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WO2025210522A1 - Système automatisé de préparation d'échantillon mis en œuvre à l'intérieur d'une cartouche jetable - Google Patents

Système automatisé de préparation d'échantillon mis en œuvre à l'intérieur d'une cartouche jetable

Info

Publication number
WO2025210522A1
WO2025210522A1 PCT/IB2025/053438 IB2025053438W WO2025210522A1 WO 2025210522 A1 WO2025210522 A1 WO 2025210522A1 IB 2025053438 W IB2025053438 W IB 2025053438W WO 2025210522 A1 WO2025210522 A1 WO 2025210522A1
Authority
WO
WIPO (PCT)
Prior art keywords
blood
chamber
cartridge
metering
blisters
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Pending
Application number
PCT/IB2025/053438
Other languages
English (en)
Inventor
Usama Ahmed Abbasi
Prakhar Jain
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Microx Labs Inc
Original Assignee
Microx Labs Inc
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Microx Labs Inc filed Critical Microx Labs Inc
Publication of WO2025210522A1 publication Critical patent/WO2025210522A1/fr
Pending legal-status Critical Current
Anticipated expiration legal-status Critical

Links

Classifications

    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01F—MEASURING VOLUME, VOLUME FLOW, MASS FLOW OR LIQUID LEVEL; METERING BY VOLUME
    • G01F23/00—Indicating or measuring liquid level or level of fluent solid material, e.g. indicating in terms of volume or indicating by means of an alarm
    • G01F23/22—Indicating or measuring liquid level or level of fluent solid material, e.g. indicating in terms of volume or indicating by means of an alarm by measuring physical variables, other than linear dimensions, pressure or weight, dependent on the level to be measured, e.g. by difference of heat transfer of steam or water
    • G01F23/28—Indicating or measuring liquid level or level of fluent solid material, e.g. indicating in terms of volume or indicating by means of an alarm by measuring physical variables, other than linear dimensions, pressure or weight, dependent on the level to be measured, e.g. by difference of heat transfer of steam or water by measuring the variations of parameters of electromagnetic or acoustic waves applied directly to the liquid or fluent solid material
    • G01F23/284—Electromagnetic waves
    • G01F23/292—Light, e.g. infrared or ultraviolet
    • G01F23/2921—Light, e.g. infrared or ultraviolet for discrete levels
    • G01F23/2922—Light, e.g. infrared or ultraviolet for discrete levels with light-conducting sensing elements, e.g. prisms
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L3/00—Containers or dishes for laboratory use, e.g. laboratory glassware; Droppers
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01F—MEASURING VOLUME, VOLUME FLOW, MASS FLOW OR LIQUID LEVEL; METERING BY VOLUME
    • G01F23/00—Indicating or measuring liquid level or level of fluent solid material, e.g. indicating in terms of volume or indicating by means of an alarm
    • G01F23/22—Indicating or measuring liquid level or level of fluent solid material, e.g. indicating in terms of volume or indicating by means of an alarm by measuring physical variables, other than linear dimensions, pressure or weight, dependent on the level to be measured, e.g. by difference of heat transfer of steam or water
    • G01F23/28—Indicating or measuring liquid level or level of fluent solid material, e.g. indicating in terms of volume or indicating by means of an alarm by measuring physical variables, other than linear dimensions, pressure or weight, dependent on the level to be measured, e.g. by difference of heat transfer of steam or water by measuring the variations of parameters of electromagnetic or acoustic waves applied directly to the liquid or fluent solid material
    • G01F23/284—Electromagnetic waves
    • G01F23/292—Light, e.g. infrared or ultraviolet

Definitions

  • this invention relates to automated sample preparation system implemented within a disposable cartridge.
  • Microfluidics is both the science which studies the behaviour of fluids through microchannels, and the technology of manufacturing microminiaturized devices containing chambers and tunnels through which fluids flow or are confined.
  • Blood testing and sample preparation procedures often require sophisticated equipment, technical expertise, and specialized laboratory settings. This presents a challenge for home patients or at low resource settings who require regular monitoring of their health conditions. Furthermore, the existing sample preparation methods involve multiple manual steps and are prone to human error during operations, readability, usability, risks in transportation and contamination risks.
  • microfluidics cartridge for use in diagnostics industry, which utilizes finger prick blood or venous blood as an input sample and performs various sample processing steps, including fluidic metering, fluidic routing, and reagent storage with an integrated biosensor.
  • An object of the invention is to provide an automated sample preparation system implemented within a disposable cartridge such that it does not require bulky storage systems.
  • Another object of the invention is to minimize the number of components requiring fabrication and assembly, resulting in a seamless manufacturing process and streamlined productization on an assembly line.
  • An additional object of the invention is to provide an automated sample preparation system implemented within a disposable cartridge such that it overcomes currently known challenges for productization of the disposable cartridge.
  • Still an additional object of the invention is to provide an automated sample preparation system implemented within a disposable cartridge such that it does not provide any contamination risk.
  • the system utilizes finger prick blood or venous blood as an input sample and performs various sample processing steps, including fluidic metering, fluidic routing, and reagent storage with an integrated biosensor.
  • the architecture of this system, enables efficient and accurate testing of blood parameters, hematology parameters such as CBC, CD4-T cells count, but not limited to, Complete Blood Cell Count (CBC) and measurement of biomarkers such as Troponin, IL-6, Procalcitonin, but not limited to, CRP, Procalcitonin, and IL-6 through ELISA and is flexible to implement other electrochemistry techniques.
  • CBC Complete Blood Cell Count
  • biomarkers such as Troponin, IL-6, Procalcitonin, but not limited to, CRP, Procalcitonin, and IL-6 through ELISA and is flexible to implement other electrochemistry techniques.
  • the invention encompasses application of this system for home monitoring of diseases, even by individuals lacking technical expertise e.g. patients themselves in haematology.
  • the present invention addresses the challenges mentioned above, in prior arts, by introducing an innovative automatic sample preparation system integrated into a disposable cartridge.
  • This system optimizes workflow of sample processing by incorporating liquid level sensing, multiple microfluidic valves, both active and passive, along with microchannels of various dimensions, a compartment for storing reagents (such as blister-packed reagents), pneumatic connectors, and motorized actuators.
  • reagents such as blister-packed reagents
  • pneumatic connectors such as blister-packed reagents
  • motorized actuators motorized actuators
  • the cartridge incorporates various elements, including but not limited to microfluidic channels of different dimensions, both active and passive microfluidic valves, metering chambers, and chambers for conducting chemistries, such as reagent mixing, sample heating, and incubation. Additionally, blister compartments are included for convenient storage of reagents.
  • the cartridge also features integrated biosensors that enable the detection and measurement of various biomarkers within the sample.
  • a flow cell sensor This sensor allows for identification, enumeration, and quantification of biological cells (or organisms like yeast and bacteria) using sensing principles such as impedance-based detection, optical detection, or fluorescence detection techniques
  • the cartridge operates in conjunction with a portable instrument that drives the necessary unit operations.
  • the instrument's functionality encompasses fluidic metering, fluidic routing, and blood dilution utilizing microfluidic valves and embedded actuators. By employing these elements, the cartridge system achieves precise and controlled sample processing.
  • the system ensures the usability of the invention for home patients, allowing them to monitor their health conditions independently.
  • the simplified operation and integration of all essential components within the cartridge eliminate the need for technical expertise in haematology.
  • a microfluidic cartridge-based blood collection and analysis system comprising: a cartridge having a linear actuator configured to be pressed to push down a cap plunger which drives blood filled inside a blood port, from a user, into a connected first metering chamber (WBC), via a microfluidic channel, and, parallelly, into a connected second metering chamber (RBC), via a microfluidic channel, in order to output metered blood, said cartridge’s bottom layer being a hydrophobic layer causing liquid meniscus surface to form a spherical shape upon contact in order to pronounce signal changes during sensing; plurality of fluid chambers including at least a first mixing chamber, a second mixing chamber, a third mixing chamber, a biosensor chamber, and plurality of blisters, said mixing chambers connected to the metering chambers via microfluidic channels, said blisters connected to metering chambers via fluidic channels, in that, o a second set of blisters configured to be pushed to dispense
  • an optical detection system disposed in a counting chamber, having optical detection windows which serve as a signal window for the measurement of liquid level detection at entrance of said counting chamber and comprising at least one laser diode and at least one PIN photodiode, the optical detection system being configured to detect a liquid level based on changes in light refraction such that when no liquid is present a flat-line electrical signal is generated and when the liquid enters the counting chamber a square pulse electrical signal is produced, o wherein the optical detection system further comprises an inclined optical window having a geometry of a right-angle prism filled with air, the inclined optical window compensating for refractive effects in a sensing region of a few microns to enhance signal strength; o wherein the optical detection system is further configured to measure a liquid meniscus formed at the optical window, the meniscus causing transient light reflection and refraction that produces a square pulse electrical signal for a short duration, the signal being used for sample volume measurement for both RBCs and white blood cells (WBCs); a
  • a pressurized first vent connected to an air solenoid valve, delivering positive pressure to drive blood from the first mixing chamber into a second RBC (third) metering chamber and waste chamber upon opening valves.
  • a counting subsystem configured to count RBCs and platelets, follows said optical detection system, in that, pressurized second vent, delivering positive pressure to drive diluted blood from the second mixing chamber into a biosensor (impedance measurement) upon opening a valve, said biosensor comprising: a fourth chamber in line with a counting chamber, configured to receive diluted blood, facilitating the counting of red blood cells (RBCs) as the diluted blood reaches said counting chamber, measurement beginning at an entry of said counting chamber and measurement ending at an exit of said fourth metering chamber; a. optical detection windows within the counting chamber, serving as signal windows for liquid level detection based on the principle of light refraction.
  • RBCs red blood cells
  • valves enable opening of a passage for transfer of diluted blook from said third mixing chamber to said optical detection system upon the application of positive pressure on a third vent connected to said third mixing chamber.
  • said cartridge comprising: a capillary tube coated with Ethylenediaminetetraacetic acid (EDTA) for collecting blood; a designated blood port for transferring the collected blood; a cap configured to be pushed until it reaches a first stopper to prevent blood flow to a downward section; a capillary valve integrated within the cap to restrict movement of the collected blood based on capillary forces and geometry of the capillary valve structure, wherein the capillary forces and the capillary valve’s geometry act in tandem to prevent blood flow through the capillary valve, thereby functioning as a reliable stopper or barrier, said geometry being defined by a channel which is narrow in its initial section and expands in its further section to create a Laplace pressure barrier to stop flow unless extra pressure is applied externally.
  • EDTA Ethylenediaminetetraacetic acid
  • an actuator activates a microfluidic first valve, that stops blood (fluid) flow from flowing into other sections, of this system, except to a designated first metering section and to a designated second metering section; a linear actuator further pushes the cap to push the blood sample into two separate metering sections:
  • ⁇ a first section is dedicated to counting of RBCs and platelets
  • ⁇ ⁇ a second section is designed for WBCs count.
  • Figure 1 illustrates, particularly, a blood port in the cartridge of this invention
  • FIG 2 illustrates blood introduction inside the blood port of Figure 1 of the cartridge of this invention
  • Figure 3 illustrates the port, of Figure 1, filled with blood sample and a capillary valve which stops blood movement into a downward section of fluid due to a capillary action and geometry of the valve;
  • Figure 5 illustrates a mechanism showing rupture of blister/s due to sharp conical pillar/s (14) placed beneath an aluminium foil, in the cartridge of this invention
  • Figure 6 illustrates the position of cap during first hard stopper
  • Figure 9 illustrates this invention’s disposable cartridge showing metering sections and buffer zone filled with blood
  • Figure 10 illustrates mixing and dilution of blood with diluent using the cartridge of this invention
  • Figure 12 illustrates blood mixing for achieving dilution ratio greater than 10000 and also Isysis of RBCs in chamber, using the cartridge of this invention
  • Figure 12 illustrates blood mixing for achieving dilution ratio greater than 10000 and also Isysis of RBCs in chamber, using the cartridge of this invention
  • Figure 13 illustrates final diluted blood pushed to an impedance sensor using the cartridge of this invention
  • Figure 14 illustrates optical detection system to monitor liquid position inside the cartridge of this invention
  • Figure 14c illustrates a top view of the cartridge showing the liquid level sensing region of this invention
  • Figure 16 illustrates leukocyte enumeration and haemoglobin detection using the cartridge of this invention
  • Figure 17 illustrates a shape memory alloy used as a linear actuator for opening and closing of the valve for the cartridge of this invention
  • Figure 18 illustrates sealing concept of an impedance sensor for the cartridge of this invention
  • Figure 19 illustrates blister protector integrated with a gripper
  • the cartridge can be broken into four sections:
  • the system comprising the blood collection and analysis subsystem further incorporates an actuator (203, Figure 22) that initially activates a microfluidic first valve (13, Figure 4), that stops blood (fluid) flow from flowing into other sections, of this system, except to a designated first metering section (11, Figure 4) and a designated second metering section (12, Figure 4). Subsequently, a linear actuator (10) further pushes the cap (7, Figure 3) to push the blood sample (5, Figure 2) into two separate metering sections: a first section (12, Figure 4) is dedicated to counting of RBCs and platelets (12, Figure 4), a second section (11, Figure 4) is designed for WBCs count (11, Figure 4).
  • the cartridge with blister-packed reagents enables controlled and precise dispensing of fluids by pressing the blisters with actuators, ensuring accurate and reliable fluid release for performing desired analytical reactions.
  • the sharp conical pillar (14) positioned within the cartridge facilitates the controlled rupture of blisters by deflecting the bottom part of the blister, allowing efficient release of fluid into the microfluidic channel for subsequent analysis.
  • Blisters are located on the cartridge. Each blister has liquid inside it.
  • the purpose of using the blister is to store the liquid and also to dispense the liquid inside the channel by applying pressure through linear actuators.
  • Third blister’s purpose is to dilute the first serially diluted blood to achieve greater dilution ratio.
  • the direct ultrasonic welding of the aluminium foil blisters onto the disposable cartridge enables efficient and rapid production, ensuring a secure and leak-proof attachment between the reagentcontaining blisters and the cartridge.
  • the integration of reagents within the disposable cartridge reduces overall reagent cost by minimizing fluid wastage, enhancing the portability and affordability of the instrument, making it suitable for resource-limited settings and enabling convenient point-of-care testing.
  • the user-friendly design of the system enables non-skilled individuals to perform self-tests at home, utilizing fingerprick blood samples, thereby facilitating convenient and accessible monitoring of blood cells for various medical applications, including chemotherapy patients requiring regular blood cell analysis.
  • the open microfluidic channel can also be thermally bonded with the COC (Cyclic olefin copolymer) and Poly styrene of thickness 100 micron to 200 microns.
  • the thin layer can be used to close the first valve (13, Figure 4) after pressing a bottom layer being a membrane wall (67, Figure 22) using small linear actuator/s (203, Figure 22) which is made of shape memory alloy.
  • the reagent storage sub-system comprises blister-packed reagents which are equipped with a blister protector (204, Figure 21), which serves to prevent blister rupture in the event of accidental user pressure.
  • a blister protector (204, Figure 21) which serves to prevent blister rupture in the event of accidental user pressure.
  • This protective mechanism minimizes risk of mishandling the microfluidic device, ensuring integrity and reliability of the system.
  • the first set of blisters (15, Figure 5) are, typically, made of aluminium foil which can be directly bonded on the disposable cartridge (1) using ultrasonic welding enabling seamless and rapid production of cartridge on an assembly line.
  • the advantage of storing the reagent in the disposable cartridge is that the reagent cost is minimized, as the bulky bottles required for automatic sample handling inside the instrument result in a significant wastage of fluid due to the larger volume needed.
  • First metering chamber is to meter WBC blood.
  • Second metering chamber is to meter RBC blood.
  • the use of ferromagnetic beads and rapid polarity changes driven by magnets mounted on motors allows for rapid and effective mixing of blood and diluent, minimizing the need for additional microfluidic valves and reducing complexity in the mixing process.
  • the inclusion of sphering reagents and fixating reagents in the dispensed liquid enhances the measurement accuracy of red blood cell volumes by transforming the cells to a spherical shape, thereby reducing impedance variations and ensuring precise assessment of cell characteristics.
  • the adjustable vent connected to a rubber gasket and controlled by an air solenoid valve, with options for atmospheric pressure or higher positive pressure, allows for regulated pressure conditions during the mixing process, ensuring optimal fluid dynamics and reliable analysis performance.
  • the system comprises a process flow and fluidic routing subsystem such that once the cartridge is loaded inside the instrument (for readings) and, thereafter, all the microfluidic valves are closed except a fourth valve (20, Figure 8) and a fifth valve (22, Figure 8) and, subsequently, the linear actuator is pressed to push down a cap plunger (7, Figure 6) which drives the blood filled inside the blood port (3) into a first metering chamber (52, Figure 9) and into a second metering chamber (53, Figure 9). Excess blood collected, inside the port, flows into a buffer zone (54, Figure 9). A first vent (48, Figure 7) is exposed to atmospheric pressure.
  • the diluent and the blood require only gentle micro mixing to preserve all cells without changing size and shape of the cells.
  • the advantage of mixing, using ferromagnetic beads, is that mixing is rapid and requires no extra microfluidic valves as in the case of back and forth mixing requiring multiple microfluidic valves actuation.
  • the dispensed liquid contains the sphering reagents and the fixating reagents also which makes the RBCs spherical in shape. This is essential since the impedance magnitude of the cells depends on the geometry of the cells, making them spherical from biconcave shape reduces the impedance variation and gives better measurement of the volume of the cells.
  • the first vent (48, Figure 7) is either exposed to atmospheric pressure or a higher positive pressure.
  • tenth valve (18) and second valve (19)] enabling the lysis of red blood cells using a lysis buffer containing formic acid and saponin as surfactants; a magnetic flea rotated at 5000 RPM to facilitate complete lysis of red blood cells upon the introduction of the metered blood and lysis buffer into the chamber; a quench buffer from fifth set of blisters (46) pushed into the third mixing chamber (35) to quench the reaction, the quenching reagent composed of sodium chloride, sodium bicarbonate, and sodium phosphate; the quench solution preserving white blood cells (WBCs) for a minimum of fifteen minutes, ensuring their stability for subsequent analysis.
  • WBCs white blood cells
  • the introduction of diluent from third set of blisters (47) into the second mixing chamber (34), followed by the lysis of red blood cells using a lysis buffer from fourth set of blisters (44), ensures the serial dilution and complete lysis of red blood cells, preparing the sample for subsequent analysis steps.
  • Fourth set of blisters (44) is the lysis buffer blister which Is connected to mixing chamber 35. These blisters are connected to the mixing chamber through a microfluidic channel and can be isolated when valve is closed.
  • the rotation of the magnetic flea at 5000 RPM within the third mixing chamber (35) enables thorough lysis of red blood cells, facilitating accurate analysis of white blood cells and ensuring reliable measurement results.
  • a quench buffer from fifth set of blisters (46) into the third mixing chamber (35) after lysis ensures the efficient quenching of the reaction, preserving the stability of white blood cells for a minimum of fifteen minutes, allowing sufficient time for subsequent analysis steps to be performed accurately.
  • Reference numeral 29 is a microfluidic valve for venting air and pressurizing the chamber 35. Quench buffer is connected to mixing chamber 35 through a microfluidic channel and this can also be isolated using valve 29. the small port close to 29 is the via which connects the quench blister to the mixing chamber.
  • the first vent (48) is pressurized with positive pressure to drive the blood from the first mixing chamber (33) into the second RBC (third) metering chamber (55, figure 11) and waste chamber (57, Figure 11) after opening the eighth valve (28, Figure 8) and seventh valve (27, Figure 8) shown in figure 11.
  • the excess of the blood from the first mixing chamber (33) flows into a waste chamber (57).
  • third set of blisters (47, Figure 7) containing diluent is pushed after opening the valves (26, 28; Figure 8) [ninth valve (26), eighth valve (28)] into the second mixing chamber (34, Figure 7) sweeping the metered blood inside the third metered chamber (55, Figure 11).
  • Reference numeral 31 is a microfluidic valve for second serial dilution.
  • fourth set of blisters (44, Figure 7) is pushed after opening the valves (19, 18) [tenth valve (19), second valve (18)] into the third mixing chamber (35) for the lysis of the red blood cells.
  • the Lysis contains formic acid and saponin as surfactant to lyse RBCs.
  • the magnetic flea is rotated at RPM of 5,000 to completely lyse the RBCs and, subsequently, a quench buffer from fifth set of blisters (46, Figure 7) is pushed into the third mixing chamber (35) to quench the reaction.
  • the composition of the quenching reagent is sodium chloride, sodium bicarbonate and sodium phosphate.
  • the quench solution preserves the WBCs for longer period of time at least for fifteen minutes.
  • Reference numeral 32 is a buffer chamber for extra blood flow.
  • Reference numeral 37 is a waste chamber as well as measurement chamber for volume dispensed for RBC count.
  • Reference numeral 38 is a second waste chamber for wash buffer volume measurement.
  • Reference numeral 39 is a third waste chamber as well as measurement chamber for volume dispensed for WBC count.
  • Reference numeral 40 is a second waste chamber for washing.
  • Reference numeral 41 is a buffer chamber to prevent overflow.
  • Reference numeral 42 is a chamber for Hemoglobin measurement.
  • Reference numeral 43 is a connection port for wash buffer blister.
  • Reference numeral 51 is a WBC metered volume.
  • Reference numeral 56 is a buffer overflow chamber.
  • the invention discloses a blood collection and analysis system comprising a cartridge (1) having: a pressurized second vent (49) ( Figure 7), delivering positive pressure to drive diluted blood from the second mixing chamber (34) into a biosensor (Impedance measurement) (36) upon opening eleventh valve (25); fourth chamber (59) and counting chamber (58) ( Figure 13) within the biosensor, facilitating the counting of red blood cells (RBCs) as the diluted blood reaches chamber (58); optical detection windows within the counting chamber (58), serving as signal windows for liquid level detection based on the principle of light refraction; an optical detection system comprising laser diodes and PIN photodiodes at each entry point of the chamber, detecting the position of the liquid through the refractive index change; a flat-line signal from the photodiodes when no liquid is present, transforming into a square pulse electrical signal as the liquid enters the entry point of the chamber, indicating the arrival of the liquid; laser diode (60) and PIN photodiode (61) (
  • Meniscus is formed inside the waste chamber.
  • the hydrophobic nature of the surface makes the surface of liquid concave towards the flow direction also the optical detection region of the measurement volume is inclined at 45 degrees.
  • the hydrophobic nature as well as this 45- degree tilt scatters more light which acts as better sensor for liquid level detection.
  • the utilization of the optical detection system employing laser diodes (60) and PIN photodiodes (61), enables efficient and accurate measurement of liquid levels within the counting chamber, ensuring precise analysis of red blood cells (RBCs) and white blood cells (WBCs) based on their respective counting principles.
  • RBCs red blood cells
  • WBCs white blood cells
  • the optical detection system is placed in front of the measurement chambers. More specifically they are located near the inclined surface which is inclined at 45 degrees.
  • the optical detection system comprises of laser diode which is placed in front of these chambers and the sensor PIN photodiode is placed opposite to it.
  • Figure 14a illustrates the angle of refraction is more when the detection region is filled with air.
  • Figure 14b illustrates the angle of refraction is minimized when the detection region is filled with liquid.
  • the system comprises a counting subsystem which is configured to count RBCs and platelets; using the system and cartridge of this invention.
  • the second vent (49, Figure 7) is pressurized to push the diluted blood from the second mixing chamber (34) into a biosensor (Impedance measurement) (36, Figure 8) after opening the eleventh valve (25, Figure 8).
  • the blood is pushed to the chambers (58, 59) [counting chamber (58), fourth chamber (59)] sequentially ( Figure 13).
  • the measurement begins at the entry of counting chamber (58) and ends at the exit of fourth metering chamber (59).
  • the counting of the RBCs starts when the diluted blood reaches counting chamber (58).
  • the counting chamber (58) has optical detection windows (207, Figure 14a and 14b) at entrance of the chamber which serve as a signal window for the measurement of liquid level detection.
  • the principle of liquid level detection is based on refraction of light. When there is no liquid, the laser beam falling on the optical window goes straight but in the presence of liquid due to change in the refractive index the light refracts. This refraction can easily be detected using PIN Photodiode.
  • the measurement is stopped once the liquid reaches the exit point of fourth metering chamber (59).
  • the optical detection system can detect entry of liquid level in counting chamber (58) and at the exit of fourth chamber (59).
  • the optical photodiode and laser are present at each entry and exit point of the chamber to detect the position of the liquid.
  • the signal from the photodiode is a flat line when there is no liquid but as the liquid enters inside the entry point of the chamber the laser beam refracts and gives a square pulse as electrical signal indicating liquid has reached.
  • Figure 14 shows a laser diode (60) and PIN photodiode (61) for measurement of the signal.
  • the liquid shown in black colour forms a meniscus with curvature which bounces most of the light from the laser diode and gives square pulse for short duration of time shown in Figure 15.
  • the same principle is adopted for WBC volume measurement.
  • Figure 14b shows a chamber which is filled with water which minimized the angle of refraction allowing more light to pass through the pin photodiode which is mounted in front of the laser diode.
  • the volume of the region where most of the laser falls has volume less than 100 nanolitre therefore measurement inaccuracy in volume is less than few hundred nanolitre.
  • Figure 14c illustrates a top view of the cartridge showing the liquid level sensing region of this invention.
  • the surface of the bottom layer of the cartridge is intentionally crafted from hydrophobic materials. This choice ensures that when liquid comes into contact with the sensing region, it exhibits distinct surface properties. Unlike conventional surfaces, the hydrophobic nature of the material causes the liquid meniscus surface to form a spherical shape (5, figure 24) upon contact. This spherical surface configuration plays a pivotal role in scattering a greater amount of light, thereby facilitating a more pronounced signal change during sensing processes.
  • the liquid sensing region is strategically inclined at an angle, as illustrated in Figures 14a and 14b.
  • This inclination serves a dual purpose, resembling a prism with angle of the prism between 30 and 60 degrees.
  • the angle of inclination is carefully selected to be greater than 30 degrees yet less than 60 degrees, optimizing its prism-like behaviour.
  • the inclined liquid sensing region functions akin to a prism, refracting light as it passes through.
  • the angle of refraction experienced by the light depends on several factors, including the wavelength of the light, the angle of the prism (in this case, the inclination of the sensing region), and the refractive index of the liquid being sensed. This prism-like behaviour enhances the interaction between light and liquid, contributing to the accuracy and sensitivity of the sensing mechanism.
  • the prism-like behaviour of the sensing region guarantees the accurate metering of both liquid and samples, while also enabling the monitoring of fluid routing in the desired direction.
  • These checkpoints are essential for ensuring precise sample preparation, and any deviations in metering accuracy or blister rupture are rigorously monitored by the liquid level sensor.
  • the measurement process for cell counting commences only when the liquid has reached the bottom sensing region of the measurement chamber, as depicted in Figure 14c. The measurement concludes when the liquid reaches the top measurement sensing region. This sequential approach ensures that the counting of cells occurs precisely within the designated measurement zones, facilitating accurate and reliable results.
  • the opening of twelfth valve (23) and thirteenth valve (24), coupled with the application of positive pressure on third vent (50), ensures controlled sample transfer from the third mixing chamber (35) to the impedance sensor, facilitating accurate leukocyte counting within the designated chambers.
  • the haemoglobin detection chamber (66), featuring an optical path length of 3mm, allows for precise absorbance measurement using a green laser or collimated green LED, enabling accurate determination of haemoglobin concentration in the blood sample.
  • the utilization of a laser diode or collimated green LED for absorbance measurement ensures accurate determination of haemoglobin concentration, enhancing the system's ability to provide reliable and precise measurement results.
  • the incubation time following the mixing of the quench into the mixing chamber 65 ensures optimal reaction and stabilization based on the saponin concentration in the lysis buffer, leading to accurate haemoglobin concentration measurement and three -part Leukocyte Count.
  • the working principle for the valve's opening and closing ensures efficient and reliable valve operation, allowing for precise control of fluid flow within the system.
  • the utilization of COC material as a membrane for valve closure provides a reliable seal, preventing unintended fluid leakage and ensuring the integrity of the system during operation.
  • FIG. 22 Another aspect of the invention is the working principle for opening and closing of the valves.
  • the working principle for the valve's opening and closing is shown in Figure 22.
  • the COC material (67, Figure 22) is used as a membrane to close the valve.
  • the shape memory alloy (203, Figure22) is actuated by applying 12V, resulting in an increase in length and directly pressing the COC membrane (67) to close the valve. When the voltage is dropped to 0, the linear actuator returns to its initial position.
  • the footprint of the shape memory alloy actuator is only 3mm, allowing multiple valves to be placed in a small footprint. It should be noted that apart from shape memory alloy valves, various other kinds of valves such as motorized motors, pneumatic valves can be used.
  • the invention discloses a blood collection and analysis system comprising a cartridge (1) having: the integration of an impedance sensor within a disposable cartridge, providing a compact and self-contained measurement solution; an impedance sensor design featuring side holes for fluid entry and exit, as illustrated in Figure 18 (36), offering improved fluid flow dynamics and optimized sensor performance; the sealing of the impedance sensor on the sides accomplished by a rubber gasket with a wedge shape (labelled as 63 in Figure 18); the placement of the rubber gasket, equipped with inlet holes, inside the slot (70) on each entry and exit side, followed by the positioning of the impedance sensor on top; the top cover (62) securely pressed and ultrasonically welded, ensuring a reliable and leak-proof flow through the system; the application of pressure during top cover placement causing the gasket to expand in all directions, effectively sealing the side holes and enhancing fluid control; the top cover featuring three holes on each side for the electrical connection of the sensor, facilitating the integration of spring connector pogo pins for reliable electrical contact; the provision of sufficient space within the
  • the impedance sensor design with side holes for fluid entry and exit offers improved fluid flow dynamics, enhancing the accuracy and reliability of measurement results.
  • the secure sealing of the impedance sensor on the sides using a rubber gasket with a wedge shape ensures a robust and leak-proof flow, preventing fluid leakage and maintaining the integrity of the system during operation.
  • the ultrasonic welding of the top cover, pressed in conjunction with the rubber gasket guarantees a reliable and durable seal, enabling consistent and accurate fluid flow through the system.
  • the integration of spring connector pogo pins for electrical connection facilitates reliable and efficient communication with the impedance sensor, ensuring precise and accurate measurement of blood samples.
  • the impedance sensor (36, Figure 18) within the disposable cartridge.
  • the impedance sensor features side openings (36, Figure 18) for the entry and exit of the fluid, as depicted in the figure 18 (36).
  • the sealing of the sensor on the sides is achieved using a rubber gasket with a wedge shape (63 in Figure 18).
  • the gasket which includes inlet holes, is placed inside the slot (70) on each entry and exit side, followed by the placement of the impedance sensor on top of it.
  • the top cover (62) is positioned and pressed for ultrasonic welding to ensure a leak-proof flow.
  • the top cover features three holes on each side for the electrical connection of the sensor, which can be achieved using spring connector pogo pins.
  • the dimension of the exposed pads inside the three holes are greater than 1mm diameter providing sufficient space for the electrical connection.
  • the biosensor is not restricted solely to impedance measurement, but it can also function as an impedance sensor, optical sensor, or both simultaneously.
  • a gripper (91, Figure 19) which is a part of the disposable cartridge having QR code or RFID code which stores the following information:
  • the cartridge is designed in such a way that it can only be inserted into the instrument in one direction.
  • the 'L'-shaped notch (90), shown in Figure 20, ensures that the cartridge can be inserted inside the instrument in only one direction.
  • the inventors present the inventive steps taken in the development of a disposable cartridge, designed for single-time use.
  • the cartridge incorporates on-board reagents, integrated sensor(s), chambers for mixing, microchannels for routing fluids (liquid and air), and a system for aligning the cartridge. It also includes a valve system that enables tight control over flow steps for fully automatic sample preparation.
  • the design aims to minimize the number of components requiring fabrication and assembly, resulting in a seamless manufacturing process and streamlined productization on an assembly line.
  • the present invention provides several notable advantages over traditional sample preparation methods.
  • the integration of multiple microfluidic valves, blister packed reagents, and actuators within the disposable cartridge streamlines the sample processing workflow, minimizing manual steps and reducing the risk of errors and contamination.
  • the portable instrument facilitates the driving of necessary unit operations, eliminating the reliance on complex laboratory setups. Consequently, the invention enables accurate and efficient blood testing, making it accessible to home patients without specialized knowledge in haematology.
  • the TECHNICAL ADVANCEMENT of this invention lies in the following:
  • the cartridge, of this invention has 4 sections: metering, liquid storage, section for running chemistry (mixing, incubation, lysis, dilution) in chambers and finally detection.
  • the entire flow is controlled with valves with feedback from liquid level sensors.

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  • Physics & Mathematics (AREA)
  • Electromagnetism (AREA)
  • Thermal Sciences (AREA)
  • Fluid Mechanics (AREA)
  • General Physics & Mathematics (AREA)
  • Health & Medical Sciences (AREA)
  • Clinical Laboratory Science (AREA)
  • Chemical & Material Sciences (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Investigating Or Analysing Biological Materials (AREA)

Abstract

La présente invention concerne un système automatisé de préparation d'échantillon mis en œuvre à l'intérieur d'une cartouche jetable. Le système utilise le sang par piqûre au doigt ou le sang veineux en tant qu'échantillon d'entrée et effectue diverses étapes de traitement d'échantillon, comprenant le dosage fluidique, le routage fluidique et le stockage de réactif avec un biocapteur intégré. L'architecture de ce système permet de tester efficacement et précisément des paramètres sanguins, des paramètres d'hématologie tels que la formule sanguine complète (CBC), le nombre de lymphocytes T CD4 -, mais sans s'y limiter, la CBC et la mesure de biomarqueurs tels que la troponine, l'IL -6, la procalcitonine, mais sans s'y limiter, la CRP, la procalcitonine et l'IL -6 par ELISA, et sa flexibilité permet de mettre en œuvre d'autres techniques d'électrochimie.
PCT/IB2025/053438 2024-04-02 2025-04-02 Système automatisé de préparation d'échantillon mis en œuvre à l'intérieur d'une cartouche jetable Pending WO2025210522A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
IN202421027377 2024-04-02
IN202421027377 2024-04-02

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WO2025210522A1 true WO2025210522A1 (fr) 2025-10-09

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PCT/IB2025/053438 Pending WO2025210522A1 (fr) 2024-04-02 2025-04-02 Système automatisé de préparation d'échantillon mis en œuvre à l'intérieur d'une cartouche jetable

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Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20150132776A1 (en) * 2012-12-17 2015-05-14 Leukodx Ltd. Kits, compositions and methods for detecting a biological condition
US20170327867A1 (en) * 2015-12-22 2017-11-16 Canon U.S. Life Sciences, Inc. Sample-to-answer system for microorganism detection featuring target enrichment, amplification and detection
US10610861B2 (en) * 2012-12-17 2020-04-07 Accellix Ltd. Systems, compositions and methods for detecting a biological condition
US20200386756A1 (en) * 2012-12-17 2020-12-10 Accellix Ltd. Systems and methods for determining a chemical state
US20230264194A1 (en) * 2020-08-14 2023-08-24 miDiagnostics NV System for analysis

Patent Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20150132776A1 (en) * 2012-12-17 2015-05-14 Leukodx Ltd. Kits, compositions and methods for detecting a biological condition
US10610861B2 (en) * 2012-12-17 2020-04-07 Accellix Ltd. Systems, compositions and methods for detecting a biological condition
US20200386756A1 (en) * 2012-12-17 2020-12-10 Accellix Ltd. Systems and methods for determining a chemical state
US20170327867A1 (en) * 2015-12-22 2017-11-16 Canon U.S. Life Sciences, Inc. Sample-to-answer system for microorganism detection featuring target enrichment, amplification and detection
US20230264194A1 (en) * 2020-08-14 2023-08-24 miDiagnostics NV System for analysis

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