypropyl cellulose, and wherein the coating comprises a cellulose derivative.

[0042] A32. The lysing cyst according to any one of embodiments A25 to A31, wherein the internal control is a whole organism, a plasmid, or a nucleic acid transcript.

[0043] Some implementation plans cover:

[0044] B1. A fluid cartridge comprising: a cartridge body including a sample chamber having an open top; and a lysis capsule disposed within the sample chamber according to any one of embodiments A1 to A24.

[0045] B2. The fluid cartridge according to embodiment B1, further comprising an injection barrel body in communication with the sample chamber, the injection barrel body being adapted to receive an injection barrel stop connected to an injection barrel plunger for actuating fluid within the fluid cartridge.

[0046] B3. The fluid cartridge according to embodiment B1 or B2, further comprising a dead space within the sample chamber located below the lysis sac-like element.

[0047] B4. The fluid cartridge according to any one of embodiments B1 to B3, wherein the sample chamber is covered by a removable seal.

[0048] B5. The fluid cartridge according to any one of embodiments B1 to B3, further comprising a cap adapted for insertion into an open top of the sample chamber.

[0049] B6. The fluid cartridge according to embodiment B5, wherein the cap includes a sleeve, and wherein when the cap is inserted into the open top of the sample chamber, the outer surface of the sleeve is in a sealing engagement with the inner surface of the sample chamber.

[0050] B7. The fluid cartridge according to embodiment B5 or B6, wherein when the cap is inserted into the open top of the sample chamber, the bottom end of the cap is arranged adjacent to the top end of the lysis sac-like member.

[0051] B8. The fluid cartridge according to embodiment B7, wherein when the cap is inserted into the open top of the sample chamber, there is a gap between the top end of the lysis sac and the bottom end of the cap.

[0052] B9. The fluid cartridge according to any one of embodiments B6 to B8, wherein the cap includes a lateral extension member, wherein a peripheral region of the lateral extension member is seated on a top surface of the cartridge body, and wherein a sleeve hangs from the lateral extension member such that an inner surface of the sleeve and a bottom surface of the lateral extension member define a recess extending upward from the bottom end of the cap.

[0053] B10. The fluid cartridge according to embodiment B9, wherein the lateral extension member has a vent extending therethrough, the vent allowing air to escape from the sample chamber when the cap is inserted into the sample chamber.

[0054] B11. The fluid box according to embodiment B10, wherein the cap includes a porous venting membrane attached to the top or bottom surface of the lateral extension member and covering the vent.

[0055] B12. The fluid box according to embodiment B11, wherein the porous venting membrane is permeable to air but impermeable to liquid.

[0056] B13. The fluid cell according to embodiment B11 or B12, wherein the porous venting membrane has a porosity of 0.2 µm to 0.4 µm.

[0057] B14. The fluid cartridge according to any one of embodiments B9 to B13, wherein the cap includes a peripheral wall extending upward from the periphery of the lateral extension member.

[0058] B15. The fluid cartridge according to embodiment B14, wherein the walls are adapted for manual gripping.

[0059] B16. The fluid cartridge according to any one of embodiments B1 to B15, further comprising an internal control contained within the lysis chamber, wherein the internal control is provided to verify assay results and / or to verify the effectiveness of the cell lysis procedure.

[0060] B17. The fluid cartridge according to embodiment B16, wherein at least a portion of the internal control is contained in an internal control reagent, the internal control reagent is disposed on at least one of the first porous membrane and the second porous membrane, and wherein the internal control reagent is adapted to dissolve upon contact with a fluid sample.

[0061] B18. The fluid cartridge according to embodiment B16 or B17, wherein at least a portion of the internal control is contained in an internal control reagent, the internal control reagent is disposed on at least a portion of the plurality of nonmagnetic beads and / or the at least one magnetic element in the lysis chamber, and wherein the internal control reagent is adapted to dissolve upon contact with a fluid sample.

[0062] B19. The fluid cartridge according to embodiment B16, wherein at least a portion of the internal control is embedded in or contained in internal control pellets contained within the lysis chamber, and wherein the internal control pellets are adapted to dissolve upon contact with a fluid sample and / or to disintegrate upon agitation of the plurality of nonmagnetic beads.

[0063] B20. The fluid cartridge according to embodiment B19, wherein the internal control pellet comprises: a core, the core comprising an excipient in which the internal control is embedded; and a coating surrounding the core and adapted to be broken by a mechanical cleavage shear force applied by movement of the plurality of nonmagnetic beads within the receiver, wherein the excipient is adapted to at least partially dissolve upon exposure to fluid after the coating is broken.

[0064] B21. The fluid cartridge according to embodiment B20, wherein the excipient comprises at least one of microcrystalline cellulose and hydroxypropyl cellulose, and wherein the coating comprises a cellulose derivative.

[0065] B22. The fluidic cartridge according to any one of embodiments B16 to B21, wherein the internal control is a whole organism, a plasmid, or a nucleic acid transcript.

[0066] Some implementation plans cover:

[0067] C1. A method for processing cells contained in a fluid sample, the method comprising: (a) dispensing the fluid sample into a sample chamber of a fluid cartridge according to any one of embodiments B1 to B3 to at least partially fill the lysis chamber of the lysis capsule with the fluid sample; (b) after (a), covering the open top of the sample chamber with a cap; and (c) after (b), subjecting the at least one magnetic element to a magnetic field, thereby causing the at least one magnetic element to move within the lysis chamber of the lysis capsule, the movement of the at least one magnetic element within the lysis chamber of the lysis capsule causing a plurality of non-magnetic beads to move within the lysis chamber of the lysis capsule, and the movement of the plurality of non-magnetic beads within the lysis chamber of the lysis capsule causing the cells contained in the fluid sample within the lysis chamber of the lysis capsule to lyse and release nucleic acids.

[0068] C2. The method according to embodiment C1, wherein the fluid sample dispensed in (a) occupies 39% to 45% of the volume of the lysis chamber of the lysis sac.

[0069] C3. The method according to embodiment C1 or C2, wherein (b) includes inserting the cap into the sample chamber such that the cap and the sample chamber are in a sealed engagement.

[0070] C4. The method according to embodiment C3, wherein after the cap is inserted into the sample chamber, a gap separates the top end of the lysis sac from the bottom end of the cap.

[0071] C5. The method according to any one of embodiments C1 to C4, wherein (b) includes transferring air from the sample chamber to the external environment through a vent in the cap.

[0072] C6. The method according to embodiment C5, wherein (b) further comprises passing the air through a porous venting membrane, the porous venting membrane being attached to the cap and covering the vent, the porous venting membrane being impermeable to liquids.

[0073] C7. The method according to any one of embodiments C1 to C6, wherein the magnetic field during (c) is generated by an electromagnet.

[0074] C8. The method according to embodiment C7, wherein (c) includes alternating the current to the electromagnet to alter the polarity of the electromagnet.

[0075] C9. The method according to embodiment C8, wherein (c) includes alternating the current at a frequency of 20 Hz to 200 Hz to alter the polarity of the electromagnet.

[0076] C10. The method according to embodiment C9, wherein (c) includes pulses of the current at two or more different frequencies to alternating the polarity of the electromagnet.

[0077] C11. The method according to any one of embodiments C1 to C10, further comprising: (d) after (c), conveying at least a portion of the fluid sample from the sample chamber to the processing chamber of the fluid cartridge.

[0078] C12. The method according to embodiment C11, further comprising: (e) during (d), retaining the lysed cell material from (c) in the lysis chamber while allowing the released nucleic acid to pass through the second porous membrane.

[0079] C13. The method according to embodiment C12, further comprising: (f) subjecting the at least one magnetic element to the magnetic field during (d), thereby moving the at least one magnetic element within the lysis chamber of the lysis capsule, the movement of the at least one magnetic element within the lysis chamber causing the plurality of non-magnetic beads within the lysis chamber to move within the lysis chamber, and the movement of the plurality of non-magnetic beads within the lysis chamber causing at least a portion of the lysed cellular material within the lysis chamber to remain suspended at least until the fluid sample has been removed from the lysis chamber.

[0080] C14. The method according to any one of embodiments C11 to C13, further comprising: (g) immobilizing at least a portion of the released nucleic acid on a solid support in the processing chamber, and removing the unimmobilized components of the fluid sample into the waste chamber of the fluid cartridge.

[0081] C15. The method according to embodiment C14 further includes: (h) after (g), eluting the immobilized nucleic acid from the solid support and delivering the eluted nucleic acid to the reaction chamber of the fluid cartridge.

[0082] C16. The method according to embodiment C15 further includes: (i) after (h), subjecting the eluted nucleic acid to the conditions of a first reaction, the first reaction providing an indication of the presence or amount of the analyte of interest.

[0083] C17. The method according to embodiment C16, further comprising: (j) releasing an internal control into the fluid sample during or after (a), the internal control being contained in the lysis chamber prior to (a); (k) immobilizing a nucleic acid (“IC nucleic acid”) associated with the internal control on the solid support during (g); (l) eluting the IC nucleic acid from the solid support and delivering the eluted IC nucleic acid to the reaction chamber after (g); and (m) subjecting the IC nucleic acid to conditions of a second reaction after (l), the results of (m) being used to verify the results of (i) and / or to verify the validity of the lysis in (c).

[0084] C18. The method according to embodiment C17, wherein at least a portion of the internal control is contained in an internal control reagent, and when (a) is initiated, the internal control reagent is disposed on at least one of the first porous membrane and the second porous membrane, and wherein during any of (a) to (c), the internal control reagent is dissolved in the fluid sample.

[0085] C19. The method according to embodiment C17, wherein at least a portion of the internal control is contained in an internal control reagent, and when (a) is initiated, the internal control reagent is disposed on at least a portion of the plurality of nonmagnetic beads and / or the at least one magnetic element, and wherein during any one of (a) to (c), the internal control reagent disposed on at least a portion of the plurality of nonmagnetic beads and / or the at least one magnetic element dissolves in the fluid sample.

[0086] C20. The method according to embodiment C17, wherein at least a portion of the internal control is contained in an internal control reagent, the internal control reagent being disposed on the inner wall of the hollow body when (a) is initiated, and wherein the internal control reagent is dissolved in the fluid sample during any of (a) to (c).

[0087] C21. The method according to embodiment C17, wherein the internal control is embedded in or contained within internal control pellets, and wherein the internal control pellets dissolve in the presence of the fluid sample and / or break apart during (c) by the movement of the plurality of nonmagnetic beads, thereby releasing the internal control into the fluid sample.

[0088] C22. The method according to any one of embodiments C17 to C21, wherein the conditions for the first reaction and the conditions for the second reaction are the same.

[0089] C23. The method according to any one of embodiments C17 to C22, wherein each of the first reaction and the second reaction is a nucleic acid amplification reaction.

[0090] C24. The method according to implementation plan C23, wherein the nucleic acid amplification reaction is a polymerase chain reaction (“PCR”).

[0091] Some implementation plans cover:

[0092] D1. A lysis container for performing cell lysis, the lysis container comprising: a lateral extension member having a vent extending therethrough; a sleeve suspending from the lateral extension member, wherein a bottom end of the sleeve defines an open bottom end of the container; a first porous membrane attached to a top or bottom surface of the lateral extension member, the first porous membrane covering the vent; a second porous membrane attached to the bottom end of the sleeve, the second porous membrane covering the open bottom end, wherein a bottom surface of the lateral extension member, an inner surface of the sleeve, and the first and second porous membranes define a lysis chamber; a plurality of non-magnetic beads contained within the lysis chamber; and at least one magnetic element contained within the lysis chamber, wherein the sizes of the first and second porous membranes are designed to retain the plurality of non-magnetic beads and the at least one magnetic element within the lysis chamber.

[0093] D2. The pyrolysis container according to embodiment D1, further comprising a peripheral wall extending upward from the periphery of the lateral extension member.

[0094] D3. The pyrolysis container according to embodiment D2, wherein the peripheral wall is suitable for manual gripping.

[0095] D4. The pyrolysis vessel according to any one of embodiments D1 to D3, wherein the second porous membrane comprises a mesh element that is liquid permeable.

[0096] D5. The pyrolysis vessel according to embodiment D4, wherein the second porous membrane is hydrophilic.

[0097] D6. The pyrolysis vessel according to embodiment D4 or D5, wherein the first porous membrane is gas-permeable but not liquid-permeable.

[0098] D7. The pyrolysis vessel according to embodiment D6, wherein the first porous membrane has a porosity of 0.2 µm to 0.4 µm, and the second porous membrane has a porosity of 30 µm to 100 µm.

[0099] D8. A pyrolysis vessel according to any one of embodiments D1 to D7, wherein each of the plurality of non-magnetic beads is composed of ceramic or glass.

[0100] D9. The pyrolysis container according to any one of embodiments D1 to D8, wherein each of the plurality of non-magnetic beads has a spherical shape.

[0101] D10. The pyrolysis container according to embodiment D9, wherein each of the plurality of nonmagnetic beads has a diameter of 100 µm to 2000 µm.

[0102] D11. The pyrolysis container according to any one of embodiments D1 to D10, wherein the at least one magnetic element is coated or encapsulated with a non-magnetic material.

[0103] D12. The pyrolysis container according to embodiment D11, wherein the non-magnetic material is metal or plastic.

[0104] D13. The pyrolysis container according to any one of embodiments D1 to D12, wherein the at least one magnetic element occupies a larger volume than any one of the plurality of non-magnetic beaded elements.

[0105] D14. A pyrolysis container according to any one of embodiments D1 to D13, wherein the at least one magnetic element comprises a plurality of edges.

[0106] D15. The pyrolysis container according to embodiment D14, wherein each of the plurality of edges is circular.

[0107] D16. The pyrolysis container according to embodiment D14 or D15, wherein the at least one magnetic element has a cubic shape.

[0108] D17. The pyrolysis container according to embodiment D16, wherein the width of each face of the cube is 2.0 mm to 4.3 mm.

[0109] D18. A pyrolysis container according to any one of embodiments D1 to D17, wherein the at least one magnetic element is composed of neodymium.

[0110] D19. The pyrolysis vessel according to embodiment D18, wherein the neodymium is N52 or N42 grade.

[0111] D20. The pyrolysis vessel according to any one of embodiments D1 to D19, wherein each of the plurality of non-magnetic beads and the at least one magnetic element are inert.

[0112] D21. The pyrolysis container according to any one of embodiments D1 to D20, wherein the plurality of non-magnetic beads occupy 50% to 75% of the volume of the pyrolysis chamber.

[0113] D22. The pyrolysis container according to embodiment D21, wherein the at least one magnetic element occupies 4.5% to 11% of the volume of the pyrolysis chamber.

[0114] D23. The lysis vessel according to any one of embodiments D1 to D22, further comprising an internal control contained within the lysis chamber, wherein the internal control is provided to verify assay results and / or to verify the effectiveness of the cell lysis procedure.

[0115] D24. The lysis vessel according to embodiment D23, wherein at least a portion of the internal control is contained in an internal control reagent disposed on the second porous membrane, and wherein the internal control reagent is adapted for dissolution upon contact with a fluid sample.

[0116] D25. The lysis vessel according to embodiment D23, wherein at least a portion of the internal control is contained in an internal control reagent, the internal control reagent is disposed on at least a portion of the plurality of nonmagnetic beads, and wherein the internal control reagent is adapted for dissolving upon contact with a fluid sample.

[0117] D26. The lysis vessel according to embodiment D23, wherein at least a portion of the internal control is contained in an internal control reagent, the internal control reagent being disposed on at least one of the bottom surface of the lateral extension member and the inner surface of the sleeve, and wherein the internal control reagent is adapted to dissolve upon contact with a fluid sample.

[0118] D27. The pyrolysis vessel according to embodiment D23, wherein at least a portion of the internal control is embedded in or contained within internal control pellets adapted to dissolve upon contact with a fluid sample and / or disintegrate upon agitation of the plurality of nonmagnetic beads, the internal control pellets being contained within the pyrolysis chamber.

[0119] Some implementation plans cover:

[0120] E1. A fluid cartridge comprising: a cartridge body including a sample chamber having an open top; and a pyrolysis vessel disposed within the sample chamber according to any one of embodiments D1 to D28.

[0121] E2. The fluid cartridge according to embodiment E1, further comprising an injection cartridge body in communication with the sample chamber, the injection cartridge body being adapted to receive an injection cartridge stop, the injection cartridge stop being connected to or connectable to an injection cartridge plunger for actuating the injection cartridge stop within the injection cartridge body to actuate fluid within the fluid cartridge.

[0122] E3. The fluid cartridge according to embodiment E1 or E2, further comprising a dead space located below the lysis vessel within the sample chamber.

[0123] E4. A fluid cartridge according to any one of embodiments E1 to E3, wherein the outer surface of the sleeve is in a sealed engagement with the inner surface of the sample chamber.

[0124] E5. A fluid cartridge according to any one of embodiments E1 to E4, wherein the peripheral region of the lateral extension member sits on the top surface of the cartridge body.

[0125] Some implementation plans cover:

[0126] F1. A method for processing cells contained in a fluid sample, the method comprising: (a) dispensing the fluid sample into a sample chamber of a fluid cartridge, the sample chamber having an open top; (b) after (a), inserting a sleeve of a lysis container according to any one of embodiments D1 to D22 into the open top of the sample chamber; (c) after (b), subjecting the at least one magnetic element to a magnetic field, thereby causing the at least one magnetic element to move within the lysis chamber of the lysis container, the movement of the at least one magnetic element within the lysis chamber of the lysis container causing a plurality of non-magnetic beads to move within the lysis chamber of the lysis container, and the movement of the plurality of non-magnetic beads within the lysis chamber of the lysis container causing the cells contained in the fluid sample within the lysis chamber of the lysis container to lyse and release nucleic acids.

[0127] F2. The method according to embodiment F1, wherein the fluid cartridge further includes an injection barrel body in communication with the sample chamber, the injection barrel body being adapted to receive an injection barrel stop, the injection barrel stop being connected to or connectable to an injection barrel plunger for actuating the injection barrel stop within the injection barrel body to actuate fluid within the fluid cartridge.

[0128] F3. The method according to embodiment F1 or F2, wherein the fluid cell further includes a dead space located below the lysis vessel within the sample chamber.

[0129] F4. The method according to any one of embodiments F1 to F3, wherein after (b), the outer surface of the sleeve is in a sealed engagement with the inner surface of the sample chamber.

[0130] F5. The method according to any one of embodiments F1 to F4, wherein the peripheral region of the lateral extension member sits on the top surface of the box body.

[0131] F6. The method according to any one of embodiments F1 to F5, wherein the fluid sample dispensed in (a) occupies 39% to 45% of the volume of the pyrolysis chamber of the pyrolysis vessel.

[0132] F7. The method according to any one of embodiments F1 to F6, wherein (b) further comprises transferring air from the sample chamber to the external environment through the vent in the first porous membrane and the lysis vessel.

[0133] F8. The method according to any one of embodiments F1 to F7, wherein the magnetic field during (c) is generated by an electromagnet.

[0134] F9. The method according to embodiment F8, wherein (c) includes alternating the current to the electromagnet to alter the polarity of the electromagnet.

[0135] F10. The method according to embodiment F9, wherein (c) includes alternating the current at a frequency of 20 Hz to 200 Hz.

[0136] F11. The method according to any one of embodiments F1 to F10, further comprising: (d) after (c), conveying at least a portion of the fluid sample from the sample chamber to the processing chamber of the fluid cartridge.

[0137] F12. The method according to embodiment F11 further includes: (e) during (d), retaining the lysed cell material from (c) in the lysis chamber while allowing the released nucleic acid to pass through the second porous membrane.

[0138] F13. The method according to embodiment F12, further comprising: (f) subjecting the at least one magnetic element to the magnetic field during (d), thereby moving the at least one magnetic element within the lysis chamber of the lysis vessel, the movement of the at least one magnetic element within the lysis chamber causing the plurality of non-magnetic beads within the lysis chamber to move within the lysis chamber, and the movement of the plurality of non-magnetic beads within the lysis chamber causing at least a portion of the lysed cell material within the lysis chamber to remain suspended at least until the fluid sample has been removed from the lysis chamber.

[0139] F14. The method according to any one of embodiments F11 to F13, further comprising: (g) immobilizing at least a portion of the released nucleic acid on a solid support in the processing chamber, and removing the unimmobilized components of the fluid sample into the waste chamber of the fluid cartridge.

[0140] F15. The method according to embodiment F14 further includes: (h) after (g), eluting the immobilized nucleic acid from the solid support and delivering the eluted nucleic acid to the reaction chamber of the fluid cartridge.

[0141] F16. The method according to embodiment F15 further includes: (i) after (h), subjecting the eluted nucleic acid to the conditions of a first reaction, the first reaction providing an indication of the presence or amount of the analyte of interest.

[0142] F17. The method according to embodiment F16, further comprising: (j) releasing an internal control into the fluid sample during or after (a), the internal control being contained in the lysis chamber prior to (a); (k) immobilizing a nucleic acid (“IC nucleic acid”) associated with the internal control on the solid support during (g); (l) eluting the IC nucleic acid from the solid support and delivering the eluted IC nucleic acid to the reaction chamber after (g); and (m) subjecting the IC nucleic acid to conditions of a second reaction after (l), the results of (m) being used to verify the results of (i) and / or to verify the validity of the lysis in (c).

[0143] F18. The method according to embodiment F17, wherein at least a portion of the internal control is contained in an internal control reagent, the internal control reagent is disposed on the second porous membrane when (a) is initiated, and wherein the internal control reagent is dissolved in the fluid sample thereafter (b).

[0144] F19. The method according to embodiment F17, wherein at least a portion of the internal control is contained in an internal control reagent, and when (a) is initiated, the internal control reagent is disposed on at least a portion of the plurality of non-magnetic beads and / or the at least one magnetic element, and wherein during (b) and / or (c), the internal control reagent disposed on at least a portion of the plurality of non-magnetic beads and / or the at least one magnetic element dissolves in the fluid sample.

[0145] F20. The method according to embodiment F17, wherein at least a portion of the internal control is embedded in or contained within an internal control pellet, and wherein the internal control pellet dissolves in the presence of the fluid sample and / or disintegrates during (c) by the movement of the plurality of nonmagnetic beads, thereby releasing the internal control into the fluid sample.

[0146] F21. The method according to any one of embodiments F17 to F19, wherein, prior to (a), before contact with the fluid sample, the internal control reagent is a dried reagent.

[0147] F22. The method according to any one of embodiments F17 to F21, wherein the conditions for the first reaction and the conditions for the second reaction are the same.

[0148] F23. The method according to any one of embodiments F17 to F22, wherein each of the first reaction and the second reaction is a nucleic acid amplification reaction.

[0149] F24. The method according to embodiment F23, wherein the nucleic acid amplification reaction is a polymerase chain reaction (“PCR”).

[0150] Some implementation plans cover:

[0151] G1. A method of manufacturing a fluid cartridge comprising a pyrolysis capsule, the method comprising: assembling the pyrolysis capsule by means of the following steps: (a) providing a hollow body having an open first end and an open second end; (b) providing a first porous membrane and a second porous membrane and attaching the second porous membrane to the hollow body, the second porous membrane covering the open second end of the hollow body; (c) introducing a plurality of non-magnetic beads into the hollow body through the open first end of the hollow body; (d) introducing at least one magnetic element into the hollow body through the open first end of the hollow body; (e) after (c) and (d), attaching the first porous membrane to the hollow body, the first porous membrane covering the open first end of the hollow body, wherein the hollow body and the attached first and second porous membranes define a pyrolysis chamber, and wherein the pore sizes of the first and second porous membranes are designed to retain the plurality of non-magnetic beads and the at least one magnetic element within the pyrolysis chamber; and (f) in (e) The lysis capsule is then fixed in the sample chamber of a fluid cell, the fluid cell having multiple chambers in fluid communication with the sample chamber.

[0152] G2. The method according to embodiment G1, wherein the first porous membrane comprises a mesh.

[0153] G3. The method according to embodiment G1 or G2, wherein the second porous membrane comprises a mesh.

[0154] G4. The method according to any one of embodiments G1 to G3, wherein the first porous membrane is hydrophilic.

[0155] G5. The method according to any one of embodiments G1 to G4, wherein the first porous membrane and the second porous membrane comprise different porosities.

[0156] G6. The method according to embodiment G5, wherein the porosity or porosity range of the first porous membrane is greater than the porosity or porosity range of the second porous membrane.

[0157] G7. The method according to embodiment G6, wherein the first porous membrane has a porosity of 70 µm to 500 µm, and the second porous membrane has a porosity of 30 µm to 100 µm.

[0158] G8. The method according to any one of embodiments G1 to G7, wherein the first porous membrane is attached to the top edge of the hollow body defining the open first end, and wherein the second porous membrane is attached to the bottom edge of the hollow body defining the open second end.

[0159] G9. The method according to any one of embodiments G1 to G8, wherein each of the plurality of non-magnetic beads is composed of ceramic or glass.

[0160] G10. The method according to any one of embodiments G1 to G9, wherein each of the plurality of non-magnetic beads has a spherical shape.

[0161] G11. The method according to embodiment G10, wherein each of the plurality of nonmagnetic beads has a diameter of 100 µm to 2000 µm.

[0162] G12. The method according to any one of embodiments G1 to G11, wherein the at least one magnetic element is coated or encapsulated with a non-magnetic material.

[0163] G13. The method according to embodiment G12, wherein the non-magnetic material is a metal or a plastic.

[0164] G14. The method according to any one of embodiments G1 to G13, wherein the at least one magnetic element occupies a larger volume than any one of the plurality of non-magnetic beads.

[0165] G15. The method according to any one of embodiments G1 to G14, wherein the at least one magnetic element comprises a plurality of edges.

[0166] G16. The method according to embodiment G15, wherein each of the plurality of edges is circular.

[0167] G17. The method according to embodiment G15 or G16, wherein the at least one magnetic element has a cubic shape.

[0168] G18. The method according to embodiment G17, wherein the width of each face of the cube is 2.0 mm to 4.3 mm.

[0169] G19. The method according to any one of embodiments G1 to G18, wherein the at least one magnetic element is composed of neodymium.

[0170] G20. The method according to implementation scheme G19, wherein the neodymium is N52 or N42 grade.

[0171] G21. The method according to any one of embodiments G1 to G20, wherein each of the plurality of non-magnetic beads and the at least one magnetic element are inert.

[0172] G22. The method according to any one of embodiments G1 to G21, wherein the plurality of non-magnetic beads occupy 50% to 75% of the volume of the pyrolysis chamber.

[0173] G23. The method according to embodiment G22, wherein the at least one magnetic element occupies 4.5% to 11% of the volume of the pyrolysis chamber.

[0174] G24. The method according to any one of embodiments G1 to G23, further comprising distributing an internal control reagent onto a component of the lysis sac, the internal control reagent comprising an internal control provided to validate assays and / or to validate the effectiveness of a cell lysis procedure performed with the plurality of nonmagnetic beads and the at least one magnetic element.

[0175] G25. The method according to embodiment G24, wherein, prior to (c), at least a portion of the internal control reagent is disposed on at least a portion of the plurality of non-magnetic beads and / or the at least one magnetic element.

[0176] G26. The method according to embodiment G24, wherein after (c), at least a portion of the internal control reagent is disposed on at least a portion of the plurality of nonmagnetic beads and / or the at least one magnetic element.

[0177] G27. The method according to embodiment G24, wherein the internal control reagent is disposed on (i) the first porous membrane, (ii) the second porous membrane, (iii) the inner surface of the hollow body, (iv) at least a portion of the plurality of non-magnetic beads and (v) on the at least one magnetic element prior to (e).

[0178] G28. The method according to any one of embodiments G24 to G27, wherein the internal control reagent is disposed in liquid form, and wherein the method further comprises drying the internal control reagent after the internal control reagent has been disposed on the lysis cyst.

[0179] G29. The method according to any one of embodiments G1 to G23, wherein an internal control is embedded in or contained within an internal control pellet adapted to dissolve upon contact with a fluid sample and / or adapted to disintegrate upon agitation of the plurality of magnetic beads in the lysis chamber, and wherein the internal control pellet is contained within the hollow body prior to (e), and wherein the internal control is provided to validate assays and / or to validate the effectiveness of a cell lysis procedure performed with the plurality of nonmagnetic beads and the at least one magnetic element.

[0180] G30. The method according to any one of embodiments G24 to G29, wherein the internal control is a whole organism, a plasmid, or a nucleic acid transcript.

[0181] G31. The method according to any one of embodiments G1 to G30, wherein each of the first porous membrane and the second porous membrane is attached to the hollow body by adhesive, heat sealing, or ultrasonic welding.

[0182] G32. The method according to any one of embodiments G1 to G31, wherein the lysis capsule is press-fitted into the sample chamber.

[0183] G33. The method according to any one of embodiments G1 to G31, wherein the outer surface of the hollow body is threadedly engaged with the inner surface of the sample chamber.

[0184] G34. The method according to any one of embodiments G1 to G33, further comprising attaching a removable protective cap to the top surface of the fluid cartridge, thereby covering the sample chamber.

[0185] Some implementation plans cover:

[0186] H1. A method for lysing cells contained in a fluid sample, the method comprising: (a) dispensing the fluid sample into a sample chamber of a fluid cartridge, the sample chamber comprising a plurality of nonmagnetic beads and at least one magnetic element, wherein at least a portion of the plurality of nonmagnetic beads and / or the at least one magnetic element has an internal control reagent deposited thereon, and wherein an internal control contained in the internal control reagent is provided to validate assay results and / or to validate the effectiveness of a cell lysis protocol; and (b) in (a) Subsequently, the fluid sample is subjected to the cell lysis procedure, which includes exposing the at least one magnetic element to a magnetic field, thereby causing the at least one magnetic element contained in the sample chamber to move, the movement of the at least one magnetic element causing the movement of the plurality of non-magnetic beads contained in the sample chamber, and the movement of the plurality of non-magnetic beads in the sample chamber causing the cells contained in the fluid sample to lyse and release nucleic acids, wherein the internal control reagent dissolves in the presence of the fluid sample, thereby releasing the internal control into the fluid sample, and wherein the movement of the at least one magnetic element and the plurality of non-magnetic beads causes the internal control contained in the dissolved internal control reagent to distribute within the fluid sample.

[0187] H2. A method for lysing cells contained in a fluid sample, the method comprising: (a) dispensing the fluid sample into a sample chamber of the fluid cartridge, the sample chamber comprising (i) a plurality of nonmagnetic beads, (ii) at least one magnetic element, and (iii) an internal control reagent contained within an internal control pellet, wherein the internal control contained in the internal control reagent is provided to validate assay results and / or to validate the effectiveness of the cell lysis protocol; and (b) in (a) Subsequently, the fluid sample is subjected to the cell lysis procedure, which includes exposing the at least one magnetic element to a magnetic field, thereby causing the at least one magnetic element contained in the sample chamber to move. The movement of the at least one magnetic element causes the plurality of non-magnetic beads contained in the sample chamber to move, and the movement of the plurality of non-magnetic beads in the sample chamber causes the cells contained in the fluid sample to lyse and release nucleic acids. The internal control pellets dissolve upon contact with the fluid sample, and / or the movement of the plurality of non-magnetic beads in the sample chamber causes the internal control pellets to disintegrate, thereby releasing the internal control into the fluid sample. The movement of the at least one magnetic element and the plurality of non-magnetic beads causes the internal control contained in the dissolved internal control reagent to distribute within the fluid sample.

[0188] H3. The method according to embodiment H1 or H2, wherein the plurality of nonmagnetic beads and the at least one magnetic element are contained within a hollow body defining a lysis chamber, the lysis chamber being arranged in the sample chamber during (a) and (b), and wherein the receiver is liquid-permeable.

[0189] H4. The method according to any one of embodiments H1 to H3, wherein each of the plurality of non-magnetic beads has a spherical shape.

[0190] H5. The method according to embodiment H4, wherein each of the plurality of nonmagnetic beads has a diameter of 100µm to 2000µm.

[0191] H6. The method according to any one of embodiments H1 to H5, wherein the at least one magnetic element is coated or encapsulated with a non-magnetic material.

[0192] H7. The method according to embodiment H6, wherein the non-magnetic material is a metal or a plastic.

[0193] H8. The method according to any one of embodiments H1 to H7, wherein the at least one magnetic element occupies a larger volume than any one of the plurality of non-magnetic beads.

[0194] H9. The method according to any one of embodiments H1 to H8, wherein the at least one magnetic element comprises a plurality of edges.

[0195] H10. The method according to embodiment H9, wherein each of the plurality of edges is circular.

[0196] H11. The method according to embodiment H9 or H10, wherein the at least one magnetic element has a cubic shape.

[0197] H12. The method according to embodiment H11, wherein the width of each face of the cube is 2.0 mm to 4.3 mm.

[0198] H13. The method according to any one of embodiments H1 to H12, wherein the at least one magnetic element is composed of neodymium.

[0199] H14. The method according to embodiment H13, wherein the neodymium is N52 or N42 grade.

[0200] H15. The method according to any one of embodiments H1 to H14, wherein each of the plurality of non-magnetic beads and the at least one magnetic element are inert.

[0201] H16. The method according to any one of embodiments H1 to H15, wherein the plurality of non-magnetic beads occupy 50% to 70% of the volume of the sample chamber.

[0202] H17. The method according to embodiment H16, wherein the at least one magnetic element occupies 4.5% to 11% of the volume of the sample chamber.

[0203] H18. The method according to any one of embodiments H1 to H17, wherein the magnetic field during (b) is generated by an electromagnet.

[0204] H19. The method according to embodiment H18, wherein (b) includes alternating the current to the electromagnet to alter the polarity of the electromagnet.

[0205] H20. The method according to embodiment H19, wherein (b) includes alternating the current at a frequency of 20 Hz to 200 Hz.

[0206] H21. The method according to any one of embodiments H1 to H20, further comprising: (c) after (b), conveying at least a portion of the fluid sample from the sample chamber to the processing chamber of the fluid cartridge.

[0207] H22. The method according to embodiment H21 further includes: (d) during (c), retaining the lysed cell material from (c) in the sample chamber while delivering the released nucleic acid to the processing chamber.

[0208] H23. The method according to embodiment H21 or H22 further comprises: (e) immobilizing at least a portion of the released nucleic acid on a solid support in the processing chamber, and removing the unimmobilized components of the fluid sample into the waste chamber of the fluid cartridge.

[0209] H24. The method according to embodiment H23 further includes: (f) after (e), eluting the immobilized nucleic acid from the solid support and delivering the eluted nucleic acid to the reaction chamber of the fluid cartridge.

[0210] H25. The method according to embodiment H24, further comprising: (g) after (f), subjecting the eluted nucleic acid to conditions of a first reaction, the first reaction providing an indication of the presence or amount of the analyte of interest.

[0211] H26. The method according to embodiment H25 further includes (h) immobilizing a nucleic acid (“IC nucleic acid”) associated with the internal control on the solid support during (e); (i) after (h), eluting the IC nucleic acid from the solid support and delivering the eluted IC nucleic acid to the reaction chamber; and (j) after (i), subjecting the IC nucleic acid to conditions of a second reaction, the second reaction providing an indication of the degree of lysis in (b).

[0212] H27. The method according to embodiment H26, wherein the conditions for the first reaction and the conditions for the second reaction are the same.

[0213] H28. The method according to implementation scheme H26 or H27, wherein each of the first reaction and the second reaction is a nucleic acid amplification reaction.

[0214] H29. The method according to implementation scheme H28, wherein the nucleic acid amplification reaction is a polymerase chain reaction (“PCR”).

[0215] Some implementation plans cover:

[0216] 11. A fluid cartridge comprising a cartridge body defining two or more pools fluidly connected or fluidly connectable, wherein at least one of the two or more pools includes an expansion pool, and wherein the cartridge body includes a first coupling structure at least partially surrounding the expansion pool, and wherein the cartridge further includes a chamber expander attached to the cartridge body and comprising: a base having a second coupling structure located on a bottom side of the base and configured to be operatively coupled to the first coupling structure to form an hermetically tight seal between the chamber expander and the cartridge body; an expansion chamber extending from the base, wherein the expansion chamber expands the volumetric capacity of the expansion pool by at least the volumetric capacity of the expansion chamber; a spout defining an opening into an interior space of the expansion chamber; and a cap configured to be coupled to the spout to close the opening.

[0217] I2. The fluid cartridge according to embodiment I1, wherein the first connection structure includes a first peripheral wall formed in the cartridge body and at least partially surrounding the expansion pool, and the second connection structure includes a second peripheral wall extending below the bottom side of the base and configured to conform to the inner or outer surface of the first peripheral wall.

[0218] 13. The fluid cartridge according to embodiment 12, wherein the second connection structure includes a third peripheral wall extending below the bottom side of the base and spaced apart from the second peripheral wall to form a peripheral groove on the bottom side of the base, and wherein the peripheral groove is configured to receive the first peripheral wall between the second peripheral wall and the third peripheral wall.

[0219] I4. The fluid box according to any one of embodiments I1 to I3, wherein the expansion chamber and the nozzle have different shapes.

[0220] I5. The fluid cartridge according to embodiment I4, wherein the expansion chamber has three straight sides and the nozzle has a circular shape.

[0221] 16. The fluid cartridge according to any one of embodiments 11 to 15, further comprising a perimeter chamfer formed in the inner surface of the nozzle around the opening.

[0222] I7. The fluid box according to embodiment I3, wherein the first peripheral wall and the peripheral groove each have three sides.

[0223] I8. The fluid box according to embodiment I7, wherein the first peripheral wall has three straight edges connected by rounded corners, and wherein the peripheral groove has three straight edges connected by rounded corners.

[0224] I9. The fluid box according to embodiment I2, wherein the first peripheral wall is attached to the second peripheral wall by adhesive or by welding.

[0225] I10. The fluid box according to embodiment I3, wherein the first peripheral wall is attached to at least one of the second peripheral wall and the third peripheral wall by means of an adhesive or by means of welding.

[0226] 111. The fluid cartridge according to any one of embodiments 11 to 110, wherein the cap comprises: an insertion sleeve configured to be inserted into the opening defined by the mouth; and a sheath wider than the insertion sleeve and including a top wall that substantially closes one end of the insertion sleeve.

[0227] I12. The fluid cartridge according to embodiment I11, wherein the insertion sleeve is hollow, and the sheath includes a vent extending through the top wall of the sheath and opening to the interior space of the insertion sleeve, and wherein the cap further includes a vent membrane disposed above the vent, wherein the vent membrane is configured to allow gas to pass through but prevent liquid from passing through.

[0228] I13. The fluid cell according to embodiment I12, wherein the venting membrane has a pore size of about 0.2 µm.

[0229] I14. The fluid box according to embodiment I12 or I13, wherein the vent includes an inner vent portion and an outer vent portion, the inner vent portion being formed on the inner surface of the top wall of the sheath, and the outer vent portion being formed on the outer surface of the top wall of the sheath, wherein the width of the inner vent is different from the width of the outer vent.

[0230] I15. The fluid box according to embodiment I14, wherein the width of the inner vent portion is smaller than the width of the outer vent portion.

[0231] 116. The fluid cartridge according to any one of embodiments 111 to 115, wherein the cap further includes radial ribs extending between the inner surface of the sheath and the outer surface of the insert sleeve, wherein when the cap is attached to the mouth to close the opening, the insert sleeve is inserted into the opening until the radial ribs contact the edge of the mouth surrounding the opening.

[0232] I17. The fluid cartridge according to any one of embodiments I11 to I16, wherein the insertion sleeve is secured within the opening defined by the nozzle by frictional engagement.

[0233] I18. The fluid cartridge according to any one of embodiments I11 to I17, wherein the cap includes circumferential ribs extending around the outer surface of the insert sleeve.

[0234] I19. The fluid cartridge according to any one of embodiments I12 to I18, further comprising at least one groove formed in the top surface of the top wall of the sheath, wherein the at least one groove extends through the vent.

[0235] I20. The fluid cartridge according to embodiment I19 further includes at least two grooves formed in the top surface of the sheath, wherein the at least two grooves intersect each other through the vent.

[0236] I21. The fluid cartridge according to any one of embodiments I1 to I20, wherein the chamber expander further includes a strut extending from the base, and wherein the cap is hingedly connected to the strut.

[0237] I22. The fluid cartridge according to embodiment I21, further comprising a tab extending from the cap, wherein the cap is connected to the support via a movable hinge that connects the free end of the support to the free end of the tab.

[0238] I23. The fluid cell according to any one of embodiments I1 to I22, wherein the expansion pool includes an inclined surface surrounding its opening.

[0239] I24. The fluid cartridge according to any one of embodiments I1 to I23, further comprising a lysis capsule disposed within the expansion chamber for performing cell lysis, wherein the lysis capsule comprises: a hollow body having an open first end and an open second end; a first porous membrane attached to the body, the first porous membrane covering the open first end; a second porous membrane attached to the body, the second porous membrane covering the open second end, wherein the hollow body defines a lysis chamber between the first porous membrane and the second porous membrane; a plurality of non-magnetic beads contained within the lysis chamber; and at least one magnetic element contained within the lysis chamber, wherein the pore sizes of the first porous membrane and the second porous membrane are designed to retain the plurality of non-magnetic beads and the at least one magnetic element within the lysis chamber.

[0240] I25. The fluid cartridge according to embodiment I24, wherein the expansion pool and hollow body of the rupture capsule have a conformal triangular shape.

[0241] I26. The fluid box according to any one of embodiments I1 to I25, wherein the chamber expander is composed of a transparent or translucent material.

[0242] I27. The fluid box according to any one of embodiments I1 to I26, wherein the chamber expander is composed of polypropylene.

[0243] I28. The fluid box according to any one of embodiments I1 to I27, wherein the box body is composed of an opaque material.

[0244] I29. The fluid box according to any one of embodiments I1 to I28, wherein the box body is composed of a thermoplastic polymer material.

[0245] I30. A fluid cartridge according to any one of embodiments I1 to I29, wherein the cartridge body is composed of a cyclic olefin copolymer (COC) or a cyclic olefin polymer (COP).

[0246] I31. The fluid cartridge according to any one of embodiments I1 to I30, wherein the cartridge body is composed of a material selected from the group consisting of polycarbonate, polyacrylamide, polyethylene, polymethyl methacrylate (PMMA), polydimethylsiloxane (PDMS), polyvinyl chloride (PVC), and polypropylene (PP).

[0247] Some implementation plans cover:

[0248] J1. In a fluid box comprising a box body defining two or more pools that are fluidly connected or fluidly connected, a method for expanding the volumetric capacity of an expansion pool of the two or more pools, the method comprising: securing a chamber expander to the box body by coupling a first coupling structure at least partially surrounding the expansion pool to a second coupling structure of a chamber expander and forming an airtight seal between the chamber expander and the box body, wherein the chamber expander comprises: a base, wherein the second coupling structure is located on a bottom side of the base; an expansion chamber extending from the base, wherein the expansion chamber expands the volumetric capacity of the expansion pool by at least the volumetric capacity of the expansion chamber; a spout defining an opening into an interior space of the expansion chamber; and a cap configured to be coupled to the spout to close the opening.

[0249] J2. The method according to embodiment J1, wherein the first connecting structure includes a first peripheral wall formed in the box body and surrounding the expansion pool, and the second connecting structure includes a second peripheral wall extending below the bottom side of the base and configured to conform to the inner or outer surface of the first peripheral wall, and wherein connecting the first connecting structure to the second connecting structure includes attaching the second peripheral wall to the inner or outer surface of the first peripheral wall.

[0250] J3. The method according to embodiment J2, wherein attaching the second peripheral wall to the inner or outer surface of the first peripheral wall includes fixing the second peripheral wall to the inner or outer surface of the first peripheral wall by means of an adhesive or by means of welding.

[0251] J4. The method according to embodiment J1, wherein the first connecting structure includes a first peripheral wall formed in the housing body and surrounding the expansion pool, and the second connecting structure includes a second peripheral wall and a third peripheral wall, the second peripheral wall extending below the bottom side of the base, the third peripheral wall extending below the bottom side of the base and spaced apart from the second peripheral wall to form a peripheral groove on the bottom side of the base, and wherein connecting the first connecting structure to the second connecting structure includes inserting the first peripheral wall into the peripheral groove between the second peripheral wall and the third peripheral wall, and attaching the first peripheral wall to at least one of the second peripheral wall and the third peripheral wall.

[0252] J5. The method according to embodiment J4, wherein attaching the first peripheral wall to at least one of the second peripheral wall and the third peripheral wall includes fixing the first peripheral wall to at least one of the second peripheral wall and the third peripheral wall by means of an adhesive or by means of welding.

[0253] J6. The method according to any one of embodiments J1 to J5, wherein the cap includes an insert sleeve and a sheath, the sheath being wider than the insert sleeve, and wherein the method includes attaching the cap to the mouth by inserting the insert sleeve into the opening defined by the mouth.

[0254] J7. The method according to any one of embodiments J1 to J6, further comprising: dispensing a quantity of liquid substance into the opening defined by the mouthpiece, wherein the quantity of liquid substance completely fills the expansion pool and at least partially fills the interior space of the expansion chamber; and attaching the cap to the opening to close the opening.

[0255] J8. The method according to any one of embodiments J1 to J7, wherein the expansion chamber and the mouth have different shapes.

[0256] J9. The method according to embodiment J8, wherein the expansion chamber has three straight sides and the mouth has a circular shape.

[0257] J10. The method according to any one of embodiments J1 to J9, wherein the chamber expander further includes a perimeter chamfer formed in the inner surface of the mouth around the opening.

[0258] J11. The method according to embodiment J4, wherein the first peripheral wall and the peripheral groove each have three sides.

[0259] J12. The method according to embodiment J11, wherein the first peripheral wall has three straight edges connected by rounded corners, and wherein the peripheral groove has three straight edges connected by rounded corners.

[0260] J13. The method according to any one of embodiments J1 to J4, wherein the cap comprises: an insertion sleeve configured to be inserted into the opening defined by the mouth portion; and a sheath wider than the insertion sleeve and including a top wall that substantially closes one end of the insertion sleeve.

[0261] J14. The method according to embodiment J13, wherein the insertion sleeve is hollow, and the sheath includes a vent hole extending through the top wall of the sheath and opening to the interior space of the insertion sleeve, and wherein the cap further includes a vent membrane disposed above the vent hole, wherein the vent membrane is configured to allow gas to pass through but prevent liquid from passing through.

[0262] J15. The method according to embodiment J13 or J14, wherein the cap further includes radial ribs extending between the inner surface of the sheath and the outer surface of the insert sleeve, wherein engaging the cap to the mouth to close the opening includes inserting the insert sleeve into the opening until the radial ribs contact the edge of the mouth surrounding the opening.

[0263] J16. The method according to embodiment J15, wherein the insertion sleeve is secured within the opening defined by the mouth by frictional engagement.

[0264] J17. The method according to embodiment J16, wherein the cap includes circumferential ribs extending around the outer surface of the insert sleeve.

[0265] J18. The method according to any one of embodiments J14 to J17, further comprising at least one groove formed in the top surface of the top wall of the sheath, wherein the at least one groove extends through the vent.

[0266] J19. The method according to embodiment J18 further includes at least two grooves formed in the top surface of the sheath, wherein the at least two grooves intersect each other through the vent.

[0267] J20. The method according to any one of embodiments J1 to J19, wherein the chamber expander further includes a strut extending from the base, and wherein the cap is hingedly connected to the strut.

[0268] J21. The method according to embodiment J20, further comprising a tab extending from the cap, wherein the cap is connected to the support via a movable hinge that connects the free end of the support to the free end of the tab.

[0269] J22. The method according to any one of embodiments J1 to J21, wherein the extended pool includes an inclined surface surrounding its opening.

[0270] J23. The method according to any one of embodiments J1 to J22, further comprising, prior to securing the chamber expander to the box body, inserting a lysis capsule into the pool opening of the expansion pool, the lysis capsule comprising: a hollow body having an open first end and an open second end; a first porous membrane attached to the hollow body and covering the open first end; a second porous membrane attached to the hollow body and covering the open second end, wherein the hollow body defines a lysis chamber between the first porous membrane and the second porous membrane; and a plurality of beads contained within the lysis chamber.

[0271] J24. The method according to embodiment J23, wherein the plurality of beads comprises a plurality of non-magnetic beads and at least one magnetic element.

[0272] Some implementation plans cover:

[0273] K1. A method of manufacturing a fluid cartridge, the method comprising: (a) providing a cartridge body including two or more pools, the two or more pools being fluidly connected or fluidly connectable; (b) inserting a lysis bladder into the pool opening of a first pool of the two or more pools, the lysis bladder including a lysis chamber and a plurality of lysis beads contained within the lysis chamber; and (c) securing a chamber expander to the cartridge body over the pool opening of the first pool, wherein the chamber expander includes: an expansion chamber that expands the volumetric capacity of the first pool by at least the volumetric capacity of the expansion chamber; a mouth that defines the expansion chamber opening into the interior space of the expansion chamber; and a cap configured to be coupled to the mouth to close the expansion chamber opening.

[0274] K2. The method according to embodiment K1, wherein (c) includes securing the chamber expander to the housing body by: connecting at least a portion of a first connection structure of the housing body around the first pool to a second connection structure of the chamber expander and forming an airtight seal between the chamber expander and the housing body.

[0275] K3. The method according to embodiment K2, wherein the first connecting structure includes a first peripheral wall formed in the housing body and surrounding the expansion pool, and the second connecting structure includes a second peripheral wall extending below the bottom side of the base of the chamber expander and configured to conform to the inner or outer surface of the first peripheral wall, and wherein connecting the first connecting structure to the second connecting structure includes attaching the second peripheral wall to the inner or outer surface of the first peripheral wall.

[0276] K4. The method according to embodiment K3, wherein attaching the second peripheral wall to the inner or outer surface of the first peripheral wall includes fixing the second peripheral wall to the inner or outer surface of the first peripheral wall by means of an adhesive or by means of welding.

[0277] K5. The method according to embodiment K2, wherein the first connecting structure includes a first peripheral wall formed on the housing body and surrounding the first pool, and the second connecting structure includes a second peripheral wall and a third peripheral wall, the second peripheral wall extending below the bottom side of the base of the chamber expander, the third peripheral wall extending below the bottom side of the base and spaced apart from the second peripheral wall to form a peripheral groove on the bottom side of the base, and wherein connecting the first connecting structure to the second connecting structure includes inserting the first peripheral wall into the peripheral groove between the second peripheral wall and the third peripheral wall, and attaching the first peripheral wall to at least one of the second peripheral wall and the third peripheral wall.

[0278] K6. The method according to embodiment K5, wherein attaching the first peripheral wall to at least one of the second peripheral wall and the third peripheral wall includes fixing the first peripheral wall to at least one of the second peripheral wall and the third peripheral wall by means of adhesive or by means of welding.

[0279] K7. The method according to any one of embodiments K1 to K6, further comprising dispensing reagent into each of one or more of the two or more pools other than the first pool, and sealing that the reagent has been dispensed into each of the pools.

[0280] K8. The method according to implementation scheme K7, wherein at least one reagent is a non-liquid reagent.

[0281] K9. The method according to any one of embodiments K1 to K8, wherein the lysis capsule comprises: a hollow body having an open first end and an open second end; a first porous membrane attached to the hollow body and covering the open first end; and a second porous membrane attached to the hollow body and covering the open second end, wherein the hollow body defines the lysis chamber between the first porous membrane and the second porous membrane.

[0282] K10. The method according to any one of embodiments K1 to K9, wherein the pyrolysis bead comprises a plurality of non-magnetic beads contained within the pyrolysis chamber; and wherein the pyrolysis capsule comprises at least one magnetic element contained within the pyrolysis chamber.

[0283] K11. The method according to any one of embodiments K1 to K10, further comprising covering the pool opening of at least one of the two or more pools other than the first pool to form a chamber that is at least partially enclosed in each covered pool.

[0284] K12. The method according to any one of embodiments K1 to K11, wherein the cap includes an insertion sleeve and a sheath, the sheath being wider than the insertion sleeve, and wherein the method includes attaching the cap to the mouth by inserting the insertion sleeve into the expansion chamber opening.

[0285] K13. The method according to any one of embodiments K1 to K11, further comprising: dispensing a certain amount of liquid substance into the opening of the expansion chamber, wherein the certain amount of liquid substance completely fills the first pool and at least partially fills the interior space of the expansion chamber; and attaching the cap to the mouth to close the opening.

[0286] K14. The method according to any one of embodiments K1 to K13, wherein the expansion chamber and the mouth have different shapes.

[0287] K15. The method according to embodiment K14, wherein the expansion chamber has three straight sides and the mouth has a circular shape.

[0288] K16. The method according to any one of embodiments K1 to K15, wherein the chamber expander further includes a perimeter chamfer formed in the inner surface of the mouth and extending around the expansion chamber opening.

[0289] K17. The method according to embodiment K5, wherein the first peripheral wall and the peripheral groove each have three sides.

[0290] K18. The method according to embodiment K17, wherein the first peripheral wall has three straight edges connected by rounded corners, and wherein the peripheral groove has three straight edges connected by rounded corners.

[0291] K19. The method according to embodiment K12, wherein the insertion sleeve is hollow, and the sheath includes a vent hole extending through the top wall of the sheath and opening to the interior space of the insertion sleeve, wherein the cap further includes a vent membrane disposed above the vent hole, and wherein the vent membrane is configured to allow gas to pass through but prevent liquid from passing through.

[0292] K20. The method according to embodiment K19, wherein the vent includes an inner vent portion and an outer vent portion, the inner vent portion being formed on the inner surface of the top wall of the sheath, the outer vent portion being formed on the outer surface of the top wall of the sheath, and wherein the width of the inner vent portion is different from the width of the outer vent portion.

[0293] K21. The method according to embodiment K20, wherein the width of the inner vent portion is smaller than the width of the outer vent portion.

[0294] K22. The method according to any one of embodiments K12 and K19 to K21, wherein the cap further includes radial ribs extending between the inner surface of the sheath and the outer surface of the insert sleeve, and wherein engaging the cap to the mouth to close the opening includes inserting the insert sleeve into the opening until the radial ribs contact the edge of the mouth surrounding the expansion chamber opening.

[0295] K23. According to the method of embodiment K22, the insertion sleeve is fixed in the expansion chamber opening by friction engagement.

[0296] K24. The method according to embodiment K23, wherein the cap includes circumferential ribs extending around the outer surface of the insert sleeve.

[0297] K25. The method according to any one of embodiments K19 to K24, further comprising at least one groove formed in the top surface of the top wall of the sheath, wherein the at least one groove extends through the vent.

[0298] K26. The method according to embodiment K25 further includes at least two grooves formed in the top surface of the sheath, wherein the at least two grooves intersect each other through the vent.

[0299] K27. The method according to any one of embodiments K1 to K26, wherein the chamber expander further includes a base and a strut extending from the base, and wherein the cap is hingedly connected to the strut.

[0300] K28. The method according to embodiment K27 further includes a tongue extending from the cap, wherein the cap is connected to the support via a movable hinge that connects the free end of the support to the free end of the tongue.

[0301] K29. The method according to any one of embodiments K1 to K28, wherein the expansion pool includes an inclined surface surrounding its opening.

[0302] Some implementation plans cover:

[0303] L1. A method for processing a fluid sample in a fluid cartridge, the fluid cartridge comprising two or more pools of uniform height and a chamber expander, the chamber expander being fixed relative to one of the two or more pools to expand the volumetric capacity of the chamber expander by increasing the volumetric capacity of the one pool, wherein the two or more pools are fluidly connected or fluidly connectable, and wherein the method comprises: (a) dispensing a quantity of the fluid sample into an opening defined by a nozzle of the chamber expander, wherein the quantity of the fluid sample completely fills the one pool and at least partially fills the interior space of the chamber expander; and (b) attaching a cap to the nozzle of the chamber expander to close the opening.

[0304] L2. The method according to embodiment L1 further includes (c) transferring air from the interior space of the chamber expander to the external environment through a vent in the cap.

[0305] L3. The method according to embodiment L2, wherein (c) includes passing air through a porous venting membrane, the porous venting membrane being attached to the cap and covering the vent holes, the porous venting membrane being impermeable to liquids.

[0306] L4. The method according to any one of embodiments L1 to L3, wherein the chamber expander is secured by connecting a first connecting structure at least partially around the one pool to a second connecting structure of the chamber expander and forming an airtight seal between the chamber expander and the one pool.

[0307] L5. The method according to any one of embodiments L1 to L4, wherein, in addition to the fluid sample being dispensed into one of the pools, at least one of the two or more pools contains a reagent for treating the fluid sample.

[0308] L6. The method according to embodiment L5, wherein the reagent is a non-liquid reagent.

[0309] L7. The method according to any one of embodiments L1 to L6, wherein the cap includes an insertion sleeve and a sheath, the sheath being wider than the insertion sleeve, and wherein (b) includes inserting the insertion sleeve into the opening.

[0310] L8. The method according to embodiment L7, wherein the insertion sleeve is fixed within the opening by friction engagement.

[0311] L9. The method according to any one of embodiments L1 to L8, wherein one of the pools includes a lysis chamber, the lysis chamber including a first porous membrane and a second porous membrane defined therebetween, and a plurality of beads contained within the lysis chamber.

[0312] L10. The method according to embodiment L9 further includes (d) agitating the plurality of beads in the lysis chamber to lyse the cells contained in the fluid sample, thereby releasing nucleic acids from the lysed cells.

[0313] L11. The method according to embodiment L10, wherein the plurality of beads comprises a plurality of non-magnetic beads and at least one magnetic element, and wherein (d) includes subjecting the at least one magnetic element to a magnetic field, thereby causing the at least one magnetic element to move in the pyrolysis chamber, the movement of the at least one magnetic element in the pyrolysis chamber causing the plurality of non-magnetic beads to move in the pyrolysis chamber.

[0314] L12. The method according to embodiment L11, wherein the magnetic field is generated by an electromagnet.

[0315] L13. The method of claim L12, further comprising alternating the current to the electromagnet to alter the polarity of the electromagnet.

[0316] L14. The method according to embodiment L13, which includes alternating the current at a frequency of 20 Hz to 200 Hz to alter the polarity of the electromagnet.

[0317] L15. The method according to embodiment L14, which includes pulsed the current at two or more different frequencies to alternate the polarity of the electromagnet.

[0318] L16. The method according to any one of embodiments L10 to L15, further comprising (e) delivering the released nucleic acid from said one pool to the processing chamber of said box.

[0319] L17. The method according to embodiment L16 further includes (f) during (e) retaining the lysed cellular material in the lysis chamber while allowing the released nucleic acid to pass through the second porous membrane.

[0320] L18. The method according to embodiment L17 further includes (g) agitating the plurality of beads in the lysis chamber during (e) to keep at least a portion of the lysed cell material in the lysis chamber suspended, at least until the fluid sample has been removed from the lysis chamber.

[0321] L19. The method according to embodiment L17 or L18, further comprising (h) immobilizing at least a portion of the released nucleic acid in the processing chamber onto a solid support, and removing the unimmobilized components of the fluid sample into the waste chamber of the cartridge.

[0322] L20. The method according to embodiment L19 further includes (i) eluting the immobilized nucleic acid from the solid support and delivering the eluted nucleic acid to the reaction chamber of the cassette.

[0323] L21. The method according to embodiment L20, further comprising (j) subjecting the eluted nucleic acid to conditions of a first reaction, the first reaction providing an indication of the presence or amount of the analyte of interest.

[0324] L22. The method according to embodiment L21, further comprising: (k) releasing an internal control into the fluid sample, the internal control being contained within the lysis chamber; (l) immobilizing a nucleic acid (“IC nucleic acid”) associated with the internal control on the solid support; eluting the IC nucleic acid from the solid support; (m) delivering the eluted IC nucleic acid to the reaction chamber; and (n) subjecting the IC nucleic acid to conditions of a second reaction, the second reaction providing an indication of the degree of lysis, wherein the internal control is provided to validate the assay and / or to validate the effectiveness of cell lysis during (d).

[0325] L23. The method according to embodiment L22, wherein at least a portion of the internal control is contained in an internal control reagent disposed on the first porous membrane, and wherein the internal control reagent is dissolved in the fluid sample.

[0326] L24. The method according to embodiment L22 or L23, wherein the internal control is contained in an internal control reagent disposed on at least a portion of the plurality of beads, and wherein the internal control reagent disposed on at least a portion of the plurality of beads is dissolved in the fluid sample.

[0327] L25. The method according to embodiment L22, wherein the internal control is embedded in or contained in an internal control pellet, the internal control pellet is contained in the lysis chamber, and wherein the internal control pellet is adapted to dissolve upon contact with the fluid sample and / or disintegrate during (d).

[0328] L26. The method according to any one of embodiments L22 to L25, wherein the conditions for the first reaction and the conditions for the second reaction are the same.

[0329] L27. The method according to any one of embodiments L22 to L26, wherein each of the first reaction and the second reaction is a nucleic acid amplification reaction.

[0330] L28. The method according to embodiment L27, wherein the nucleic acid amplification reaction is a polymerase chain reaction (“PCR”).

[0331] Some implementation plans cover:

[0332] M1. A bead delivery cap for dispensing a pyrolysis agent into a sample cell of a fluid cartridge, the cap comprising: a cap body including a deformable wall defining a chamber; a lower sleeve located below the deformable wall and defining a recess opening into the chamber; a brittle membrane attached to an open bottom end of the lower sleeve and closing the recess and the chamber; and a pyrolysis agent comprising a plurality of nonmagnetic beads and at least one magnetic element, wherein the pyrolysis agent is contained within the chamber and the recess in an amount such that deformation of the deformable wall causes the pyrolysis agent to rupture the brittle membrane, thereby releasing the pyrolysis agent from the chamber and the recess.

[0333] M2. The bead delivery cap according to embodiment M1, wherein the lower sleeve includes at least one sealing rib extending around the outer surface of the lower sleeve.

[0334] M3. A beaded delivery cap according to embodiment M1 or M2, wherein the lower sleeve has a cylindrical shape.

[0335] M4. A beaded delivery cap according to any one of embodiments M1 to M3, wherein the cap body includes a lateral extension member, the upper end of the lower sleeve is connected to the lateral extension member, and wherein the deformable wall extends upward from the lateral extension member.

[0336] M5. A beaded cap is delivered according to any one of embodiments M1 to M4, wherein the deformable wall is dome-shaped when not in a deformable state.

[0337] M6. The beaded delivery cap according to embodiment M4 or M5, further comprising a circumferential perimeter wall spaced apart from the deformable wall and projecting upward from the outer perimeter of the lateral extension member.

[0338] M7. The beaded delivery cap according to embodiment M6, wherein the top of the upper peripheral wall is located above the deformable wall.

[0339] M8. A beaded delivery cap according to any one of embodiments M1 to M7, wherein the cap body is an integral structure made of polymer material.

[0340] M9. The beaded delivery cap according to embodiment M8, wherein the polymer material is a thermoplastic elastomer.

[0341] M10. A beaded delivery cap according to any one of embodiments M1 to M9, wherein the fragile membrane comprises a porous film.

[0342] M11. A bead delivery cap according to any one of embodiments M1 to M10, further comprising a peelable covering film covering the outer surface of the fragile film.

[0343] M12. A beaded delivery cap according to any one of embodiments M1 to M11, wherein the fragile membrane includes one or more rupture lines configured to make the fragile membrane more prone to breakage.

[0344] M13. The beaded delivery cap according to embodiment M12, wherein the one or more rupture lines consist of a single line having a C-shape or U-shape.

[0345] M14. A beaded delivery cap according to any one of embodiments M1 to M13, wherein the at least one magnetic element is arranged adjacent to the inner surface of the deformable wall.

[0346] M15. A beaded delivery cap according to any one of embodiments M1 to M14, wherein the deformable wall includes a vent formed in the deformable wall and in fluid communication with the chamber.

[0347] M16. The bead delivery cap according to embodiment M15, further comprising a porous membrane covering the vent, the porous membrane being attached to the top or bottom surface of the deformable wall.

[0348] M17. A beaded delivery cap according to any one of embodiments M1 to M16, wherein each of the plurality of non-magnetic beads is composed of ceramic or glass.

[0349] M18. A beaded delivery cap according to any one of embodiments M1 to M17, wherein each of the plurality of non-magnetic beads has a spherical shape.

[0350] M19. A beaded delivery cap according to embodiment M18, wherein each of the plurality of non-magnetic beads has a diameter of 100 µm to 2000 µm.

[0351] M20. A beaded delivery cap according to any one of embodiments M1 to M19, wherein the at least one magnetic element is coated or encapsulated with a non-magnetic material.

[0352] M21. The beaded delivery cap according to embodiment M20, wherein the non-magnetic material is metal or plastic.

[0353] M22. A beaded delivery cap according to any one of embodiments M1 to M21, wherein the at least one magnetic element occupies a larger volume than any one of the plurality of non-magnetic beads.

[0354] M23. A beaded delivery cap according to any one of embodiments M1 to M22, wherein the at least one magnetic element has a cubic shape.

[0355] M24. The beaded delivery cap according to embodiment M23, wherein the width of each face of the cube is 2.0 mm to 4.3 mm.

[0356] M25. A beaded delivery cap according to any one of embodiments M1 to M24, wherein the at least one magnetic element is composed of neodymium.

[0357] M26. The beaded delivery cap according to embodiment M25, wherein the neodymium is N52 or N42 grade.

[0358] M27. A beaded delivery cap according to any one of embodiments M1 to M26, wherein each of the at least one magnetic element and each of the plurality of non-magnetic beads is inert.

[0359] M28. A beaded delivery cap according to any one of embodiments M1 to M27, wherein a force of 1.0 lb to 5.0 lb is required to be applied to the deformable wall to break the fragile membrane.

[0360] M29. A beaded delivery cap according to any one of embodiments M1 to M28, further comprising an internal control contained within the cap body, wherein the internal control is provided to verify assay results and / or to verify the effectiveness of a cell lysis procedure performed with a lysis agent.

[0361] M30. A bead delivery cap according to embodiment M29, wherein the internal control is contained in an internal control reagent, wherein at least a portion of the internal control reagent is disposed on at least a portion of the plurality of nonmagnetic beads and / or the at least one magnetic element, and wherein the internal control reagent is adapted to dissolve upon contact with a fluid sample.

[0362] M31. A bead delivery cap according to embodiment M29, wherein the internal control is embedded in or contained in internal control pellets adapted to dissolve upon contact with a fluid sample and / or disintegrate upon agitation of the plurality of magnetic beads, the internal control pellets being contained within the cap body.

[0363] Some implementation plans cover:

[0364] N1. A fluid cartridge comprising: a cartridge body including a sample chamber having an open top; and a beaded delivery cap inserted into the open top of the sample chamber according to any one of embodiments M1 to M32.

[0365] N2. The fluid cartridge according to embodiment N1, further comprising an injection barrel body in communication with the sample chamber, the injection barrel body being adapted to receive an injection barrel stop connected to an injection barrel plunger for actuating fluid within the fluid cartridge.

[0366] Some implementation plans cover:

[0367] O1. A method for lysing cells contained in a sample, the method comprising: (a) providing the sample to a sample chamber of a fluid cartridge; (b) inserting a beaded delivery cap according to any one of embodiments M1 to M32 into the sample chamber such that (i) the outer surface of the lower sleeve is sealed to the inner surface of the sidewall of the sample chamber, and (ii) the fragile membrane is positioned above the sample in the sample chamber; (c) applying a force to the deformable wall to deform the deformable wall to the extent that the lysing agent ruptures the fragile membrane and is released from the chamber and the recess into the sample chamber; and (d) subjecting the sample and the lysing agent to a magnetic field, the magnetic field causing the at least one magnetic element to agitate the plurality of nonmagnetic beads to lyse the cells contained in the sample.

[0368] O2. The method according to embodiment O1, wherein (c) includes moving the bead-shaped delivery cap actuator to apply the force to the deformable wall.

[0369] O3. The method according to embodiment O1, wherein (c) includes manually applying the force to the deformable wall.

[0370] O4. The method according to any one of embodiments O1 to O3, wherein the pyrolysis agent occupies at least 90% of the volume of the chamber and the recess of the cap.

[0371] O5. The method according to any one of embodiments O1 to O4, wherein (a) includes providing the sample to the sample chamber using a pipette.

[0372] O6. The method according to any one of embodiments O1 to O5, wherein (c) includes applying a force of 1.0 lb to 5.0 lb to the deformable wall.

[0373] Some implementation plans cover:

[0374] P1. A method for lysing cells contained in a fluid sample, the method comprising: (a) providing the fluid sample to a sample chamber of a fluid cartridge; (b) securing a cap to an open end of the sample chamber, the cap including a deformable wall defining a chamber containing a lysing agent and closed by a fragile membrane, the lysing agent comprising a plurality of nonmagnetic beads and at least one magnetic element, the at least one magnetic element being larger than any one of the plurality of nonmagnetic beads; (c) applying a force to the top of the deformable wall to cause the chamber to collapse to the extent that the lysing agent contained within the chamber ruptures the fragile membrane, thereby releasing the lysing agent from the chamber into the sample chamber; and (d) subjecting the fluid sample and the lysing agent to a magnetic field, the magnetic field causing the at least one magnetic element to agitate the plurality of nonmagnetic beads to lyse the cells contained within the fluid sample.

[0375] P2. The method according to embodiment P1, wherein the deformable wall is dome-shaped when not in a deformable state.

[0376] P3. The method according to embodiment P1 or P2, wherein the cap is an integral structure composed of polymer material.

[0377] P4. The method according to embodiment P3, wherein the polymer material is a thermoplastic elastomer.

[0378] P5. The method according to any one of embodiments P1 to P4, wherein the fragile membrane comprises a porous membrane.

[0379] P6. The method according to any one of embodiments P1 to P5, further comprising removing the peelable covering film from the outer surface of the fragile film prior to (b).

[0380] P7. The method according to any one of embodiments P1 to P6, wherein the fragile membrane includes one or more rupture lines, the one or more rupture lines being configured to make the fragile membrane more prone to rupture.

[0381] P8. The method according to embodiment P7, wherein the one or more rupture lines consist of a single line having a C-shape or a U-shape.

[0382] P9. The method according to any one of embodiments P1 to P8, wherein the at least one magnetic element is arranged adjacent to the inner surface of the deformable wall.

[0383] P10. The method according to any one of embodiments P1 to P9, wherein the deformable wall includes a vent formed in the deformable wall and in fluid communication with the chamber.

[0384] P11. The method according to embodiment P10 further includes a porous membrane covering the vent, the porous membrane being attached to the top or bottom surface of the deformable wall.

[0385] P12. The method according to any one of embodiments P1 to P11, wherein each of the plurality of non-magnetic beads is composed of ceramic or glass.

[0386] P13. The method according to any one of embodiments P1 to P12, wherein each of the plurality of non-magnetic beads has a spherical shape.

[0387] P14. The method according to embodiment P13, wherein each of the plurality of nonmagnetic beads has a diameter of 100 µm to 2000 µm.

[0388] P15. The method according to any one of embodiments P1 to P14, wherein the at least one magnetic element is coated or encapsulated with a non-magnetic material.

[0389] P16. The method according to embodiment P15, wherein the non-magnetic material is a metal or a plastic.

[0390] P17. The method according to any one of embodiments P1 to P16, wherein the at least one magnetic element occupies a larger volume than any one of the plurality of non-magnetic beads.

[0391] P18. The method according to any one of embodiments P1 to P17, wherein the at least one magnetic element has a cubic shape.

[0392] P19. The method according to embodiment P18, wherein the width of each face of the cube is 2.0 mm to 4.3 mm.

[0393] P20. The method according to any one of embodiments P1 to P19, wherein the at least one magnetic element is composed of neodymium.

[0394] P21. The method according to embodiment P20, wherein the neodymium is N52 or N42 grade.

[0395] P22. The method according to any one of embodiments P1 to P21, wherein each of the plurality of non-magnetic beads and the at least one magnetic element are inert.

[0396] P23. The method according to any one of embodiments P1 to P22, wherein (c) includes applying a force of 1.0 lb to 5.0 lb to the deformable wall.

[0397] P24. The method according to any one of embodiments P1 to P23, further comprising arranging an internal control in the chamber, wherein the internal control is provided to verify the determination of the fluid sample performed after (d) and / or to verify the effectiveness of (d) in lysing the cells.

[0398] P25. The method according to embodiment P24, wherein at least a portion of the internal control is contained in an internal control reagent, the internal control reagent is disposed on at least a portion of the plurality of non-magnetic beads and / or the at least one magnetic element, wherein the internal control reagent is adapted to dissolve upon contact with the fluid sample.

[0399] P26. The method according to embodiment P24, wherein at least a portion of the internal control is embedded in or contained within an internal control pellet, the internal control pellet being adapted to dissolve upon contact with the fluid sample and / or disintegrate upon agitation of the plurality of magnetic beads, the internal control pellet being contained within the chamber.

[0400] Some implementation plans cover:

[0401] Q1. A method for lysing cells contained in a fluid sample, the method comprising: (a) positioning an electromagnet at a predetermined distance from a sample chamber of a microfluidic device, the sample chamber containing the fluid sample and a lysing agent, the lysing agent comprising a plurality of nonmagnetic beads and at least one magnetic element; (b) generating a magnetic field through the electromagnet, the magnetic field targeting the sample and the lysing agent contained in the sample chamber; and (c) reversing the polarity of the magnetic field, causing the at least one magnetic element to move randomly within the sample chamber to agitate the plurality of nonmagnetic beads to lyse the cells contained in the fluid sample, such that nucleic acids are released from the lysed cells, wherein the electromagnet is spatially separated from the sample chamber and remains stationary while generating the magnetic field targeting the fluid sample and the lysing agent contained within the sample chamber.

[0402] Q2. The method according to implementation scheme Q1, wherein (c) includes reversing the polarity of the magnetic field at a predetermined frequency of 60 Hz to 200 Hz.

[0403] Q3. The method according to embodiment Q1, wherein (b) includes driving the electromagnet with a predetermined voltage of 10 volts to 50 volts.

[0404] Q4. The method according to implementation scheme Q1, wherein (c) includes charging the electromagnet by reversing the polarity of the magnetic field using a switching amplifier.

[0405] Q5. The method according to any one of embodiments Q1 to Q4, further comprising, before (a), (d) manually or robotically dispensing the fluid sample into the sample chamber, optionally wherein (d) is performed with a pipette, and wherein the sample chamber contains the lysis agent before (d).

[0406] Q6. The method according to embodiment Q5 further includes (e) covering the open top of the sample chamber with a cap after (d) and before (b) and (c).

[0407] Q7. The method according to any one of embodiments Q1 to Q6, further comprising: (f) after (c), conveying at least a portion of the fluid sample from the sample chamber to the processing chamber of the fluid cartridge.

[0408] Q8. The method according to embodiment Q7 further includes: (g) during (e), retaining the lysed cell material from (c) in the sample chamber while allowing the released nucleic acids from the lysed cells to pass through a porous membrane.

[0409] Q9. The method according to embodiment Q8 further comprises: (h) reversing the polarity of the magnetic field during (g) to move the at least one magnetic element in the sample chamber, the movement of the at least one magnetic element in the sample chamber causing the plurality of non-magnetic beads to move in the sample chamber, and the movement of the plurality of non-magnetic beads in the sample chamber causing at least a portion of the lysed cellular material in the sample chamber to remain suspended at least until the fluid sample has been removed from the sample chamber.

[0410] Q10. The method according to any one of embodiments Q8 to Q9, further comprising: (i) immobilizing at least a portion of the released nucleic acid on a solid support in the processing chamber, and removing the unimmobilized components of the fluid sample into the waste chamber of the microfluidic device.

[0411] Q11. The method according to embodiment Q10 further includes: (j) after (i), eluting the immobilized nucleic acid from the solid support and delivering the eluted nucleic acid to the reaction chamber of the microfluidic device.

[0412] Q12. The method according to embodiment Q11 further includes: (k) after (j), subjecting the eluted nucleic acid to the conditions of a first reaction, the first reaction providing an indication of the presence or amount of the analyte of interest.

[0413] Q13. The method according to embodiment Q12 further comprises: (l) releasing an internal control into the fluid sample during or after (a), wherein the internal control is contained in the sample chamber prior to (a); (m) immobilizing a nucleic acid (“IC nucleic acid”) associated with the internal control on the solid support during (i); (n) eluting the IC nucleic acid from the solid support and delivering the eluted IC nucleic acid to the reaction chamber after (i); and (o) subjecting the IC nucleic acid to the conditions of a second reaction after (n), wherein the results of (o) are used to verify the results of (k) and / or to verify the effectiveness of the lysis in (c).

[0414] Q14. The method according to embodiment Q13, wherein at least a portion of the internal control is contained in an internal control reagent, and when (a) is initiated, the internal control reagent is disposed on at least a portion of the sample chamber, and wherein the internal control reagent dissolves in the fluid sample during any of (a) to (c).

[0415] Q15. The method according to embodiment Q13, wherein at least a portion of the internal control is contained in an internal control reagent, and when (a) is activated, the internal control reagent is disposed on at least a portion of the plurality of non-magnetic beads and / or the at least one magnetic element, and wherein during any one of (a) to (c), the internal control reagent disposed on at least a portion of the plurality of non-magnetic beads and / or the at least one magnetic element dissolves in the fluid sample.

[0416] Q16. The method according to embodiment Q13, wherein at least a portion of the internal control is contained in an internal control reagent, and when (a) is initiated, the internal control reagent is disposed on the inner wall of the sample chamber, and wherein the internal control reagent dissolves in the fluid sample during any of (a) to (c).

[0417] Q17. The method according to embodiment Q13, wherein the internal control is embedded in or contained within internal control pellets, and wherein the internal control pellets dissolve in the presence of the fluid sample and / or break apart during (c) by the movement of the plurality of nonmagnetic beads, thereby releasing the internal control into the fluid sample.

[0418] Q18. The method according to any one of embodiments Q13 to Q17, wherein the conditions for the first reaction and the conditions for the second reaction are the same.

[0419] Q19. The method according to any one of embodiments Q13 to Q18, wherein each of the first reaction and the second reaction is a nucleic acid amplification reaction.

[0420] Q20. The method according to implementation plan Q19, wherein the nucleic acid amplification reaction is a polymerase chain reaction (“PCR”).

[0421] Some implementation plans cover:

[0422] R1. A fluid cartridge comprising: a sample chamber having an open top and a sample outlet port; and a pyrolysis chamber within the sample chamber, wherein the pyrolysis chamber is defined by an inner wall of the sample chamber and a first porous membrane attached to the sample chamber, the first porous membrane overlapping the open top; and a second porous membrane spaced apart from the first porous membrane attached to the sample chamber, the second porous membrane overlapping the sample outlet port; a plurality of non-magnetic beads contained within the pyrolysis chamber; and at least one magnetic element contained within the pyrolysis chamber, wherein the pore sizes of the first porous membrane and the second porous membrane are designed to retain the plurality of non-magnetic beads and the at least one magnetic element within the pyrolysis chamber.

[0423] R2. The fluid cartridge according to embodiment R1, wherein at least one of the first porous membrane and the second porous membrane includes a mesh.

[0424] R3. The fluid cartridge according to embodiment R1, wherein at least one of the first porous membrane and the second porous membrane comprises a filter matrix.

[0425] R4. A fluid cell according to any one of embodiments R1 to R3, wherein the porosity or porosity range of the first porous membrane is greater than the porosity or porosity range of the second porous membrane.

[0426] R5. The fluid cell according to embodiment R4, wherein the first porous membrane has a porosity of 70 µm to 500 µm, and wherein the second porous membrane has a porosity of 30 µm to 100 µm.

[0427] R6. The fluid box according to any one of embodiments R1 to R5, wherein each of the plurality of non-magnetic beads is composed of ceramic or glass.

[0428] R7. The fluid cartridge according to any one of embodiments R1 to R6, wherein each of the plurality of nonmagnetic beads has a spherical shape, and wherein each of the plurality of nonmagnetic beads optionally has a diameter of 100 µm to 2000 µm.

[0429] R8. The fluid cartridge according to any one of embodiments R1 to R7, wherein the at least one magnetic element is coated or encapsulated with a non-magnetic material.

[0430] R9. The fluid box according to any one of embodiments R1 to R8, wherein the at least one magnetic element occupies a larger volume than any one of the plurality of non-magnetic beads.

[0431] R10. A fluid box according to any one of embodiments R1 to R9, wherein the at least one magnetic element has a cubic shape, and wherein each face of the cube optionally has a width of 2.0 mm to 4.3 mm.

[0432] R11. The fluid cell according to any one of embodiments R1 to R10, wherein the at least one magnetic element is composed of neodymium, and wherein the neodymium is optionally of N52 or N42 grade.

[0433] R12. The fluid cartridge according to any one of embodiments R1 to R11, further comprising an internal control contained within the lysis chamber, wherein the internal control is provided to verify assay results and / or to verify the effectiveness of the cell lysis procedure performed with the plurality of nonmagnetic beads and the at least one magnetic element.

[0434] R13. The fluid cartridge according to embodiment R12, wherein at least a portion of the internal control is contained in an internal control reagent, the internal control reagent is disposed on at least one of the first porous membrane and the second porous membrane, and wherein the internal control reagent is adapted to dissolve upon contact with a fluid sample.

[0435] R14. The fluid cartridge according to embodiment R12, wherein at least a portion of the internal control is contained in an internal control reagent, the internal control reagent is disposed on at least a portion of the plurality of nonmagnetic beads and / or the at least one magnetic element, and wherein the internal control reagent is adapted to dissolve upon contact with a fluid sample.

[0436] R15. The fluid cartridge according to embodiment R12, wherein at least a portion of the internal control is contained in an internal control reagent disposed on the inner wall of the sample chamber, and wherein the internal control reagent is adapted to dissolve upon contact with a fluid sample.

[0437] R16. The fluid cartridge according to embodiment R12, wherein at least a portion of the internal control is embedded in or contained in internal control pellets contained within the lysis chamber, and wherein the internal control pellets are adapted to dissolve upon contact with a fluid sample and / or to disintegrate upon agitation of the plurality of magnetic beads.

[0438] R17. The fluid cartridge according to any one of embodiments R1 to R16, further comprising an injection barrel body in communication with the sample chamber, the injection barrel body being adapted to receive an injection barrel stop connected to an injection barrel plunger for actuating fluid within the fluid cartridge.

[0439] R18. A fluid cartridge according to any one of embodiments R1 to R17, wherein the sample chamber is covered by a removable seal.

[0440] R19. The fluid cartridge according to any one of embodiments R1 to R18, further comprising a cap adapted for insertion into an open top of the sample chamber.

[0441] R20. A fluid cell according to any one of embodiments R1 to R19, wherein the first porous membrane is press-fitted within a portion of the sample chamber.

[0442] R21. A fluid cartridge according to any one of embodiments R1 to R19, wherein the first porous membrane is heat-sealed within a portion of the sample chamber, and optionally wherein the sample chamber includes a first lateral wall with an energy director, the first porous membrane being heat-sealed onto the first lateral wall.

[0443] R22. The fluid cell according to any one of embodiments R1 to R21, wherein the second porous membrane is press-fitted within a portion of the sample chamber.

[0444] R23. A fluid cartridge according to any one of embodiments R1 to R21, wherein the second porous membrane is heat-sealed within a portion of the sample chamber, and optionally wherein the sample chamber includes a second lateral wall with an energy guide, the second porous membrane being heat-sealed onto the second lateral wall.

[0445] Some implementation plans cover:

[0446] S1. A method for lysing cells contained in a fluid sample, the method comprising: (a) dispensing the fluid sample into a sample chamber of a fluid cartridge according to embodiment R1 to at least partially fill the lysis chamber with the fluid sample; (b) after (a), covering the open top of the sample chamber with a cap; and (c) after (b), subjecting the at least one magnetic element to a magnetic field, thereby causing the at least one magnetic element to move within the lysis chamber, the movement of the at least one magnetic element within the lysis chamber causing the movement of a plurality of non-magnetic beads within the lysis chamber, and the movement of the non-magnetic beads within the lysis chamber causing the cells contained in the fluid sample within the lysis chamber to lyse and release nucleic acids.

[0447] S2. The method according to embodiment S1, further comprising (d) prior to (c), contacting the fluid sample with an internal control, the internal control being disposed in the sample chamber and optionally in the lysis chamber, wherein the internal control is provided to verify the assay results and / or to verify the effectiveness of (c) in lysing cells and releasing nucleic acids within the fluid sample contained in the lysis chamber.

[0448] S3. The method according to embodiment S2, wherein at least a portion of the internal control is contained within an internal control reagent, the internal control reagent being disposed in the sample chamber and optionally in the lysis chamber.

[0449] S4. The method according to embodiment S3, wherein at least a portion of the internal control reagent is disposed on at least one of the first porous membrane and the second porous membrane, and wherein the internal control reagent is adapted to dissolve upon contact with the fluid sample.

[0450] S5. The method according to embodiment S3, wherein at least a portion of the internal control reagent is disposed on at least a portion of the plurality of non-magnetic beads and / or the at least one magnetic element, and wherein the internal control reagent is adapted to dissolve upon contact with the fluid sample.

[0451] S6. The method according to embodiment S3, wherein at least a portion of the internal control reagent is disposed on the inner wall of the sample chamber, and wherein the internal control reagent is adapted to dissolve upon contact with the fluid sample.

[0452] S7. The method according to embodiment S2, wherein at least a portion of the internal control is embedded in or contained in an internal control pellet, the internal control pellet being adapted to dissolve upon contact with the fluid sample and / or to disintegrate during (c).

[0453] S8. The method according to any one of embodiments S1 to S7, wherein the fluid cartridge includes an injection barrel body in communication with the sample chamber, the injection barrel body being adapted to receive an injection barrel stop connected to an injection barrel plunger for actuating fluid within the fluid cartridge.

[0454] S11. The method according to any one of embodiments S1 to S8, further comprising covering the sample chamber with a removable seal.

[0455] S10. The method according to any one of embodiments S1 to S9, further comprising inserting a cap into the open top of the sample chamber.

[0456] S11. The method according to any one of embodiments S1 to S10, comprising press-fitting the first porous membrane within a portion of the sample chamber.

[0457] S12. The method according to any one of embodiments S1 to S10, comprising heat-sealing the first porous membrane within a portion of the sample chamber, and optionally wherein the sample chamber includes a first lateral wall with an energy director, the first porous membrane being heat-sealed onto the first lateral wall.

[0458] S13. The method according to any one of embodiments S1 to S12, comprising press-fitting the second porous membrane within a portion of the sample chamber.

[0459] S14. The method according to any one of embodiments S1 to S12, comprising heat-sealing the second porous membrane within a portion of the sample chamber, and optionally wherein the sample chamber includes a second lateral wall with an energy director, the second porous membrane being heat-sealed onto the second lateral wall.

[0460] S15. The method according to any one of embodiments S1 to S14, wherein the magnetic field during (c) is generated by an electromagnet.

[0461] S16. The method according to embodiment S15, wherein (c) includes alternating the current to the electromagnet to alter the polarity of the electromagnet.

[0462] S17. The method according to embodiment S16, wherein (c) includes alternating the current at a frequency of 20 Hz to 200 Hz to alter the polarity of the electromagnet.

[0463] S18. The method according to embodiment S17, wherein (c) includes pulses of the current at two or more different frequencies to alter the field polarity of the electromagnet.

[0464] S19. The method according to any one of embodiments S1 to S18, further comprising: (e) after (c), conveying at least a portion of the fluid sample from the sample chamber to the processing chamber of the fluid cartridge.

[0465] S20. The method according to embodiment S19 further includes: (f) during (e), retaining the lysed cell material in the lysis chamber while allowing the released nucleic acid to pass through the second porous membrane.

[0466] S21. The method according to embodiment S20, further comprising: (g) during (e), subjecting the at least one magnetic element to the magnetic field, thereby moving the at least one magnetic element in the lysis chamber, the movement of the at least one magnetic element in the lysis chamber causing the plurality of non-magnetic beads to move in the lysis chamber, and the movement of the non-magnetic beads in the lysis chamber causing at least a portion of the lysed cell material in the lysis chamber to remain suspended until the fluid sample is removed from the lysis chamber.

[0467] S22. The method according to any one of embodiments S19 to S21, further comprising: (h) immobilizing at least a portion of the released nucleic acid on a solid support in the processing chamber, and removing the unimmobilized components of the fluid sample into the waste chamber of the fluid cartridge.

[0468] S23. The method according to embodiment S22 further includes: (i) after (h), eluting the immobilized nucleic acid from the solid support and delivering the eluted nucleic acid to the reaction chamber of the fluid cartridge.

[0469] S24. The method according to embodiment S23 further includes: (j) after (i), subjecting the eluted nucleic acid to reaction conditions that provide an indication of the presence or amount of the analyte of interest.

[0470] S25. The method according to embodiment S24, wherein the reaction is a nucleic acid amplification reaction, and optionally the nucleic acid amplification reaction is a polymerase chain reaction (“PCR”).

[0471] Some implementation plans cover:

[0472] T1. A method of manufacturing a fluid cartridge, the method comprising: (a) providing a cartridge body including a sample chamber and one or more chambers, the sample chamber having an open first end and a sample outlet port at a second end of the sample chamber, the one or more chambers being fluidly connected to or fluidly connectable to the sample chamber; (b) providing a first porous membrane and a second porous membrane and attaching the second porous membrane to the sample chamber, the second porous membrane covering the sample outlet port; (c) introducing a plurality of non-magnetic beads into the sample chamber through the open first end of the sample chamber; (d) introducing at least one magnetic element into the sample chamber through the open first end of the sample chamber; and (e) in (c) and (d) Subsequently, the first porous membrane is attached to the sample chamber, the first porous membrane overlapping the open first end of the sample chamber, wherein the inner wall of the sample chamber and the attached first and second porous membranes define a pyrolysis chamber, and wherein the size of the pores of the first and second porous membranes is designed to retain the plurality of non-magnetic beads and the at least one magnetic element in the pyrolysis chamber.

[0473] T2. The method according to embodiment T1, wherein the first porous membrane comprises a mesh or a filter matrix.

[0474] T3. The method according to embodiment T1 or T2, wherein the second porous membrane comprises a mesh or a filter matrix.

[0475] T4. The method according to any one of embodiments T1 to T3, wherein the first porous membrane is hydrophilic.

[0476] T5. The method according to any one of embodiments T1 to T4, wherein the porosity or porosity range of the first porous membrane is greater than the porosity or porosity range of the second porous membrane.

[0477] T6. The method according to embodiment T5, wherein the first porous membrane has a porosity of 70 µm to 500 µm, and the second porous membrane has a porosity of 30 µm to 100 µm.

[0478] T7. The method according to any one of embodiments T1 to T6, wherein the first porous membrane is press-fitted into the sample chamber.

[0479] T8. The method according to any one of embodiments T1 to T6, wherein the first porous membrane is heat-sealed in the sample chamber.

[0480] T9. The method according to any one of embodiments T1 to T8, wherein the second porous membrane is press-fitted into the sample chamber.

[0481] T10. The method according to any one of embodiments T1 to T8, wherein the second porous membrane is heat-sealed in the sample chamber.

[0482] T11. The method according to any one of embodiments T1 to T10, wherein each of the plurality of non-magnetic beads is composed of ceramic or glass.

[0483] T12. The method according to any one of embodiments T1 to T11, wherein each of the plurality of nonmagnetic beads has a spherical shape, and wherein each of the plurality of nonmagnetic beads optionally has a diameter of 100 µm to 2000 µm.

[0484] T13. The method according to any one of embodiments T1 to T12, wherein the at least one magnetic element is coated or encapsulated with a non-magnetic material.

[0485] T14. The method according to any one of embodiments T1 to T13, wherein the at least one magnetic element occupies a larger volume than any one of the plurality of non-magnetic beads.

[0486] T15. The method according to any one of embodiments T1 to T14, wherein the at least one magnetic element has a cubic shape, and wherein the width of each face of the cubic is optionally 2.0 mm to 4.3 mm.

[0487] T16. The method according to any one of embodiments T1 to T15, wherein the at least one magnetic element is composed of neodymium, and wherein the neodymium is optionally of N52 or N42 grade.

[0488] T17. The method according to any one of embodiments T1 to T16, wherein each of the plurality of non-magnetic beads and the at least one magnetic element are inert.

[0489] T18. The method according to any one of embodiments T1 to T18, further comprising arranging an internal control on a component of the lysis sac-like element, wherein the internal control is provided to verify assay results and / or to verify the effectiveness of the cell lysis procedure performed with the plurality of nonmagnetic beads and the at least one magnetic element.

[0490] T19. The method according to embodiment T18, comprising arranging at least a portion of the internal control onto at least a portion of the plurality of nonmagnetic beads and / or the magnetic element before or after (c), wherein the internal control is contained in an internal control reagent adapted for dissolving upon contact with a fluid sample.

[0491] T20. The method according to embodiment T18, comprising arranging the internal control onto at least one of: (i) the first porous membrane, (ii) the second porous membrane, (iii) the inner wall of the sample chamber, (iv) at least a portion of the plurality of nonmagnetic beads, and (v) the at least one magnetic element preceding (e), wherein the internal control is contained in an internal control reagent adapted to dissolve upon contact with a fluid sample.

[0492] T21. The method according to any one of embodiments T18 to T20, wherein arranging the internal control comprises arranging the internal control reagent in liquid form and drying the internal control reagent after it has been arranged.

[0493] T22. The method according to any one of embodiments T1 to T17, wherein an internal control is embedded in or contained in internal control pellets, the internal control pellets being adapted to dissolve upon contact with a fluid sample and / or to disintegrate upon agitation of the plurality of magnetic beads in the lysis chamber, and wherein the internal control pellets are arranged in the lysis chamber prior to (e).

[0494] T23. The method according to any one of embodiments T18 to T22, wherein the internal control is a whole organism, a plasmid, or a nucleic acid transcript.

[0495] Some implementation plans cover:

[0496] U1. A method for performing cell lysis on a fluid sample, the method comprising: (a) dispensing the fluid sample into a lysis chamber, wherein the lysis chamber includes sidewalls, a first porous membrane, a second porous membrane spaced apart from the first porous membrane, a plurality of nonmagnetic beads, and at least one magnetic element, the plurality of nonmagnetic beads being contained within a space between the first porous membrane and the second porous membrane, the at least one magnetic element being contained within the space, wherein the pore sizes of the first porous membrane and the second porous membrane are designed to retain the plurality of nonmagnetic beads and the at least one magnetic element within the space, and wherein the fluid sample is dispensed into the lysis chamber through the first porous membrane; and (b) subjecting the at least one magnetic element to a changing magnetic field of varying polarity to move the magnetic element as it seeks to realign with the changing polarity of the changing magnetic field, wherein the movement of the magnetic element imparts motion to the plurality of nonmagnetic beads to achieve mechanical lysis (“cell lysis”) of cells present in the fluid sample contained within the lysis chamber.

[0497] U2. The method according to implementation scheme U1, wherein the changing magnetic field is generated by an electromagnet.

[0498] U3. The method according to implementation scheme U2, wherein (b) includes alternating the current to the electromagnet to alter the polarity of the electromagnet.

[0499] U4. The method according to embodiment U3, wherein (b) includes alternating the current at a frequency of 20 Hz to 200 Hz to alter the polarity of the electromagnet.

[0500] U5. The method according to embodiment U4, wherein (b) includes pulses of the current at two or more different frequencies to alter the polarity of the electromagnet.

[0501] U6. The method according to any one of embodiments U1 to U5, further comprising: (c) after (b), conveying at least a portion of the fluid sample from the pyrolysis chamber to a processing chamber, the processing chamber being fluidly connected to or connectable to the pyrolysis chamber, wherein the portion of the fluid sample conveyed from the pyrolysis chamber passes through the second porous membrane.

[0502] U7. The method according to embodiment U6 further includes: (d) during (c), retaining the lysed cell material in the lysis chamber while allowing the released nucleic acid to pass through the second porous membrane.

[0503] U8. The method according to embodiment U7 further comprises: (e) during (d), subjecting the at least one magnetic element to a changing magnetic field of varying polarity to move the magnetic element in the lysis chamber, the movement of the at least one magnetic element in the lysis chamber imparting movement to the plurality of non-magnetic beads, and the movement of the plurality of non-magnetic beads in the lysis chamber of the lysis sac causing at least a portion of the lysed cellular material in the lysis chamber to remain suspended at least until the fluid sample has been removed from the lysis chamber.

[0504] U9. The method according to embodiment U7 or U8 further includes: (f) immobilizing at least a portion of the released nucleic acid on a solid support in the processing chamber, and removing the unimmobilized components of the fluid sample from the processing chamber.

[0505] U10. The method according to embodiment U9 further includes: (g) after (h), eluting the immobilized nucleic acid from the solid support and delivering the eluted nucleic acid to the reaction chamber of the fluid cartridge.

[0506] U12. The method according to embodiment U11 further comprises: (i) releasing an internal control into the fluid sample during or after (a); (j) immobilizing a nucleic acid (“IC nucleic acid”) associated with the internal control on the solid support during (f); (k) eluting the IC nucleic acid from the solid support and delivering the eluted IC nucleic acid to the reaction chamber after (f); and (l) subjecting the IC nucleic acid to the conditions of a second reaction after (k), the results of (l) being used to verify the results of (h) and / or to verify the effectiveness of the cell lysis in (b).

[0507] U13. The method according to embodiment U12, wherein at least a portion of the internal control is contained in an internal control reagent, and when (a) is initiated, the internal control reagent is disposed on at least one of the first porous membrane and the second porous membrane, and wherein during either (a) or (c), the internal control reagent is dissolved in the fluid sample.

[0508] U14. The method according to embodiment U12, wherein at least a portion of the internal control is contained in an internal control reagent, and when (a) is activated, the internal control reagent is disposed on at least a portion of the plurality of non-magnetic beads and / or the at least one magnetic element, and wherein during either (a) or (b), the internal control reagent disposed on at least a portion of the plurality of non-magnetic beads and / or the at least one magnetic element dissolves in the fluid sample.

[0509] U15. The method according to embodiment U12, wherein at least a portion of the internal control is contained in an internal control reagent, the internal control reagent being disposed on the sidewall of the lysis chamber when (a) is initiated, and wherein the internal control reagent dissolves in the fluid sample during either (a) or (b).

[0510] U16. The method according to embodiment U12, wherein at least a portion of the internal control is embedded in or contained within an internal control pellet, and wherein the internal control pellet dissolves in the presence of the fluid sample and / or breaks down during (c) by the movement of the plurality of nonmagnetic beads, thereby releasing the internal control into the fluid sample.

[0511] U17. The method according to any one of embodiments U12 to U16, wherein the conditions for the first reaction and the conditions for the second reaction are the same.

[0512] U18. The method according to any one of embodiments U12 to U17, wherein each of the first reaction and the second reaction is a nucleic acid amplification reaction.

[0513] U19. The method according to implementation plan U18, wherein the nucleic acid amplification reaction is a polymerase chain reaction (“PCR”).

[0514] Some implementation plans cover: continuing

[0515] V1. A pellet containing an internal control for use in a nucleic acid amplification reaction, wherein the internal control is provided to verify assay results and / or to verify the effectiveness of a cell lysis protocol, and wherein the pellet is adapted to be destroyed upon exposure to a collision force required for cell rupture, thereby releasing the internal control.

[0516] V2. The pellet according to embodiment V1, wherein the internal control pellet comprises: a core, the core comprising an excipient in which the internal control is embedded; and a coating surrounding the core and adapted to be broken by a mechanical pyrolysis shear force applied by movement of the plurality of nonmagnetic beads within the receiver, wherein the excipient is adapted to at least partially dissolve upon exposure to a fluid after the coating is broken.

[0517] V3. The pellets according to embodiment V2, wherein the excipient comprises at least one of microcrystalline cellulose and hydroxypropyl cellulose, and wherein the coating comprises a cellulose derivative.

[0518] V4. A pellet according to any one of embodiments V1 to V3, wherein the internal control is a whole organism, a plasmid, or a nucleic acid transcript.

[0519] Some implementation plans cover:

[0520] W1. An apparatus for performing cell lysis thereunder, the apparatus comprising: a fluid acceptor; a plurality of nonmagnetic beads and at least one magnetic element within the acceptor; and an internal control disposed within a pellet located within the acceptor, wherein the pellet is adapted to be destroyed upon being subjected to a force exerted by movement of the plurality of nonmagnetic beads within the acceptor, thereby releasing the internal control from the pellet, and wherein the internal control is provided to verify assay results and / or to verify the effectiveness of the cell lysis procedure within the acceptor.

[0521] W2. The apparatus according to claim W1, comprising at least one porous membrane disposed in a flow path through the receiver.

[0522] X3. The apparatus according to embodiment X1 or X2, wherein the internal control pellet comprises: a core, the core comprising an excipient, the internal control being embedded within the excipient; and a coating surrounding the core and adapted to be broken by a mechanical cleavage shear force applied by movement of the plurality of nonmagnetic beads within the receiver, wherein the excipient is adapted to at least partially dissolve upon exposure to a fluid after the coating is broken.

[0523] W4. The apparatus according to embodiment W3, wherein the excipient comprises at least one of microcrystalline cellulose and hydroxypropyl cellulose, and wherein the coating comprises a cellulose derivative.

[0524] W5. The apparatus according to any one of embodiments W1 to W4, wherein the internal control is a whole organism, a plasmid, or a nucleic acid transcript.

[0525] Other features and characteristics of the subject matter of this disclosure, as well as the methods of operation, the functions and combinations of related elements and components of the structure, and the economics of manufacture, will become more apparent when considering the following description with reference to the accompanying drawings, all of which form part of this specification, wherein like reference numerals designate corresponding portions in the various drawings. Attached Figure Description

[0526] The accompanying drawings, which are incorporated herein and form part of the specification, illustrate various embodiments of the subject matter of this disclosure. In the drawings, the same reference numerals indicate the same or similarly functional elements.

[0527] Figure 1 This is a rear perspective view of the instrument used to process fluid cartridges as described in this article.

[0528] Figure 2 This is a frontal perspective view of the instrument.

[0529] Figure 3 This is an exploded top perspective view of an example of a fluid cell processed in the instrument described in this article.

[0530] Figure 4 This is a top plan view of the fluid box body.

[0531] Figure 5 This is a bottom plan view of the box body.

[0532] Figure 6 This is a top perspective view of the box itself.

[0533] Figure 7 This is a bottom perspective view of the box body.

[0534] Figure 8 This is a schematic cross-section of the fluid cell through the reaction / detection chamber of the fluid cell.

[0535] Figure 9 This is a bottom perspective view of the protective vent of the fluid chamber.

[0536] Figure 10 and Figure 10 "Detail A" shows the protective ventilator along... Figure 9 The cross section of line AA in the diagram.

[0537] Figure 11 This is an exploded top perspective view of the fluid cartridge's stopper, stopper ring, and syringe stopper.

[0538] Figure 12This is the top perspective view of the blocking component.

[0539] Figure 13 This is the top view of the blocking component.

[0540] Figure 14 This is the bottom view of the blocking component.

[0541] Figure 15 This is a top perspective view of the sample chamber cap of the fluid cartridge.

[0542] Figure 16 This is a side view of the sample chamber cap.

[0543] Figure 17 It is along the sample chamber cap Figure 15 The cross section of line AA in the diagram.

[0544] Figure 18 It is along the fluid box Figure 4 A cross-section of a portion of line BB in the diagram shows the body of the box.

[0545] Figure 19 It is along the fluid box Figure 4 A cross-section of a portion of line AA in the diagram shows the body of the box.

[0546] Figure 20 This is a perspective view of the syringe driver of the instrument.

[0547] Figure 21 It is a longitudinal section of a portion of the fluid cartridge, which passes through the sample chamber and syringe barrel of the cartridge body and through a portion of the instrument, and shows the syringe plunger, the contact detector plunger, the pressure plate, and the upper block of the instrument in an elevated position relative to the cartridge.

[0548] Figure 22 It is a longitudinal section of a portion of the fluid cartridge, which passes through the sample chamber and syringe barrel of the cartridge body and through a portion of the instrument, and shows the syringe plunger, the plunger of the contact detector, the pressure plate, and the upper block of the instrument in a lowered position engaged with the cartridge.

[0549] Figure 23 It is a graph showing the motor current requirements and stop travel for four different fluid boxes.

[0550] Figure 24 The flowchart illustrates a method for controlling the position of a syringe stop using the motor requirements of a syringe drive module and the output of an encoder connected to the motor, and thus the volume of fluid drawn into the syringe body of the fluid cartridge.

[0551] Figure 25This is a partial top perspective view showing the instrument's box support frame.

[0552] Figure 26 This is a partial top perspective view showing the box support frame that supports the fluid box.

[0553] Figure 27 It is a schematic cross-section passing through the first and second thermal modules of the instrument and through the reaction / detection chamber of the box, wherein the first thermal module is in an elevated position relative to the second thermal module and the box.

[0554] Figure 28 It is a schematic cross-section passing through the first and second thermal modules of the instrument and through the reaction / detection chamber of the box, wherein the first thermal module is in a lowered position relative to the second thermal module and the box.

[0555] Figure 29 This is a top perspective view of the instrument's upper frame.

[0556] Figure 30 This is a side view of the upper rack.

[0557] Figure 31 This is a bottom perspective view of the upper rack.

[0558] Figure 32 This is a partial perspective view of the top of the instrument, showing the first (top) thermal module and the second (bottom) thermal module.

[0559] Figure 33 This is a top partial perspective view of the first (top) thermal module and the second (bottom) thermal module.

[0560] Figure 34 This is a partial bottom perspective view of the first (top) thermal module and the second (bottom) thermal module.

[0561] Figure 35 This is a top perspective view of the first (top) thermal module.

[0562] Figure 36 This is a bottom perspective view of the first (top) thermal module.

[0563] Figure 37 It is through the first (top) thermal module Figure 35 The cross-sectional view of line AA in the diagram.

[0564] Figure 38 This is a perspective view of the first thermal module, showing the first thermal component of the first thermal module in an exploded view.

[0565] Figure 39 This is an exploded perspective view of the second thermal component of the second thermal module.

[0566] Figure 40 This is a front view of the first and second thermal components of the second thermal module.

[0567] Figure 41 This is a left-side view of the second thermal component of the second thermal module.

[0568] Figure 42 This is a right-side view of the first thermal component of the second thermal module.

[0569] Figure 43 This is a top perspective view of the second thermal component of the second thermal module.

[0570] Figure 44 This is a partial top left perspective view of the instrument's contact detector when the first thermal module is in the raised position relative to the box.

[0571] Figure 45 This is a partial top left perspective view of the instrument's contact detector when the first thermal module is in a lowered position relative to the box and the contact detector is in contact with the box.

[0572] Figure 46 This is a partial top perspective view of the contact detector of the instrument.

[0573] Figure 47 A flowchart illustrating an implementation scheme for performing molecular determinations using the instruments and fluidic chambers described herein is shown.

[0574] Figure 48 This is a curve showing the temperature distribution of the thermal cycler as described in this article.

[0575] Figure 49 This is a perspective view of a first embodiment of a rupture capsule for use in a fluid cartridge.

[0576] Figure 50 It is along the ruptured sac-like structure Figure 49 The cross section of line AA in the diagram.

[0577] Figure 51 It is a cross-section of a second embodiment of a lysis sac located within the sample chamber of a fluid cell, wherein the sample chamber is closed by a cap.

[0578] Figure 52 It is a cross-section of a third embodiment of a lysis sac located within the sample chamber of a fluid cell, wherein the sample chamber is closed by a cap.

[0579] Figure 53 This is a perspective view of the implementation scheme of the pyrolysis vessel.

[0580] Figure 54 It is along the pyrolysis vessel Figure 53 The cross section of line AA in the diagram.

[0581] Figure 55 It is a cross-section of the pyrolysis vessel located in the sample chamber of the fluid cell.

[0582] Figure 56 It is a perspective view of a fluid box supported in a box holder having a magnet housing that houses a variable magnet (e.g., an electromagnet) within it.

[0583] Figure 57 This is a schematic diagram showing an electromagnet inside a magnet housing and connected to an oscillating circuit configured to repeatedly change the polarity of the electromagnet.

[0584] Figure 58 It is an exploded perspective view of the electromagnet assembly.

[0585] Figure 59 This is a side view of the electromagnet assembly.

[0586] Figure 60 It is along the electromagnet assembly Figure 59 The cross section of line AA in the diagram.

[0587] Figure 61 This is a flowchart of a method for performing cell lysis and reactions using instruments and a fluidic chamber including a lysis capsule.

[0588] Figure 62 This is a flowchart of a method for using an internal control in a reaction using instruments and fluid cartridges.

[0589] Figure 63 This is a flowchart of an alternative method for introducing fluid samples into the lysis chamber of a lysis capsule in a fluid cell.

[0590] Figure 64 This is a flowchart of a method for manufacturing a fluid cartridge containing a rupture capsule.

[0591] Figure 65 It is a partial top perspective view of a fluid cell with a chamber expander and the cap of the chamber expander being in the open position.

[0592] Figure 66 It is along the fluid box and chamber expander Figure 65 A partial cross-sectional view of line XX in the diagram.

[0593] Figure 67 It is a fluid box along which includes a rupture capsule and a chamber expander. Figure 65 The exploded section view of line XX in the diagram.

[0594] Figure 68 It is along the fluid box, rupture capsule, and chamber expander Figure 65 The cross-sectional view of line YY in the diagram.

[0595] Figure 69 This is a top perspective view of a chamber expander that can be attached to a fluid cartridge, with the cap of the chamber expander in the open position.

[0596] Figure 70 This is a bottom perspective view of the chamber expander, with the cover of the chamber expander in the open position.

[0597] Figure 71 This is a top plan view of the chamber expander, with the cover of the chamber expander in the open position.

[0598] Figure 72 This is a bottom plan view of the chamber expander, with the cover of the chamber expander in the open position.

[0599] Figure 73 It is along the cap of the chamber expander Figure 69 The top perspective partial section view of line ZZ in the middle.

[0600] Figure 74 It is used in Figure 65 Top perspective view of the hollow body of the rupture capsule used in the fluid box.

[0601] Figure 75 It is a top perspective view of a bead delivery cap having a ruptureable chamber containing a pyrolysis agent (i.e., magnetic elements and non-magnetic pyrolysis beads).

[0602] Figure 76 This is a bottom view of the beaded delivery cap.

[0603] Figure 77 It is a beaded delivery cap with an intact beaded component containing a chamber along the beaded component. Figure 75 The cross-sectional view of line AA in the diagram.

[0604] Figure 78 It is the edge of the beaded delivery cap. Figure 75 A cross-sectional view of line AA in the diagram, where the beaded elements contain chambers that rupture to release the pyrolyte.

[0605] Figure 79 This is a partial cross-section of the sample chamber through the box body, showing the beaded delivery cap inserted into the sample chamber.

[0606] Figure 80 It is a partial cross-section of a fluid cartridge, an instrument pressure plate, and a contact detector with a bead delivery cap actuator, wherein the pressure plate and the contact detector are in an elevated position relative to the fluid cartridge, such that the bead delivery cap actuator does not engage with the bead delivery cap.

[0607] Figure 81It is a partial cross-section of a fluid cartridge, a pressure plate, and a contact detector having a bead delivery cap actuator, wherein the pressure plate and the contact detector are in a lowered position relative to the fluid cartridge, such that the bead delivery cap actuator engages with the bead delivery cap.

[0608] Figure 82 It is a partial cross-section of the fluid cell passing through the mechanical pyrolysis sample chamber, which provides the pyrolysis chamber and the pyrolysis agent for performing mechanical pyrolysis within the sample chamber.

[0609] Figure 83 It is a local cross section of the fluid cell passing through the alternative mechanical pyrolysis sample chamber.

[0610] Figure 84 It is a local cross section of the fluid cell passing through the alternative mechanical pyrolysis sample chamber.

[0611] Figure 85 This is a schematic cross-sectional view of a coated microsphere containing an internal reference material (“IC microsphere”).

[0612] Figure 86 This is a flowchart of a method for manufacturing a pyrolysis chamber within the sample chamber of a fluid cell. Detailed Implementation

[0613] While the subject matter of this disclosure may be embodied in various forms, the following description and figures are intended only to disclose some of these forms as specific examples of the subject matter. Accordingly, the subject matter of this disclosure is not intended to be limited to the forms or embodiments described and shown. definition

[0614] Unless otherwise defined, all field terms, indexes, and other technical terms or terminology used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this disclosure pertains. All patents, applications, published applications, and other publications mentioned herein are incorporated herein by reference in their entirety. If any definition set forth in this section is contrary to or otherwise inconsistent with the definitions set forth in the patents, applications, published applications, and other publications incorporated herein by reference, the definitions set forth in this section shall prevail over those incorporated herein by reference.

[0615] Unless otherwise stated or implied by the context, as used herein, “a” or “an” means “at least one” or “one or more”.

[0616] References to "an implementation," "implementation," "further implementation," "example," "some aspects," "further aspects," "various aspects," etc., in the specification indicate that the described implementation, example, or aspect may include a particular feature, structure, or characteristic; however, each implementation covered by this disclosure may not necessarily include that particular feature, structure, or characteristic. Furthermore, such phrases do not necessarily refer to the same implementation, example, or aspect. Moreover, when a particular feature, structure, or characteristic is described in connection with an implementation, whether explicitly stated or not, such feature, structure, or characteristic is also a description in connection with other implementations, examples, or aspects.

[0617] This specification may use various terms describing relative spatial arrangement and / or orientation or direction when describing the position and / or orientation or direction of movement, force or other dynamic action of components, devices, locations, features, or a portion thereof. Unless specifically stated or otherwise indicated by the context of the specification, such terms, including but not limited to top, bottom, above, below, under, top of, upper, lower, left, right, front, back, below, beside, adjacent, between, horizontal, vertical, diagonal, longitudinal, transverse, radial, axial, clockwise, counterclockwise, etc., are used to conveniently refer to such components, devices, locations, features, or a portion thereof or movement, force or other dynamic action as shown in the drawings, and are not intended to be limiting.

[0618] Unless otherwise stated or implied by the context, the terms used herein to describe the physical and / or spatial relationship between a first component, structure or part thereof and a second component, structure or part thereof (such as attachment, connection, fixation, joining, linking, coupling or similar terms or variations thereof) shall cover the direct relationship of direct contact between the first component, structure or part thereof and the second component, structure or part thereof, or the presence of one or more intermediate components, structures or parts thereof between the first component, structure or part thereof and the second component, structure or part thereof.

[0619] Unless otherwise stated, any specific dimensions mentioned in this specification are merely examples of implementations of apparatus embodying various aspects of this disclosure and are not intended to be limiting.

[0620] Within the scope of use herein, whether explicitly indicated or not, the terms “about” or “approximately” apply to all numerical values ​​and terms indicating specific physical orientations or relationships such as horizontal, vertical, parallel, perpendicular, concentric, or similar terms specified herein. The term generally refers to a series of numbers, orientations, and relationships that, in the context of this disclosure, would be considered by one of ordinary skill in the art to be a reasonable amount of deviation from the enumerated numerical values, orientations, and relationships (i.e., having an equivalent function or result). For example, and not intended to be restrictive, the term may be interpreted to include a deviation of ±10% from a given numerical value, orientation, or relationship, provided that such deviation does not alter the final function or result of said value, orientation, or relationship. Thus, in some cases, as would be understood by one of ordinary skill in the art, a value of about 1% or approximately 1% may be interpreted as a range from 0.9% to 1.1%.

[0621] Within the scope of this document, the term "proximity" means close (spatial proximity) or adjacent. Proximity objects or portions thereof may be spaced apart from each other, or may be in actual or direct contact with each other. In some instances, proximity objects or portions thereof may be connected to each other or may be integrally formed with each other.

[0622] Within the scope of use herein, the terms “substantially” and “basically” refer to a considerable degree or extent. When used in conjunction with, for example, an event, situation, characteristic, or property, the terms may refer to instances in which the event, situation, characteristic, or property occurs precisely as described herein, and instances in which the event, situation, characteristic, or property is close to approximately occurring, taking into account typical tolerance levels or variability of the embodiments described herein.

[0623] Within the scope of use herein, the terms “optional” and “optionally” or the term “may” (e.g., in phrases such as “may include,” “may include,” “may produce,” “may provide,” or similar phrases) mean that the components, structures, elements, events, situations, characteristics, properties, etc., subsequently described may be included or present, or may not be included or present, and describes instances of components, structures, elements, events, situations, characteristics, properties included or present, as well as instances of components, structures, elements, events, situations, characteristics, properties, etc., not included or present therein.

[0624] Within the scope of this document, the terms “first,” “second,” and similar terms preceding the names of elements (e.g., components, devices, positions, features, or a portion thereof, or the direction of movement, force, or other dynamic action) are used for identification purposes to distinguish between similar elements and are not intended to necessarily imply order or hierarchy. Furthermore, the terms “first” and “second” are not intended to exclude the inclusion of additional similar elements. Moreover, unless the context otherwise requires, the use of the term “first” preceding the name of an element (e.g., components, devices, positions, features, or a portion thereof, or the direction of movement, force, or other dynamic action) does not necessarily imply or require the presence of additional (e.g., “second,” “third,” etc.) such elements.

[0625] Within the scope of this document, the terms or phrases “configured to,” “adapted to,” “operable as,” “constructed and arranged as,” and similar terms mean that the subject of the terms or phrases includes, constitutes, or otherwise covers the necessary (multiple) structures, (multiple) mechanisms, (multiple) arrangements, (multiple) components, (multiple) materials, (multiple) algorithms, (multiple) circuits, programming, etc., to automatically or permanently, or selectively when required to do so, perform one or more specified tasks or achieve specified outputs or characteristics.

[0626] Within the scope of this document, the term "amplification reaction" or "nucleic acid amplification reaction" refers to a procedure used to produce multiple copies of a specific segment of a nucleic acid. Amplification reactions can be isothermal or require repeated cycles between different temperatures, such as those required by polymerase chain reaction (PCR).

[0627] The term "amplification reagent" refers to materials containing one or more components required for an amplification reaction. In nucleic acid amplification, such components may include primers, nucleotide triphosphates, and / or coenzyme factors (e.g., divalent cations such as Mg++) required for the amplification of target nucleic acids.

[0628] Within the scope of this document, the term "analyte" refers to a molecule or substance that is detected or analyzed in an assay. Examples of analytes include nucleic acids, proteins (e.g., antibodies, peptides, and prions), and antigens.

[0629] Within the scope of this document, the term "determination" refers to a procedure for detecting and / or quantifying an analyte in a sample. This involves contacting a sample containing or suspected of containing an analyte with one or more reagents and subjecting it to conditions that allow the generation of a detectable signal, which provides useful information about the presence of the analyte in the sample or the amount of the analyte (e.g., mass or concentration).

[0630] Within the scope of this document, the term "analyst" refers to an automated instrument capable of performing one or more steps of determination, including the step of determining the presence or absence of one or more analytes suspected of being present in a fluid sample.

[0631] Within the scope of this document, the term "molecular assay" refers to a procedure for specifically detecting and / or quantifying a target molecule, such as a specific nucleic acid. A sample containing or suspected of containing a target molecule is contacted with one or more reagents comprising at least one reagent specific to said target molecule and subjected to conditions that allow the generation of a detectable signal providing useful information about the presence of the target molecule. For example, in cases where the molecular assay includes an amplification reaction (such as polymerase chain reaction (PCR)), the reagents include primers that can be specifically targeted at the target nucleic acid, and the generation of a detectable signal can be at least partially achieved by providing a labeled probe (e.g., a fluorescently labeled probe) that hybridizes in a target-specific manner with an amplicon generated by the primers in the presence of the target. Alternatively, the reagents may include an intercalation dye (e.g., SYBR® Green) for detecting the formation of double-stranded nucleic acids.

[0632] Within the scope of this article, the term "point-of-care testing" (POCT), sometimes referred to as near-patient testing, refers to testing performed at a site close to the patient's care or treatment. This can be in a hospital, a doctor's office, or a field testing setting. Unlike high-throughput systems, POCT systems are generally small and easily portable. Most POCT systems are capable of running measurements on a single sample or a limited number of samples simultaneously.

[0633] Within the scope of this document, the term "reagent" refers to any substance or mixture involved in the assay, other than the sample material and the product being measured. Examples of reagents used in molecular assays may include nucleotides, enzymes, primers, probes, and salts.

[0634] Within the scope of use herein, the term "receiver" or "fluid receiver" refers to any type of fluid container configured to contain a sample or another fluid (collectively referred to herein as "fluid"), including, for example, tubes, vials, cuvettes, pools or boxes, or other articles having one or more pools or chambers formed therein or attached thereto, microtiter plates, etc. Tubes may be cylindrical (i.e., with a circular cross-section) or non-cylindrical and may have flattened or rounded closed ends. A non-limiting example of such a receiver is the Aptima® Multiplex Detection Swab Collection Kit (Hologic, Inc., Marlborough, Massachusetts).

[0635] Within the scope of this document, the term "sample" means any substance suspected of containing at least one analyte of interest. Analytes of interest can be, for example, nucleic acids, proteins, chemicals, etc. Substances can be derived from any source, including animals, industrial processes, the environment, water sources, food products, or solid surfaces (e.g., surfaces in medical facilities). Substances obtained from animals can include, for example, blood or blood products, urine, mucus, sputum, saliva, semen, tears, pus, feces, nasopharyngeal or genitourinary specimens obtained by swabs or other collection devices, and other bodily fluids or materials. The term "sample" will be understood to mean a specimen in its natural form or at any stage of processing.

[0636] Within the scope of this document, the terms “thermal contact” or “thermal connectivity” mean the ability to allow the transfer of thermal energy between two systems or bodies at different temperatures. The two systems or bodies may be in direct physical contact, allowing thermal energy transfer to occur directly from one system or body to the other, or an intermediate material comprising air may be arranged between the two systems or bodies, allowing thermal energy transfer to occur through said intermediate material.

[0637] Within the scope of this document, the term "unit dosage form" means an amount sufficient to perform a single assay. That is, in contrast to bulk reagents provided in amounts that can be used to perform multiple assays, a "unit dose" or "unitized" reagent is an amount of reagent that can be used for a single assay (which may be designed to determine the presence of one or more analytes).

[0638] A "fluid cartridge" is an apparatus comprising a fluid network of two or more chambers or pools containing fluids, the fluids being fluidly interconnected or fluidly interconnectable via one or more fluid channels. The apparatus is configured to interface with a processing instrument or analyzer for handling the fluid cartridge. A fluid cartridge may include one or more of the following: a sample chamber for receiving and introducing a fluid sample into the fluid cartridge; multiple storage chambers containing one or more materials (such as reagents, buffers, or probes); multiple processing chambers in which one or more treatments (such as combination / mixing, filtration, purification, etc.) are performed on the fluid materials; multiple waste chambers in which one or more waste materials are stored; and multiple reaction chambers in which a chemical or biochemical reaction occurs.

[0639] Within the scope of this document, "processing" a fluid cartridge means performing one or more processes on a fluid or other material contained within the cartridge, including, for example, one or more of the following: applying positive or negative pressure to the cartridge; applying physical pressure to at least one chamber of the cartridge to cause the chamber to collapse at least partially; or actuating a pump mechanism operatively coupled to the cartridge to enable fluid movement between chambers within the cartridge's fluid network; actuating or otherwise altering a flow control mechanism (such as a valve) to change the flow control mechanism between an open state that allows fluid to flow through the flow control mechanism and a closed state that prevents fluid from flowing through the flow control mechanism; combining two or more materials within the cartridge chambers; filtering or otherwise purifying a fluid sample within the cartridge; heating and / or cooling the fluid in one or more chambers of the cartridge; and detecting and recording signals based on optical emission from the fluid contained in one or more chambers of the cartridge.

[0640] Within the scope of this document, "internal control" refers to a molecule detected to validate assay results (such as a negative assay result in which the analyte is not detected). Amplification reactions (e.g., PCR) can be affected by, for example, the presence of inhibitors in the sample (e.g., hemoglobin), errors in the sample extraction process, or malfunctions in the thermal cycler. In the case of amplification reactions, internal controls are used to demonstrate that reagents and conditions should allow the target analyte (if present in the sample) to be successfully amplified and detected during the assay. In amplification reactions where the target analyte is a nucleic acid, internal controls typically have at least a partially different sequence from the target analyte, but may have properties that result in similar amplification and detection characteristics (e.g., similar GC content). Nucleic acid internal controls can be amplified using dedicated amplification primers or using the same amplification primers as the target analyte. Internal control nucleic acids may lack the sequence targeted by a probe against the target analyte and may contain the sequence targeted by a probe against a probe specific to the internal control nucleic acid. Internal nucleic acid controls can be in the form of nucleic acid transcripts, or they can be nucleic acids contained within plasmids or cells (such as yeast). In this case, in addition to carrying internal nucleic acid controls, cells can also be used as “extraction controls” to monitor the effectiveness of lysis procedures for releasing nucleic acids from target microorganisms and other extraction procedures (such as filtration, target capture, purification, etc.).

[0641] Within the scope of this document, the term "porous membrane" refers to a selective barrier that controls the passage of substances—depending on the size of the substances, allowing some substances to pass through the barrier while preventing others from passing through. A porous membrane can be, for example, a woven mesh, a thin porous substrate, or a filter matrix (e.g., spun or sintered); however, unless specifically indicated for a particular example or application, the term "porous membrane" is not intended to imply, and should not be construed as implying, a particular composition, structure, shape factor, or thickness. Detailed description of the attached figures

[0642] Instrument Overview

[0643] Figure 1 and Figure 2 The internal components of an instrument 10 as described herein are shown. This instrument receives and operates a test platform (such as a fluid cartridge (i.e., a device configured to be placed within and interfaced with a processing instrument and including reagent and sample storage and fluid disposal components such as fluid flow channels and flow control valves)) to process samples on or within the test platform (e.g., to perform assays (such as molecular assays) and collect data on the results of the assays). Instrument 10 includes components for applying thermal energy to one or more reaction / detection regions of the test platform, components for transmitting optical signals to and / or from the reaction / detection regions, and components for actuating a syringe pump within the test platform. Figure 1 This is the rear perspective view of instrument 10, and Figure 2 This is a front perspective view of instrument 10. Instrument 10 can be an on-the-fly test system for providing sample-to-result testing using a disposable fluid cartridge, the disposable fluid cartridge comprising interconnected chambers (or pools) and a reaction chamber that can be pre-packaged in unit doses containing all the reagents required to perform the desired test. The fluid cartridge can be a closed system that minimizes the chance of contamination.

[0644] Typically, such an instrument will include a housing, within which the internal components will be enclosed, but from Figure 1 The casing is omitted, allowing the internal components to be seen.

[0645] like Figure 1 and Figure 2 As shown, the test platform (e.g., fluid chamber 500) is located within instrument 10, and the internal components of the instrument can generally be divided into a first frame or upper frame 300 (referring to those internal components located above the chamber 500) and a second frame or lower frame 400 (referring to those internal components located below the fluid chamber 500 and supported thereon). The fluid chamber 500 can be a microfluidic chamber, meaning that at least a portion of any fluid pathways, channels, chambers, pools, reaction chambers, etc., in which fluid flows and / or is retained is geometrically constrained to a small scale (e.g., submillimeter), at which the surface forces acting on the fluid satisfy or exceed volume forces. The upper frame 300 may include a syringe actuator 360 configured to actuate a syringe plunger coupled to a syringe stop within the chamber 500, as will be described herein.

[0646] Fluid box

[0647] exist Figures 3 to 19 An embodiment of the fluid cartridge 500 and its components is shown in the figure. Figure 3 An exploded top perspective view of cartridge 500 is shown. The fluid cartridge 500 includes a cartridge body 502, a first (e.g., top) membrane 512, a second (e.g., bottom) membrane 530, an elastomeric syringe stop 540, a stop ring 550, a stop 570, a filter 538, a purification column insert 536 that can be in disc form for positioning and holding the purification column (e.g., a silica column, GF51 grade Hahnemühle Life Science (product number GF51RL01550) from Dasser, Germany), a cap 516, and a protective vent 560. For convenience and consistency with the example shown in the figures, membrane 512 will be referred to herein as the top membrane, and membrane 530 as the bottom membrane. A syringe driver 360 is coupled to instrument 10 (see description below). Figure 20The plunger 362 includes a plunger head 364 and a plunger rib 366, the plunger head being received within a recess formed in a stop 540, the plunger rib engaging a stop 570 as described below. The cartridge body 502 of the fluid cartridge 500 includes a plurality of chambers or pools W1 to W12 and SB, the plurality of chambers or pools including: (i) sample chambers and storage chambers containing or configured to receive materials (e.g., sample materials, reagents, buffer solutions, etc.) used in performing assays (e.g., molecular assays); (ii) chambers or pools in which two or more materials can be combined and mixed; (iii) chambers or pools for receiving and retaining waste materials; and (iv) reaction / detection chambers 510a1, 510a2, 510b1, 510b2 (i.e., detection areas), in which reactions can occur and / or detectable signals emitted by reactions within the chambers can be detected. In the context of this disclosure, although the terms “pool” and “chamber” may be used interchangeably in some descriptions, generally, the term “pool” refers to, but is not limited to, an open reservoir or recess (such as pools W1 to W12 and SB) formed in the housing body 502, and the term “chamber” refers to, but is not limited to, a pool in the housing body 502 that is at least partially closed, for example by a first membrane 512, a second membrane 530, and / or a vent 560, to form a at least partially closed compartment or space. More than one of the functions of containing, combining, reacting, and detecting may occur in one or more functional chambers of the housing 500. As described below, the functional chambers within the housing may be fluidly interconnected by fluid channels or conduits, and the housing includes one or more fluid flow control valves that may be selectively actuated, for example, by a valve actuator (not shown) of the instrument 10, to controllably permit or prevent fluid flow within the fluid channels operatively associated with the valves. The example shown has four reaction / detection chambers 510a1, 510a2, 510b1, and 510b2, which are arranged in two pairs (or sets or groups) 510a1, 510a2 and 510b1, 510b2. In other examples, the box has fewer or more than four reaction / detection chambers. For example, the box may have one or more sets or three aggregated reaction / detection chamber sets.

[0648] The box body 502 has a first (e.g., top) surface 501 ( Figure 4 ) and the second (e.g., bottom) surface 503 ( Figure 5For convenience and consistency with the examples shown in the accompanying drawings, surface 501 will be referred to herein as the top surface, and surface 503 will be referred to herein as the bottom surface. The box body 502 may be injection molded from a thermoplastic polymer material, and preferably from polypropylene (PP), such as cyclic olefin copolymers (COC) or cyclic olefin polymers (COP), including polycarbonate, polyacrylamide, polyethylene, polymethyl methacrylate (PMMA), polydimethylsiloxane (PDMS), and polyvinyl chloride (PVC). In some embodiments, the box body 502 is formed by stereolithography or by sintering. The box body 502 may be made of an opaque material.

[0649] like Figure 4 (Top plan view of box body 502) and Figure 5 As shown in the bottom plan view of the housing body 502, the housing body 502 includes a plurality of through holes H1 to H32 extending between the top surface 501 and the bottom surface 503 to fluidly connect components from one surface to the other. To avoid confusion in the drawings, the through holes H1 to H32 are only... Figure 4 The box body 502 includes a plurality of bottom recesses G1 to G20 formed in the bottom surface 503 and a plurality of top recesses G21 to G32 formed in the top surface 501. Each of the recesses G1 to G32 may have a depth between 0.01 mm and 0.5 mm, preferably between 0.2 mm and 0.4 mm, and most preferably about 0.3 mm, and may have a width of about 0.5 mm. Each of the through holes H1 to H18 is associated with a corresponding valve V1 to V18, the corresponding valve comprising a cylindrical recess formed in the bottom surface 503 of the box body 502 and the cylindrical recess is generally coaxially arranged relative to the associated through hole and has a diameter larger than that of the associated through hole. In a non-limiting example, the recess associated with each of valves V1 to V18 may have a diameter between 1 mm and 10 mm, preferably between 2 mm and 8 mm, preferably about 4 mm, and a depth between 0.02 mm and 0.4 mm, preferably between 0.05 mm and 0.15 mm, and most preferably about 0.1 mm. One or both of the recesses G1 to G32 terminate at the associated valves V1 to V18. Through holes H1 to H10 are also associated with chambers W1 to W10, through holes H11 and H12 are associated with chamber W6, through hole H19 is associated with chamber W11, and through hole H20 is associated with chamber W12. Through holes H21 to H32 are not directly associated with valves or chambers and provide connection between the recess or other feature on the top surface 501 and the recess or other feature on the bottom surface 503. The cartridge body 502 also includes central through holes H1c to H10c arranged in a circular pattern within the pool SB (injection cartridge body).

[0650] The through-holes H1 to H32 and H1c to H10c, valves V1 to V18 and associated recesses formed in the housing body 502, as well as the bottom recesses G1 to G21 and top recesses G22 to G32, form a network of fluid channels and fluid control valves in these channels. For this purpose, it is necessary to close the through-holes, recesses, and grooves that are open to the top surface 501 or bottom surface 503 of the housing body 502. The bottom film 530 is fixed to the bottom surface 503 of the housing body 502 to cover the bottom recesses G1 to G20 to form corresponding channels (which may be microfluidic channels), to cover the recesses of valves V1 to V18 to form corresponding valves, to cover the central through-holes H1c to H10c, and the through-holes H19 to H32 flush with the bottom surface 503. The bottom film 530 may comprise a material similar to the housing body 502, including, for example, polypropylene (PP). The bottom film 530 may include a thermoplastic film having a thickness between 0.1 mm and 0.2 mm (100 µm to 200 µm), which is bonded or welded to the surface of the bottom surface 503 by thermal welding technology (e.g., by laser welding), bonding, adhesive or chemical bonding methods.

[0651] Valves V1 to V18 are formed of a bottom diaphragm 530, which may be deformable and extends across each opposing recess of an annular valve seat, the annular valve seat being defined respectively between the recess of each valve V1 to V18 and the associated through-hole H1 to H18 of the valve. A single valve seat 505 between the recess of valve V2 and the associated through-hole H2 is... Figure 7 The image is marked in the image. In a non-limiting example, the surface of the deformable bottom film 530, disposed opposite the recesses of valves V1 to V18, is approximately planar and parallel to the bottom surface 503 of the housing body 502 when undeformed, and is spaced apart from the valve seats between the recesses and the through-holes. The bottom film 530 is deformable by an external actuator (not shown) with a width greater than the width (diameter) of the through-hole but not greater than the width (diameter) of the recess forming the valve seat, and partially pushes the film into the recess. The deformed bottom film 530 contacts each valve seat of valves V1 to V18, blocking the associated through-holes H1 to H18, the diameter of which is smaller than the diameter of each associated recess, such that the film contacts the valve seat and seals the associated through-hole.

[0652] The top film 512 can be fixed to the top surface 501 of the housing body 502, for example, by thermal welding, adhesive bonding, or chemical bonding, to close the top grooves G21 to G32 flush with the top surface 501 in the same way that the bottom film 530 closes the bottom grooves G1 to G20 to form corresponding channels, thus forming corresponding channels (which may be microfluidic channels). The top film 512 can be made of a material similar to the housing body 502 (e.g., polypropylene) and can have a thickness of about 0.1 mm.

[0653] Fluid cartridge 500 may include auxiliary detection areas 594a, 594b (see Figure 4 , Figure 5 , Figure 7 In a non-limiting example, each auxiliary detection region 594a, 594b includes a microarray slide (or biochip) adhered to the bottom surface 503 of the cartridge body 502 within a recessed cavity, which, when covered, for example, by a bottom film 530, forms a detection chamber for nucleic acid analysis. The instrument 10 may include means (not shown) for optically exciting the microarray slide and means for optically detecting a microarray image (not shown) representing an analyte of interest (e.g., nucleic acid) of the sample being analyzed in the cartridge. For further description of the microarray slide, see, for example, U.S. Patent No. 10,654,039.

[0654] refer to Figure 4 and Figure 5Bottom recesses G1 to G10 extend between central through-holes H1c to H10c and recesses associated with each of valves V1 to V10, respectively. Each of valves V1 to V10 is associated with through-holes H1 to H10. Each of the through-holes H1 to H10 associated with valves V1 to V10 connects chambers W1 to W10 to bottom recesses G1 to G10, respectively. In this context, connection to or through a top or bottom recess means connection of a corresponding channel formed by or through each recess when covered by a top or bottom film 512 or 530. Through-hole H11 associated with valve V11 connects chamber W6 to bottom recess G12. Through-hole H12 associated with valve V12 connects chamber W6 to bottom recess G13. The through-hole H13 associated with valve 13 connects to the bottom recess G15 of reaction / detection chambers 510b1 and 510b2, which in turn connects to the top recess G21. The through-hole H14 associated with valve V14 connects to the bottom recess G16 of reaction / detection chambers 510a1 and 510a2, which in turn connects to the top recess G22. The through-hole H15 associated with valve V15 connects the bottom recess G17 to the top recess G29, which merges with the top recess G30. The through-hole H16 associated with valve V16 connects the bottom recess G18 to the top recess G30. The through-hole H17 associated with valve V17 connects the bottom recess G19 to the top recess G31. The through-hole H18 associated with valve V18 connects the bottom recess G20 to the top recess G32, which merges with the top recess G31. The through-hole H19 connects the bottom recess G11 to chamber W11. The through-hole H20 connects the bottom recess G14 to chamber W12. Through hole H21 connects bottom recess G11 to top recess G23. Through hole H22 connects bottom recess G12 to top recess G21. Through hole H23 connects bottom recess G13 to top recess G22. Through hole H24 connects bottom recess G14 to top recess G24. Through hole H25 connects bottom recess G17 to top recess G25 connected to chamber 510b2. Through hole H26 connects bottom recess G18 to top recess G26 connected to chamber 510b1. Through hole H27 connects bottom recess G19 to top recess G27 connected to chamber 510a2. Through hole H28 connects bottom recess G20 to top recess G28 connected to chamber 510a1. Through hole H29 connects top recess G30 to auxiliary detection area 594b of the box. Through hole H31 connects auxiliary detection area 594b to top recess G23, which is connected via through hole H21 to bottom recess G11 of chamber W11 (e.g., waste chamber) via through hole H19. Through hole H30 connects top recess G31 to auxiliary detection area 594a of the box.Through-hole H32 connects auxiliary detection area 594a to top recess G24, which is connected via through-hole H24 to bottom recess G14, which is connected via through-hole H20 to chamber W12 (e.g., waste chamber). Therefore, when valve V8 is opened, reaction chamber 510a1 is connected to auxiliary detection area 594a via grooves G28, G20, G32, and G31. When valve V17 is opened, reaction chamber 510a2 is connected to auxiliary detection area 594a via grooves G27, G19, and G31. When valve V15 is opened, reaction chamber 510b2 is connected to auxiliary detection area 594b via channels G25, G17, G29, and G30. When valve V16 is opened, reaction chamber 510b1 is connected to auxiliary detection area 594b via channels G26, G18, and G30.

[0655] Such as a schematic cross-sectional view of box 500. Figure 8 As shown, chambers 510a1, 510a2, 510b1 and 510b2 are defined by openings formed in the box body 502, which extend between the top surface 501 and the bottom surface 503 and are closed by the bottom film 530 and the top film 512. Reaction / detection chambers 510a1, 510a2, 510b1, and 510b2 receive reaction mixtures prepared from the contents of one or more of chambers W1 to W10. The reaction mixtures are exposed to heat (e.g., isothermal or thermal cycling distribution) within chambers 510a1, 510a2, 510b1, and 510b2 by contacting the top portions of adjacent chambers 510a1, 510a2, 510b1, and 510b2 of fluid cartridge 500 with a top heater, and by contacting the bottom portions of adjacent chambers 510a1, 510a2, 510b1, and 510b2 of fluid cartridge 500 with a bottom heater. Reactions (e.g., amplification reactions) occur within chambers 510a1, 510a2, 510b1, and 510b2. The reaction mixture within chambers 510a1, 510a2, 510b1, and 510b2 may include a detectable probe that emits a detectable optical signal during the reaction upon hybridization with a molecule of interest, for example, a fluorescence signal with an emission wavelength when emitted by light of a certain excitation wavelength. For this purpose, at least one wall of chambers 510a1, 510a2, 510b1, and 510b2 may be transparent or translucent. For example, if the housing body 502 is made of an opaque material, the top film 512 may be transparent or translucent, or at least a portion of the top film 512 covering chambers 510a1, 510a2, 510b1, and 510b2 may be transparent or translucent to allow excitation signals to be delivered to the chambers from above and to allow emission signals to be detected from above the chambers.

[0656] To promote uniform heat distribution in chambers 510a1, 510a2, 510b1, and 510b2, the bottom film 530 may include a layer of thermally conductive material, such as a metal foil (e.g., aluminum), which is disposed on the bottom surface 503 of the housing body 502, at least adjacent to chambers 510a1, 510a2 and adjacent to chambers 510b1 and 510b2. Figure 8 As shown, the lower film 530 may have cutouts 531a and 531b on chambers 510a1, 510a2 and chambers 510b1, 510b2, respectively. A thermally conductive laminated seal 532a is disposed within the cutout 531a and attached to the bottom surface 503 of the housing body 502 on chambers 510a1, 510a2, and a thermally conductive laminated seal 532b is disposed within the cutout 531b and attached to the bottom surface 503 of the housing body 502 on chambers 510b1, 510b2. The positions of the thermally conductive laminates 532a and 532b are... Figure 5 The cutouts 531a, 531b and the associated thermally conductive laminated seals 532a, 532b are shown in dashed lines. These cutouts can be rectangular (e.g., ...). Figure 3 and Figure 5 (as shown in the diagram), circular, elliptical, or any desired shape. When the reaction / detection chambers are arranged as spatially separated groups of chambers (where a “group” may include one or more chambers), discrete thermally conductive laminated seals may be provided to cover each group. For example, when the chambers 510a1, 510a2 and 510b1, 510b2 of the fluid cartridge 500 are arranged as spatially separated groups (e.g., pairs), two separate thermally conductive laminates are provided: a laminated seal 532a for covering groups 510a1, 510a2 and a laminated seal 532b for covering groups 510b1, 510b2.

[0657] In a non-limiting example, each thermally conductive laminated seal 532a, 532b includes a plastic layer 533 (e.g., polypropylene) to which a conductive foil layer 534 is laminated. Suitable commercially available products include Thermo-Fisher AB 3599, available from Thermo-Fisher Scientific in Waltham, Massachusetts. The conductive foil layer 534 may also be optically reflective (e.g., aluminum or metallized PET film). The plastic layer 533 and the conductive foil layer 534 can be bonded together by a suitable adhesive or other means suitable for securing the plastic to the foil. In a non-limiting example, for a total thickness of 70 µm to 100 µm for each thermally conductive laminated seal 532a, 532b, the conductive foil layer 534 has a thickness of 60 µm to 80 µm, and the plastic layer 533 has a thickness of 10 µm to 20 µm. As noted above, the bottom film 530 may have a thickness of approximately 0.1 mm to 0.2 mm (100 µm to 200 µm). In another example, each thermally conductive laminated seal 532a, 532b includes a second plastic layer (not shown) attached to the opposite side of the conductive foil layer 534.

[0658] Each thermally conductive laminated seal 532a, 532b is attached to the housing body 502 by heat sealing, ultrasonic welding, adhesives, or other suitable methods for bonding the plastic layer 533 of each thermally conductive laminated seal 532a, 532b to the housing body 502 to prevent fluid leakage from chambers 510a1, 510a2, 510b1, 510b2. In this regard, for heat sealing or ultrasonic welding, the housing body 502 may include an energy director to facilitate the heat sealing or ultrasonic welding process. An energy director is a component or feature in heat sealing applications that helps focus and control the flow of energy (heat or vibration) to the area where a seal is being created. Examples of energy directors include raised features (e.g., ribs) adjacent to or surrounding each of chambers 510a1, 510a2, 510b1, 510b2 to form narrow edges (e.g., dome-shaped or knife-edge (triangular) sections) that focus energy at the edges and promote localized material melting at the edges to facilitate sealing of laminated seals 532a, 532b. Conductive laminated seals 532a, 532b are heat-sealed by melting and fusing the energy directors surrounding chambers 510a1, 510a2, 510b1, 510b2 with the plastic layer 533 of each of the laminated seals 532a, 532b. International application PCT / US2025 / 026844, filed on April 29, 2025, entitled "Fluidic Cartridge and Apparatuses for Processing Fluidic Cartridges", describes an example of a fluid cartridge using an energy director to facilitate heat sealing or ultrasonic welding of the seal to the cartridge body.

[0659] The conductive foil layer 534 of each thermally conductive laminated seal 532a, 532b is an effective thermal conductor, which, combined with a relatively thin plastic layer (such as polypropylene) used as an insulator, promotes rapid conductive heat transfer from the heaters arranged below the chambers 510a1, 510a2, 510b1, 510b2, thereby rapidly heating the chambers by the heaters arranged below the chambers, and promotes uniform heat distribution to minimize the thermal gradient across the chambers 510a1, 510a2, 510b1, 510b2.

[0660] In some examples, the conductive foil layer 534 can improve the intensity and accuracy of optical emission signal detection from chambers 510a1, 510a2, 510b1, and 510b2. The conductive foil layer 534 of each thermally laminated seal 532a, 532b can provide a reflective surface to increase the intensity of the optical emission signal. An optical excitation signal introduced from above each of chambers 510a1, 510a2, 510b1, and 510b2 passes through the reaction mixture within the chamber and excites the probe-associated mark. Then, as the optical excitation signal is reflected away from the conductive foil layer 534 at the bottom of each chamber, the reflected excitation signal again passes through the reaction mixture within the chamber and again excites the probe-associated mark. Furthermore, the optical emission signals collected from above chambers 510a1, 510a2, 510b1, and 510b2 will be enhanced because both optical signals emitted directly toward the top of each chamber and optical signals emitted toward the bottom of each chamber and reflected toward the top of the chamber by the conductive foil layer 534 at the bottom of the chamber can be collected.

[0661] Furthermore, the laminated seals can increase the accuracy of the emitted signals collected from chambers 510a1, 510a2, 510b1, and 510b2. A relatively thick transparent or translucent thin film layer (e.g., a thickness of 100 µm to 200 µm, such as the bottom film 530) directly covering chambers 510a1, 510a2, 510b1, and 510b2 can serve as an optical emitter (i.e., a light tube) that can laterally transmit optical signals from one chamber to adjacent chambers (e.g., between chambers 510a1 and 510a2, and between chambers 510b1 and 510b2). This inter-chamber optical transmission is reduced or eliminated by thermally conductive laminated seals 532a and 532b having a plastic layer 533 that can be as thin as 10 µm to 20 µm, directly covering chambers 510a1, 510a2, 510b1, and 510b2. In addition, metal foils such as aluminum foil are impermeable to water, thereby preventing vapor from being transferred to or from chambers 510a1, 510a2, 510b1, 510b2, to enhance the stability of the dried (dehydrated or lyophilized) reagents stored in the chambers.

[0662] The reagents required to perform a specified reaction within reaction chambers 510a1, 510a2, 510b1, and 510b2 may be pre-applied in wet form and then dried onto the interior surfaces of laminated seals 532a and 532b (i.e., the outer surfaces of the plastic layer 533 of laminated seals 532a and 532b), as described in International Application No. PCT / US 2025 / 026844, filed April 29, 2025, entitled “Fluidic Cartridge and Apparatuses for Processing Fluidic Cartridges”.

[0663] The laminated seals 532a and 532b are separated from the bottom film 530—that is, the laminated seals 532a and 532b are structurally and functionally isolated from the bottom film 530. Accordingly, depending on the specific operational, functional, and / or structural requirements (such as defined channels) for the bottom film, different formulations and configurations of the bottom film 530 can be employed without requiring changes to the laminated seals. In other examples, the bottom film covers a portion of the surface of the cartridge, which is spatially separated or isolated from one or more reaction / detection chambers covered by one or more laminated seals; in this case, cutouts formed in the bottom film are unnecessary.

[0664] The functional chambers W1 to W12 and SB of the cartridge body 502 contain or are configured to receive, during use of the fluid cartridge 500, a fluid sample, various reagent products, a purification column, and at least one of a fluid or solid intended for the preparation, amplification, and analysis of the sample. Other chambers may serve as mixing chambers to temporarily hold two or more different materials combined therein, or may serve as waste chambers. Examples of the functions of chambers W1 to W12 and CW and / or their contents are illustrated in Table 1 below:

[0665] Table 1

[0666] As explained above, chambers W1 to W5 and W7 to W10 each include through-holes H1 to H5 and H7 to H10 formed through the bottom wall of the respective chamber, and functional chamber W6 includes three through-holes H6, H11, and H12 formed through the bottom wall of the chamber. Through-hole H1 forms the sample outlet port from sample chamber W1. The syringe barrel SB includes central through-holes H1c, H2c, H3c, H4c, H5c, H6c, H7c, H8c, H9c, and H10c formed through the bottom wall of the barrel. Each of chambers W1 to W10 is in independent fluid communication with the central chamber SB via channels formed by grooves G1, G2, G3, G4, G5, G6, G7, G8, G9, and G10, respectively, controlled by valves V1, V2, V3, V4, V5, V6, V7, V8, V9, and V10. Fluid can flow between these different functional chambers in one direction or the other (i.e., from each of chambers W1 to W10 to the syringe barrel SB or vice versa).

[0667] Details of the example of cap 516 are in Figures 15 to 17 As shown in the figure, and Figure 18 This is a partial cross-section of the box body 502, showing the cap 516 inserted into the sample chamber W1. The desired properties of the cap 516 include: it effectively seals the sample chamber W1; it is easily pressed into the sample chamber by the user and will not fall off; it is vented to prevent pressurization during insertion; and it includes material covering the vent(s) to prevent liquid leakage, and has sufficiently small openings (orifices) (e.g., 0.2 µm) to prevent viral particles or aerosols from escaping from the vent(s). The cap 516 can be positioned about axis Z (see...). Figure 17 The device is rotationally symmetric and includes an upper portion 518 with laterally extending members, which in the illustrated example take the form of a radial wall 522 oriented radially relative to axis Z and a peripheral wall 520 surrounding the radial wall 522 and extending axially relative to axis Z. The peripheral wall 520 may include a flat gripping portion 517 or may otherwise be adapted for an improved manual grip.

[0668] The cap 516 also includes a lower portion 519 defined by a sleeve or peripheral wall 525, which hangs (e.g., extends below) from the radial wall 522 and extends axially relative to the axis Z. The inner surface of the sleeve or peripheral wall 525, the bottom surface of the laterally extending member, or the radial wall 522 defines a recess or cavity 528 extending upward from the bottom end of the cap 516. The upper portion 518 of the cap 516 is wider than the lower portion 519, thereby defining a radially annular shoulder 524 at the peripheral region of the lower surface of the radial wall 522. The peripheral wall 525 is inserted into the sample chamber W1; for this purpose, the wall 525 may be tapered, and the radial shoulder 524 contacts the top edge surface 514 of the wall of the sample chamber W1 (see...). Figure 6 and Figure 18 The outer surface 527 of the peripheral wall 525 is sealed to the inner surface 515 of the sample chamber W1 (see...). Figure 18 The lower portion 519 may also include radially extending annular ribs 526a, 526b that protrude from the outer surface 527 of the peripheral wall 525 to contact the inner surface 515 of the sample chamber W1 (see...). Figures 16 to 18 This enhances the seal between the peripheral wall 525 of the cap 516 and the sample chamber W1. The outer surface 527 of the peripheral wall 525 can be frictionally engaged with the inner surface 515 of the wall of the sample chamber W1, or the peripheral wall 525 can be connected to the wall of the sample chamber W1 by mating threads (not shown) on the peripheral wall 525 and the wall of the sample chamber W1.

[0669] Vent holes 523 are formed in the radial wall 522, and side vent holes 521a and 521b are formed in the peripheral wall 520. Vent holes 523 may have a width (e.g., diameter) of about 2 mm, and are preferably formed by a porous venting membrane 529 attached to the top surface or bottom surface of the radial wall 522 (only when...). Figure 17 (As shown in the image) Coverage. Suitable materials for membrane 529 include Traketch® PET / PET0.2 Vent R300, part number 063390, from SABEU GmbH & Co. KG of Nordheim, Germany, which comprises a 23 µm thick PET membrane material with 60 g / m 2 The non-woven PET backing has a pore size of 0.2 ± 0.4 µm and a pore density of 320 ± 50 x 10⁻⁶. 6 / cm 2 The total thickness is 140 ± 50 µm.

[0670] The cap 516 can be formed from a thermoplastic elastomer (such as TPE Thermolast® M TM6MHD from KRAIBURG TPE GmbH & Co. KG in Waldkreiburg, Germany) (e.g., injection molding).

[0671] The fluid cartridge 500 may include two functional sections. For example... Figure 6 and Figure 7As shown, the sample preparation section 504 of the fluid cartridge 500 includes multiple chambers (e.g., chambers W1 to W12) that contain or can receive various materials (which may include liquids or other fluids) used in sample preparation for performing assays or other procedures on the sample within the cartridge during operation of the cartridge by the instrument 10. The sample preparation section 504 is configured to receive sample specimens in a sample chamber (e.g., chamber W1) (which may include or be connected to a fluid inlet port at which fluid samples are introduced into the sample chamber), and is configured to process the sample using materials contained in one or more other chambers within the sample preparation section 504, for example, to isolate target molecules (e.g., lysing and purifying nucleic acids using silica-based purification) from other components of the sample, and is configured to combine the isolated molecules with materials used in performing assays (such as amplification reagents and / or detection probes) to form a reaction mixture. Amplification reagents and / or detection probes may be provided in a dried (e.g., lyophilized or speckled) form in one or more chambers W2 to W10 of the sample preparation section 504, and reconstruction fluids for combination and reconstruction with reagents and / or probes may be contained in one or more chambers W2 to W10 of the sample preparation section 504. Valves V1 to V10, which control the flow of fluid into and out of chambers W1 to W10 respectively, and valves V11 and V12, which control the flow of fluid into and out of chamber W6, may be referred to as sample preparation (or processing) valves because they are located in chambers W1 to W10 within the sample preparation section 504 of the fluid cartridge 500 and control the flow of fluid in chambers W1 to W10.

[0672] refer to Figure 6 and Figure 7 The reaction / detection section 506 of the fluid cartridge 500 is configured to receive a processed sample (reaction mixture) from the sample preparation section 504 and is configured to provide a platform for one or more reactions to occur, for example, to amplify and detect target molecules (e.g., real-time PCR). The reaction / detection section 506 includes one or more reaction chambers (e.g., reaction / detection chambers 510a1, 510a2, 510b1, 510b2), each of which defines a enclosure capable of containing a fluid substance, and within the enclosure, a reaction can occur, and a detectable signal emitted during the reaction can be detected from the enclosure. The detectable signal may be an optical signal (such as fluorescence), and the detection of the detectable signal can indicate the presence and / or amount of target molecules in the sample. Valves V13 to V18 that control the flow of fluid into and out of reaction chambers 510a1, 510a2, 510b1, and 510b2 can be referred to as reaction valves because they are located within reaction chambers 510a1, 510a2, 510b1, and 510b2 in the reaction / detection section 506 of housing 500 and control the flow of fluid into reaction chambers 510a1, 510a2, 510b1, and 510b2.

[0673] Mechanical pyrolysis on the plate

[0674] The fluid cartridge 500 can be configured to facilitate mechanical lysis on the execution plate to break down cells (e.g., pathogenic microorganisms including bacteria, viruses, parasites, etc.) contained in the sample material dispensed into the sample chamber W1, to release nucleic acids (DNA or RNA) from the cells of the sample material for downstream molecular assays. In such an embodiment, in the sample chamber W1 (see...) Figure 3 The lysis chamber provides a lysis chamber containing components (including a lysis agent) that can be used to perform mechanical lysis of sample material. The lysis chamber may include a variety of shape factors, examples of which are described herein.

[0675] Figure 49 This is a perspective view of a first embodiment of the rupture capsule 600, and Figure 50 It is along the ruptured sac-like element 600 Figure 49 The cross-section of line AA in the fluid cartridge 500. The capsule 600 is placed in the sample chamber W1 of the fluid cartridge 500, which is closed by the cap 516. The capsule 600 can be press-fitted into the sample chamber W1, or the capsule 600 can be threaded into the inner surface of the sample chamber W1.

[0676] refer to Figure 49 and Figure 50 The capsule 600 includes a hollow body 602, which, in the illustrated example, includes: a first (e.g., upper) portion 604, which may be cylindrical or generally cylindrical; a second (e.g., lower) portion 610, which may be cylindrical or generally cylindrical, centered or coaxial with respect to the first portion 604, and having a smaller width (diameter) than the first portion 604; and a tapered transition portion 616 between the first portion 604 and the narrower second portion 610. In another example, the transition between the first portion 604 and the narrower second portion 610 is not tapered. The segment 610 may include raised sealing ribs 611 surrounding the segment 610 to provide a sealing interface connection between the capsule 600 and the sample chamber W1. The hollow body 602 may include an integral component molded (e.g., injection molded) from a plastic material such as polypropylene, polyethylene, acrylonitrile butadiene styrene (“ABS”), or polyethylene terephthalate (“PET”).

[0677] A first edge 606 surrounds an open first end 608 at one end of the hollow body 602 (the top end in the shown embodiment), and a second edge 612 surrounds an open second end 614 at the opposite end of the hollow body 602 (the bottom end in the shown embodiment).

[0678] For example, a first porous membrane or barrier 618 is attached to a first edge 606 by an adhesive, by heat sealing, or by ultrasonic welding. A second porous membrane 620 is attached to a second edge 612 by an adhesive, by heat sealing, or by ultrasonic welding. A first membrane 618 covering an open first end 608 and a second membrane or barrier 620 covering an open second end 614 define a lysis chamber 622 within a hollow body 602 between membranes 618 and 620, and include lysis elements, such as beads, as described below. The first and second membranes may be filters, wherein the first membrane 618 filters out larger sample components (e.g., undigested food particles and mucus found in feces or other gastrointestinal samples), thereby inhibiting such larger components from entering the lysis chamber 622, and the second membrane 620 filters out lysed cell debris, thereby inhibiting such cell debris from leaving the lysis chamber 622, while allowing targets of interest (e.g., DNA and RNA) to exit the lysis chamber.

[0679] The first porous membrane 618 may be a mesh and is preferably hydrophilic (naturally hydrophilic or treated to be hydrophilic) to facilitate the passage of fluid sample material through the first membrane 618. Suitable materials include polyamide, polypropylene, polyethylene terephthalate (PETP), ethylene tetrafluoroethylene (ETFE), or polyether ether ketone (PEEK). The porosity (pore size) of the first porous membrane 618 may be, for example, from 70 µm to 500 µm (e.g., about 300 µm), with the maximum size limited by the size of the pyrolysis beads to be retained within the pyrolysis chamber 622. Suitable meshes for the first porous membrane 618 are available from Sefar, Inc., Buffalo, NY, part number 03-300 / 51 HPL, with a pore size of 300 µm.

[0680] Vent 523 covering cap 516 (see Figure 17 The venting membrane 529 is preferably thinner than the first porous membrane 618 to allow air to escape from the sample chamber W1 through the vent 523 without allowing liquid to pass through.

[0681] The second porous membrane 620 may be a mesh or filter matrix (e.g., a sintered or spun filter) and is preferably hydrophilic (naturally hydrophilic or treated to be hydrophilic). Suitable materials include polyamide, polypropylene, polyethylene terephthalate (PETP), ethylene tetrafluoroethylene (ETFE), or polyether ether ketone (PEEK). The porosity (pore size) of the second porous membrane 620 is preferably smaller than that of the first porous membrane 618, because the first porous membrane is designed to allow sample fluid to pass through the lysis chamber 622, and the second porous membrane 620 is designed to capture the lysed cell material. The porosity (pore size) of the second porous membrane 620 may be, for example, from 30 µm to 100 µm, for example, about 70 µm. The pore size of the second porous membrane 620 should be small enough to capture the lysed cell material, but not too small to avoid easy clogging. Suitable mesh for the second porous membrane 620 is available from Sefar, Inc., Buffalo, NY, part number 03-70 / 33 HPL, with a pore size of 70 µm.

[0682] Protective meshes and supports (not shown) may be added to either side of the first porous membrane 618 and / or the second porous membrane 620 to help maintain membrane integrity during pyrolysis. Because membrane materials (e.g., polyethersulfone (PES)) can be fragile, they are prone to breakage during pyrolysis by beads and magnets. Mechanically robust supporting meshes (such as woven nylon or polyester meshes) or structures (such as injection-molded or 3D-printed meshes) may be layered on top and / or at the bottom of the membranes for support and protection. The supporting meshes may have porosities up to about 300 µm to 350 µm, with the maximum size limited by the size of the pyrolysis beads within the pyrolysis chamber 622.

[0683] Optionally, the filter element 630 (e.g., a sintered filter) may be provided within the pyrolysis chamber 622, with or without the second porous membrane 620, for example at a location covering the open second end 614, such as... Figure 50As shown in the diagram. In some applications, this filter element will trap additional lysed cell debris that was not trapped by the second porous membrane 620. The porosity (pore size) of the filter element 630 may be the same as that of the second porous membrane 620 (e.g., in the range of 30 µm to 100 µm or about 70 µm). The porosity of the filter element 630 may vary throughout its thickness, for example, having a pore size that gradually decreases from a larger pore size (e.g., 100 µm) at the top surface of the filter element 630 to a smaller pore size (e.g., 30 µm) at the bottom surface of the filter element 630. In a non-limiting example, if the filter element 630 is included within the lysis chamber 622, the pore size of the second porous membrane 620 may be much larger than described above, for example, up to 300 µm, because an additional filter element can be used to capture the lysed cell material. In another example, the second porous membrane 620 can be omitted. In this case, the filter element 630 can be retained within the open second end 614 by interference fit, friction fit, or by a wall or shelf extending inward from the open second end 614 below the filter element 630. In another example, the filter element can be located outside the lysis chamber 622 within the sample chamber W1 between the second membrane 620 and the through-hole H1. In another embodiment, the filter element is omitted because while the filter element can effectively trap cell debris, it may also trap lysed nucleic acids intended to be expelled from the sample chamber.

[0684] The lysis chamber 622 comprises a plurality of non-magnetic beads 624 and a magnetic element 626 filling a portion of the volume of the lysis chamber 622. The non-magnetic beads 624 and the magnetic element 626 may be collectively referred to as “lysis beads” or “lysis agent”. A fluid sample is provided to the lysis chamber 622, and, as described below, the magnetic element 626 is agitated to give the non-magnetic beads 624 movement to achieve mechanical lysis (referred to as bead milling) of the cells present in the sample contained within the lysis chamber 622. The pore sizes of the first porous member 618 and the second porous member 620 are small enough to retain the non-magnetic beads 624 and the magnetic element 626 within the lysis chamber 622. In a non-limiting example, the lysis chamber 622 has a volume of approximately 600 μl, the non-magnetic bead 624 has a volume of approximately 300 μl (i.e., approximately 50% of the lysis chamber volume), and the magnetic element 626 has a volume of approximately 27 μl to 64 μl (i.e., approximately 4.5% to approximately 11% of the lysis chamber volume), thereby leaving approximately 236 μl to 273 μl (39% to 45% of the 600 μl lysis chamber) of space in the lysis chamber 622 for the fluid sample material. In this example, the non-magnetic bead 624 occupies 50% to 75% of the usable volume of the lysis chamber 622 (i.e., the total volume of the lysis chamber 622 minus the volume occupied by the magnetic element 626). Factors affecting the amount of nonmagnetic beads 624 to be provided relative to the total volume of the lysis chamber 622 include (1) providing enough beads to efficiently and effectively grind (lyse) the sample molecules within the lysis chamber 622, and (2) not providing too many nonmagnetic beads 624, which would excessively restrict the movement of the magnetic element 626. In another embodiment, there is an available open volume in the sample chamber above the capsule 600 (i.e., above the first membrane 618), and the volume of fluid sample material added to the sample chamber W1 exceeds the available volume within the lysis chamber 622.

[0685] Suitable nonmagnetic beads 624 include beads made of ceramic, glass, silica, or zirconium, and can be spherical or substantially spherical in shape, wherein the size (e.g., diameter) ranges from 100 µm to 2000 µm (e.g., about 500 µm (0.5 mm)) depending on the intended application (i.e., the intended lysis target). In a non-limiting example, the nonmagnetic beads are inert relative to the sample material (i.e., the beads do not react with the cellular material or bind to released nucleic acids). Suitable beads include those with product numbers 055-0120 available from Final Advanced Materials SARL in Dudenheim, France. ZrO2 beads, cerium-stabilized. ZrO2: 83% - CeO2: 17%, Ø 0.40 - 0.70: 3.75 + / - 0.05 kg / L.

[0686] Magnetic element 626 is a permanent magnet made of a magnetic material such as N52 or N42 grade neodymium (NdFeB), and is preferably axially magnetized (i.e., the north and south poles (“N” and “S”) are located at two points 180º apart). As described below, magnetic element 626 is exposed to a changing magnetic field, which in turn causes magnetic element 626 to move within lysis chamber 622, thereby giving non-magnetic bead-like elements 624 motion to mechanically lyse or destroy cells contained within the sample provided to lysis chamber 622, thereby releasing their internal components (e.g., DNA, RNA, proteins, and organelles). Magnetic element 626 can be of any shape, including cubes, spheres, rods, disks, etc. Magnetic element 626 with an edge 628 (e.g., a cube or other parallelepiped) exhibits better performance because it more effectively gives non-magnetic bead-like elements 624 the desired motion. Edge 628 can be circular. In a non-limiting example, the magnetic element 626 is cubic in shape, with sides having a width of about 2 mm to 3 mm (e.g., up to about 1 / 8 inch). One factor to consider when determining the size of the magnetic element 626 is that, while larger and stronger magnets may be preferred in some applications, the size of the magnet will be limited by the available volume within the pyrolysis chamber 622 to allow for sufficient movement of the magnetic element 626.

[0687] The magnetic element 626 may be coated or encapsulated (e.g., molded) with a non-magnetic material that does not react with the sample solution to prevent reaction between the magnetic element 626 and the sample solution and / or to protect the magnetic element 626 from abrasion by the non-magnetic bead 624. Coating materials may include Teflon® (polytetrafluoroethylene), polypropylene, epoxy resin, urethane, nickel, or gold. Coating thickness (e.g., a total thickness of up to about 4.3 mm) may increase the width of the facet of the magnetic element 626.

[0688] In a non-limiting example, the magnetic element 626 and / or its coating are inert relative to the sample material (i.e., the element will not react with the cellular material or bind to released nucleic acids).

[0689] Figure 51 This is a cross-section of the sample chamber W1 of the box body 502 with a cap 516, and an alternative embodiment of the capsule 600ʹ disposed within the sample chamber W1 below the cap 516. The capsule 600ʹ can be press-fitted into the sample chamber W1, or the capsule 600ʹ can be threaded into the inner surface of the sample chamber W1.

[0690] The capsule-shaped member 600ʹ has a hollow body 602ʹ, the hollow body having a first edge 606ʹ surrounding an open first end 608ʹ (the top end in the shown embodiment) and a second edge 612ʹ surrounding an open second end 614ʹ (the bottom end in the shown embodiment). A first porous membrane 618ʹ, identical to the first porous membrane 618 described above, can be attached to the first edge 606ʹ, for example by adhesive, heat sealing, or ultrasonic welding. A second porous membrane 620ʹ, identical to the second porous membrane 620 described above, can be attached to the second edge 612ʹ, for example by adhesive, heat sealing, or ultrasonic welding.

[0691] The first membrane 618ʹ and the second membrane 620ʹ define a pyrolysis chamber 622ʹ between membranes 618ʹ and 620ʹ within the hollow body 602ʹ. Protective mesh and support members (not shown) may be added to either side of the first porous membrane 618ʹ and / or the second porous membrane 620ʹ to help maintain the integrity of the membranes during pyrolysis.

[0692] Similar to the hollow body 602 of the capsule 600 described above, the hollow body 602ʹ includes: a first portion 604ʹ, which may be cylindrical or generally cylindrical; a second portion 610ʹ, which may be cylindrical or generally cylindrical and has a smaller width (diameter) than the first portion 604ʹ; and a transition portion 616ʹ between the first portion 604ʹ and the narrower second portion 610ʹ, which may be tapered or non-tapered as shown. The segment 610ʹ may include raised sealing ribs surrounding the segment 610ʹ for providing a sealed interface connection between the capsule 600ʹ and the sample chamber W1. Figure 51 For unmarked text, see [link / reference]. Figure 50 (Sealing rib 611 in the middle). The hollow body 602ʹ differs from the hollow body 602 described above in that the second part 610ʹ is not centered or coaxial with respect to the first part 604ʹ. The hollow body 602ʹ may include an integral part molded (e.g., injection molded) from a plastic material such as polypropylene, polyethylene, acrylonitrile butadiene styrene (“ABS”), or polyethylene terephthalate (“PET”).

[0693] The lysis chamber 622ʹ of the capsule 600ʹ comprises a plurality of non-magnetic beads 624ʹ and a magnetic element 626ʹ. The plurality of non-magnetic beads may be identical to the non-magnetic beads 624 described above, filling a portion of the volume of the lysis chamber 622ʹ. The magnetic element may be identical to the magnetic element 626 described above and may include an edge 628ʹ. The non-magnetic beads 624ʹ and the magnetic element 626ʹ may be collectively referred to as “lysis beads” or “lysis agent”. A fluid sample is supplied to the lysis chamber 622ʹ, and the magnetic element 626ʹ is agitated as described below to impart motion to the non-magnetic beads 624ʹ, thereby achieving mechanical lysis of cells present in the sample contained within the lysis chamber 622ʹ. The relative volumes of the lysis chamber 622ʹ, the non-magnetic beads 624ʹ, and the magnetic element 626ʹ can be as described above for the lysis chamber 622, the non-magnetic beads 624, and the magnetic element 626.

[0694] The lysis capsule 600ʹ may include an optional filter element 630ʹ within the lysis chamber 622ʹ, which may be the same as the filter element 630 described above. Alternatively, the filter element 630ʹ may be arranged within the sample chamber W1 outside the hollow body 602ʹ, or the filter element 630ʹ may be omitted.

[0695] Figure 52 This is a cross-section of the sample chamber W1 of the box body 502 with a cap 516, and an alternative embodiment of the capsule 600ʹʹ disposed within the sample chamber W1 below the cap 516. A gap 632 may be provided between the bottom end of the cap 516 and the top end of the capsule 600ʹʹ. A similar gap may be provided between the cap 516 and the capsule 600ʹ described herein and / or between the cap 516 and the capsule 600ʹ described herein. The capsule 600ʹʹ may be press-fitted into the sample chamber W1, or the capsule 600ʹʹ may be threaded into the inner surface of the sample chamber W1.

[0696] The capsule-like member 600ʹʹ has a hollow body 602ʹʹ, the hollow body having a first edge 606ʹʹ surrounding an open first end 608ʹʹ (the top end of the illustrated embodiment) and a second edge 612ʹʹ surrounding an open second end 614ʹʹ (the bottom end of the illustrated embodiment). The hollow body 602ʹʹ may comprise an integral part molded (e.g., injection molded) from a plastic material such as polypropylene, polyethylene, acrylonitrile butadiene styrene (“ABS”), or polyethylene terephthalate (“PET”). The hollow body 602ʹʹ includes a sleeve that may be cylindrical or substantially cylindrical, and the sleeve has a constant width (e.g., diameter) between the first edge 606ʹʹ and the second edge 612ʹʹ. For example, a first porous membrane 618ʹʹ, identical to the first porous membrane 618 described above, can be attached to a first edge 606ʹʹ by adhesive, heat sealing, or ultrasonic welding. Similarly, a second porous membrane 620ʹʹ, identical to the second porous membrane 620 described above, can be attached to a second edge 612ʹʹ by adhesive, heat sealing, or ultrasonic welding.

[0697] The first membrane 618ʹʹ and the second membrane 620ʹʹ define a pyrolysis chamber 622ʹʹ within the hollow body 602ʹʹ, between membranes 618ʹʹ and 620ʹʹ. Protective mesh and support members (not shown) may be added to either side of the first porous membrane 618ʹʹ and / or the second porous membrane 620ʹʹ to help maintain membrane integrity during pyrolysis.

[0698] The lysis chamber 622 of the capsule 600ʹʹ contains a plurality of non-magnetic beads 624ʹʹ and a magnetic element 626ʹʹ. The plurality of non-magnetic beads may be identical to the non-magnetic beads 624 described above, filling a portion of the volume of the lysis chamber 622ʹʹ. The magnetic element may be identical to the magnetic element 626 described above and may include an edge 628ʹʹ. The non-magnetic beads 624ʹʹ and the magnetic element 626ʹʹ may be collectively referred to as “lysis beads” or “lysis agent.” A fluid sample is supplied to the lysis chamber 622ʹʹ, and the magnetic element 626ʹʹ is agitated as described below to impart movement to the non-magnetic beads 624ʹʹ, thereby achieving mechanical lysis of cells present in the sample contained within the lysis chamber 622ʹʹ. The relative volumes of the pyrolysis chamber 622ʹʹ, the nonmagnetic bead 624ʹʹ, and the magnetic element 626ʹʹ can be described above for the pyrolysis chamber 622, the nonmagnetic bead 624, and the magnetic element 626.

[0699] An optional filter element 630, identical to the filter element 630 described above, may be provided within the sample chamber W1 below the lysis capsule 600ʹʹ. Alternatively, the filter element 630ʹʹ may be omitted.

[0700] Dead space 634 can be provided in the sample chamber W1 between the bottom wall of the sample chamber W1 and the lysis sac 600ʹʹ for collecting lysed cell material that can pass through a second membrane 620ʹʹ but not through an additional filter element 630ʹʹ.

[0701] The lysis vesicles 600, 600ʹ, or 600ʹʹ may include a multi-stage filtration system. A pre-filter (not shown) is integrated into the lysis vesicles 600, 600ʹ, or 600ʹʹ to replace or supplement the first porous membrane 618, 618ʹ, or 618ʹʹ (e.g., where the first porous membrane includes a mesh element) and is configured to remove large (generally greater than 50 µm) gastrointestinal samples, such as feces, particles, undigested food, and mucus. The pre-filter may have a sufficiently large pore size to allow target cells of interest to pass through. For example, if the target of interest is a parasite of about 40 µm, the pore size of the pre-filter should be greater than 40 µm to allow said target to enter the lysis vesicle. The pre-filter is preferably hydrophilic and may be a woven mesh filter (such as nylon or polyester or similar) or a membrane filter. The filter can be supported and protected by a protective layer (e.g., by woven nylon or polyester mesh as described above) to prevent it from being damaged during mechanical pyrolysis.

[0702] In another example, a pre-filter (not shown) may be integrated into the sample cap assembly, for example, through the open bottom end of the sleeve or the peripheral wall 525 of the lower portion 519 of the cap 516 (see [reference]). Figure 16 , 17 ).

[0703] Following the pre-filter of a multi-stage filtration system may be one or more enhanced post-filters (not shown) to supplement or replace a second porous membrane 620, 620, or 620,000 (e.g., where the second porous membrane includes a mesh). The post-filters capture finer particles in the micrometer range and allow targets of interest (DNA and RNA molecules) to pass through for downstream processes. During lysis, microorganisms and other components in the sample are mechanically ground into smaller particles (submicrometer scale). The purpose of the finer post-filters is to selectively remove smaller particles without affecting the transport of target molecules and sample volume from the lysis chambers 622, 622, or 622,000. In the presence of more than one post-filter, the filters may have progressively smaller porosities, such as a first post-filter nylon mesh with approximately 30 µm porosity and a second post-filter nylon mesh with approximately 6 µm porosity.

[0704] The filter pore size (from 0.22 µm to 500 µm) and the type of filter can be selected based on the sample type and filtration requirements.

[0705] Figure 53 This is a top perspective view of the alternative lysis vessel 700, including an integral cap for closing the sample chamber W1 and a lysis capsule for containing the material used to perform lysis. Figure 54 It is along the pyrolysis vessel 700 Figure 53 The cross section of line AA, and Figure 55 It is a cross-section of the sample chamber W1 of the box body 502, in which the lysis container 700 is placed inside the sample chamber W1.

[0706] Details of the pyrolysis vessel 700 are in Figures 53 to 55 The lysis container 700, which combines the lysis capsule and the cap for closing the sample chamber W1, as described above, has the following desired properties: it effectively seals the sample chamber W1; it is easily pressed into the sample chamber by the user and will not be accidentally dislodged; it is ventilated to prevent pressurization during insertion; and it includes a material covering the vent(s) to prevent liquid leakage and has sufficiently small openings (orifices) to prevent viral particles or aerosols from escaping from the vent(s).

[0707] The pyrolysis container 700 includes a hollow body 702, which is rotationally symmetrical about an axis Y and, in the illustrated example, includes a first (e.g., upper) portion 704, which may be cylindrical or generally cylindrical, and a second (e.g., lower) portion 710, which may be cylindrical or generally cylindrical, centered or coaxial with respect to the first portion 704 and having a smaller width (diameter) than the first portion 704. The first portion 704 has a laterally extending member in the form of a radial wall 708 oriented radially relative to the axis Y, while a peripheral wall 706 surrounds the radial wall 708 and extends axially relative to the axis Y. The lower portion 710 is defined by a sleeve 725 extending below the radial wall 708, which may be cylindrical or generally cylindrical, extends axially relative to the axis Y, and has a constant width (e.g., diameter) between the radial wall 708 and a bottom edge 712 surrounding a lower open end 714. The width difference between the first portion 704 and the second portion 710 defines an annular shoulder 716 at the peripheral region of the lower surface of the radial wall 708. The second portion 710 may also include radially extending annular ribs 727a, 727b projecting from the outer surface of the sleeve 725.

[0708] Hollow body 702 may include integral parts molded (e.g., injection molded) from plastic materials such as polypropylene, polyethylene, acrylonitrile butadiene styrene (“ABS”), or polyethylene terephthalate (“PET”).

[0709] Vent holes 723 are formed in the radial wall 708, and side vent holes (not shown) may be formed in the peripheral wall 706. Vent holes 723 may have a width (e.g., diameter) of about 2 mm and may be covered by a membrane 729, which may be porous, such as the Traketch® Pet / Pet0.2 Vent R300 (part number 063390) from SABEU GmbH & Co. KG in Nordheim, Germany, comprising a 23 µm thick membrane material PET with 60 g / m³. 2 The non-woven PET backing has a pore size of 0.2 ± 0.4 µm and a pore density of 320 ± 50 x 10⁻⁶. 6 / cm 2 The total thickness is 140 ± 50 µm.

[0710] The hollow body 702 can be formed from a thermoplastic elastomer (such as TPE Thermolast® M TM6MHD from KRAIBURG TPE GmbH & Co. KG in Waldkreiburg, Germany) (e.g., injection molding).

[0711] For example, the porous membrane 720 can be attached to the second edge 712 by adhesive, heat sealing, or ultrasonic welding. Protective mesh and support members (not shown) can be added to either side of the porous membrane 720 to help maintain the integrity of the membrane during pyrolysis.

[0712] Radial wall 708 and membrane 729, along with porous membrane 720, define a pyrolysis chamber 722 within the hollow body 702 between radial wall 708 / membrane 729 and porous membrane 720.

[0713] The lysis chamber 722 of the lysis vessel 700 includes a plurality of non-magnetic beads 724 and a magnetic element 726. The plurality of non-magnetic beads 724 may be identical to the non-magnetic beads 624 described above, filling a portion of the volume of the lysis chamber 722. The magnetic element 724 may be identical to the magnetic element 626 described above and may include an edge 728. The non-magnetic beads 724 and the magnetic element 726 may be collectively referred to as "lysis beads" or "lysis agent". A fluid sample is placed within the lysis chamber 722, and the magnetic element 726 is agitated as described below to give the non-magnetic beads 724 movement, thereby achieving mechanical lysis of cells present in the sample contained within the lysis chamber 722.

[0714] In a non-limiting example, the lysis chamber 722 has a volume of about 870 μl, the non-magnetic bead 724 has a volume of about 300 μl, and the magnetic element 726 has a volume of about 27 μl to 64 μl, so that if the sample is filled into the radial wall 708, about 506 μl to 543 μl of space is left for the sample in the lysis chamber 622.

[0715] Figure 55 The image shows a cross-section through the sample chamber W1, illustrating the insertion of the lysis vessel 700 into the sample chamber W1. A sleeve 725 is inserted into the sample chamber W1; for this purpose, the sleeve 725 may be tapered, and a radial shoulder 716 contacts the top edge surface 514 of the wall of the sample chamber W1. The outer surface 731 of the sleeve 725 seals against the inner surface 515 of the sample chamber W1. Ribs 727a and 727b contact the inner surface 515 to enhance the seal between the sleeve 725 of the vessel 700 and the sample chamber W1. The outer surface of the sleeve 725 may frictionally engage with the inner surface of the inner wall of the sample chamber W1, or the sleeve 725 may be connected to the inner wall of the sample chamber W1 via mating threads (not shown) on the sleeve 725 and the inner wall of the sample chamber W1.

[0716] After the sample is dispensed into chamber W1, the lysis vessel 700 is inserted into sample chamber W1. Accordingly, the porosity of the porous membrane 720 should be large enough to allow unlysplit sample to pass through the membrane 720, and may be greater than the porosity of the second porous membrane 620 described above. Accordingly, an optional filter element 730, which may be the same as the filter element 630 described above, may be provided in sample chamber W1 below the lysis vessel 700 to capture the lysed cellular material that will pass through the porous membrane 720.

[0717] Dead space 734 can be provided within sample chamber W1 between the bottom wall of sample chamber W1 and lysis container 700 for collecting lysed cell material.

[0718] In an alternative embodiment, the pyrolysis bead is delivered from a cap having a ruptureable compartment or chamber for containing the bead to a sample contained in a sample chamber, and the bead can be released from the cap into the sample chamber. Figure 75 It is a top perspective view of a bead delivery cap 900 having a fractured chamber containing magnetic and non-magnetic pyrolysis beads, and Figure 76 This is a bottom view of the beaded delivery cap. Figure 77 It is a cross-sectional view of a bead delivery cap 900 containing an intact bead chamber, and Figure 78 It is a cross-sectional view of the bead delivery cap 900, wherein the bead contains a chamber that ruptures to release the pyrolyzed bead.

[0719] The bead delivery cap 900 is rotationally symmetrical about axis Z (see [link]). Figure 77 , 78 The cap body includes an upper portion 902 and a lower portion 908. The upper portion 902 of the cap body includes a laterally extending member, which in the illustrated example takes the form of a radial wall 904 oriented radially relative to axis Z and a peripheral wall 906 surrounding the radial wall 904 and extending axially relative to axis Z. The lower portion 908 is defined by a lower sleeve or lower peripheral wall 910 that hangs (e.g., extends below) from the radial wall 904 and extends axially relative to axis Z. The lower sleeve 910 surrounds an open space 918 extending upward from the bottom end 916 of the cap body. The upper portion 902 of the cap body is wider than the lower portion 908, thereby defining a radially annular shoulder 920 at the peripheral region of the lower surface of the radial wall 904.

[0720] The cap body of the bead delivery cap 900 includes a collapsible chamber defined by a deformable wall 930, which initially bulges outward and extends over a radial wall 904 to define an inner chamber 932 adjacent to (open to) an open space 918. At least one magnetic element 926 and a plurality of non-magnetic beads 924 are arranged in the open space 918 and the inner chamber 932, and are retained by a fragile membrane 934, attached to the bottom end 916 of the cap body, for example by adhesive, by heat sealing, or by ultrasonic welding, to close the open space 918. The chamber 932 and the open space 918 together define a pyrolysis bead compartment, which is at least partially collapsible and closed by the fragile membrane 934. The non-magnetic beads 924 may be the same as the non-magnetic beads 624 described above, and the magnetic element 926 may be the same as the magnetic element 626 described above and may include an edge 928. The non-magnetic bead 924 and the magnetic element 926 can be collectively referred to as "pyrolysis beads" or "pyrolysis agent". For example... Figure 77 As shown, the pyrolysis beads can be arranged within the bead delivery cap 900, wherein the magnetic element 926 is arranged within a chamber 932 defined by a deformable wall 930, and the non-magnetic beads 924 are arranged within an open space 918 defined by a sleeve 910 and a fragile membrane 934. In another configuration, a portion of the plurality of non-magnetic beads 924 may be included together with the magnetic element 926 within the chamber 932, or the magnetic element 926 may be included within the open space 918.

[0721] like Figure 77 As shown, the deformable wall 930 may include vents 938, which may be covered by a porous venting membrane 940 attached to the lower surface of the deformable wall 930, surrounding a portion of the vents 938. Figure 77As shown, the ventilated membrane 940 can be fixed to the lower (inner) surface of the deformable wall 930, or the ventilated membrane 940 can be fixed to the upper (outer) surface of the deformable wall 930. Suitable materials for the membrane 940 include Traketch® PET / PET 0.2 Vent R300, part number 063390 from SABEU GmbH & Co. KG of Nooteheim, Germany, which comprises a 23 µm thick PET membrane material with 60 g / m³. 2 The non-woven PET backing has a pore size of 0.2 ± 0.4 µm and a pore density of 320 ± 50 x 10⁻⁶. 6 / cm 2 The total thickness is 140 ± 50 µm. The vent membrane 940 covering the vent 938 of the beaded delivery cap 900 preferably allows air to escape from the sample chamber W1 through the vent 938, but does not allow liquid to pass through.

[0722] The cap body of the bead delivery cap 900 can be an integral structure (i.e., a single piece) made of a flexible polymer material. For example, the polymer material of the bead delivery cap 900 can be formed of a thermoplastic elastomer material.

[0723] Figure 79 This is a cross-section through the sample chamber W1 of the cartridge body 502, showing a bead delivery cap 900 inserted into the sample chamber W1. After the sample is dispensed into the chamber W1, the bead delivery cap 900 is inserted into the sample chamber W1. A sleeve 910 is inserted into the sample chamber W1; for this purpose, the sleeve 910 may be tapered, and a radial shoulder 920 contacts the top edge surface 514 of the wall of the sample chamber W1. The sleeve 910 is sealingly engaged with the inner surface 515 of the wall of the sample chamber W1. Sealing ribs 914a, 914b contact the inner surface 515 to enhance the seal between the sleeve 910 of the bead delivery cap 900 and the wall of the sample chamber W1. The sleeve 910 may be frictionally engaged with the inner surface 515 of the wall of the sample chamber W1, or the sleeve 910 may be coupled to the wall of the sample chamber W1 by means of mating threads (not shown) on the sleeve 910 and the wall of the sample chamber W1.

[0724] Filter element 942 and / or porous membrane 944 can be provided in the bottom of sample chamber W1 above outlet port H1. The porous membrane 944 can be attached to the bottom of sample chamber W1, for example, by adhesive, heat sealing, or ultrasonic welding. The porous membrane 944 can be a mesh or filter matrix, and is preferably hydrophilic (naturally hydrophilic or treated to be hydrophilic). Suitable materials include polyamide, polypropylene, polyethylene terephthalate (PETP), ethylene tetrafluoroethylene (ETFE), or polyether ether ketone (PEEK). The porosity (pore size) of the porous membrane 944 can be, for example, from 30 µm to 100 µm, such as about 70 µm. The pore size of the porous membrane 944 should be small enough to capture lysed cellular material, but not too small to avoid easy clogging. Suitable mesh elements for the second porous membrane 620 are available from Sefar, Inc., Buffalo, NY, part number 03-70 / 33 HPL, with a pore size of 70 µm. Filter element 942 may comprise a sintered filter having the same porosity (pore size) as the porous membrane 944 (e.g., a range of 30 µm to 100 µm or approximately 70 µm). The porosity of filter element 942 may vary throughout its thickness, for example, having a pore size that gradually decreases from a larger pore size (e.g., 100 µm) at the top surface of filter element 942 to a smaller pore size (e.g., 30 µm) at the bottom surface of filter element 942.

[0725] The deformable wall 930 is configured to remain in an undeformed state when a required amount of force is applied to the deformable wall 930. Figure 77 (As shown in the image) collapsed into a deformed state ( Figure 78 (As shown in the image). In the undeformed state, such as... Figure 77 As shown, the deformable wall 930 is located above the open space 918 of the sleeve 910. In the deformed state, as... Figure 78As shown, the deformable wall 930 is recessed outward and protrudes into the open space 918 of the sleeve 910. When the deformable wall 930 deforms from its undeformed state to its deformed state, the sleeve 910 maintains its shape, ensuring a sealed engagement against the inner surface 515 of the sample chamber W1. Although the vent 938 allows gas to escape from the chamber 932 while the deformable wall 930 is deforming to prevent pressure buildup within the chamber 932 and thus allow the deformable wall 930 to deform more effectively, the collapsing wall 930 pushes the magnetic element 926 (and any non-magnetic bead 924 that may be contained within the chamber 932) into the non-magnetic bead 924 contained within the open space 918. Collapsing the deformable wall 930 into the open space 918 eliminates the chamber 932 and reduces the volume of the open space 918. When the combined volume of the magnetic element 926 and the non-magnetic bead 924 (i.e., the fractured bead) exceeds the remaining volume of the recess 918, the fragile membrane 934 ruptures (e.g., as shown in the image). Figure 78 (As shown in the diagram, segments 934a and 934b of the fragile membrane) further release the pyrolysis beads into the sample chamber W1. The pyrolysis beads pass through filter element 942 and / or porous membrane 944 (see...). Figure 79 The sample is retained in sample chamber W1, and sample chamber W1 functions as a lysis chamber.

[0726] To facilitate controlled rupture of the fragile membrane 934, the membrane can be configured to rupture in response to the application of a certain amount of force to the fragile membrane 934. The fragile membrane 934 may be a porous membrane and / or may be composed of easily ruptured materials such as aluminum, polymeric materials, or combinations thereof. In some embodiments, the fragile membrane 934 may include rupture lines (e.g., Figure 76 The rupture line 936 (represented by the dashed line 936) is designed to make the membrane 934 more prone (or easier) to rupture when a required force is applied. The rupture line 936 can be a scribe line or a series of perforations or other partial cuts, indentations, etc., formed in the membrane 934. The arrangement of the rupture lines 936 can be adjusted to change the required amount of force needed to rupture the fragile membrane 934. For example, the rupture line 936 can be as follows: Figure 76 The single line shown has, for example, a "U" shape or a "C" shape, or the break line may include multiple lines that may intersect each other in an "X", a cross "+" shape, or a star "*" shape.

[0727] The top of the peripheral wall 906 may be higher than the top of the deformable wall 930 to prevent the user or instrument from accidentally deforming the deformable wall 930.

[0728] The deformable wall 930 can have a thickness smaller than that of the sleeve 910, making it more easily deformable when an external force is applied. For example, the deformable wall 930 can have a thickness of 0.5 mm to 1.0 mm, and the sleeve 910 can have a thickness of 1.0 mm to 2.0 mm. The deformable wall 930 can have any shape suitable for allowing it to deform. For example, in the illustrated embodiment, the deformable wall 930 can be circular, such as generally hemispherical or dome-shaped. In other embodiments, the deformable wall 930 can have other shapes suitable for deformation, such as, for example, cylindrical.

[0729] In some implementation schemes, such as Figure 77 As shown, the bead delivery cap 900 may include a peelable bottom cover film 922 covering the outer surface of the fragile film 934. For illustrative purposes, in Figure 77 The diagram shows a bottom cover film 922 with an exaggerated thickness and a slight gap between the bottom cover film 922 and the fragile film 934. In practice, the bottom cover film will adhere to the fragile film 934 in face-to-face contact. The bottom cover film 922 is configured to peel off from the fragile film 934 without affecting the fragile film 934 or damaging the sealing connection between the bottom end 916 of the sleeve 910 and the fragile film 934. In some embodiments, the bottom cover film 922 may be composed of a material (such as aluminum) with sufficient flexibility for peeling. The user removes the bottom film 922 from the bead delivery cap 900 before the bead delivery cap 900 is inserted into the sample chamber.

[0730] In some implementation schemes, such as Figure 77 As shown, the bead delivery cap 900 may include a peelable top cover film 923 that is adhered to the top edge of the peripheral wall 906 and covers the deformable wall 930. The top cover film 923 is configured to peel off from the peripheral wall 906. In some embodiments, the bottom cover film 923 may be composed of a material (such as aluminum) that is flexible enough for peeling. The user removes the top film 923 from the bead delivery cap 900 before it is inserted into the sample chamber.

[0731] In some embodiments, the sample chamber cap may include a pouch (not shown), instead of the fragile membrane 934 or being contained within the fragile membrane 934, to enclose the pyrolysis beads (at least one magnetic element 926 and a plurality of non-magnetic beads 924) within the open space 918. The pouch may consist of a liquid-soluble film (e.g., a PVA film or a PVOH film). The pouch may be secured to the inner surface of the sleeve 910 and configured to displace from the open space 918 when the deformable wall 930 collapses into its deformed state, and configured to dissolve within the sample cell W1 prior to performing the pyrolysis operation.

[0732] In the process of loading and containing at least one magnetic element 926 and a plurality of non-magnetic beads 924 into the cap body of the bead delivery cap 900, the bottom end 916 of the lower sleeve 910 is initially open to allow loading of the pyrolysis agent (or pouch) into the chamber 932 and the open space 918. The cap body is inverted such that the open bottom end 916 faces upward, and in some embodiments, the magnetic element 926 is first loaded into the chamber 932 such that the magnetic element 926 is arranged adjacent to the inner surface of the deformable wall 930. The positioning of the larger magnetic element 926 adjacent to the deformable wall 930 allows the deformable wall 930 to more effectively deflect the smaller non-magnetic beads 924 against the fragile membrane 934.

[0733] A plurality of non-magnetic beads 924 are then loaded into the open space 918 of the cap body and partially into the chamber 932, such that the magnetic element 926 and the plurality of non-magnetic beads 924 substantially fill the chamber 932 and the open space 918. The amount (e.g., volume) of the pyrolysis beads (magnetic element 926 and / or non-magnetic beads 924) loaded into the chamber 932 and the open space 918 is adjusted to ensure that deformation of the deformable wall 930 causes the non-magnetic beads 924 to press against and apply a sufficient amount of force to rupture the fragile membrane 934. In a non-limiting example, the collective volume of the chamber 932 and the open space 918 may be 700 μl to 800 μl (e.g., about 733.5 μl), and the volume below the bead delivery cap 900 of the sample chamber W1 may be 700 μl to 800 μl (e.g., about 718.10 μl). In a non-limiting example, the pyrolysis beads 926, 924 are densely packed in the chamber 932 and the open space 918, for example occupying at least 90% of the volume of the chamber 932 and the open space 918.

[0734] After the cap body is filled with pyrolysis beads, a fragile membrane 934 is attached to the bottom end 916 of the sleeve 910 to close the open space 918 and retain the magnetic element 926 and the plurality of non-magnetic beads 924 in the chamber 932 and the open space 918. The fragile membrane 934 can be secured to the bottom end 916 by any suitable means, such as adhesive, heat sealing, or ultrasonic welding. As discussed above, the fragile membrane 934 is configured to rupture when the deformable wall 930 deforms from an undeformed state to a deformed state. When the deformable wall 930 is deformed to displace the pyrolysis beads in the chamber 932 and the open space 918, a sufficient amount of force is applied to rupture the fragile membrane 934. The amount of force required to rupture the fragile membrane 934 is greater than the amount of force that would be applied to the fragile membrane 934 by accidental contact by a user or machine. In some embodiments, the amount of force required to rupture the fragile membrane 934 by applying force to the deformable wall 930 is about 1.0 lb to about 5.0 lb. The amount of force required to break the fragile membrane 934 can be adjusted by modifying the following: (1) the size and shape of the deformable wall 930; (2) the volume of the chamber 932 and / or the open space 918; (3) the volume of the rupture beads loaded into the chamber 932 and the open space 918; and (4) the composition and structure of the fragile membrane 934, including the arrangement of one or more rupture lines 936 on the fragile membrane 934. The deformable wall 930 can be collapsed manually or by means of a bead delivery cap actuator of the instrument, as described below.

[0735] After the fragile membrane 934 is attached to the bottom end 916 of the lower sleeve 925, the bottom cover film 922 can preferably be adhered to the fragile membrane 934 using a releasable adhesive that leaves no residue. The bottom cover film 922 can be peeled off immediately before the cap 900 is inserted into the sample cell W1 to maintain shielding and protection of the fragile membrane 934 during pre-treatment disposal. By shielding the fragile membrane 934, the bottom cover film 922 helps prevent accidental breakage of the fragile membrane 934, for example, through exposure to accidental contact. In some embodiments, the bottom cover film 922 and the fragile membrane 934 can be secured together to the bottom end 221 of the sleeve 220, such that the fragile membrane 934 and the bottom cover film 922 are applied simultaneously. In some embodiments, the bottom cover film 922 can be applied to the fragile membrane 934 after the fragile membrane 934 has been secured to the bottom end 916 of the sleeve 910.

[0736] In each embodiment of the lysis capsule 600, 600ʹ, 600ʹʹ or lysis vessel 700 described above, subjecting the magnetic element 626, 626ʹ, 626ʹʹ or 726 to a magnetic field of varying polarity will cause the magnetic element to move as it continuously seeks to realign with the changing north and south poles of the changing magnetic field. This movement of the magnetic element 626, 626ʹ, 626ʹʹ or 726 will cause the non-magnetic bead 624, 624ʹ, 624ʹʹ or 724 to move accordingly within the corresponding lysis chamber 622, 622ʹ, 622ʹʹ or 722, and the movement of the non-magnetic bead within the lysis chamber will achieve mechanical lysis of the sample material contained within the lysis chamber along with the magnetic element and the non-magnetic bead.

[0737] In an alternative implementation, the lysis agent can be pre-placed in the sample chamber without requiring a separate containing capsule or container. This arrangement eliminates the need for a separate lysis capsule (e.g., one of lysis capsules 600, 600ʹ, 600ʹʹ that needs to be inserted into the sample chamber) or a sample chamber cap containing the lysis agent (such as a bead delivery cap 900 that may further require a bead delivery cap actuator).

[0738] Figure 82 This is a partial cross-section of the fluid cartridge 1000 through the mechanical pyrolysis sample chamber 1002, which provides a pyrolysis chamber and pyrolysis agent for performing mechanical pyrolysis directly within the sample chamber. Except for including the mechanical pyrolysis sample chamber 1002, the fluid cartridge 1000 may be substantially the same as the fluid cartridge 500 described herein. Like the fluid cartridge 500, the fluid cartridge 1000 may include a cartridge body made of injection-molded thermoplastic polymer material.

[0739] Generally, the mechanical pyrolysis sample chamber 1002 may include a fluid sample chamber 1009 and a pyrolysis chamber 1022. The fluid sample chamber has an open first (e.g., upper) end 1014 and a sample outlet port 1028. The pyrolysis chamber is defined by an inner wall 1018 of the fluid sample chamber 1009, an upper or first porous membrane 1030 fixed within the sample chamber 1009, and a lower or second porous membrane 1032 spaced apart from the first porous membrane 1030 and fixed within the sample chamber 1009. A plurality of non-magnetic beads 1024 and at least one magnetic element 1026 are arranged within the pyrolysis chamber 1022. The pore sizes of the first porous membrane 1030 and the second porous membrane 1032 are designed to retain the plurality of non-magnetic beads 1024 and at least one magnetic element 1026 within the pyrolysis chamber 1022.

[0740] More specifically, in a non-limiting example, the mechanical lysis sample chamber 1002 may include a fluid sample chamber 1009, which is composed of a lower section 1008, an intermediate section 1006 wider than the lower section 1008, and an upper section 1004 wider than the intermediate section 1006. The open upper end 1014 of the sample chamber 1002 is surrounded by the inner wall 1016 of the upper section 1004. The sample chamber 1002 may include an inclined transition section 1012 between the intermediate section 1006 and the lower section 1008. A sample outlet port 1028 extends from the lower section 1008 to connect the mechanical lysis sample chamber 1002 to a syringe barrel (such as...) Figure 82 Channel 1029 of the syringe body (not shown). An inclined transition surface 1027 may be provided between the lower section 1008 and the sample outlet port 1028.

[0741] A transverse ledge 1010 between the upper section 1004 and the intermediate section 1006 supports a first porous membrane 1030 that overlaps with the open upper end 1014. The first porous membrane 1030 can be press-fitted into the upper section 1002, wherein the outer periphery of the first porous membrane 1030 is interference-fitted with the inner wall 1016 of the upper section 1002. For this purpose, the first porous membrane 1030 can be a compressible material that is slightly wider than the upper section 1004 and has a thickness sufficient to allow the material to be press-fitted into the upper section 1002.

[0742] The lower or second porous membrane 1032 overlaps with the sample outlet port 1028 and the inclined transition surface 1027, and can be press-fitted into the lower section 1008, wherein the outer periphery of the second porous membrane 1032 is interference-fitted with the inner wall 1020 of the lower section 1008. For this purpose, the second porous membrane 1032 can be a compressible material that is slightly wider than the width of the lower section 1008, and has a thickness sufficient to allow the material to be press-fitted into the lower section 1008.

[0743] The lysis chamber 1022 is defined by an inner wall 1018 between a first porous membrane 1030 and a second porous membrane 1032 in the intermediate section 1006. The lysis chamber 1022 includes a plurality of non-magnetic beads 1024 and a magnetic element 1026, the plurality of non-magnetic beads being identical to the non-magnetic beads 624 described above, thereby filling a portion of the volume of the lysis chamber 1022, and the magnetic element being identical to the magnetic element 626 described above. A fluid sample is supplied to the lysis chamber 1022, and the magnetic element 1026 is agitated as described below to impart movement to the non-magnetic beads 1024, thereby achieving mechanical lysis of cells present in the sample contained within the lysis chamber 1022. The relative volumes of the lysis chamber 1022, the non-magnetic beads 1024, and the magnetic element 1026 can be as described above for the lysis chamber 622, the non-magnetic beads 624, and the magnetic element 626.

[0744] The first porous membrane 1030 can be positioned at a certain distance below the top edge 1015 of the chamber 1002, such as... Figure 82 As shown in the diagram, a portion above the first porous membrane 1030 of the upper section 1004 (the top space) can provide a sample loading reservoir to retain a volume of fluid sample as it passes through the first porous membrane 1030. The volumes of the upper section 1004 and the lysis chamber 1022 will depend on the volume of the sample to be processed in the mechanically lysed sample chamber 1002. In a non-limiting example, the volume of the upper section 1004 is about 1000 µL, and the volume of the lysis chamber 1022 can be about 800 µL. In a non-limiting example, the volumes of the non-magnetic beads 1024 and the magnetic element 1026 can be about 200 μl to 300 μl (the volume of the magnetic element 1026 can be about 64 μl (4 mm × 4 mm × 4 mm)), thus leaving about 500 μl to 600 μl of fluid in the lysis chamber 1022. After the sample is dispensed into chamber 1002, the chamber can be closed by inserting a cap (such as cap 516 described herein) into the open upper end 1014 of the upper section 1004 or by securing a cover or other covering to the open upper end 1014. The distance between the first porous membrane 1030 and the top edge 1015 can be varied by changing the position of the shelf 1010, depending on factors such as the desired volume of the lysis chamber 1022, the desired volume of the sample loading reservoir, and / or the shape factor of the covering used to close the open upper end 1014.

[0745] Figure 83This is a partial cross-section of the fluid cartridge 1000 passing through the alternative mechanical pyrolysis sample chamber 1002ʹ. The mechanical pyrolysis sample chamber 1002ʹ is substantially identical to the mechanical pyrolysis sample chamber 1002ʹ, except that it uses a heat sealer having a size and shape corresponding to the first porous membrane 1030 to heat seal the first porous membrane 1030 to the ledge 1010ʹ, and a heat sealer having a size and shape corresponding to the second porous membrane 1034 to heat seal the second porous membrane 1034, which may be a post-filter, to the ledge 1038 arranged above the lower section 1008. To facilitate heat sealing, the ledges 1010ʹ and 1038 may respectively include energy directors 1036 and 1040. An energy director is a component or feature in a heat sealing application that helps focus and control the flow of energy to the area where a seal is being created. Examples of energy directors include raised features (e.g., ribs) protruding above ledges 1010ʹ and 1038 to form narrow edges (e.g., dome-shaped or blade-shaped (triangular) sections), which will concentrate energy at the edges and promote localized melting of the material at the edges to facilitate a seal. The heat-sealing process melts the energy directors 1036 and 1040, and the molten plastic penetrates the first porous membrane 1030 and the second porous membrane 1034, respectively, thereby forming a strong bond upon cooling and hardening. For this application, a woven mesh is well-suited for use as the first porous membrane 1030 and the second porous membrane 1034 because the open mesh will allow the penetration of the molten plastic.

[0746] Can be combined Figure 82 and Figure 83 Variations are shown in the diagram. For example, the first porous membrane 1030 may be press-fitted and the second porous membrane 1034 may be heat-sealed to the shelf 1038, or the first porous membrane 1030 may be heat-sealed and the second porous membrane 1032 may be press-fitted into the lower section 1008. Some variations may include both the porous membrane 1034 heat-sealed to or otherwise supported on the shelf 1038 and the porous membrane 1032 disposed within the lower section 1008 of the sample chamber. For variations having only one second porous membrane 1034 heat-sealed to the shelf 1038, the lower section 1008 of the sample chamber may be omitted.

[0747] Figure 84This is a partial cross-section of the fluid chamber 1000 passing through the alternative mechanical lysis sample chamber 1002ʹʹ. The mechanical lysis sample chamber 1002ʹʹ is substantially identical to the mechanical lysis sample chamber 1002, except for the protective layer 1042 disposed below the first porous membrane 1030 and the protective layer 1044 disposed above the second porous membrane 1032. The first porous membrane 1030 and the second porous membrane 1032 must remain structurally intact throughout the mechanical lysis process. If either filter is prone to damage, protective layer 1042 or protective layer 1044 may be added. Each of the protective layers 1042 and 1044 may be a mechanically robust supporting mesh (e.g., a woven nylon or polyester mesh) or a structure (such as an injection-molded or 3D-printed mesh), and may have a porosity at least as large as that of the first and second porous membranes, and up to about 300 µm to 350 µm, with the maximum size limited by the size of the pyrolysis beads within the pyrolysis chamber 1022.

[0748] The protective layer 1042 can be used in conjunction with a first porous membrane 1030 press-fitted into the upper section 1004, or with a first porous membrane 1030 heat-sealed to a wall shelf 1010ʹ. For heat-sealing applications, the protective layer 1042 can be attached to the first porous membrane 1030, for example, by an adhesive, such that molten plastic penetrating the pores of the protective layer 1042 secures both the protective layer 1042 and the first porous membrane 1030 to the wall shelf 1010ʹ. Similarly, the protective layer 1044 can be used in conjunction with a second porous membrane 1032 press-fitted into the lower section 1008, or with a second porous membrane 1034 heat-sealed to a wall shelf 1038. For either application, the protective layer 1044 can be placed on top of or on the second porous membrane 1032, wherein the protective layer is held in place by gravity. Alternatively, before or after press-fitting the second porous membrane 1032 into the lower section 1008, or before or after heat-sealing the second porous membrane 1034 to the wall shelf 1038, a protective layer 1044 may be attached to the second porous membrane 1032 or the second porous membrane 1034, for example, by an adhesive.

[0749] The pore size of the first porous membrane 1030 for mechanical lysis sample chambers 1002, 1002ʹ, and 1002ʹʹ should be larger than the pore size of the second porous membrane 1032 or 1034. The first porous membrane 1030 should be large enough to allow pathogens (such as 40 µm parasites) to pass through, and the second porous membrane 1032 or 1034 should provide sufficient filtration to prevent downstream blockage. The pore size of both the first and second porous membranes should be small enough to retain the lysis beads within the lysis chamber 1022.

[0750] The first porous membrane 1030 may be a mesh or a filter matrix, and is preferably hydrophilic (naturally hydrophilic or treated to be hydrophilic) to facilitate the passage of fluid sample material through the first porous membrane 1030. Suitable materials include polyamide, polypropylene, polyethylene terephthalate (PETP), ethylene tetrafluoroethylene (ETFE), or polyether ether ketone (PEEK). Other suitable materials include woven mesh filters (such as nylon or polyester or similar) or membrane filters. The porosity (pore size) of the first porous membrane 1030 may be, for example, from 70 µm to 500 µm (e.g., about 300 µm), with the maximum size limited by the size of the pyrolysis beads to be retained within the pyrolysis chamber 622. Suitable mesh for the first porous membrane 1030 is available from Sefar, Inc., Buffalo, NY, part number 03-50 / 37, with a pore size of 50 µm, and suitable filters for the first porous membrane 1030 are available from Porex Filtration Group, product number 4899 or 3677.

[0751] The second porous membrane 1032 or 1034 can be a mesh or a filter matrix, and is preferably hydrophilic (naturally hydrophilic or treated to be hydrophilic). Suitable materials include polyamide, polypropylene, polyethylene terephthalate (PETP), ethylene tetrafluoroethylene (ETFE), or polyether ether ketone (PEEK). The porosity (pore size) of the porous membrane 944 can be, for example, from 30 µm to 100 µm, such as about 70 µm. The pore size of the porous membrane 944 should be small enough to capture lysed cellular material, but not too small to avoid easy clogging. Suitable meshes for the second porous membrane 1032 or 1034 are available from Sefar, Inc., Buffalo, NY, part number 03--1 / 1, with a pore size of 1 µm. Alternatively, the second porous membrane 1032 or 1034 may comprise a sintered filter having a porosity (pore size) in the range of 30 µm to 100 µm or about 70 µm. The porosity of the second porous membrane 1032 or 1034 may vary throughout its thickness, for example, having a pore size that gradually decreases from a larger pore size (e.g., 100 µm) at the top surface of the second porous membrane 1032 or 1034 to a smaller pore size (e.g., 30 µm) at the bottom surface of the second porous membrane 1032 or 1034. Suitable filters for the second porous membrane 1032 or 1034 are available from Sterlitech PES 0.65 – 1.2 µm.

[0752] Internal control

[0753] In some applications, such as when performing molecular assays on a testing platform (e.g., a fluidic chamber 500, 800, or 1000), it may be desirable to combine an internal control with the reaction mixture. The internal control (e.g., nucleic acids (DNA and / or RNA), such as nucleic acid transcripts, plasmids, or nucleic acids extracted from a whole organism (e.g., yeast)) will be exposed to the same assay conditions as the sample, such as lysis (in the case of a whole organism containing the internal control or a ruptureable encapsulated pellet containing the internal control, as described below), sample purification, combination with amplification reagents and / or detection probes, thermal cycling, emission signal detection, etc. The internal control nucleic acid may respond to the same amplification reagents as the sample target, but a different detection probe will be provided to bind the internal control nucleic acid and the sample target to distinguish between them. If the amplification and detection procedures are performed correctly, a signal indicating the presence of the internal control (i.e., a positive result for the internal control nucleic acid) can be expected. On the other hand, failure to detect a signal indicating the presence of an internal control (i.e., a negative result for the internal control nucleic acid), or detection of less internal control than expected, can indicate an error or malfunction in one or more steps of the sample preparation process (e.g., lysis or analyte purification), material delivery, amplification, and / or detection. Such errors or malfunctions can be system-based (e.g., the instrument or a module within the instrument has malfunctioned) and / or material-based (e.g., one or more reagents and / or probes have become unstable). Therefore, internal controls are provided to validate assay results and / or to validate the effectiveness of the cell lysis protocol, for example, to confirm that all steps of the assay (including extraction, amplification, and detection) have been performed as intended.

[0754] One way to introduce an internal control into the reaction mixture is to simply dispense a certain amount of the reagent containing the internal control (“internal control reagent” or “ICR”) together with the sample into sample chamber W1. Alternatively, the internal control reagent can be pre-positioned in the cartridge such that it binds to the sample after it has been dispensed into the sample chamber, eliminating the need for a technician to add the internal control to the sample before or at the time the sample is introduced into the fluid cartridge.

[0755] On-plate mechanical lysis provides flexibility in how internal controls are added to the reaction mixture by offering additional mechanisms for introducing internal controls. Internal controls can be provided in non-liquid form, with dried reagents being an example. For instance, a fluid form of the internal control reagent can be applied to and dried on a portion of a lysis capsule, lysis vessel, or sample chamber, such that when that portion of the lysis capsule, lysis vessel, or sample chamber comes into contact with a fluid sample, the internal control reagent will dissolve and bind to the fluid sample. For applications not involving on-plate mechanical lysis, a fluid form of the internal control reagent can be applied to and dried on a portion of the sample chamber, such that when that portion of the sample chamber comes into contact with a fluid sample, the internal control reagent will dissolve and bind to the fluid sample.

[0756] More specifically, for applications involving mechanical lysis capsules 600, 600ʹ, 600ʹʹ on a plate, a fluid form of internal control reagent can be applied to and dried on first porous membranes 618, 618ʹ, 618ʹʹ, respectively (sometimes referred to as plaque formation), such that when a fluid sample is dispensed into the capsules through the first porous membrane, the dried internal control reagent dissolves and is “washed” from the first porous membrane and combined with the fluid sample entering the lysis chambers 622, 622ʹ, 622ʹʹ. Alternatively or additionally, a fluid form of internal control reagent can be applied to and dried on second porous membranes 620, 620ʹ, or 620ʹʹ of the lysis capsules 600, 600ʹ, 600ʹʹ.

[0757] For the lysis capsule 700, an internal control reagent in fluid form can be applied to the porous membrane 720 and dried thereon into a non-liquid form. When the fluid sample is dispensed into the sample chamber W1 and the capsule 700 is inserted into the sample chamber W1, the sample will pass through the porous membrane 720, and the dried internal control reagent will dissolve and combine with the fluid sample entering the lysis chamber 722.

[0758] For mechanical lysis processes performed directly within a sample chamber without a lysis capsule (e.g., mechanical lysis sample chambers 1002, 1002ʹ, 1002ʹʹ), an internal control reagent in fluid form can be applied to and dried on a first porous membrane 1030, such that when a fluid sample is dispensed into the sample chamber through the first porous membrane, the dried internal control reagent dissolves and is “washed” from the first porous membrane and combined with the fluid sample entering the lysis chamber 1022. Alternatively or additionally, an internal control reagent in fluid form can be applied to and dried on a second porous membrane 1032 or 1034.

[0759] Alternatively or additionally, the internal control reagent may be applied to and dried on the nonmagnetic beaded parts 624, 624, 624, 624, of the lysis capsules 600, 600ʹ, 600ʹʹ, the nonmagnetic beaded part 724 of the lysis capsule 700, the nonmagnetic beaded part 924 of the beaded delivery cap 900, or the nonmagnetic beaded part 1024 of the mechanical lysis sample chambers 1002, 1002ʹ, 1002ʹʹ. When a fluid sample is introduced into the lysis chambers 622, 622, 622, 622, 722 of the lysis vessel 700, the mechanically lysed sample chambers 1002, 1002, 1002, 1022, or when the non-magnetic bead 924 with the bead delivery cap is released into the sample chamber W1, the dried internal control reagent dissolves and (particularly when the non-magnetic bead moves throughout the lysis chamber via a magnetic element) dissolves from the non-magnetic bead and is combined with the fluid sample.

[0760] Alternatively or additionally, the internal control reagent may be applied to the inner walls of the hollow bodies 602, 602ʹ, 602ʹʹ, 702 of the lysis capsules / containers 600, 600ʹ, 600ʹʹ, 700, respectively, or applied to and dried on the inner walls of the lysis chamber 1022 of the mechanically lysed sample chambers 1002, 1002ʹ, 1002ʹʹ, respectively, such that when fluid samples are introduced into the lysis chambers 622, 622ʹ, 622ʹʹ, 722, 1022, respectively, the dried internal control reagent dissolves and is “washed” from the inner walls and combined with the fluid samples in the lysis chambers 622, 622ʹ, 622ʹʹ, 722, 1022.

[0761] Alternatively or additionally, the internal control may be embedded in or contained in internal control pellets (microspheres) or capsules adapted to dissolve in the presence of a fluid sample or to disintegrate upon mechanical lysis (e.g., by bead milling) to release the internal control nucleic acid into the sample. In the case of releasing the internal control by disintegrating the internal control pellets through mechanical lysis, the internal control pellets are subjected to the same shear-impact forces exerted by a lysis agent (e.g., the non-magnetic beads or other lysis agents described above), and the nucleic acid released from the pellets can be used to verify the assay results against a process used to confirm that mechanical lysis has occurred, since the internal control nucleic acid, if released from the lysed internal control pellets, is nearly ready for amplification and detection. In this respect, the use of internal control pellets to verify the lysis process is not limited to the specific lysis system or process described herein, thereby imparting motion to a magnetic element surrounded by multiple non-magnetic elements by a varying magnetic field. Internal control pellets configured to release nucleic acids when subjected to shear-impact forces imposed by a mobile lysing agent can be used to verify the lysis process in any system or process in which cells are disrupted (i.e., lysed) by shear-impact forces imposed by a mobile lysing agent, regardless of what such lysing agents are or how they impart motion to the lysing agents.

[0762] Figure 85This is a schematic cross-sectional view of a coated microsphere or capsule 750 (“IC microsphere”) containing an internal control. The IC microsphere 750 includes a core 752 comprising nucleic acid (e.g., DNA and / or RNA) 754 embedded in an excipient 756 and surrounded by a coating 758. Each IC microsphere may be generally spherical with a diameter of about 0.5 mm to 1 mm, or rod-shaped with a length of about 1 mm. Desired performance parameters for the IC microsphere may include maintaining the stability of the internal control nucleic acid for certain specified shelf lives under certain environmental conditions, such as temperature and relative humidity. For example, the design parameters for the IC microsphere may include maintaining the stability of the internal control nucleic acid for up to 18 months in an environment of 70% to 95% relative humidity at 30ºC and / or during exposure to uncontrolled product transport (e.g., 90 hours at 55ºC). Other desired design parameters for the IC microsphere 750 may include the ability of the microsphere, and particularly the protective coating 758, to withstand mechanical stress during product transport. It is also expected that the coating 758 can be mechanically cleaved (i.e., at least partially destroyed or fragmented to expose the core 752) within a short time range (e.g., less than or equal to five minutes) during an on-plate cleavage procedure, and that the core 752 will dissolve upon exposure to a liquid sample after coating cleavage to release (multiple) internal control nucleic acids. In some applications, it is expected that the coating 758 can be chemically cleaved (i.e., at least partially destroyed or fragmented to expose the core 752) upon exposure to chemical cleavage conditions (such as solutions with a pH greater than 7 (e.g., between 7.5 and 8.5)) or other chemical cleavage conditions. Excipients are adapted to at least partially dissolve upon exposure to a fluid after coating cleavage to release internal controls. Other design parameters may include the core 752 comprising a material capable of binding IC nucleic acids as a matrix / medium / excipient, and the pellets minimizing reaction with the sample target analyte under turbulent conditions (e.g., during mechanical cleavage).

[0763] In a non-restrictive example, DNA internal control 754 is a plasmid internal control, and the mass ratio of the DNA internal control is 6.10E-14 (mass / microsphere). The internal control may be contained in an internal control reagent comprising molecular-level water free of nucleases. RNA internal control 754 is an in vitro transcript (IVT), and the mass ratio of the RNA internal control is 9.33E-15 (mass / microsphere).

[0764] Suitable materials for core excipient 756 include Avicel® PH101 (microcrystalline cellulose) and Klucel Fusion X™ (hydroxypropyl cellulose), available from Ashland, Inc., Wilmington, Delaware. Suitable materials for coating 758 include Aquarius™ Protect Moisture Barrier VAA (a blend of polyvinyl alcohol-based cellulose derivatives and natural waxes), also available from Ashland, Inc.

[0765] Cores 752 can be formed by wet granulation or extrusion from a “dough” formed by combining cellulose or carbohydrate powder with a liquid internal control formulation. Individual core pellets can be formed by spheroidization (also known as pelletizing), which is a process of shaping the extrudate (output from an extruder) into small, round or spherical pellets. The extruded pellets are then dried, for example, in a fluidized bed, and a coating 758 can be applied by a spraying process, such as a top spraying process, a bottom spraying process (also known as the Wurster process or Wurster coating), or a tangential spraying process (also known as a rotor or HP coating)).

[0766] In the examples described herein, the IC microparticles 750 may be combined with non-magnetic beads 624, 624, 624, 624, respectively, within the lysis chambers 622, 622, 622, 624, of the lysis capsules 600, 600ʹ, 600ʹʹ; with non-magnetic beads 724 within the lysis chamber 722 of the lysis container 700; with non-magnetic beads 1024 within the lysis chamber 1022 of the mechanically lysed sample chambers 1002, 1002ʹ, 1002ʹʹ; or with non-magnetic beads 924 contained in the bead delivery cap 900.

[0767] Pellets with embedded nucleic acids and protective coatings (such as enteric protective coatings) can be used to provide a stable form of nucleic acid for delivery in applications other than as internal controls to validate assay results or to validate the effectiveness of lysis protocols. For example, nucleic acid-coated pellets can be used for oral delivery of nucleic acids, wherein the pellet coating is adapted to dissolve upon exposure to a liquid environment or a liquid environment at a specific pH level (such as the acidic environment of the stomach (low pH) or the hypoacidic environment of the small intestine (high pH)).

[0768] Manufacturing fluid boxes

[0769] The following description presents an example of a method for manufacturing a fluid cartridge (such as fluid cartridge 500) containing a rupture capsule (such as one of rupture capsules 600, 600ʹ, 600ʹʹ). Figure 64A flowchart illustrating an embodiment of method S840 for manufacturing a fluid cartridge is shown. In various embodiments, some of the method steps shown may be performed simultaneously in a different order than shown, or may be omitted. Additional method steps may also be performed as needed. The workflow begins at step S842.

[0770] As in step S842, a hollow body with open first and second ends (e.g., a hollow body 602 with open ends 608, 614) is provided. Figure 50 ), with an open end 608ʹ, 614ʹ and a hollow body 602ʹ ( Figure 51 ), or a hollow body 602ʹʹ with open ends 608ʹʹ, 614ʹʹ ( Figure 52 To assemble the ruptured cystic components.

[0771] Step S844 includes providing first and second porous membranes, such as first and second membranes 618, 620, first and second membranes 618ʹ, 620ʹ, and first and second membranes 618ʹʹ, 620ʹʹ. The second membrane is attached to the hollow body to cover the open second end of the hollow body. For example, the second porous membrane 620 is attached to the second edge 612 of the hollow body 602 to cover the open end 614 by means of adhesive, heat sealing, or ultrasonic welding. Figure 50 The second porous membrane 620ʹ is attached to the second edge 612ʹ of the hollow body 602ʹ to cover the open end 614ʹ. Figure 51 Alternatively, a second porous membrane 620ʹʹ can be attached to the second edge 612ʹʹ of the hollow body 602ʹʹ to cover the open end 614ʹʹ. Figure 52 ).

[0772] Step S846 includes introducing non-magnetic beads into the hollow body, where they are retained by a second (lower) porous membrane. For example, non-magnetic beads 624 are introduced into the hollow body 602, where they are retained by a second porous membrane 620 covering the second ends 614. Figure 50 Non-magnetic beads 624ʹ are introduced into the hollow body 602ʹ, where they are retained by a second porous membrane 620ʹ covering the second end 614ʹ. Figure 51 Alternatively, non-magnetic beads 624ʹʹ can be introduced into the hollow body 602ʹʹ, where they are retained by a second porous membrane 620ʹʹ covering the second end 614ʹʹ. Figure 52 ).

[0773] Step S848 includes introducing at least one magnetic element into the hollow body, which is retained by a second (lower) porous membrane. For example, magnetic element 626 is introduced into the hollow body 602, wherein it is retained by a second porous membrane 620 covering the second end 614. Figure 50 ); introduce the magnetic element 626ʹ into the hollow body 602ʹ, where it is retained by a second porous membrane 620ʹ covering the second end 614ʹ ( Figure 51 Alternatively, the magnetic element 626ʹʹ can be introduced into the hollow body 602ʹʹ, where it is retained by a second porous membrane 620ʹʹ covering the second end 614ʹʹ. Figure 52 ).

[0774] Step S850 includes attaching a first porous membrane to the hollow body to cover the open first end of the hollow body and form a pyrolysis chamber. For example, the first porous membrane 618 is attached to the first edge 606 of the hollow body 602 to cover the open end 618 by means of an adhesive, by heat sealing, or by ultrasonic welding. Figure 50 ); The first porous membrane 618ʹ is attached to the first edge 606ʹ of the hollow body 602ʹ to cover the open end 618ʹ ( Figure 51 ); or attach the first porous membrane 618ʹʹ to the first edge 606ʹʹ of the hollow body 602ʹʹ to cover the open end 618ʹʹ ( Figure 52 ).

[0775] Optionally, a non-liquid internal reference reagent may be applied to the first porous membrane before step S850, to the inner surface of the hollow body before step S846, or to at least a portion of the non-magnetic beads before step S846. The internal reference reagent may be applied in liquid form to the first porous membrane, the inner surface of the hollow body, or the non-magnetic beads, and then dried. Optionally, after step S844, an internal reference capsule containing or embedded with the internal reference reagent may be introduced into the hollow body.

[0776] In step S852, the lysis capsule constructed in steps S842 to S850 is fixed in the sample chamber of the fluid cartridge, for example, in sample chamber W1 of fluid cartridge 500. In a non-limiting example, the lysis capsule (e.g., lysis capsule 600, 600ʹ, 600ʹʹ) can be press-fitted into the sample chamber W1, and in another example, the lysis capsule can be threaded into the inner surface of the sample chamber W1.

[0777] The following description presents an example of a method for manufacturing a fluid cell (such as fluid cell 1000) containing a mechanically fractured sample chamber (such as one of mechanically fractured sample chambers 1002, 1002ʹ, 1002ʹʹ). Figure 86A flowchart illustrating an embodiment of method S860 for manufacturing such a fluid cartridge is shown. In various examples, some of the method steps shown may be performed simultaneously in a different order than shown, or may be omitted. Additional method steps may also be performed as needed. The workflow begins at step S862.

[0778] In step S862, a cartridge body is provided. The cartridge body includes a sample chamber and one or more other chambers, the sample chamber having an open first end and a sample outlet port at a second end of the sample chamber, the one or more chambers being fluidly connected to or potentially fluidly connected to the sample chamber. For example, the cartridge body may include a sample chamber 1009 having an open first end 1014 and a sample outlet port 1028, and being connected to an injection cartridge body SB ( ) via a channel 1029. Figure 82 The syringe body SB Figure 82 (Not shown in the image) connection. The sample chamber 1009 may include a lower section 1008, an intermediate section 1006 wider than the lower section 1008, and an upper section 1004 wider than the intermediate section 1006. Transverse shelves 1010, 1010ʹ may be provided between the upper section 1004 and the intermediate section 1006. Figure 82 and 83 ), and the wall shelf 1038 can be arranged above the lower section 1008 ( Figure 83 ).

[0779] Step S864 includes providing a first porous membrane and a second porous membrane, for example, a first porous membrane 1030 and a second porous membrane 1032 and / or a second porous membrane 1034, and attaching one of the second membranes 1032 and 1034 to the sample chamber to cover the sample outlet port 1028. For example, the second porous membrane 1032 may be press-fitted into the lower section 1008 of the sample chamber 1009, wherein the outer periphery of the second porous membrane 1032 is press-fitted with the inner wall 1020 of the lower section 1008 to cover the sample outlet port 1028. Figure 82 Alternatively or additionally, the second porous membrane 1034 may be heat-sealed to the shelf 1038, which may include melting energy guides 1040 on the shelf 1038 to allow molten plastic to penetrate the second porous membrane 1034. Figure 83 ).

[0780] In optional step S865, a protective layer may be disposed above the second porous membrane. For example, protective layer 1044 may be disposed above the second porous membrane 1032. Figure 84The protective layer 1044 may be placed on the second membrane 1032 and / or the second porous membrane 1034 after step S864, or the protective layer 1044 may be attached to the second porous membrane 1032 and / or the second porous membrane 1034, for example by an adhesive, before or after step S864.

[0781] In step S866, non-magnetic beads are introduced into the sample chamber through the open first end and retained by the second (lower) porous membrane. For example, non-magnetic beads 1024 are introduced into the sample chamber 1009 through the open first end 1014, and they are retained by the second porous membrane 1032. Figure 82 ) or the second porous membrane 1034 ( Figure 83 It is retained in the sample chamber.

[0782] In step S868, at least one magnetic element is introduced into the sample chamber through an open first end and retained by a second (lower) porous membrane. For example, magnetic element 1026 is introduced into sample chamber 1009 through an open first end 1014, and it is retained by a second porous membrane 1032. Figure 82 ) or the second porous membrane 1034 ( Figure 83 It is retained in the sample chamber.

[0783] In step S870, the first porous membrane is attached to the sample chamber at a position overlapping the open first end of the sample chamber, such that the first porous membrane, the second porous membrane, and the inner wall of the sample chamber form a lysis chamber. For example, the first porous membrane 1030 can be press-fitted into the upper section 1004 against the wall holder 1010, wherein the outer periphery of the first porous membrane 1030 is interference-fitted with the inner wall 1016 of the upper section 1004 to overlap with the open first end 1014. Figure 82 Alternatively, the first porous membrane 1030 may be heat-sealed to the shelf 1010ʹ, which may include melting energy guides 1036 on the shelf 1010ʹ, allowing molten plastic to penetrate the first porous membrane 1030. Figure 83 ).

[0784] In an optional step S869, the protective layer may be disposed above the non-magnetic beads and magnetic elements beneath the first porous membrane. Step S869 may be performed before step S870, for example by disposing the protective layer 1042 within the upper segment 1004 before press-fitting the first porous membrane 1030 into the upper segment 1004. Figure 84 Alternatively, step S869 can be performed before step S870, for example by heat-sealing the first porous membrane 1030 to the shelf 1010. Figure 83The protective layer 1042 is attached to the first porous membrane 1030, which may include a plurality of energy guides 1036 on the melting wall 1010ʹ, allowing the molten plastic to penetrate the protective layer 1042.

[0785] Electromagnet

[0786] Figure 56 This is a perspective view of the fluid cartridge 500 supported in the cartridge holder 412 of the instrument 10, which will be described in more detail below. A variable magnet (e.g., an electromagnet) is housed within the magnet housing 450 of the cartridge holder 412, adjacent to the sample chamber located below the cap 516. Figure 57 This is a schematic diagram showing an electromagnet 452 within a magnet housing 450 and connected to a circuit 454 (e.g., an oscillating circuit), the circuit being configured to (e.g., with a sinusoidal wave) alternate the current to the electromagnet 452 to repeatedly change the polarity of the electromagnet 452. A fluid cartridge 500 is positioned within a cartridge holder such that the sample chamber W1, the pyrolysis chamber 622, and the magnetic element 626 are as close as possible to the electromagnet 452, such that the magnetic element 626 will be affected by the magnetic field generated by the electromagnet 452.

[0787] The characteristics of electromagnet 452 are as follows: Figures 58 to 60 As shown in the diagram. The electromagnet or electromagnet assembly 452 includes a coil 460, a can-shaped cylinder 456, and a can-shaped base plate 458 connected to a circuit 454. The coil 460 includes a core 462 surrounded by a wire winding 464. The core 462 may be made of a material with high permeability and low conductivity to allow current to flow through the core 462 while minimizing thermally induced eddy currents. Suitable core materials include laminates or powdered metals commonly used in electromagnet applications, such as the 200C series mix-70 iron powder core available from Micrometals, Inc. (Anaheim, California). The wire winding 464 may include conductive materials such as copper, or 28 US gauge "AWG" magnet wire, acrylic varnish. The can-shaped cylinder 456 and the can-shaped base plate 458 may be formed of annealed 410 stainless steel. An opening 466 may be formed in the can-shaped cylinder 456 to receive a thermostat switch (not shown) for use in closing the electromagnet 452 if it overheats. The can-shaped cylinder 456 and the can-shaped base plate 458 together form a housing that partially encloses the coil 460, leaving only one end of the coil 460 facing the fluid box 500 exposed to contain and amplify the magnetic field emitted from the coil 460.

[0788] In a non-limiting example, a Hall effect sensor (not shown) may be positioned adjacent to the fluid cell 500—for example, within the cell holder 412 below the sample chamber W1—to confirm that an oscillating magnetic field is being generated and that the magnet 426 is moving. In another non-limiting example, the Hall effect sensor will detect two magnetic fields: an oscillating electromagnetic field with a regular signal (such as a sine wave) from the electromagnet 452, and a magnetic field from the magnetic element 626 that can have a larger signal (if the magnetic element 626 in the sample chamber W1 is closer to the sensor than the electromagnet 452) and a more chaotic signal (due to the chaotic movement of the magnetic element 626). By detecting both magnetic fields, the Hall effect sensor confirms that the electromagnet 452 is operating and that the magnetic element 626 is present and moving.

[0789] Figure 57 The examples shown refer to those corresponding to Figure 49 and Figure 50 The components of the embodiment (i.e., the pyrolysis capsule 600). However, it should be noted that the following description of the interaction between the electromagnet and the magnetic element in the pyrolysis chamber applies to any embodiment of the pyrolysis capsule, pyrolysis container, or bead delivery cap described herein.

[0790] In the context of this disclosure, "close proximity" means that the electromagnet and the magnetic element are close enough that a change in the polarity of the electromagnet causes a desired movement of the magnetic element. The degree of proximity considered "close" can vary with the strength of the electromagnet, the strength of the magnetic element, the thickness of the sample chamber wall, the thickness of the magnet housing wall, and the materials of the magnet housing and the sample chamber. In a non-limiting example, the electromagnet 452 is spaced approximately 1.5 mm from the sample chamber W1 and approximately 7 mm from the surface of the magnetic element 626. The sample chamber W1 has a diameter of approximately 13 mm, and the wall of the sample chamber is approximately 0.5 mm thick.

[0791] Electromagnet 452 may include at least one individual electromagnet, for example, driven by a switching amplifier. The frequencies of electromagnet 452 and oscillation circuit 454 (i.e., the frequencies at which the electromagnet reverses its polarity) may be in the range of 60 Hz to 200 Hz or 20 Hz to 120 Hz, and the driving voltage of the oscillation circuit may be in the range of 10 V to 50 V.

[0792] Protective ventilation mask

[0793] refer to Figure 9 and including detail A Figure 10The protective vent 560 may include at least two components: a vent membrane 562, which is hermetically sealed to the top of the cartridge body 502 to cover chambers W1 to W12 of the sample preparation section 504; and a protective cover 566, which is thermally laminated to the top surface of the vent membrane 562 and can be peeled off by the user from the vent membrane before use of the cartridge. A plunger orifice 563, formed at least in the vent membrane 562 (and optionally also provided in the protective cover 566), is positioned above the syringe barrel SB and provided with access to the syringe barrel SB via a syringe plunger. The sample chamber orifice 561 is aligned with the sample chamber W1 and is covered by the protective cover 566 until the protective cover 566 is removed to allow access to the sample chamber W1. Thus, the protective co...

Claims

1. A lysis capsule for performing a cell lysis procedure on a fluid sample, wherein the lysis capsule comprises: A hollow body having an open first end and an open second end; A first porous membrane is attached to the body, the first porous membrane covering the open first end; A second porous membrane is attached to the body, the second porous membrane covering the open second end, wherein the hollow body defines a pyrolysis chamber between the first porous membrane and the second porous membrane; Multiple non-magnetic beads are arranged in the pyrolysis chamber; as well as At least one magnetic element is arranged in the pyrolysis chamber, wherein the pore sizes of the first porous membrane and the second porous membrane are designed to retain the plurality of non-magnetic beads and the at least one magnetic element in the pyrolysis chamber.

2. A fluid cartridge, comprising: The box body includes a sample chamber having an open top. as well as The lysis capsule according to claim 1 is arranged in the sample chamber.

3. A method for processing cells contained in a fluid sample, the method comprising: (a) Dispensing the fluid sample into the sample chamber of the fluid cartridge according to claim 2, so as to at least partially fill the lysis chamber of the lysis capsule with the fluid sample; (b) After (a), cover the open top of the sample chamber with a cap; as well as (c) Following (b), the at least one magnetic element is subjected to a magnetic field, thereby causing the at least one magnetic element to move within the lysis chamber of the lysis capsule. The movement of the at least one magnetic element within the lysis chamber of the lysis capsule causes the plurality of non-magnetic beads to move within the lysis chamber of the lysis capsule, and the movement of the plurality of non-magnetic beads within the lysis chamber of the lysis capsule causes the cells contained in the fluid sample within the lysis chamber of the lysis capsule to lyse and release nucleic acids.

4. A lysis container for performing cell lysis, the lysis container comprising: A lateral extension member having a vent extending therethrough; A sleeve, the sleeve hanging from the lateral extension member, wherein the bottom end of the sleeve defines an open bottom end of the container; A first porous membrane is attached to the top or bottom surface of the lateral extension member and covers the vent; a second porous membrane is attached to the bottom end of the sleeve and covers the open bottom end, wherein the bottom surface of the lateral extension member, the inner surface of the sleeve, and the first and second porous membranes define a pyrolysis chamber. Multiple non-magnetic beads contained within the pyrolysis chamber; as well as At least one magnetic element is contained within the pyrolysis chamber, wherein the size of the first porous membrane and the second porous membrane is designed to retain the plurality of non-magnetic beads and the at least one magnetic element within the pyrolysis chamber.

5. A fluid cartridge, comprising: The box body includes: Sample chamber, the sample chamber having an open top; and The lysis vessel according to claim 4 is arranged in the sample chamber.

6. A method for processing cells contained in a fluid sample, the method comprising: (a) Dispensing the fluid sample into the sample chamber of the fluid cartridge, the sample chamber having an open top; (b) After (a), insert the sleeve of the pyrolysis vessel according to any one of claims 4 into the open top of the sample chamber; as well as (c) Following (b), the at least one magnetic element is subjected to a magnetic field, thereby causing the at least one magnetic element to move within the lysis chamber of the lysis container. The movement of the at least one magnetic element within the lysis chamber of the lysis container causes the plurality of non-magnetic beads to move within the lysis chamber of the lysis container, and the movement of the plurality of non-magnetic beads within the lysis chamber of the lysis container causes the cells contained in the fluid sample within the lysis chamber of the lysis container to lyse and release nucleic acids.

7. A method of manufacturing a fluid cartridge comprising a rupture capsule, the method comprising: The ruptured cystic structure is assembled using the following steps: (a) Provide a hollow body having an open first end and an open second end; (b) Providing a first porous membrane and a second porous membrane and attaching the second porous membrane to the hollow body, the second porous membrane covering the open second end of the hollow body; (c) Introduce a plurality of non-magnetic beads into the hollow body through the open first end of the hollow body; (d) Introduce at least one magnetic element into the hollow body through the open first end of the hollow body; (e) After (c) and (d), the first porous membrane is attached to the hollow body, the first porous membrane covering the open first end of the hollow body, wherein the hollow body and the attached first and second porous membranes define a pyrolysis chamber, and wherein the size of the pores of the first and second porous membranes is designed to retain the plurality of non-magnetic beads and the at least one magnetic element in the pyrolysis chamber; as well as (f) Following (e), the lysis capsule is secured to the sample chamber of a fluid cell having a plurality of chambers in fluid communication with the sample chamber.

8. A method for lysing cells contained in a fluid sample, the method comprising: (a) Dispensing the fluid sample into a sample chamber of a fluid cartridge, the sample chamber comprising a plurality of non-magnetic beads and at least one magnetic element, wherein at least a portion of the plurality of non-magnetic beads and / or the at least one magnetic element has an internal control reagent deposited thereon, and wherein the internal control contained in the internal control reagent is provided to validate the assay results and / or to validate the effectiveness of the cell lysis protocol. as well as (b) Following (a), the fluid sample is subjected to the cell lysis procedure, which includes exposing the at least one magnetic element to a magnetic field, thereby causing the at least one magnetic element contained in the sample chamber to move, the movement of the at least one magnetic element causing the movement of the plurality of non-magnetic beads contained in the sample chamber, and the movement of the plurality of non-magnetic beads in the sample chamber causing the cells contained in the fluid sample to lyse and release nucleic acids, wherein the internal control reagent dissolves in the presence of the fluid sample, thereby releasing the internal control into the fluid sample, and wherein the movement of the at least one magnetic element and the plurality of non-magnetic beads causes the internal control contained in the dissolved internal control reagent to be distributed within the fluid sample.

9. A method for lysing cells contained in a fluid sample, the method comprising: (a) Dispensing the fluid sample into the sample chamber of the fluid cartridge, the sample chamber comprising (i) a plurality of non-magnetic beads, (ii) at least one magnetic element, and (iii) an internal control reagent contained within an internal control pellet, wherein the internal control contained in the internal control reagent is provided to validate the assay results and / or to validate the effectiveness of the cell lysis protocol. as well as (b) Following (a), the fluid sample is subjected to the cell lysis procedure, which includes exposing the at least one magnetic element to a magnetic field, thereby causing the at least one magnetic element contained in the sample chamber to move, the movement of the at least one magnetic element causing the plurality of non-magnetic beads contained in the sample chamber to move, and the movement of the plurality of non-magnetic beads in the sample chamber causing the cells contained in the fluid sample to lyse and release nucleic acids, wherein the internal control pellets dissolve upon contact with the fluid sample and / or the movement of the plurality of non-magnetic beads in the sample chamber causes the internal control pellets to break down, thereby releasing the internal control into the fluid sample, and wherein the movement of the at least one magnetic element and the plurality of non-magnetic beads causes the internal control contained in the dissolved internal control reagent to distribute within the fluid sample.

10. A fluid cartridge comprising a cartridge body defining two or more pools fluidly connected or fluidly connectable, wherein at least one of the two or more pools includes an expansion pool, and wherein the cartridge body includes a first coupling structure at least partially surrounding the expansion pool, and wherein the cartridge further includes a chamber expander attached to the cartridge body and comprising: The base has a second connecting structure located on the bottom side of the base and configured to be operatively connected to the first connecting structure to form an airtight seal between the chamber expander and the box body; An expansion chamber extending from the base, wherein the expansion chamber expands the volumetric capacity of the expansion pool by at least the volumetric capacity of the expansion chamber; Mouth portion, the mouth portion defining an opening into the interior space of the expansion chamber; And a cap, which is configured to attach to the mouth to close the opening.

11. In a fluid cartridge comprising a cartridge body defining two or more fluidly connected or fluidly connectable pools, a method for expanding the volumetric capacity of an expanded pool of the two or more pools, the method comprising: The chamber expander is secured to the housing body by connecting a first connecting structure, at least partially surrounding the expansion pool, to a second connecting structure of the chamber expander and forming an airtight seal between the chamber expander and the housing body, wherein the chamber expander comprises: The base, wherein the second connecting structure is located on the bottom side of the base; An expansion chamber extending from the base, wherein the expansion chamber expands the volumetric capacity of the expansion pool by at least the volumetric capacity of the expansion chamber; a mouth portion defining an opening for access to the interior space of the expansion chamber; and A cap, configured to attach to the mouth to close the opening.

12. A method of manufacturing a fluid cartridge, the method comprising: (a) Provide a housing body comprising two or more pools that are fluidly connected or fluidly connectable; (b) Inserting a lysis capsule into the pool opening of the first pool of the two or more pools, the lysis capsule comprising a lysis chamber and a plurality of lysis beads contained within the lysis chamber; as well as (c) Attaching a chamber expander to the housing body above the opening of the first pool, wherein the chamber expander comprises: an expansion chamber that expands the volume capacity of the first pool by at least the volume capacity of the expansion chamber; a mouth that defines the opening of the expansion chamber into the interior space of the expansion chamber; and a cap that is configured to engage with the mouth to close the opening of the expansion chamber.

13. A method for processing a fluid sample in a fluid cartridge, the fluid cartridge comprising: Two or more pools of uniform height; A chamber expander, wherein the chamber expander is fixed relative to one of the two or more pools to expand the volume capacity of the chamber expander by increasing the volume capacity of the one pool, wherein the two or more pools are fluidly connected or fluidly connectable, and wherein the method comprises: (a) Dispensing a certain amount of the fluid sample into an opening defined by the nozzle of the chamber expander, wherein the certain amount of the fluid sample completely fills the pool and at least partially fills the interior space of the chamber expander; as well as (b) Attach the cap to the mouth of the chamber expander to close the opening.

14. A beaded delivery cap for dispensing pyrolysis agent into a sample cell of a fluid cartridge, the cap comprising: A cap body, the cap body including a deformable wall defining a chamber; A lower sleeve, the lower sleeve being located below the deformable wall and defining a recess opening into the chamber; A fragile membrane is attached to the open bottom end of the lower sleeve and seals the recess and the chamber; as well as A pyrolysis agent comprising a plurality of non-magnetic beads and at least one magnetic element, wherein the pyrolysis agent is contained within the chamber and the recess in such an amount that deformation of the deformable wall causes the pyrolysis agent to rupture the fragile membrane, thereby releasing the pyrolysis agent from the chamber and the recess.

15. A fluid cartridge, comprising: The box body includes a sample chamber having an open top. as well as According to claim 14, the bead delivery cap is inserted into the open top of the sample chamber.

16. A method for lysing cells contained in a sample, the method comprising: (a) The sample is provided to the sample chamber of the fluid cartridge; (b) Insert the bead delivery cap according to claim 14 into the sample chamber such that (i) the outer surface of the lower sleeve is in a sealed engagement with the inner surface of the side wall of the sample chamber, and (ii) the fragile membrane is located above the sample in the sample chamber; (c) Applying a force to the deformable wall to deform it to the extent that the pyrolysis agent breaks the fragile membrane and is released from the chamber and the recess into the sample chamber; as well as (d) Exposing the sample and the lysis agent to a magnetic field, the magnetic field causing the at least one magnetic element to agitate the plurality of non-magnetic beads to lyse the cells contained in the sample.

17. A method for lysing cells contained in a fluid sample, the method comprising: (a) Providing the fluid sample to the sample chamber of the fluid cartridge; (b) Secure a cap to the open end of the sample chamber, the cap including a deformable wall defining a chamber containing a pyrolysis agent and sealed by a fragile membrane, the pyrolysis agent including a plurality of non-magnetic beads and at least one magnetic element, and the at least one magnetic element being larger than any one of the plurality of non-magnetic beads. (c) Apply a force to the top of the deformable wall to cause the chamber to collapse to the extent that the pyrolysis agent contained in the chamber breaks the fragile membrane, thereby releasing the pyrolysis agent from the chamber into the sample chamber; as well as (d) subjecting the fluid sample and the lysis agent to a magnetic field, the magnetic field causing the at least one magnetic element to agitate the plurality of non-magnetic beads to lyse the cells contained in the fluid sample.

18. A method for lysing cells contained in a fluid sample, the method comprising: (a) The electromagnet is positioned at a predetermined distance from the sample chamber of the microfluidic device, the sample chamber containing the fluid sample and the pyrolysis agent, the pyrolysis agent comprising a plurality of non-magnetic beads and at least one magnetic element; (b) A magnetic field is generated by the electromagnet, the magnetic field being targeted at the sample and the pyrolysis agent contained within the sample chamber; as well as (c) Reverse the polarity of the magnetic field to move the at least one magnetic element in the sample chamber in a random manner to agitate the plurality of nonmagnetic beads to lyse the cells contained in the fluid sample, thereby releasing nucleic acids from the lysed cells, wherein the electromagnet is spatially separated from the sample chamber and remains stationary, while generating the magnetic field targeting the fluid sample and the lysing agent contained in the sample chamber.

19. A fluid cartridge, comprising: The sample chamber has an open top and a sample outlet port; as well as A lysis chamber within the sample chamber, wherein the lysis chamber is defined by the inner wall of the sample chamber and a first porous membrane attached to the sample chamber, the first porous membrane overlapping the open top; And a second porous membrane, the second porous membrane being spaced apart from the first porous membrane attached to the sample chamber, the second porous membrane overlapping the sample outlet port; Multiple non-magnetic beads contained within the pyrolysis chamber; And at least one magnetic element contained within the pyrolysis chamber, wherein the pore sizes of the first porous membrane and the second porous membrane are designed to retain the plurality of non-magnetic beads and the at least one magnetic element within the pyrolysis chamber.

20. A method for lysing cells contained in a fluid sample, the method comprising: (a) Dispensing the fluid sample into the sample chamber of the fluid cartridge according to claim 19, so as to at least partially fill the pyrolysis chamber with the fluid sample; (b) After (a), cover the open top of the sample chamber with a cap; as well as (c) After (b), the at least one magnetic element is subjected to a magnetic field, thereby causing the at least one magnetic element to move in the lysis chamber. The movement of the at least one magnetic element in the lysis chamber causes the plurality of non-magnetic beads to move in the lysis chamber, and the movement of the non-magnetic beads in the lysis chamber causes the cells contained in the fluid sample in the lysis chamber to lyse and release nucleic acids.

21. A method for manufacturing a fluid cartridge, the method comprising: (a) A box body is provided, the box body including a sample chamber having an open first end and a sample outlet port at a second end of the sample chamber, and one or more chambers fluidly connected to or fluidly connectable to the sample chamber; (b) Providing a first porous membrane and a second porous membrane and attaching the second porous membrane to the sample chamber, the second porous membrane covering the sample outlet port; (c) Introduce a plurality of non-magnetic beads into the sample chamber through the open first end of the sample chamber; (d) Introduce at least one magnetic element into the sample chamber through the open first end of the sample chamber; as well as (e) Following (c) and (d), the first porous membrane is attached to the sample chamber, the first porous membrane overlapping the open first end of the sample chamber, wherein the inner wall of the sample chamber and the attached first and second porous membranes define a pyrolysis chamber, and wherein the pore sizes of the first and second porous membranes are designed to retain the plurality of non-magnetic beads and the at least one magnetic element within the pyrolysis chamber.

22. A method for performing cell lysis on a fluid sample, the method comprising: (a) Dispensing the fluid sample into a lysis chamber, wherein the lysis chamber includes a sidewall, a first porous membrane, a second porous membrane spaced apart from the first porous membrane, a plurality of non-magnetic beads contained in a space between the first porous membrane and the second porous membrane, and at least one magnetic element contained in the space, wherein the pore sizes of the first porous membrane and the second porous membrane are designed to retain the plurality of non-magnetic beads and the at least one magnetic element in the space, and wherein the fluid sample is dispensed into the lysis chamber through the first porous membrane; as well as (b) subjecting the at least one magnetic element to a changing magnetic field of varying polarity to move the magnetic element as it seeks to realign with the changing polarity of the changing magnetic field, wherein the movement of the magnetic element imparts motion to the plurality of nonmagnetic beads to achieve mechanical lysis ("cell lysis") of cells present in the fluid sample contained in the lysis chamber.

23. A pellet comprising an internal control for use in a nucleic acid amplification reaction, wherein the internal control is provided to verify assay results and / or to verify the effectiveness of a cell lysis protocol, and wherein the pellet is adapted to be destroyed upon exposure to a collision force required for cell rupture, thereby releasing the internal control.

24. An apparatus for performing cell lysis, the apparatus comprising: Fluid receiver; The receiver contains a plurality of non-magnetic beads and at least one magnetic element; as well as An internal control is disposed within a pellet located within the receptor, wherein the pellet is adapted to be destroyed upon being subjected to a force exerted by movement of the plurality of nonmagnetic beads within the receptor, thereby releasing the internal control from the pellet, and wherein the internal control is provided to verify assay results and / or to verify the effectiveness of cell lysis protocols within the receptor.

Images