WO2018085678A1 - Cartouche fluidique pour cytométrie et analyse supplémentaire - Google Patents

Cartouche fluidique pour cytométrie et analyse supplémentaire Download PDF

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Publication number
WO2018085678A1
WO2018085678A1 PCT/US2017/059965 US2017059965W WO2018085678A1 WO 2018085678 A1 WO2018085678 A1 WO 2018085678A1 US 2017059965 W US2017059965 W US 2017059965W WO 2018085678 A1 WO2018085678 A1 WO 2018085678A1
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WIPO (PCT)
Prior art keywords
sample
flow cell
fluidic
flow
μιη
Prior art date
Application number
PCT/US2017/059965
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English (en)
Inventor
Wendian Shi
Yuzhe DING
Original Assignee
Cytochip 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 Cytochip Inc. filed Critical Cytochip Inc.
Priority to CN201780080470.5A priority Critical patent/CN110177619B/zh
Priority to EP17866884.4A priority patent/EP3535056A4/fr
Priority claimed from US15/803,133 external-priority patent/US10634602B2/en
Publication of WO2018085678A1 publication Critical patent/WO2018085678A1/fr

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Definitions

  • the disclosure relates to medicine and cytometry. Background
  • Flow cytometry is a popular tool for cellular analysis of biological samples.
  • Typical cytometry analyses involve two parts.
  • the first part is sample preparation.
  • some cytometry analyses label target cells with a specific fluorophore, so that these cells can be detected by an optical measurement of fluorescence signals.
  • some cytometry analyses require selectively lysing cells in samples, leaving only target cells intact for cytometry measurement.
  • the second part is sample analysis.
  • the sample stream is focused into a narrow stream when flowing through a flow cell, where the target cells are measured one by one for optical or other signals. This narrow sample stream is usually obtained by hydrodynamic focusing of sheath flow.
  • the signal measured in flow cytometry can be used to evaluate individual target cells' characteristics, such as cell size and cell surface roughness. With the help of fluorescent labeling, additional cellular characteristics can also be evaluated such as the existence of a cellular nucleus, the amount of DNA inside the cell, antigens on a cellular membrane, and many other characteristics. As the cells are measured one by one, the total number of target cells detected can also be determined by counting the number of measured signal peaks. Additionally, some cytometry analyses also require measuring particle density in the sample, meaning the number of target particles per sample volume, which is also known as the absolute count in cytometry analyses. For this measurement, not only the total number of detected particles needs to be determined, but also the corresponding volume of the sample needs to be determined. These two pieces of information can be used together to calculate the number of particles per sample volume, e.g., the absolute count.
  • the sample preparation steps are usually carried out by manual operation.
  • the preparation steps are often performed in different containers, such as centrifugal tubes or vials, and only the final prepared sample is then loaded into a commercial cytometer for optical or other measurement.
  • These manual steps of sample preparation require precise fluid handling by trained technicians, and are thus not suitable for applications where users are minimally trained.
  • the cytometry analyses are performed in a non- laboratory environment, such as in emergency rooms or physician offices. Therefore, it is important that the biological sample is self- contained and not exposed to environment causing biological contaminations. For this purpose, it is advantageous that both the sample preparation step and the measurement step are carried out in a self-contained manner such as inside a non-exposed container.
  • the absolute count measurement requires that the total number of detected target cells and corresponding sample volume be known.
  • a fixed amount of sample with a known volume is injected into the system to determine the absolute count.
  • the fluidic system often introduces dead volumes, meaning that some portion of the sample does not go through the cytometer measurement. These dead volumes cause the real sample volume being measured to be different from the known volume being injected into the system, and therefore introduce inaccuracy to the absolute count.
  • the present disclosure provides various fluidic cartridges and methods of using and making such fluidic cartridges. These fluidic cartridges can perform both the sample preparation and the cytometer analysis. These fluidic cartridges can be used for various types of cytometry analyses. In various embodiments, the fluidic cartridges as disclosed herein can be used to determine the absolute count. In various embodiments, the fluidic cartridges as disclosed herein can be used for DNA analysis of cell populations in tumor diagnosis. In various embodiments, the fluidic cartridges as disclosed herein can be used for CD4+/CD8+ lymphocyte subtype analyses in AIDS diagnosis. In various embodiments, the fluidic cartridges as disclosed herein can be used for cell analysis in complete blood count (CBC). These fluidic cartridges can also be used for other types of analyses including, but not limited to, analyzing analytes, proteins, enzymes, nucleic acids and other biological markers in samples.
  • CBC complete blood count
  • the device comprises a cartridge device.
  • the cartridge device comprises: an inlet configured for receiving the sample into the cartridge device; a fluidic structure fluidly connected to the inlet and configured for mixing at least a portion of the sample with at least a portion of a reagent to form one or more sample mixtures; a flow cell fluidly connected to the fluidic structure and configured for forming one or more sample streams from the one or more sample mixtures, wherein the sample streams are formed in the flow cell without a sheath flow, and wherein the flow cell comprises an optically transparent area configured for measuring an optical signal from the sample streams to detect the target particles in the sample; and a flow sensor fluidly connected to the flow cell and configured for measuring a sensing signal from the sample streams that enter the flow sensor.
  • a cartridge device as disclosed herein further comprises a reagent.
  • a device as disclosed herein further comprises a reader instrument device, wherein the reader instrument device is configured for receiving, operating, and/or actuating the cartridge device.
  • the reader instrument device neither receives any liquid from the cartridge device nor transfers any liquid into the cartridge device.
  • Various embodiments of the present disclosure provide a method for analyzing target particles in a sample.
  • the method comprises: applying the sample to a cartridge device as disclosed herein, which is configured for collecting a predetermined sample volume into the cartridge device; transferring the cartridge device into a reader instrument device as disclosed herein; mixing at least a portion of the collected sample and at least a portion of a reagent to form one or more sample mixtures inside the cartridge device; forming one or more sample streams from the one or more sample mixtures in a flow cell inside the cartridge device, wherein the sample streams are formed in the flow cell without a sheath flow; measuring an optical signal from the sample streams at the flow cell to detect the target particles in the sample streams; and using the reader instrument device to analyze the measured optical signal to quantify the target particles in the sample.
  • Various embodiments of the present disclosure provide a method for analyzing particles in a sample.
  • the method comprises: applying the sample to a cartridge device as disclosed herein, which is configured for collecting a predetermined sample volume into the cartridge device; transferring the cartridge device into a reader instrument device as disclosed herein; mixing at least a portion of the collected sample and at least a portion of a reagent to form one or more sample mixtures inside the cartridge device; forming one or more sample streams from the one or more sample mixtures in a flow cell inside the cartridge device, wherein at least two separate sample mixtures are transferred into the same flow cell to form at least two separate sample streams without a sheath flow; measuring an optical signal from the sample streams at the flow cell to detect the target particles in the sample streams; and using the reader instrument device to analyze the measured optical signal to quantify the target particles in the sample.
  • a method as disclosed herein further comprises: flowing the sample streams through a flow sensor that is fluidly connected to the flow cell; measuring a sensing signal from the sample streams at the flow sensor to detect the entrance of the sample streams into the flow sensor and/or the exit of the sample streams out of the flow sensor; and using the reader instrument device to analyze the measured optical signal and sensing signal to determine the concentration of the target particles in the sample.
  • FIG. 1 illustrates, in accordance with various embodiments of the disclosure, one non- limiting example of the basic fluidic unit used in a cartridge device as disclosed herein.
  • FIGs. 2A-2D illustrate, in accordance with various embodiments of the disclosure, a few non- limiting examples of passive valves.
  • FIGs. 3A-3C illustrate, in accordance with various embodiments of the disclosure, a few non-limiting examples of active valves.
  • FIGs. 4A-4C illustrate, in accordance with various embodiments of the disclosure, one non-limiting example of implementing a passive valve in a basic fluidic unit.
  • FIG. 5 illustrates, in accordance with various embodiments of the disclosure, a symbol drawing that represents a basic fluidic unit as described herein.
  • FIGs. 6A-6B illustrate, in accordance with various embodiments of the disclosure, one non-limiting example of a sheathless flow cell as described herein and its symbolic drawing.
  • FIGs. 7A-7B illustrate, in accordance with various embodiments of the disclosure, one non-limiting example of a flow sensor as described herein, which has two sensing zones along the length of a fluidic channel, and its symbolic drawing.
  • FIGs. 8A-8B illustrate, in accordance with various embodiments of the disclosure, another non-limiting example of a flow sensor as described herein, which has only one sensing zone along the length of a fluidic channel, and its symbolic drawing.
  • FIGs. 9A-9C illustrate, in accordance with various embodiments of the disclosure, one exemplary configuration of a cartridge device as disclosed herein, where a basic fluidic unit 9001, a sheathless flow cell 9007 and a flow sensor 9009 with two sensing zones 9011 and 9012 are connected in serial by fluidic conduits 9006 and 9008.
  • FIGs. 10A-10B illustrate, in accordance with various embodiments of the disclosure, another exemplary configuration of a cartridge device as disclosed herein, where a flow sensor 10007 is connected to the micro fluidic channel 10004 of a basic fluidic unit 10001 with a fluidic conduit 10006.
  • FIGs. 11A-11B illustrate, in accordance with various embodiments of the disclosure, another exemplary configuration of a cartridge device as disclosed herein, where a basic fluidic unit 11001, a sheathless flow cell 11007 and a flow sensor 11009 with one sensing zone 11012 are connected in serial by fluidic conduits 11006 and 11008.
  • FIGs. 12A-12G illustrate, in accordance with various embodiments of the disclosure, another exemplary configuration of a cartridge device as disclosed herein, where two basic fluidic units 12101 and 12201 are used in serial with a sheathless flow cell 12301 and a flow sensor 12401.
  • FIGs. 13A-13C illustrate, in accordance with various embodiments of the disclosure, another exemplary configuration of a cartridge device as disclosed herein, where three basic fluidic units 13101, 13201 and 13301 are used in serial with a sheathless flow cell 13401 and a flow sensor 13501.
  • FIGs. 14A-14B illustrate, in accordance with various embodiments of the disclosure, another exemplary configuration of a cartridge device as disclosed herein, where four basic fluidic units 14101, 14201, 14301 and 14401 are used in serial with a sheathless flow cell 14501 and a flow sensor 14601.
  • FIGs. 15A-15D illustrate, in accordance with various embodiments of the disclosure, the top view (in x-y plane) of a few examples of a flow cell as described herein.
  • FIGs. 16A-16B illustrate, in accordance with various embodiments of the disclosure, an example where a plurality of particles flow through a flow cell for detection.
  • FIGs. 17A-17D illustrate, in accordance with various embodiments of the disclosure, exemplary designs for determining the absolute count of particles, where the outlet 17103 of the flow cell 17101 is coupled to the inlet 17202 of the flow sensor 17201 by a fluidic conduit 17001.
  • FIGs. 18A-18B illustrate, in accordance with various embodiments of the disclosure, another exemplary design for determining the absolute count of particles, where the inlet 18102 of the flow cell 18101 is coupled to the outlet 18203 of the flow sensor 18201 by a fluidic conduit 18001
  • FIG. 19 illustrates, in accordance with various embodiments of the disclosure, one non-limiting example of an analyzer having a cartridge device and a reader instrument device.
  • the cartridge 19101 having the fluidic structure 19102 can be inserted into a docking slot 19202 on the reader instrument 19201.
  • FIGs. 20A-20B illustrate, in accordance with various embodiments of the disclosure, exemplar processes for building a sheathless flow cell as described herein.
  • the term “comprising” or “comprises” is used in reference to compositions, methods, and respective component(s) thereof, that are useful to an embodiment, yet open to the inclusion of unspecified elements, whether useful or not. It will be understood by those within the art that, in general, terms used herein are generally intended as “open” terms (e.g., the term “including” should be interpreted as “including but not limited to,” the term “having” should be interpreted as “having at least,” the term “includes” should be interpreted as “includes but is not limited to,” etc.).
  • the device comprises a cartridge device.
  • the cartridge device comprises: an inlet configured for receiving the sample into the cartridge device; a fluidic structure fluidly connected to the inlet and configured for mixing at least a portion of the sample with at least a portion of a reagent to form one or more sample mixtures; a flow cell fluidly connected to the fluidic structure and configured for forming one or more sample streams from the one or more sample mixtures, wherein the sample streams are formed in the flow cell without a sheath flow, and wherein the flow cell comprises an optically transparent area configured for measuring an optical signal from the sample streams to detect the target particles in the sample; and a flow sensor fluidly connected to the flow cell and configured for measuring a sensing signal from the sample streams that enter the flow sensor.
  • the cartridge device has a size in the range of about 0.1-1 cm 3 , 1-5 cm 3 , 5-25 cm 3 , 25- 50 cm 3 , or 50- 200 cm 3 .
  • a device as disclosed herein further comprises a reader instrument device, wherein the reader instrument device is configured for receiving, operating, and/or actuating the cartridge device.
  • the reader instrument device is configured for measuring the optical signal at the flow cell to quantify the target particles in the sample.
  • the reader instrument device is configured for measuring the sensing signal at the flow sensor to quantify the volume of the sample streams.
  • the reader instrument device is configured for measuring the optical signal at the flow cell and the sensing signal at the flow sensor to determine the concentration of the target particles in the sample.
  • the reader instrument device comprises a control unit configured for measuring the optical signal at the flow cell.
  • the reader instrument device comprises a control unit configured for measuring the optical signal at the flow cell and the sensing signal at the flow sensor. In various embodiments, the reader instrument device neither receives any liquid from the cartridge device nor transfers any liquid into the cartridge device.
  • a cartridge device as disclosed herein further comprises a reagent.
  • the reagent comprises a fluorescent labeling agent that selectively labels the target particles in the sample with fluorescence, and wherein the optical signal from the sample streams comprises fluorescence.
  • a cartridge device as disclosed herein further comprises a first reagent, which is mixed with a portion of the received sample to form a first sample mixture, and a second reagent, which is mixed with another portion of the received sample to form a second sample mixture; and the two sample mixtures are separately transferred into the flow cell to form two separate sample streams.
  • the two sample mixtures are separately formed in a chamber or separately transferred into a chamber before being separately transferred into the flow cell.
  • the chamber has a volume in the range of about 0.01-0.1 ml, 0.1-0.2 ml, 0.2-0.4 ml, 0.4-0.8 ml, 0.8-2 ml, or 2- 10 ml.
  • a cartridge device as disclosed herein further comprises a fluidic conduit fluidly connected to the inlet and configured for receiving or collecting the sample.
  • the fluidic conduit is closed by a valve and/or sealed by an external structure after the sample is collected into the fluidic conduit. In accordance with various embodiments of the present disclosure, closing by the value and/or sealing by the external structure prevents the collected sample from exiting the cartridge device.
  • the fluidic conduit is configured for collecting a predetermine sample volume in the range of about ⁇ . ⁇ - ⁇ , 1-5 ⁇ , 5-10 ⁇ , 10-20 ⁇ , or 20-50 ⁇ .
  • at least a portion of the reagent is transferred into the fluidic conduit to flush a portion of the collected sample into a chamber to form a sample mixture.
  • the sample, reagent, sample mixtures, or sample streams are enclosed inside the cartridge device to prevent or limit their exposure to the environment outside the cartridge.
  • the fluidic structure is inside the cartridge device to prevent or limit exposing the sample, reagent or sample mixtures to the environment outside the cartridge.
  • the flow cell is inside the cartridge device to prevent or limit exposing the sample streams to the environment outside the cartridge.
  • the flow sensor is inside the cartridge device to prevent or limit exposing the sample streams to the environment outside the cartridge.
  • the fluidic structure comprises one or a plurality of fluidic conduits. In various embodiments, the fluidic structure comprises one or a plurality of chambers. In various embodiments, each chamber has a volume in the range of about 0.01- 0.1 ml, 0.1-0.2 ml, 0.2-0.4 ml, 0.4-0.8 ml, 0.8-2 ml, or 2-10 ml.
  • the fluidic structure comprises one or a plurality of chambers; each chamber has a volume in the range of about 0.01-0.1 ml, 0.1-0.2 ml, 0.2-0.4 ml, 0.4-0.8 ml, 0.8-2 ml, or 2-10 ml; and the fluidic structure is configured for transferring the sample mixtures from one of the chambers to the flow cell to form the sample streams.
  • a cartridge device as disclosed here comprises one flow cell. In some embodiments, a cartridge device as disclosed comprises two, three, four, five, or more flow cells. In some embodiments, a cartridge device as disclosed comprises a plurality of flow cells. In various embodiments, the flow cell is configured for allowing a flow rate in the range of 0.001-0.01, 0.01-0.1, 0.1-1, 1-50, 50-200, or 200-1000 ⁇ /min. In various embodiments, the flow cell has a cross section in the shape of a rectangular, trapezoid, oval, circle, or half circle, or any other shape, or a combination thereof.
  • the flow cell has a width in the range of about 1-10 ⁇ , 10-40 ⁇ , 40-100 ⁇ , or 100-200 ⁇ . In various embodiments, the flow cell has a depth in the range of about 1-10 ⁇ , 10-40 ⁇ , 40-100 ⁇ , or 100-200 ⁇ . In various embodiments, the flow cell has a length in the range of about of 1-10 ⁇ , 10-100 ⁇ , 100-1,000 ⁇ , 1,000-10,000 ⁇ , or 10,000-50,000 ⁇ . In various embodiments, the sample streams formed in the flow cell have a cross section of the same size as the flow cell.
  • the flow cell has a width in the range of about 1-10 ⁇ , 10- 40 ⁇ , 40-100 ⁇ , or 100-200 ⁇ and a depth in the range of about 1-10 ⁇ , 10-40 ⁇ , 40- 100 ⁇ , or 100-200 ⁇ ; and the sample streams have a cross section of the same size as the flow cell.
  • the optically transparent area on the flow cell has a transmission rate of 50-60%, 60-70%, 70-80%, 80-90%, 90-96%, or 96-99.9% for the optical signal from the sample streams.
  • the optical signal comprises scattered light, reflected light, transmitted light, fluorescence, light absorption, light extinction, or white light image, or a combination thereof.
  • the optically transparent area on the flow cell has a transmission rate of 50-60%, 60-70%, 70- 80%, 80-90%, 90-96%, or 96-99.9% for the optical signal from the sample streams, and the optical signal comprises scattered light, reflected light, transmitted light, fluorescence, light absorption, light extinction, or white light image, or a combination thereof.
  • the optically transparent area on the flow cell is made of a plastic material.
  • the plastic material is cyclic olefin copolymer, cyclo-olefin polymer, poly-methyl methacrylate, polycarbonate, polystyrene, or poly-chloro- tri-fluoro-ethylene, or a combination thereof.
  • the flow sensor comprises a fluidic channel and a sensing zone on the fluidic channel; the fluidic channel is fluidly connected to the flow cell to allow the sample streams to flow through; and a sensing signal is measured when the sample streams enter the sensing zone.
  • the sensing signal comprises an optical signal.
  • the optical signal comprises light transmission through and/or light reflection from the sample streams.
  • the fluidic channel in the flow sensor has a channel width in the range of about 0.001-0.05mm, 0.05-1 mm, or 1-5 mm, and a channel depth in the range of about 0.001-0.01 mm, 0.01-0.5 mm, 0.5-1 mm, or 1-2 mm.
  • the flow cell and the flow sensor are configured to have the same flow rate for the sample streams flowing through.
  • the fluidic connection between the flow cell and the flow sensor is configured for a sample stream to have the same flow rate flowing through the flow cell and the flow sensor.
  • the sensing zone comprises an optically transparent area configured for measuring an optical signal that changes levels between the absence and presence of the sample streams in the sensing zone.
  • the optically transparent area on the sensing zone has a transmission rate of 50-60%, 60-70%, 70-80%, 80- 90%, 90-96%, or 96-99.9% for the optical signal from the sample streams.
  • the optical signal comprises scattered light, reflected light, transmitted light, fluorescence, light absorption, light extinction, or white light image, or a combination thereof.
  • the optically transparent area on the sensing zone is made of a plastic material.
  • the plastic material is cyclic olefin copolymer, cyclo-olefin polymer, poly-methyl methacrylate, polycarbonate, polystyrene, or poly-chloro- tri-fluoro-ethylene, or a combination thereof.
  • the flow sensor comprises one sensing zone on the fluidic channel. In some embodiments, the flow sensor comprises two, three, four, five, or more sensing zones on the fluidic channel. In some embodiments, the flow sensor comprises a plurality of sensing zones on the fluidic channel.
  • the fluidic structure comprises at least one basic fluidic unit that comprises: a chamber configured to accommodate a fluid; a venting port connected to the chamber, wherein the venting port is connected to a pneumatic pressure source, an ambient pressure, or the atmosphere pressure; a microfluidic channel connected to the chamber; and a valve on the microfluidic channel.
  • the cartridge device is configured for transferring the sample mixtures from the chamber into the flow cell to form the sample streams when an external actuation mechanism is applied to the cartridge device.
  • the cartridge device is configured for transferring the sample mixtures from the chamber into the flow cell to form the sample streams when an external actuation mechanism is applied to the cartridge device.
  • the external actuation mechanism comprises a pneumatic pressure source.
  • the external actuation mechanism is configured for forming the sample streams with a flow rate in the range of 0.001-0.01, 0.01-0.1, 0.1-1, 1-50, 50-200, or 200-1000 ⁇ /min.
  • the cartridge device is configured for transferring the sample mixtures from the chamber into the flow cell to form the sample streams when an external actuation mechanism is applied to the cartridge device, and the external actuation mechanism comprises a pneumatic pressure source.
  • the chamber of the basic fluidic unit has a volume in the range of about 0.01-0.1 ml, 0.1-0.2 ml, 0.2-0.4 ml, 0.4-0.8 ml, 0.8-2 ml, or 2-10 ml.
  • the microfluidic channel of the basic fluidic unit has a cross section of a size in the range of about 0.001-0.01 mm 2 , 0.01-0.1 mm 2 , 0.1-0.25 mm 2 , 0.25-0.5 mm 2 , 0.5- 1 mm 2 , 1-2 mm 2 , or 2-10 mm 2 .
  • the chamber of the basic fluidic unit has a volume in the range of about 0.01-0.1 ml, 0.1-0.2 ml, 0.2-0.4 ml, 0.4-0.8 ml, 0.8-2 ml, or 2-10 ml
  • the microfluidic channel of the basic fluidic unit has a cross section of a size in the range of about 0.001-0.01 mm 2 , 0.01-0.1 mm 2 , 0.1-0.25 mm 2 , 0.25-0.5 mm 2 , 0.5-1 mm 2 , 1-2 mm 2 , or 2-10 mm 2 .
  • the chamber of the basic fluidic unit when the cartridge device is in use, is so positioned that the at least a portion of the fluid inside the chamber is pulled by gravity towards the microfluidic channel and/or away from the venting port. In various embodiments, when the cartridge device is in use, the chamber of the basic fluidic unit has a volume larger than the volume of the fluid accommodated therein and an air gap exists between the venting port and the fluid accommodated therein.
  • the valve of the basic fluidic unit is a passive valve that is configured for allowing a fluid flow to pass through the microfluidic channel when a pneumatic pressure is applied to the fluid flow and stopping the fluid flow when no pneumatic pressure is applied to the fluid flow.
  • the valve of the basic fluidic unit is a passive valve that comprises one of the following structures: (i) a channel with a hydrophilic inner surface embedded with a patch of a hydrophobic surface, (ii) a channel with a hydrophobic inner surface embedded with a patch of a hydrophilic surface, (iii) an enlargement of the channel cross section along the flow direction in a channel with a hydrophilic inner surface, and (iv) a contraction of the channel cross section along the flow direction in a channel with a hydrophobic inner surface.
  • the valve of the basic fluidic unit is an active valve operated by an actuation mechanism external to the cartridge device.
  • Various embodiments of the present disclosure provide a method for analyzing particles in a sample.
  • the method comprises: providing a cartridge device as disclosed herein and a reader instrument device as disclosed herein; applying the sample to the cartridge device; transferring the cartridge device into the reader instrument device; operating the reader instrument device to actuate the cartridge device; and analyzing the target particles in the sample.
  • Various embodiments of the present disclosure provide a method for analyzing target particles in a sample.
  • the method comprises: applying the sample to a cartridge device as disclosed herein, which is configured for collecting a predetermined sample volume into the cartridge device; transferring the cartridge device into a reader instrument device as disclosed herein; mixing at least a portion of the collected sample and at least a portion of a reagent to form one or more sample mixtures inside the cartridge device; forming one or more sample streams from the one or more sample mixtures in a flow cell inside the cartridge device, wherein the sample streams are formed in the flow cell without a sheath flow; measuring an optical signal from the sample streams at the flow cell to detect the target particles in the sample streams; and using the reader instrument device to analyze the measured optical signal to quantify the target particles in the sample.
  • Various embodiments of the present disclosure provide a method for analyzing target particles in a sample.
  • the method comprises: applying the sample to a cartridge device as disclosed herein, which is configured for collecting a predetermined sample volume into the cartridge device; transferring the cartridge device into a reader instrument device as disclosed herein; mixing at least a portion of the collected sample and at least a portion of a reagent to form one or more sample mixtures inside the cartridge device; forming one or more sample streams from the one or more sample mixtures in a flow cell inside the cartridge device, wherein at least two separate sample mixtures are transferred into the same flow cell to form at least two separate sample streams without a sheath flow; measuring an optical signal from the sample streams at the flow cell to detect the target particles in the sample streams; and using the reader instrument device to analyze the measured optical signal to quantify the target particles in the sample.
  • a portion of the collected sample is mixed with a first reagent to form a first sample mixture and another portion of the collected sample is mixed with a second reagent to form a second sample mixture; and the two sample mixtures are separately transferred into the flow cell to form two separate sample streams.
  • the two sample mixtures are separately formed in a chamber or separately transferred into a chamber before being separately transferred into the flow cell.
  • the chamber has a volume in the range of about 0.01-0.1 ml, 0.1-0.2 ml, 0.2- 0.4 ml, 0.4-0.8 ml, 0.8-2 ml, or 2-10 ml.
  • the sample is collected into a fluidic conduit.
  • the fluidic conduit is closed by a valve and/or sealed by an external structure after the sample is collected into the fluidic conduit. In accordance with various embodiments of the present disclosure, closing by the value and/or sealing by the external structure prevents the collected sample from exiting the cartridge device.
  • the fluidic conduit is configured for collecting a predetermine sample volume in the range of about ⁇ . ⁇ - ⁇ , 1-5 ⁇ , 5-10 ⁇ , 10-20 ⁇ , or 20-50 ⁇ .
  • at least a portion of the reagent is transferred into the fluidic conduit to flush a portion of the collected sample into a chamber to form a sample mixture.
  • a method as disclosed herein further comprises: flowing the sample streams through a flow sensor that is fluidly connected to the flow cell; measuring a sensing signal from the sample streams at the flow sensor to detect the entrance of the sample streams into the flow sensor and/or the exit of the sample streams out of the flow sensor; and using the reader instrument device to analyze the measured optical signal and sensing signal to determine the concentration of the target particles in the sample.
  • a method as disclosed herein further comprises: flowing the sample streams through a flow sensor that is fluidly connected to the flow cell; measuring a sensing signal from the sample streams at the flow sensor to detect the sample streams entering and/or exiting the flow sensor; and using the reader instrument device to analyze the measured optical signal and sensing signal to determine the concentration of the target particles in the sample.
  • the sample streams in the flow cell and the flow sensor have the same flow rate.
  • a method as disclosed herein further comprises: flowing the sample streams through a flow sensor that is fluidly connected to the flow cell; measuring a sensing signal from the sample streams at the flow sensor to quantify the volume of the sample streams; and using the reader instrument device to analyze the measured optical signal and sensing signal to determine the concentration of the target particles in the sample.
  • the sample streams in the flow cell and the flow sensor have the same flow rate.
  • the optical signal and sensing signal are measured by the reader instrument device.
  • the collected sample, reagent, sample mixtures, or sample streams are enclosed inside the cartridge device to prevent or limit their exposure to the environment outside the cartridge.
  • the mixing step is performed in a fluidic structure.
  • the fluidic structure comprises one or a plurality of fluidic conduits.
  • the fluidic structure comprises one or a plurality of chambers.
  • each chamber has a volume in the range of about 0.01-0.1 ml, 0.1-0.2 ml, 0.2-0.4 ml, 0.4-0.8 ml, 0.8-2 ml, or 2-10 ml.
  • the mixing step is performed in a fluidic structure comprising one or a plurality of chambers, and each chamber has a volume in the range of about 0.01-0.1 ml, 0.1-0.2 ml, 0.2-0.4 ml, 0.4-0.8 ml, 0.8-2 ml, or 2-10 ml.
  • the fluidic structure is inside the cartridge device to prevent or limit exposing the sample, reagent or sample mixtures to the environment outside the cartridge.
  • the flow cell is inside the cartridge device to prevent or limit exposing the sample streams to the environment outside the cartridge.
  • the flow sensor is inside the cartridge device to prevent or limit exposing the sample streams to the environment outside the cartridge.
  • the flow cell has a width in the range of about 1-10 ⁇ , 10- 40 ⁇ , 40-100 ⁇ , or 100-200 ⁇ . In various embodiments, the flow cell has a depth in the range of about 1-10 ⁇ , 10-40 ⁇ , 40-100 ⁇ , or 100-200 ⁇ . In various embodiments, the flow cell has a length in the range of about of 1-10 ⁇ , 10-100 ⁇ , 100-1,000 ⁇ , 1,000- 10,000 ⁇ , or 10,000-50,000 ⁇ . In various embodiments, the sample streams formed in the flow cell have a cross section of the same size as the flow cell.
  • the flow cell has a width in the range of about 1-10 ⁇ , 10- 40 ⁇ , 40-100 ⁇ , or 100-200 ⁇ and a depth in the range of about 1-10 ⁇ , 10-40 ⁇ , 40- 100 ⁇ , or 100-200 ⁇ ; and the sample streams have a cross section of the same size as the flow cell.
  • the sample streams in the flow cell have a flow rate in the range of 0.001-0.01, 0.01-0.1, 0.1-1, 1-50, 50-200, or 200-1000 ⁇ /min when the optical signal is measured from the sample streams.
  • the optical signal measured from the sample streams at the flow cell comprises scattered light, reflected light, transmitted light, fluorescence, light absorption, light extinction, or white light image, or a combination thereof.
  • the sample streams in the flow sensor have a flow rate in the range of 0.001-0.01, 0.01-0.1, 0.1-1, 1-50, 50-200, or 200-1000 ⁇ /min when the sensing signal is measured from the sample streams.
  • the sensing signal measured from the sample streams at the flow sensor comprises an optical signal.
  • the optical signal comprises light transmission through and/or light reflection from the sample streams.
  • the sample streams have the same flow rate in the flow cell and the flow sensor.
  • the reagent comprises a fluorescent labeling agent that selectively labels the target particles in the sample with fluorescence, and wherein the optical signal from the sample streams comprises fluorescence.
  • each of the sample streams is separately formed and measured in the flow cell.
  • at least two separate sample mixtures are transferred into the same flow cell to form at least two separate sample streams.
  • the at least two separate sample streams are formed consecutively (i.e., immediately one after another). In other embodiments, the at least two separate sample streams are formed nonconsecutively (i.e., not immediately one after another).
  • at least one sample stream comprises white blood cells as the target particles detected in the flow cell and at least another sample stream comprises red blood cells and/or platelet cells as the target particles detected in the flow cell.
  • the fluidic channel in the flow sensor has a channel width in the range of about 0.001-0.05mm, 0.05-1 mm, or 1-5 mm, and a channel depth in the range of about 0.001-0.01 mm, 0.01-0.5 mm, 0.5-1 mm, or 1-2 mm; and wherein the sample streams in the flow cell and the flow sensor have the same flow rate.
  • mixing is performed in at least one basic fluidic unit that comprises: a chamber configured to accommodate a fluid; a venting port connected to the chamber, wherein the venting port is connected to a pneumatic pressure source, an ambient pressure, or the atmosphere pressure; a microfluidic channel connected to the chamber; and a valve on the microfluidic channel.
  • the chamber of the basic fluidic unit has a volume in the range of about 0.01-0.1 ml, 0.1-0.2 ml, 0.2-0.4 ml, 0.4-0.8 ml, 0.8-2 ml, or 2-10 ml.
  • the microfluidic channel of the basic fluidic unit has a cross section of a size in the range of about 0.001-0.01 mm 2 , 0.01-0.1 mm 2 , 0.1-0.25 mm 2 , 0.25-0.5 mm 2 , 0.5-1 mm 2 , 1-2 mm 2 , or 2-10 mm 2 .
  • mixing is performed in at least one basic fluidic unit that comprises: a chamber configured to accommodate a fluid, wherein the chamber has a volume in the range of about 0.01-0.1 ml, 0.1-0.2 ml, 0.2-0.4 ml, 0.4-0.8 ml, 0.8-2 ml, or 2-10 ml; a venting port connected to the chamber, wherein the venting port is connected to a pneumatic pressure source, an ambient pressure, or the atmosphere pressure; a microfluidic channel connected to the chamber, wherein the microfluidic channel has a cross section of a size in the range of about 0.001-0.01 mm 2 , 0.01-0.1 mm 2 , 0.1-0.25 mm 2 , 0.25-0.5 mm 2 , 0.5-1 mm 2 , 1-2 mm 2 , or 2-10 mm 2 ; and a valve on the microfluidic channel.
  • the sample mixtures are transferred from the chamber into the flow cell to form the sample streams when an external actuation mechanism is applied to the cartridge device.
  • the external actuation mechanism comprises a pneumatic pressure source.
  • the external actuation mechanism is configured for forming the sample streams with a flow rate in the range of 0.001-0.01, 0.01- 0.1, 0.1-1, 1-50, 50-200, or 200-1000 ⁇ /min.
  • the sample mixtures are transferred from the chamber into the flow cell to form the sample streams when an external actuation mechanism is applied to the cartridge device, and the external actuation mechanism comprises a pneumatic pressure source.
  • the target particles have a size in the range of 0.1-1 ⁇ , 1-10 ⁇ , 10-15 ⁇ , 15-30 ⁇ , 30-50 ⁇ , or 50-100 ⁇ . In various embodiments, the target particles have a concentration in the range of 1-100, 100-1000, 1000-5000, 5000-20,000, or 20,000-50,000 target particles per ⁇ sample steam. In certain embodiments, the target particles have a size in the range of 0.1-1 ⁇ , 1-10 ⁇ , 10-15 ⁇ , 15-30 ⁇ , 30-50 ⁇ , or 50-100 ⁇ ; and the target particles have a concentration in the range of 1-100, 100-1000, 1000-5000, 5000-20,000, or 20,000-50,000 target particles per ⁇ sample steam.
  • the target particles comprise cells, plant cells, animal cells, blood cells, white blood cells, red blood cells, platelet cells, viruses, bacteria, fungi, yeasts, beads, fluorescent beads, or non-fluorescent beads; or other particles of proteins, enzymes, nucleic acids, polysaccharides, or polypeptides; or other particles bound with biological markers; or their combinations.
  • FIG. 1 illustrates one non-limiting example of the basic fluidic unit to be used in the cartridge.
  • the basic fluidic unit 1001 has a chamber 1002, a venting port 1003 and at least one microfluidic channel 1004 that accesses the chamber and has a valve 1005 on the microfluidic channel.
  • the operation of basic fluidic unit 1001 depends on gravity or any other force serving as the replacement for gravity (e.g., centrifugal force) to keep fluid in position. Additionally, the basic fluidic unit 1001 uses another force such as pneumatic pressure to transfer fluid. More information regarding the design, operation and manufacturing of fluid unit 1001 can be found in U.S.
  • the valve 1005 can be a passive valve. In other embodiments, the valve 1005 can be an active valve. In certain embodiments, the valve 1005 can be a hybrid or combination of passive and active valves. In other embodiments, the valve 1005 can be any design known to one of ordinary skill in the art.
  • FIGs. 2A-2D illustrate a few non-limiting examples of passive valves. Other passive valve designs known to persons skilled in the art can also be used.
  • FIG. 2A is a passive valve design having a channel with a hydrophilic inner surface and a patch of a hydrophobic surface.
  • FIG. 2B is a passive valve design having a channel with a hydrophobic inner surface and a patch of a hydrophilic surface.
  • FIG. 2C is a passive valve design having an enlargement of the channel cross-section along the flow direction and the channel has a hydrophilic surface.
  • FIG. 2D is a passive valve design having a narrow down of the channel cross-section along the flow direction and the channel has a hydrophobic surface.
  • FIGs. 3A-3C illustrate a few non-limiting examples of active valves. Other active valve designs known to persons skilled in the art can also be used.
  • FIG. 3A shows a valve design that includes a flexible membrane 3001 and a substrate 3002. When the flexible membrane 3001 is bent away from the substrate 3002, the valve is in an "open” status to allow fluid flow to pass through. When the flexible membrane 3001 is bent towards the substrate 3002 leaving no gap, the valve is in the "close” status and fluid flow is not able to pass through.
  • FIG. 3B shows a valve design that has a movable membrane 3003 and a substrate 3004. When the movable membrane 3003 is away from the substrate 3004, there is a fluid path 3005 between the inlet and outlet, and the valve is in "open” status.
  • FIG. 3C shows a valve design that has a plug 3008 on the channel.
  • the plug 3008 is pulled away from the channel leaving the substrate 3009, the channel is in the "open” status allowing fluid flow from the inlet 3010 to the outlet 3011.
  • the plug 3008 is inserted into the channel contacting the substrate 3009, the channel is in the "close” status and there is no fluid path between the inlet 3010 and the outlet 3011.
  • the plug 3008 can be made of solid material, a polymer, an elastomer, a gel, a wax, a silicon oil or other materials.
  • FIG. 4A illustrates another non- limiting example of implementing a passive valve in a basic fluidic unit 4001.
  • the transition area 4005 from the chamber 4002 to the channel 4004 provides a narrowing of the flow channel cross section.
  • this transition area 4005 is equivalent to the sudden narrow down of a hydrophobic channel, and acts as a passive valve to stop fluid in the chamber 4002 from entering the channel 4004.
  • both the channel inner surface and the chamber inner surface within this transition area 4005 are hydrophilic, as shown in FIG. 4C, this transition area is equivalent to the sudden enlargement of a hydrophobic channel, and acts as a passive valve to stop fluid in the channel 4004 from entering the chamber 4002. Additional designs of passive valves known to person skilled in the art can also be implemented.
  • FIG. 5 illustrates a symbol drawing that represents a basic fluidic unit as described herein, where the basic fluidic unit 5001 includes a chamber 5002, a venting port 5003 and at least one microfluidic channel 5004 that accesses the chamber and has a valve 5005 on the microfluidic channel.
  • the valve 5005 can be either a passive valve, an active valve or a hybrid or combination of both.
  • the basic fluidic unit 5001 can have one or a plurality of microfluidic channels (each having a valve) accessing the chamber 5002 (see, e.g., U.S. Application 15/176,729 and PCT Application PCT/US 16/36426, which are incorporated herein by reference in their entirety as if fully set forth).
  • present disclosure provides fluidic cartridges having at least one basic fluidic unit as described herein.
  • the fluidic cartridges may have additional fluidic structures.
  • additional fluidic structure is one or more flow cells for a cytometer analysis.
  • the flow cell With conventional flow cytometers, the flow cell usually has a core diameter of several hundreds of micrometers. To achieve a sample stream of a smaller core diameter, e.g., a few to dozens of micrometers, the flow cell utilizes sheath flow to focus the sample stream.
  • a fluidic cartridge as described herein includes a conventional flow cell with sheath flow.
  • a fluidic cartridge as described herein includes a sheathless flow cell instead of the conventional flow cell with sheath flow.
  • the sheathless flow cell has a fluidic channel having a core diameter chosen according to the target sample stream diameter.
  • a fluidic channel having a diameter of 30 ⁇ can be used to achieve a target sample stream having a diameter of 30 ⁇ .
  • the channel of the flow cell can be transparent to certain excitation light and emission light wavelengths, so that optical signals can be measured from samples in the flow cell (FIG. 6A).
  • FIG. 6B illustrates a symbolic drawing that represents a sheathless flow cell as described herein. Since the flow cell does not utilize sheath flow, the sample stream has a cross section in the same size as the flow cell.
  • FIG. 7A illustrates one non-limiting example of the flow sensor, which has two sensing zones along the length of a fluidic channel.
  • the sensor detects whether there is fluid inside the channel overlapping with the sensing zones.
  • the volume of fluid filling up the channel between the two sensing zones can be determined by the known geometry of the channel.
  • FIG. 7B is a symbolic drawing to represent this design.
  • FIG. 8A illustrates another non-limiting example of the flow sensor, which has only one sensing zone along the length of a fluidic channel.
  • FIG. 8B is a symbolic drawing to represent this design.
  • Described herein are various fluidic units and additional fluidic structures that can be used together in various configurations to achieve functions of a flow cytometer analysis integrating sample preparation and performing absolute count in a self-contained cartridge.
  • FIG. 9 A shows one exemplary configuration, where a basic fluidic unit 9001, a sheathless flow cell 9007 and a flow sensor 9009 with two sensing zones 9011 and 9012 are connected in serial by fluidic conduits 9006 and 9008.
  • the upstream end of the flow cell 9007 is connected to the microfluidic channel 9004 of the basic fluidic unit 9001, and the downstream end of the flow cell 9007 is connected with the flow sensor.
  • sample in the chamber 9002 of the unit 9001 will pass through the flow cell first and then through the flow sensor for a cytometer analysis.
  • a fluid sample 9101 can first be loaded into the chamber 9002. Pneumatic pressures can then be applied to the vent 9003 of the basic fluidic unit 9001 and to the outlet port 9010 of the flow sensor.
  • the pneumatic pressure at vent 9003 is higher than the pneumatic pressure at port 9010, it creates a pressure difference that pumps sample 9101 from the chamber 9002 into the flow cell 9007 for the cytometer analysis, and then into the flow sensor 9009 for volume measurement.
  • the valve 9005 is a passive valve, a sufficiently-high pressure difference can pump the fluid sample to pass the valve 9005.
  • the valve 9005 is an active valve, the valve 9005 can be switched to open status before the pressure can pump the fluid sample 9101 to pass the valve 9005.
  • the recorded data includes the measured physical signal A (optical emission, electrical impedance, etc.) along time T as an array (A, 7).
  • FIG. 9C shows one example of the recorded data, where the amplitude of the signal A is plotted against the time T. The number of particles detected in the cytometer is determined by the number of peaks in the signal A. Meanwhile, as the sample continues to pass through the flow sensor 9009, the time point Tj of the sample reaching the first sensing zone 9011 is recorded, and the time point T 2 of the sample reaching the second zone 9012 is also recorded.
  • the number of particles N detected between 77 and 72 can be determined from the record signal (A, 7) as shown in the example of FIG. 9C.
  • the fluid volume Vo for filling up the channel between the sensing zone 9011 and the sensing zone 9012 is a known parameter from the flow sensor design (see, e.g., US Application 15/209,226 and PCT Application PCT/US 16/42089, which are incorporated herein by reference in their entirety as if fully set forth). Because the sheathless flow cell contains only the fluid sample for analysis (no sheath flow), the fluid volume between the two sensing zones can be used to determine the volume of sample analyzed in the flow cell 9007. Therefore, the absolute count can then be calculated as:
  • FIGs. 9A-9C also has the feature that the whole fluidic structure can be implemented in a self-contained cartridge device.
  • the cartridge device can be a molded piece of plastic with additional sealing layers.
  • FIG. 10A shows another exemplary configuration, where a flow sensor 10007 is connected to the micro fluidic channel 10004 of a basic fluidic unit 10001 with a fluidic conduit 10006. Meanwhile, a sheathless flow cell 10009 is connected downstream of the flow sensor 10007 by a fluidic conduit 10008.
  • the number of cells counted N is determined by the signal (A, 7) between time points Tj+AT and ⁇ 2 + ⁇ , as shown in FIG. 10B, where AT can be any empirical value to compensate the time delay between the sample reaching the first sensing zone 10011 and sample reaching the flow cell 10009.
  • FIG. 11 A shows another exemplary configuration, where a basic fluidic unit 11001, a sheathless flow cell 11007 and a flow sensor 11009 with one sensing zone 11012 are connected in serial by fluidic conduits 11006 and 11008.
  • the upstream end of the flow cell is connected to the micro fluidic channel 11004 of the basic fluidic unit 11001, and the downstream end of the flow cell is connected to the flow sensor.
  • sample in the chamber 11002 of the unit 11001 passes through the flow cell first and then through the flow sensor for a cytometer analysis.
  • FIG. 11 A shows another exemplary configuration, where a basic fluidic unit 11001, a sheathless flow cell 11007 and a flow sensor 11009 with one sensing zone 11012 are connected in serial by fluidic conduits 11006 and 11008.
  • the upstream end of the flow cell is connected to the micro fluidic channel 11004 of the basic fluidic unit 11001, and the downstream end of the flow cell is connected to the flow sensor.
  • sample in the chamber 11002 of the unit 11001 passes through
  • the time points (when the flow cell starts to detect particles) and T2 (when the sample reaches the sensing zone 11012) are used to determine the total particle count N from the recorded signal (A, 7).
  • the fluid volume Vi to obtain the particle count N includes the total fluid conduit volume between the flow sensor 11007 and the sensing zone 11012 of the flow sensor 11009.
  • the fluid volume Vj is a parameter known from the fluidic design.
  • the configuration of FIG. 10 can also be used for cytometer analysis with absolute count:
  • FIG. 12A shows another exemplary configuration, where two basic fluidic units 12101 and 12201 are used in serial with a sheathless flow cell 12301 and a flow sensor 12401.
  • a fluidic conduit 12001 connects the basic fluidic unit 12101's microfluidic channel 12104 (having a valve 12105) with the basic fluidic unit 12201's microfluidic channel 12204 (having a valve 12205).
  • the unit 12201 has a second microfluidic channel 12206 (having a valve 12207), which connects to the upstream end of the flow cell 12301 by a fluid conduit 12002.
  • the downstream end of the flow cell 12301 is further connected to the flow sensor 12401 by a fluid conduit 12003.
  • the flow sensor 12401 has two sensing zones 12402 and 12403.
  • pneumatic pressures are applied to three ports, including the vent 12103 of unit 12101 (Pi), the vent 12203 (P 2 ) of unit 12201, and at the downstream port 12404 (P 3 ) of the flow sensor 12401.
  • a fluid sample can be transferred between the chamber 12102 and the chamber 12202, and further transferred into the flow cell for a cytometer analysis with the absolute count.
  • FIG. 12B An exemplary method of controlling the pneumatic pressures (Pi, P2 and P 3 ) and the corresponding fluid transfer is shown in the diagram of FIG. 12B.
  • the fluid sample can be transferred from the first chamber into the second chamber (chamber 12202).
  • the fluid sample can be transferred from the second chamber into the first chamber.
  • a sample can be transferred in the fluidic configuration for the cytometer analysis with absolute count.
  • a fluid sample can be initially introduced into the first chamber (chamber 12102). The sample can then be transferred to the second chamber (chamber 12202). The sample can then be driven through the flow cell and the flow sensor for the cytometer analysis with absolute count as described above.
  • the fluid sample be can initially introduced into the second chamber and next driven through the flow cell and the flow sensor for the cytometer analysis with absolute count.
  • a fluid sample A can be initially introduced into the first chamber and a fluid sample B can be initially introduced to the second chamber, and then the two samples can be transferred between the two chambers for a plurality of mixing cycles, before being delivered to the flow cell for the cytometer analysis with the absolute count.
  • This mixing action involves the fluid sample moving along one direction from the first chamber into the second chamber and then along an opposite direction from the second chamber into the first chamber, and vice versa.
  • the fluid sample A has a predetermined volume.
  • the fluid sample B has a predetermined volume.
  • a fluid sample A can be initially introduced to the first chamber 12102 and a fluid sample C can be initially loaded in the fluid conduit 12001.
  • the fluid A and the fluid C can be mixed together, and then delivered to the flow cell for the cytometer analysis with the absolute count.
  • the sample exiting the outlet of the flow sensor can be disposed or collected in a reservoir.
  • the fluidic configuration of this example achieves the sample preparation, cytometer analysis and the absolute count function in a self-contained manner, without an exchange of fluid sample between the fluidic structure and the outside environment.
  • Such an embodiment can be used for a cytometer analysis of different biological samples.
  • the fluid sample C can be a biological sample such as whole blood from human body.
  • the fluid sample A can be a reagent containing a fluorophore-conjugated antibody targeting specific cell types, e.g. CD4+ lymphocytes, in the whole blood. By mixing these two fluids together and then measuring the mixture in the flow cell, it achieves the absolute count of the CD4+ Lymphocyte.
  • the fluid sample C can be a human whole blood
  • the fluid sample A can be a lysing solution selectively targeting Red Blood Cells (RBCs). After mixing the two fluids together and incubating the mixture for a period of time, the mixture is measured in the flow cell for a cytometer analysis such as the absolute count of the white blood cells (WBCs).
  • WBCs white blood cells
  • FIG. 12C shows an exemplary fluidic configuration, where an additional fluid conduit 12004 is connected to the fluid conduit 12001 to introduce the initial sample C.
  • the sample can be introduced via the port 12006 into the fluidic conduit 12001.
  • a valve 12005 can then be closed to seal the conduit 12004, preventing the sample from exiting the port 12006.
  • the fluid conduit 12001 can be used to collect a predetermined volume of the initial sample.
  • the valve 12005 can be a blood clotting valve (see, e.g., U.S. Patent 8,845,979, which incorporated herein by reference in its entirety as if fully set forth). Other methods known to persons skilled in the art can also be used to introduce the initial samples.
  • a reservoir chamber can be connected to outlet port of the flow sensor to collect the fluid sample after the cytometer analysis.
  • a reservoir 12501 with a venting port 12502 can be connected to the outlet port 12404 of the flow sensor by a fluidic conduit 12503.
  • the pneumatic pressure P3 can be adjusted by controlling the pneumatic pressure P4 at the venting port 12502 of the reservoir.
  • FIG. 12E An exemplary operation of this fluidic configuration is illustrated in FIG. 12E.
  • FIG. 12F shows one example of these embodiments.
  • the second basic fluidic unit 12201 has a third microfluidic channel 12208 (with a valve 12209), which connects by a fluidic conduit 12007 to a reservoir structure 12601 with a venting port 12602.
  • the venting port 12602 corresponds to a pneumatic pressure P5.
  • a fluid sample D is initially stored in the reservoir 12601.
  • FIG. 12G shows a pneumatic control for operating this configuration.
  • the sample D initially stored in the reservoir 12601 is transferred into the second chamber 12202.
  • the rest of the pneumatic operation for the cytometer analysis can be the same as the example in the FIG. 12B.
  • FIG. 13 A there are three basic fluidic units 13101, 13201 and 13301.
  • the units 13101 and 13201 have microfluidic channels 13104 (with the valve 13105) and 13204 (with the valve 13205), respectively.
  • the unit 13301 has three microfluidic channels 13304 (with the valve 13305), 13306 (with the valve 13307) and 13308 (with the valve 13309).
  • a fluidic conduit 13001 connects the channels 13104 and 13304, whereas another fluidic conduit 13002 connects the channel 13204 and 13306.
  • the fluid unit 13302 is also connected to an upstream port of a sheathless flow cell 13401 by a fluidic conduit 13003, while the downstream port of the flow cell 13401 is connected to a flow sensor 13501 by a fluidic conduit 13004.
  • This fluidic configuration is operated by controlling the pneumatic pressures at the venting ports of the basic fluidic units 13103 (PI), 13203 (P2) and 13303 (P3), and by controlling the pneumatic pressure of the outlet port 13504 of the flow sensor (P4).
  • This fluidic configuration and the fluid transfer diagram can be used to perform more complex sample preparation and cytometer analysis.
  • a fluid sample Al is initially introduced into the first chamber and a fluid sample A2 is initially introduced into the second chamber.
  • fluid samples B l and B2 are each introduced into the fluidic conduit 13001 and 13002, respectively.
  • the fluid samples Al and B l are transferred into the third chamber.
  • the pneumatic control P4 ⁇ Po and the sample mixture of Al and Bl is transferred into the sheathless flow cell and the flow sensor for the cytometer analysis with the absolute count.
  • This step can be repeated to achieve desirable mixing uniformity.
  • the fluid samples A2 and B2 do not move. This is achieved by keeping the pneumatic pressure
  • steps of repeated transfer between the second chamber and the third chamber can also be carried out similar to the repeated transfer steps between the first and the third chamber.
  • the fluid sample Al has a predetermined volume.
  • the fluid sample A2 has predetermined volume.
  • the fluid sample B l has predetermined volume.
  • the fluid sample B2 has predetermined volume.
  • the fluid sample Al can be a dilution buffer for RBC analysis and the sample B l can be whole blood
  • the sample A2 can be a lysing buffer for WBC analysis and the sample B2 can be whole blood.
  • Al and B 1 can be transferred into the third chamber to form a mixture, and then into the flow cell for counting and analyzing of RBCs and platelets in the blood.
  • A2 and B2 can then be transferred into the third chamber to form a mixture, and then into the flow cell for counting and analyzing WBCs in the blood.
  • Different dilution buffers and lysing buffers known to persons skilled in the art of hematology analyzers can be used. In this way, the fluid configuration can be used to achieve the Complete Blood Count (CBC) analysis widely used in clinical tests.
  • CBC Complete Blood Count
  • FIGs. 12A-12G and FIGS. 13A-13C show examples having two and three basic fluidic units, respectively. In other embodiments, more basic fluidic units can be used in the configuration to achieve additional complexity.
  • the fluidic conduits for connecting the basic fluidic units e.g. the fluid conduit 12001, the fluid conduit 13001 and the fluid conduit 13002
  • the fluidic conduits for connecting the basic fluidic units are fluid channels. In other embodiments, fluid structures with additional complexity can be used as the fluidic conduits.
  • FIG. 14A shows an example with four basic fluidic units, 14101, 14201, 14301 and 14401.
  • Each of the four units 14010, 14201 and 14401 has a microfluidic channel with a valve.
  • the unit 14301 has four microfluidic channels including channel 14304 (with valve 14305), channel 14306 (with valve 14307), channel 14308 (with valve 14309), and channel 14310 (with valve 14311).
  • the basic fluidic unit 14101 is connected to the basic fluidic unit 14301 by a fluidic conduit 14001.
  • the basic fluidic unit 14201 is connected to the basic fluidic unit 14301 by a fluidic conduit 14002.
  • the basic fluidic unit 14401 is connected to the basic fluidic unit 14301 by a fluidic conduit 14003.
  • the upstream of a sheathless flow cell 14501 is connected to the basic fluidic unit 14301 by a fluidic conduit 14004, while the downstream of the flow cell 14501 is connected by a fluidic conduit 14005 to a flow sensor 14601 that has two sensing zones 14602 and 14603.
  • the flow sensor 14601 is then connected to a reservoir chamber 14701 that has a venting port 14702.
  • a sample Al can be initially stored in the chamber 14102 of the basic fluidic unit 14101
  • a sample A2 can be initially stored in the chamber 14202 of the basic fluidic unit 14201
  • a sample A3 can be initially stored in the chamber 14402 of the basic fluidic unit 14401.
  • a sample Bl can be induced into the fluidic conduit 14001 by an inlet port 14801 through a fluidic conduit 14801 with a valve 14803.
  • the valve 14803 can be closed after inducing the sample.
  • the fluidic conduit 14001 can be used to collect a predetermined volume of the sample.
  • a sample B2 can be induced into the fluidic conduit 14002 by an inlet port 14901 through a fluidic conduit 14902 with a valve 14903.
  • the valve 14903 can be closed after inducing the sample.
  • the fluidic conduit 14002 can be used to collect a predetermined volume of the sample.
  • FIG. 14B An exemplary method of controlling the pneumatic pressures (Pi, P2, P 3 , P4 and P5) is described below and the corresponding fluid transfer is shown in the diagram of FIG. 14B.
  • the pneumatic pressure P5 at the venting port 14702 of the reservoir 14701 balances with the pressure P5' at the downstream port 14604 of the flow sensor 14601, when there is a pneumatic path between these two ports (e.g., when there is air path in the reservoir 14701 to balance the venting port 14702 and the port 14604).
  • a fluid sample in the third chamber can be transferred into the fourth chamber by applying a pneumatic control (P4 ⁇ Po and
  • a fluid sample in the third chamber can be transferred into the flow cell and the flow sensor for the cytometer analysis, by applying a pneumatic control (Ps ⁇ Po and
  • the sheathless flow cell is where the target particles in a fluid sample flow are detected and measured by different signals such as fluorescence, light scattering, light absorption, and light extinction, white light imaging, etc.
  • An excitation light (EL) beam from a light source can be shaped and used to illuminate a designated sensing area of the flow cell, and trigger the above signals from the target particles.
  • the sheathless flow cell can be a fluidic channel that has various geometry shapes.
  • FIGS. 15A-15D show the top view (in x-y plane) of a few examples of the flow cell.
  • the top view (x-y plane) is defined as the plane perpendicular to the direction of the excitation light (z-axis).
  • the length is defined the as channel dimension along the sample flow (x-axis)
  • the width is defined as the dimension along the y-axis.
  • the depth is defined as the channel dimension along the z-axis.
  • FIG. 15B shows an example of a flow cell that has a gradually decreased width, where the maximum width is Wj and the minimum width is W2. In other embodiments, the flow cell can have a gradually increased width.
  • FIG. 15C shows an example of a flow cell that has a non-gradually changing width, where the maximum width is Wj and the minimum width is W2.
  • FIG. 15D shows an example of a flow cell that has a fixed width at various positions along channel length.
  • the difference of the maximum width Wj and the minimum width W2 are within a designated difference.
  • a non-limiting example of the range of the width difference is (Wj-W2)AV2 ⁇ 20%.
  • the ranges of the channel width and the depth are chosen to be large enough so that target particles (e.g., cells in biological samples), can pass through the flow cell without blocking it. Meanwhile, they are chosen to be small enough to minimize the coincidence error in the flow cytometer analysis.
  • the minimum width Wj can be in the range of 1-10 ⁇ , 10-40 ⁇ , 40 to 100 ⁇ , or 100 to 200 ⁇ .
  • the depth of the channel can be in the range of 1-10 ⁇ , 10-40 ⁇ , 40 to 100 ⁇ , or 100 to 200 ⁇ .
  • the cross section of the channel (in y-z plane) can have the shape of a rectangular, a trapezoid, a circle, a half circle, or any other shapes.
  • the length of the flow cell should be long enough for the optical detection of particles in the sample stream, and meanwhile, short enough to reduce the flow resistance of the sample stream flowing through. In various embodiments, the length of the flow cell can be in the range of about 1-10 ⁇ , 10-100 ⁇ , 100-1,000 ⁇ , 1,000-10,000 ⁇ , or 10,000-50,000 ⁇ .
  • the sheathless flow cell is used for optical measurement
  • at least one surface of the channel is transparent to the light wavelength involved in the measurement.
  • the material for forming the channel surface can be any transparent material such as glass, quartz, and plastics including, but not limited to, Cyclic Olefin Copolymer (COC), Cyclo-olefin Polymer (COP), Poly-Methyl methacrylate (PMMA), polycarbonate (PC), Polystyrene (PS), and Poly-chloro- tri-fluoro-ethylene (PCTFE) materials such as Aclar, etc.
  • COC Cyclic Olefin Copolymer
  • COP Cyclo-olefin Polymer
  • PMMA Poly-Methyl methacrylate
  • PC polycarbonate
  • PS Polystyrene
  • PCTFE Poly-chloro- tri-fluoro-ethylene
  • the fluid sample for analysis in the flow cell can be a fluid suspension of a plurality of particles.
  • the fluid sample can be a blood sample containing different cells such as white blood cells, red blood cells and platelets.
  • the fluidic sample can be a blood sample, in which certain types of cells remain intact, such as white blood cells, while other types of cells have been lysed, such as red blood cells.
  • the fluid sample can be a blood sample in which certain types of cells have been labeled with a fluorophore.
  • the fluidic sample can be a mixture of the cells and other particles, such as non-fluorescent beads and/or fluorescent beads.
  • the fluid samples can also be other biological samples such as cerebrospinal fluid, urine, saliva, semen, etc.
  • FIG. 16A shows an example where a plurality of particles flow through the flow cell for detection. All particles in the sample flow through in a one-by-one manner. Under the illumination of the excitation light (EL), each cell can be characterized for optical signals that include but are not limited to fluorescence (FL) and light scattering (LS).
  • FIG. 16B shows another example where a plurality of particles flow through the flow cell for detection. Some particles are flowing through while overlapping with each other.
  • the target particles are flowing through still in a one-by-one manner without overlapping with other target particles.
  • the light scattering from the target particles may be blocked by other particles overlapping with them.
  • other signals that include but are not limited to fluorescence signal can still be measured to detect these target particles, if these particles are treated with specific fluorophore to distinguish from the other particles beforehand.
  • the other particles can be non-fluorescent or treated with different fluorophore distinguishable from the target particles.
  • the fluid sample can be a blood sample in which the white blood cells are labeled with fluorophore and the red blood cells are not labeled with fluorophore.
  • the corresponding fluorescence signal can be measured to detect and characterize the white blood cell even when there are red blood cells overlapping with them.
  • the fluid sample can be a blood sample in which the white blood cells are labeled with fluorophore and the red blood cells are lysed.
  • the corresponding fluorescence signal and light scattering signals can be measured simultaneously from these cells for detection and characterization.
  • the fluid sample can be a blood sample in which there are fluorophore-labeled white blood cells and fluorescent beads. When these white blood cells and the beads pass through the flow cell one-by-one, they can be detected by the corresponding fluorescence signal. Other cells having no fluorescence or a different wavelength of fluorescence do not impede the measurement.
  • the fluid sample can be a blood sample in which there are red blood cells and beads among other cells. When the cells pass through one-by-one, light scattering signals can be measured to detect and characterize the red blood cells and the beads.
  • the bead can be label with a fluorophore, so they can be distinguished from the red blood cells by the light scattering signals or by fluorescence signals, or by both signals.
  • FIG. 17A shows one exemplary design, where the outlet 17103 of the flow cell 17101 is coupled to the inlet 17202 of the flow sensor 17201 by a fluidic conduit 17001.
  • the outlet of the flow cell 17101 can be coupled to the inlet 17202 of the flow sensor 17201 directly and without additional fluidic conduit.
  • the flow sensor 17201 has two sensing zones 17204 and 17205. A fluid sample flows into the inlet 17102 of the flow cell 17101 and then out of the outlet 17203 of the flow sensor 17201. The signal measured in the flow cell 17101 is recorded, as illustrated in FIG. 17B.
  • Time To is when the sample starts being detected in the flow cell 17101, Ti is when the fluid sample passes the sensing zone 17204, and T2 is when the fluid sample passes the sensing zone 17205. From time Ti to T 2 , the total number of target particles detected in the flow cell 17101 is N.
  • the fluid volume Vo between the two sensing zones 17204, 17205 is a known parameter from the design of the flow sensor. Because the flow cell 17101 has a sheathless design, the volume of fluid flows through the flow cell 17101 is only the fluid sample. Therefore, the sample volume being measured in the flow cell 17101 between Ti and T2 equals to Vo. In this design, the absolute count is determined as:
  • FIG. 17C shows another exemplary design, where the flow sensor 17201 has only one sensing zone 17205.
  • FIG. 17D is the signal measured from this design, where time To is when the sample starts being detected in the flow cell, and T2 is when the fluid sample passes the sensing zone 17205. From time To to T 2 , the total number of target particles detected in the flow cell is N'.
  • the volume Vo' is the total fluid volume filling up the fluidic conduit from the flow cell 17101 to the sensing zone 17205.
  • the absolute count is determined as:
  • FIG. 18A shows another exemplary design, where the inlet 18102 of the flow cell 18101 is coupled to the outlet 18203 of the flow sensor 18201 by a fluidic conduit 18001.
  • the flow sensor 18201 has two sensing zones 18204 and 18205.
  • a fluid sample flows into the inlet 18102 of the flow sensor 18201 and then out of the outlet 18203 of the flow sensor 18201.
  • the signal measured in the flow cell 18101 is recorded, as illustrated in FIG. 18B.
  • Ti is when the fluid sample passes the sensing zone 18204
  • T2 is when the fluid sample passes the sensing zone 18205.
  • the number of cells counted N" is determined by the signal (A, T) between time points Tj+AT and ⁇ 2+ ⁇ , as shown in FIG.
  • the fluid volume Vo between the two sensing zones 18204, 18205 is a known parameter from the design of the flow sensor 18201. In this design, the absolute count is determined as:
  • This combination of the flow cell 18101 and the flow sensor 18201 can be used for measurement of particles or cells.
  • the size of the target particles can be in the range of 0.1-1 ⁇ , 1-10 ⁇ , 10-15 ⁇ , 15-30 ⁇ , 30-50 ⁇ , or 50-100 ⁇ depending on the size of the flow cell 18101.
  • the size of the particles being measured should be smaller than size of the flow cell 18101, and the size difference can range from 1-5 ⁇ , 5-10 ⁇ , 10-20 ⁇ , or 20-50 ⁇ .
  • the concentration of the target particles in the fluid sample can be in the range of 1-100, 100-1000, 1000-5000, 5000-20,000, or 20,000- 50,000 particles or cells per ⁇ sample.
  • the flow rate can be in the range of 0.001-1, 1-50, 50-200, or 200-1000 microliters per minute ( ⁇ /min). In certain embodiments, the ranges can be 1-50 or 50-200 ⁇ /min.
  • the range of the fluid sample volume can be constrained by the cartridge size.
  • the volume of the flow sensor and the total volume of the sample can be in the range of 0.1-1 ⁇ , 1-200 ⁇ , 200-1000 ⁇ , 1 -5 ml, or 5-30 ml. In certain embodiments, the range can be 1-200 ⁇ , 200-1000 ⁇ , or 1- 5 ml. In certain embodiments, by considering both the sample volume and the flow rate, the measurement is completed in less than 10 minutes.
  • sample preparation steps can be further integrated with the cytometer analysis including the absolute count.
  • the integration of the above functions enables the fluidic configurations to be operated as a self-contained structure for a cytometer analysis, without fluid exchange with the outside environment after the fluid samples having been loaded into the cartridge.
  • pneumatic pressures are applied to the venting ports of the basic fluidic units and additional venting ports of other fluidic structures such as a reservoir (see, e.g., FIG. 12D and FIG. 14A).
  • the fluid sample is a biological sample containing cells
  • a high flow rate in a confined channel may induce a large shear force to lyse the cells.
  • the pressure difference between any two of the applied pressures can be in the range of 0-1, 1-5, 5-15, or 15-30 psi. In certain embodiments, the range can be 0-1, 1-5, or 5-15 psi.
  • At least one of the venting ports can be connected to the atmosphere pressure of the environment.
  • another pressure higher than the atmosphere pressure applied introduces a positive pressure difference in comparison to the atmosphere pressure.
  • This positive pressure difference can be in the range of 0-1, 1-5, 5-15, or 15-30 psi. In certain embodiments, the range can be 0-1, 1-5, or 5-15 psi.
  • another pressure lower than the atmosphere pressure applied introduces a negative pressure difference.
  • This negative pressure difference can be in a range of 0-1, 1-5, 5-15, or 15-30 psi. In certain embodiments, the range can be 0-1, 1-5, or 5-15 psi.
  • the flow rate achieved for transferring the sample via the channel between any two of the basic fluidic units can be in the range of 0-1, 1-50, 50-200, or 200-1000 ⁇ /min, or 1-10 ml/min. In certain embodiments, the range can be 1 microliter to 1-50, 50-200, or 200-1000 ⁇ /min.
  • Various fluidic configurations incorporating a plurality of basic fluidic units and a plurality of the combinations of the sheathless flow cell and the flow sensor can be implemented in various manufacturing structures to form a fluidic cartridge.
  • this cartridge can be inserted into a reader instrument for operation, as shown in the example of FIG. 19.
  • the cartridge 19101 having the fluidic structure 19102 can be inserted into a docking slot 19202 on the reader instrument 19201.
  • a control unit of the reader instrument records the signals from the cytometer analysis.
  • the signals include but are not limited to the optical signals such as fluorescence, light scattering, light absorption, etc.
  • the reader instrument has alignment mechanisms and features to align the sheathless flow cell with the optics in the instrument for optical signal measurement.
  • the control unit of the reader instrument also detects the signals from the flow sensor to determine the absolute count.
  • the control unit of the reader instrument also applies the pneumatic pressure source to the cartridges to drive the fluid transfer.
  • the control unit of the reader instrument also supports additional actuations such as opening or closing a valve structure in the cartridge fluidics.
  • the cartridge is self-contained and there is no exchange of liquid samples between the cartridge and the reader instrument.
  • the cartridge is not self-contained, and the reader instrument has on-board liquid storage and there is liquid exchange between the reader instrument and the cartridge, such as liquid infusion from the reader instrument into the cartridge.
  • the cartridge stays stationary after being inserted into the reader, whereas the interface for external connections such as the pneumatic pressure source moves to make contact with the cartridge.
  • the cartridge can be movable after being inserted into the reader, and is moved to make contact with the interface for external connections such as pneumatic pressure sources.
  • the sheathless flow cell in the fluidic structures can be built with various manufacturing processes.
  • an open fluidic channel for the flow cell can be built by injection molding, embossing, etching, CNC, laser cutting, or die cutting, etc.
  • a cover can then be added onto the patterns to form enclosed fluidic channel to be the flow cell.
  • the cover can be added by various manufacturing process, such as thermal fusion bonding, thermal lamination, adhesive bonding, solvent assisted bonding, laser wielding, and ultrasonic wielding, etc.
  • Non-limiting examples of building the sheathless flow cell are described here.
  • optical signals are detected from particles flowing inside the sheathless flow cell. Smooth surface of the flow cell is useful to achieve acceptable optical signals.
  • FIG. 20A shows one example of the sheathless flow cell 20101 having two pieces.
  • the cross-section view (y-z plane) is perpendicular to the direction of sample flow (x-axis).
  • the bottom piece 20102 forms three sides of a channel without a cover.
  • the top piece 20103 adds a cover side to the channel, which then forms an enclosed channel.
  • the bottom and the top surfaces 20104 and 20105 can achieve smoothness for optical measurement in the two pieces 20102 and 20103, respectively.
  • FIG. 20B shows another example of building the sheathless flow cell 20201 having three pieces.
  • the 20203 and a top piece 20204 are added separately.
  • the three pieces together forms an enclosed channel as the flow cell.
  • the surface 20205 and 20206 can achieve smoothness for optical measurement in the two pieces 20203 and 20204, respectively.
  • the cartridge device for the cytometer analysis can be of any size.
  • the cartridge device is received in the reader instrument device for the measurement and analysis and has a size in the range of about 0.1-1 cm 3 , 1-5 cm 3 , 5-25 cm 3 , 25-50 cm 3 , or 50-200 cm 3 .
  • the numbers expressing quantities of ingredients, properties such as concentration, reaction conditions, and so forth, used to describe and claim certain embodiments of the disclosure are to be understood as being modified in some instances by the term "about.”
  • the numerical parameters set forth in the written description and attached claims are approximations that can vary depending upon the desired properties sought to be obtained by a particular embodiment.
  • the numerical parameters should be construed in light of the number of reported significant digits and by applying ordinary rounding techniques.
  • a device for analyzing target particles in a sample includes a cartridge device, wherein the cartridge device comprises: an inlet configured for receiving the sample into the cartridge device; a fluidic structure fluidly connected to the inlet and configured for mixing at least a portion of the sample with at least a portion of a reagent to form one or more sample mixtures; a flow cell fluidly connected to the fluidic structure and configured for forming one or more sample streams from the one or more sample mixtures, wherein the sample streams are formed in the flow cell without a sheath flow, and wherein the flow cell comprises an optically transparent area configured for measuring an optical signal from the sample streams to detect the target particles in the sample; and a flow sensor fluidly connected to the flow cell and configured for measuring a sensing signal from the sample streams that enter the flow sensor.
  • the fluidic structure comprises one or a plurality of chambers, wherein each chamber has a volume in the range of about 0.01-0.1 ml, 0.1-0.2 ml, 0.2-0.4 ml, 0.4-0.8 ml, 0.8-2 ml, or 2-10 ml; and wherein the fluidic structure is configured for transferring the sample mixtures from one of the chambers to the flow cell to form the sample streams.
  • the flow cell has a width in the range of about 1-10 ⁇ , 10-40 ⁇ , 40-100 ⁇ , or 100-200 ⁇ and a depth in the range of about 1-10 ⁇ , 10-40 ⁇ , 40-100 ⁇ , or 100-200 ⁇ ; and wherein the sample streams have a cross section of the same size as the flow cell.
  • the flow cell has a length in the range of about of 1-10 ⁇ , 10-100 ⁇ , 100-1,000 ⁇ , 1,000-10,000 ⁇ , or 10,000-50,000 ⁇ .
  • the optically transparent area on the flow cell has a transmission rate of 50-60%, 60-70%, 70-80%, 80- 90%, 90-96%, or 96-99.9% for the optical signal from the sample streams, and wherein the optical signal comprises scattered light, reflected light, transmitted light, fluorescence, light absorption, light extinction, or white light image, or a combination thereof.
  • the optically transparent area on the flow cell is made of a plastic material.
  • the cartridge device further comprises a reagent.
  • the reagent comprises a fluorescent labeling agent that selectively labels the target particles in the sample with fluorescence, and wherein the optical signal from the sample streams comprises fluorescence.
  • the flow sensor comprises a fluidic channel and a sensing zone on the fluidic channel, wherein the fluidic channel is fluidly connected to the flow cell to allow the sample streams to flow through; and wherein a sensing signal is measured when the sample streams enter the sensing zone.
  • the fluidic channel in the flow sensor has a channel width in the range of about 0.001-0.05mm, 0.05-1 mm, or 1- 5 mm, and a channel depth in the range of about 0.001-0.01 mm, 0.01-0.5 mm, 0.5-1 mm, or 1-2 mm.
  • the sensing zone comprises an optically transparent area configured for measuring an optical signal that changes levels between the absence and presence of the sample streams in the sensing zone.
  • the fluidic connection between the flow cell and the flow sensor is configured for a sample stream to have the same flow rate flowing through the flow cell and the flow sensor.
  • the fluidic structure comprises at least one basic fluidic unit that comprises: a chamber configured to accommodate a fluid; a venting port connected to the chamber, wherein the venting port is connected to a pneumatic pressure source, an ambient pressure, or the atmosphere pressure; a microfluidic channel connected to the chamber; and a valve on the microfluidic channel.
  • the chamber has a volume in the range of about 0.01-0.1 ml, 0.1-0.2 ml, 0.2-0.4 ml, 0.4-0.8 ml, 0.8-2 ml, or 2- 10 ml, and wherein the microfluidic channel has a cross section of a size in the range of about 0.001-0.01 mm 2 , 0.01-0.1 mm 2 , 0.1-0.25 mm 2 , 0.25-0.5 mm 2 , 0.5-1 mm 2 , 1-2 mm 2 , or 2-10 mm 2 .
  • the cartridge device is configured for transferring the sample mixtures from the chamber into the flow cell to form the sample streams when an external actuation mechanism is applied to the cartridge device, and wherein the external actuation mechanism comprises a pneumatic pressure source.
  • the chamber when the cartridge device is in use, the chamber is so positioned that the at least a portion of the fluid inside the chamber is pulled by gravity towards the microfluidic channel and/or away from the venting port.
  • the chamber when the cartridge device is in use, has a volume larger than the volume of the fluid accommodated therein and an air gap exists between the venting port and the fluid accommodated therein.
  • the valve is a passive valve that is configured for allowing a fluid flow to pass through the microfluidic channel when a pneumatic pressure is applied to the fluid flow and stopping the fluid flow when no pneumatic pressure is applied to the fluid flow.
  • the valve is a passive valve that comprises one of the following structures: (i) a channel with a hydrophilic inner surface embedded with a patch of a hydrophobic surface, (ii) a channel with a hydrophobic inner surface embedded with a patch of a hydrophilic surface, (iii) an enlargement of the channel cross section along the flow direction in a channel with a hydrophilic inner surface, and (iv) a contraction of the channel cross section along the flow direction in a channel with a hydrophobic inner surface.
  • the device further comprises a reader instrument device, wherein the reader instrument device is configured for receiving, operating, and/or actuating the cartridge device.
  • the reader instrument device is configured for measuring the optical signal at the flow cell to quantify the target particles in the sample.
  • the reader instrument device is configured for measuring the optical signal at the flow cell and the sensing signal at the flow sensor to determine the concentration of the target particles in the sample.
  • a method for analyzing target particles in a sample includes applying the sample to a cartridge device, which is configured for collecting a predetermined sample volume into the cartridge device; transferring the cartridge device into a reader instrument device; mixing at least a portion of the collected sample and at least a portion of a reagent to form one or more sample mixtures inside the cartridge device; forming one or more sample streams from the one or more sample mixtures in a flow cell inside the cartridge device, wherein the sample streams are formed in the flow cell without a sheath flow; measuring an optical signal from the sample streams at the flow cell to detect the target particles in the sample streams; and using the reader instrument device to analyze the measured optical signal to quantify the target particles in the sample.
  • the method includes flowing the sample streams through a flow sensor that is fluidly connected to the flow cell; measuring a sensing signal from the sample streams at the flow sensor to detect the entrance of the sample streams into the flow sensor and/or the exit of the sample streams out of the flow sensor; and using the reader instrument device to analyze the measured optical signal and sensing signal to determine the concentration of the target particles in the sample.
  • the target particles have a size in the range of 0.1-1 ⁇ , 1-10 ⁇ , 10-15 ⁇ , 15-30 ⁇ , 30-50 ⁇ , or 50-100 ⁇ ; and wherein the target particles have a concentration in the range of 1-100, 100- 1000, 1000-5000, 5000-20,000, or 20,000-50,000 target particles per ⁇ sample steam.
  • the reagent comprises a fluorescent labeling agent that selectively labels the target particles in the sample with fluorescence, and wherein the optical signal from the sample streams comprises fluorescence.
  • the mixing step is performed in a fluidic structure comprising one or a plurality of chambers, wherein each chamber has a volume in the range of about 0.01-0.1 ml, 0.1-0.2 ml, 0.2-0.4 ml, 0.4-0.8 ml, 0.8-2 ml, or 2-10 ml.
  • the flow cell has a width in the range of about 1-10 ⁇ , 10-40 ⁇ , 40-100 ⁇ , or 100-200 ⁇ and a depth in the range of about 1-10 ⁇ , 10-40 ⁇ , 40-100 ⁇ , or 100-200 ⁇ ; and wherein the sample streams have a cross section of the same size as the flow cell.
  • the sample streams in the flow cell have a flow rate in the range of 0.001-0.01, 0.01-0.1, 0.1-1, 1-50, 50- 200, or 200-1000 ⁇ /min when the optical signal is measured from the sample streams.
  • the optical signal measured from the sample streams at the flow cell comprises scattered light, reflected light, transmitted light, fluorescence, light absorption, light extinction, or white light image, or a combination thereof.
  • the flow sensor comprises a fluidic channel and a sensing zone on the fluidic channel, wherein the fluidic channel is fluidly connected to the flow cell to allow the sample streams to flow through; and wherein a sensing signal is measured when the sample streams enter the sensing zone.
  • the fluidic channel in the flow sensor has a channel width in the range of about 0.001-0.05mm, 0.05-1 mm, or 1-5 mm, and a channel depth in the range of about 0.001-0.01 mm, 0.01-0.5 mm, 0.5- 1 mm, or 1-2 mm; and wherein the sample streams in the flow cell and the flow sensor have the same flow rate.
  • the sensing zone comprises an optically transparent area configured for measuring an optical signal that changes levels between the absence and presence of the sample streams in the sensing zone.
  • mixing is performed in at least one basic fluidic unit that comprises: a chamber configured to accommodate a fluid, wherein the chamber has a volume in the range of about 0.01-0.1 ml, 0.1-0.2 ml, 0.2-0.4 ml, 0.4-0.8 ml, 0.8-2 ml, or 2-10 ml; a venting port connected to the chamber, wherein the venting port is connected to a pneumatic pressure source, an ambient pressure, or the atmosphere pressure; a microfluidic channel connected to the chamber, wherein the microfluidic channel has a cross section of a size in the range of about 0.001-0.01 mm 2 , 0.01-0.1 mm 2 , 0.1-0.25 mm 2 , 0.25-0.5 mm 2 , 0.5-1 mm 2 , 1-2 mm 2 , or 2-10 mm 2 ; and a valve on the microfluor unit that comprises: a chamber configured to accommodate a fluid, wherein the chamber has a volume in the range of about 0.01-0.
  • the sample mixtures are transferred from the chamber into the flow cell to form the sample streams when an external actuation mechanism is applied to the cartridge device, and wherein the external actuation mechanism comprises a pneumatic pressure source.
  • the chamber when the cartridge device is in use, the chamber is so positioned that the at least a portion of the fluid inside the chamber is pulled by gravity towards the microfluidic channel and/or away from the venting port.
  • the chamber when the cartridge device is in use, has a volume larger than the volume of the fluid accommodated therein and an air gap exists between the venting port and the fluid accommodated therein
  • At least two separate sample mixtures are transferred into the same flow cell to form at least two separate sample streams.
  • a method for analyzing particles in a sample includes applying the sample to a cartridge device, which is configured for collecting a predetermined sample volume into the cartridge device; transferring the cartridge device into a reader instrument device; mixing at least a portion of the collected sample and at least a portion of a reagent to form one or more sample mixtures inside the cartridge device; forming one or more sample streams from the one or more sample mixtures in a flow cell inside the cartridge device, wherein at least two separate sample mixtures are transferred into the same flow cell to form at least two separate sample streams without a sheath flow; measuring an optical signal from the sample streams at the flow cell to detect the target particles in the sample streams; and using the reader instrument device to analyze the measured optical signal to quantify the target particles in the sample.
  • a portion of the collected sample is mixed with a first reagent to form a first sample mixture and another portion of the collected sample is mixed with a second reagent to form a second sample mixture; and wherein the two sample mixtures are separately transferred into the flow cell to form two separate sample streams.
  • the two sample mixtures are separately formed in a chamber or separately transferred into a chamber before being separately transferred into the flow cell.
  • the chamber has a volume in the range of about 0.01-0.1 ml, 0.1-0.2 ml, 0.2-0.4 ml, 0.4-0.8 ml, 0.8-2 ml, or 2- 10 ml.
  • the sample is collected into a fluidic conduit.
  • the fluidic conduit is closed by a valve and/or sealed by an external structure after the sample is collected into the fluidic conduit.
  • the fluidic conduit is configured for collecting a predetermine sample volume in the range of about 0.1-1 ⁇ , 1- 5 ⁇ , 5-10nL, 10-20 ⁇ , or 20-50 ⁇ .
  • At least a portion of the reagent is transferred into the fluidic conduit to flush a portion of the collected sample into a chamber to form a sample mixture.
  • At least one sample stream comprises white blood cells as the target particles detected in the flow cell and at least another sample stream comprises red blood cells and/or platelet cells as the target particles detected in the flow cell.

Abstract

L'invention concerne des dispositifs et des procédés pour analyser des particules dans un échantillon. Dans divers modes de réalisation, la présente invention concerne des dispositifs et des procédés de cytométrie et d'analyse supplémentaire. Dans divers modes de réalisation, la présente invention concerne un dispositif de cartouche et un dispositif d'instrument de lecture, le dispositif d'instrument de lecture recevant, fonctionnant, et/ou actionnant le dispositif de cartouche. Dans divers modes de réalisation, la présente invention concerne un procédé d'utilisation d'un dispositif tel que décrit ici pour analyser des particules dans un échantillon.
PCT/US2017/059965 2016-11-07 2017-11-03 Cartouche fluidique pour cytométrie et analyse supplémentaire WO2018085678A1 (fr)

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CN201780080470.5A CN110177619B (zh) 2016-11-07 2017-11-03 用于细胞计数和附加分析的流体卡盒
EP17866884.4A EP3535056A4 (fr) 2016-11-07 2017-11-03 Cartouche fluidique pour cytométrie et analyse supplémentaire

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Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20120140205A1 (en) * 2007-04-02 2012-06-07 Life Technologies Corporation Methods and systems for controlling the flow of particles for detection
US20130137135A1 (en) * 2011-11-28 2013-05-30 California Institute Of Technology Compositions and methods for leukocyte differential counting
US20140016131A1 (en) * 2009-01-23 2014-01-16 University Of Washington Virtual core flow cytometry

Family Cites Families (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP3307670B1 (fr) * 2015-06-12 2020-12-09 Cytochip Inc. Unités fluidiques et cartouches pour une analyse de multiples analytes
US10634602B2 (en) * 2015-06-12 2020-04-28 Cytochip Inc. Fluidic cartridge for cytometry and additional analysis

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20120140205A1 (en) * 2007-04-02 2012-06-07 Life Technologies Corporation Methods and systems for controlling the flow of particles for detection
US20140016131A1 (en) * 2009-01-23 2014-01-16 University Of Washington Virtual core flow cytometry
US20130137135A1 (en) * 2011-11-28 2013-05-30 California Institute Of Technology Compositions and methods for leukocyte differential counting

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
See also references of EP3535056A4 *

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