# US 12,005,441 B1 · Automated microscopic cell analysis > Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt - Number: US 12,005,441 B1 - Document ID: US12005441B1 - Status: Granted - Kind: B1 - Application number: 16/235,099 - Filed: 2018-12-28 - Granted: 2024-06-11 - Earliest claimed: 2014-11-26 - Assignees: Medica Corporation - Applicants: Medica Corporation - Inventors: Ronald Jones; Adrian Gropper; Robert Hagopian; Charles Rogers; Thomas Vitella; Tyler Cote; Donald Barry; Dirk Osterloh; Chen Yi - Web page: https://patentgrep.com/patent/US12005441B1 ## CPC - B01L 3/502715: {characterised by interfacing components, e.g. fluidic, electrical, optical or mechanical interfaces} - B01L 3/502738: {characterised by integrated valves (throttle valves in microfluidic sample containers B01L3/502746)} - G01N 15/1433: using image recognition - G01N 15/1459: {the analysis being performed on a sample stream} - G01N 15/1484: {microstructural devices} - G01N 33/56966: {Animal cells} - B01L 2200/0605: Metering of fluids - B01L 2200/0684: Venting, avoiding backpressure, avoid gas bubbles - B01L 2200/10: Integrating sample preparation and analysis in single entity, e.g. lab-on-a-chip concept - B01L 2200/16: Reagents, handling or storing thereof - B01L 2300/0816: Cards, e.g. flat sample carriers usually with flow in two horizontal directions - B01L 2300/0867: Multiple inlets and one sample wells, e.g. mixing, dilution - B01L 2300/0883: Serpentine channels - B01L 2400/0406: capillary forces - B01L 2400/0481: squeezing of channels or chambers - B01L 2400/0487: fluid pressure, pneumatics - B01L 2400/0622: distribution valves, valves having multiple inlets and/or outlets, e.g. metering valves, multi-way valves - B01L 2400/0644: rotary valves - G01N 15/0227: using imaging; using holography - G01N 2015/012: {Red blood cells} - G01N 2015/016: {White blood cells} - G01N 2015/018: {Platelets} - G01N 2015/0294: {Particle shape} - G01N 2015/1006: {for cytology} - G01N 2015/1486: {Counting the particles} - G01N 2015/1493: {Particle size} - B01L: CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE - G01N: INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES (measuring or testing apparatus or processes other than immunoassay, involving enzymes or microorganisms C12M, C12Q) ## Abstract This disclosure describes single-use test cartridges, cell analyzer apparatus, and methods for automatically performing microscopic cell analysis tasks, such as counting and analyzing blood cells in biological samples. A small measured quantity of a biological sample, such as whole blood, is placed in a mixing bowl on the disposable test cartridge after being inserted into the cell analyzer. The analyzer also deposits a known amount of diluent/stain in the mixing bowl and mixes it with the blood. The analyzer takes a measured amount of the mixture and dispenses in a sample cup on the cartridge in fluid communication with an imaging chamber. The geometry of the imaging chamber is chosen to maintain the uniformity of the mixture, and to prevent cells from crowding or clumping as it is transferred into the imaging chamber by the analyzer. Images of all of the cellular components within the imaging chamber are counted and analyzed to obtain a complete blood count. ## Claims ### Claim 1 A test cartridge for use with a cell analyzer, which includes an automatic microscope with image processing software, for analyzing and counting biological particles in a sample of whole blood, including cells and platelets, comprising: a) a sample collection port for receiving the sample, b) a metering mechanism operatively connected to the sample collection port and operative to separate a known volume of the sample, wherein the metering mechanism consists of at least one of a valve, a pump, a capillary tube or camera, c) a chamber containing diluent and/or stain, and d) an imaging chamber operatively connected to the metering mechanism and operatively connected to the diluent and/or stain chamber, wherein the imaging chamber is configured to receive the sample and diluent and/or stain, and is configured to allow one or more images that are statistically representative of the number and distribution of at least one type of the biological particles in the imaging chamber to be captured by the automated microscope with imaging software. ### Claim 2 (depends on 1) The test cartridge of claim 1 further including a photometric chamber operatively connected to the metering mechanism from which a photometric measurement can be taken to measure hemoglobin. ### Claim 3 (depends on 1) The test cartridge of claim 1 further including a mixing chamber that is operatively connected to the imaging chamber and the metering mechanism and the diluent and/or stain chamber, to receive the known amount of sample and diluent and/or stain, and in which the sample and the diluent and/or stain can be mixed to make a substantially uniform mixture of sample and diluent and/or stain. ### Claim 4 (depends on 1) The cartridge of claim 1 wherein the metering mechanism is defined by a volume in a rotary valve. ### Claim 5 (depends on 1) The test cartridge of claim 1 wherein the chamber containing the diluent/and or stain is operatively connected to the metering mechanism. ### Claim 6 (depends on 5) The test cartridge of claim 5 wherein the metering mechanism is a rotary valve which is operatively connected between at least the sample collection port, the chamber containing diluent and/or stain, and the imaging chamber. ### Claim 7 (depends on 5) The test cartridge of claim 5 wherein the rotary valve is also operatively connected to the mixing chamber. ### Claim 8 (depends on 1) The test cartridge of claim 1 wherein the chamber containing diluent and/or stain includes a known amount of diluent and/or stain. ### Claim 9 (depends on 1) The test cartridge of claim 1 wherein the imaging chamber is of sufficient volume to contain all of the known volume of sample from the metering mechanism when mixed with an unknown amount of diluent and/or stain. ### Claim 10 (depends on 9) The test cartridge of claim 9 wherein the imaging chamber has a geometry such that the biological particles do not overlap or crowd, when the particles settle to the bottom of the imaging chamber. ### Claim 11 (depends on 1) The cartridge of claim 1 wherein the imaging chamber is of sufficient volume to contain all of the predetermined volume of sample from the metering mechanism when mixed with an unknown amount of diluent and/or stain. ### Claim 12 (depends on 1) The test cartridge of claim 1 wherein the metering mechanism is a rotary valve having a cylindrical stem. ### Claim 13 (depends on 12) The test cartridge of claim 12 wherein the metering mechanism is a pass-through conduit within the cylindrical stem. ### Claim 14 (depends on 1) The test cartridge of claim 1 wherein the metering mechanism is a recessed volume on the face of the cylindrical stem. ### Claim 15 (depends on 1) The test cartridge of claim 1 wherein the diluent and/or stain is prepackaged in a blister in the cartridge. ### Claim 16 (depends on 1) The test cartridge of claim 1 where the diluent and/or stain includes a sphering agent such as zwitterionic detergent to provide isovolumetric reshaping of the red blood cells. ### Claim 17 (depends on 1) The test cartridge of claim 1 where the diluent and/or stain includes an antibody conjugated to a detectable label that targets specific cells or specific antigens associated with cells. ### Claim 18 (depends on 1) The test cartridge of claim 1 wherein the shape of the imaging chamber in planar view is serpentine. ### Claim 19 (depends on 18) The test cartridge of claim 18 wherein an outside turning radius of the serpentine imaging chamber is about twice an inside turning radius of the serpentine imaging chamber. ### Claim 20 (depends on 18) The test cartridge of claim 18 wherein the total volume of the imaging chamber is less than 100 microliters and the depth is less than 200 μm. ### Claim 21 (depends on 1) A cell analyzer for holding the cartridge of claim 1 in a field of view of the microscope, and further including a processor responsive to the microscope and operative to count at least some of the particles in the imaging chamber of the cartridge. ### Claim 22 (depends on 21) The analyzer of claim 21 wherein the automated microscope with image processing software captures images that include all of the particles in the imaging chamber. ### Claim 23 (depends on 21) The analyzer of claim 21 wherein the automated microscope with image processing software captures images of all of at least one or more type of biological particle in the captured images. ### Claim 24 (depends on 21) The analyzer of claim 21 further including analytical software for calculating the number of one or more type of biological particles per unit volume of sample by dividing the total number of the one or more type of biological particle by the known volume of sample. ### Claim 25 (depends on 21) The analyzer of claim 21 further including analytical software for counting one or more type of biological particle in a predetermined percentage of all of the captured images to obtain the total number of the one or more type of biological particle in the predetermined percentage of all the captured images. ### Claim 26 (depends on 25) The analyzer of claim 25 further including analytical software for calculating the number of the one or more type of biological particles per unit volume in the sample, by dividing the number of particles of the one or more type counted in the predetermined percentage of all the captured images by the predetermined percentage and further dividing by the known volume of sample. ### Claim 27 (depends on 21) The analyzer of claim 21 further including image processing software for capturing images of a portion of the mixture in the imaging chamber that is selected to be statistically representative of the number and distribution of the one or more types of biological particles in the imaging chamber. ### Claim 28 (depends on 27) The analyzer of claim 27 further including analytical software for calculating the number of the one or more types of biological particle per unit volume of sample, by dividing the total number of the one or more type of biological particle counted in the images of the portion of the mixture by the percentage of the portion of the imaging chamber of which the images are captured and further dividing by the known volume of sample. ### Claim 29 (depends on 28) The analyzer of claim 28 further including analytical software for calculating the number of the one or more type of biological particles per unit volume in the sample based on a known dilution ratio resulting from mixing the known amount of sample with a known amount of diluent and/or stain from the chamber containing diluent and/or stain. ### Claim 30 (depends on 21) The analyzer of claim 21 further including image processing and analytical software for counting all of five different types of white cells in the captured images and calculating a five-part differential of the white cells based on the statistical representation of the captured images and the known volume of sample. ### Claim 31 (depends on 21) The analyzer of claim 21 further including image processing and analytical software for counting all of three different types of white cells in the captured images and calculating a three-part differential of the white cells based on the statistical representation of the captured images in the known volume of sample. ### Claim 32 (depends on 21) The analyzer of claim 21 further including image processing and analytical software for counting at least one of cells, platelets, sperm, bacteria, spores, and inorganic particles. ### Claim 33 (depends on 21) The analyzer of claim 21 further comprising a screen to display the digital images of the particles. ### Claim 34 (depends on 1) The test cartridge of claim 1 further including a mixing chamber that is operatively connected to the imaging chamber and operatively connected to the metering mechanism, and wherein the chamber containing the diluent and/or stain is operatively connected to the metering mechanism, whereby the mixing chamber can receive the sample from the metering mechanism and diluent and/or stain from the metering mechanism, and in which the sample and diluent and/or stain can be mixed to make a substantially uniform mixture of sample and diluent and/or stain. ### Claim 35 A test cartridge for use with a cell analyzer, which includes an automatic microscope with image processing software, for analyzing and counting biological particles in a sample of whole blood, including cells and platelets, comprising: a) a sample collection port for receiving the sample, b) means for metering operatively connected to the sample collection port and operative to separate a known volume of the sample, c) a chamber containing diluent and/or stain, and d) an imaging chamber operatively connected to the metering mechanism and operatively connected to the diluent and/or stain chamber, wherein the imaging chamber is configured to receive the sample and diluent and/or stain, and is configured to allow one or more images that are statistically representative of the number and distribution of at least one type of the biological particles in the imaging chamber to be captured by the automated microscope with imaging software. ### Claim 36 (depends on 35) The test cartridge of claim 35 further including a photometric chamber operatively connected to the metering mechanism from which a photometric measurement can be taken to measure hemoglobin. ### Claim 37 (depends on 35) The test cartridge of claim 35 further including a mixing chamber that is operatively connected to the imaging chamber and the metering mechanism and the diluent and/or stain chamber, to receive the known amount of sample and diluent and/or stain, and in which the sample and the diluent and/or stain can be mixed to make a substantially uniform mixture of sample and diluent and/or stain. ### Claim 38 (depends on 35) The cartridge of claim 35 wherein the metering mechanism is defined by a volume in a rotary valve. ### Claim 39 (depends on 35) The test cartridge of claim 35 wherein the chamber containing the diluent/and or stain is operatively connected to the metering mechanism. ### Claim 40 (depends on 39) The test cartridge of claim 39 wherein the metering mechanism is a rotary valve which is operatively connected between at least the sample collection port, the chamber containing diluent and/or stain, and the imaging chamber. ### Claim 41 (depends on 39) The test cartridge of claim 39 wherein the rotary valve is also operatively connected to the mixing chamber. ### Claim 42 (depends on 35) The test cartridge of claim 35 wherein the chamber containing diluent and/or stain includes a known amount of diluent and/or stain. ### Claim 43 (depends on 35) The test cartridge of claim 35 wherein the imaging chamber is of sufficient volume to contain all of the known volume of sample from the metering mechanism when mixed with an unknown amount of diluent and/or stain. ### Claim 44 (depends on 43) The test cartridge of claim 43 wherein the imaging chamber has a geometry such that the biological particles do not overlap or crowd, when the particles settle to the bottom of the imaging chamber. ### Claim 45 (depends on 35) The cartridge of claim 35 wherein the imaging chamber is of sufficient volume to contain all of the predetermined volume of sample from the metering mechanism when mixed with an unknown amount of diluent and/or stain. ### Claim 46 (depends on 35) The test cartridge of claim 35 wherein the metering mechanism is a rotary valve having a cylindrical stem. ### Claim 47 (depends on 46) The test cartridge of claim 46 wherein the metering mechanism is a pass-through conduit within the cylindrical stem. ### Claim 48 (depends on 35) The test cartridge of claim 35 wherein the metering mechanism is a recessed volume on the face of the cylindrical stem. ### Claim 49 (depends on 35) The test cartridge of claim 35 wherein the diluent and/or stain is prepackaged in a blister in the cartridge. ### Claim 50 (depends on 35) The test cartridge of claim 35 where the diluent and/or stain includes a sphering agent such as zwitterionic detergent to provide isovolumetric reshaping of the red blood cells. ### Claim 51 (depends on 35) The test cartridge of claim 35 where the diluent and/or stain includes an antibody conjugated to a detectable label that targets specific cells or specific antigens associated with cells. ### Claim 52 (depends on 35) The test cartridge of claim 35 wherein the shape of the imaging chamber in planar view is serpentine. ### Claim 53 (depends on 52) The test cartridge of claim 52 wherein an outside turning radius of the serpentine imaging chamber is about twice an inside turning radius of the serpentine imaging chamber. ### Claim 54 (depends on 52) The test cartridge of claim 52 wherein the total volume of the imaging chamber is less than 100 microliters and the depth is less than 200 μm. ### Claim 55 (depends on 35) A cell analyzer for holding the cartridge of claim 35 in a field of view of the microscope, and further including a processor responsive to the microscope and operative to count at least some of the particles in the imaging chamber of the cartridge. ### Claim 56 (depends on 55) The analyzer of claim 55 wherein the automated microscope with image processing software captures images that include all of the particles in the imaging chamber. ### Claim 57 (depends on 55) The analyzer of claim 55 wherein the automated microscope with image processing software captures images of all of at least one or more type of biological particle in the captured images. ### Claim 58 (depends on 55) The analyzer of claim 55 further including analytical software for calculating the number of one or more type of biological particles per unit volume of sample by dividing the total number of the one or more type of biological particle by the known volume of sample. ### Claim 59 (depends on 55) The analyzer of claim 55 further including analytical software for counting one or more type of biological particle in a predetermined percentage of all of the captured images to obtain the total number of the one or more type of biological particle in the predetermined percentage of all the captured images. ### Claim 60 (depends on 59) The analyzer of claim 59 further including analytical software for calculating the number of the one or more type of biological particles per unit volume in the sample, by dividing the number of particles of the one or more type counted in the predetermined percentage of all the captured images by the predetermined percentage and further dividing by the known volume of sample. ### Claim 61 (depends on 55) The analyzer of claim 55 further including image processing software for capturing images of a portion of the mixture in the imaging chamber that is selected to be statistically representative of the number and distribution of the one or more types of biological particles in the imaging chamber. ### Claim 62 (depends on 61) The analyzer of claim 61 further including analytical software for calculating the number of the one or more types of biological particle per unit volume of sample, by dividing the total number of the one or more type of biological particle counted in the images of the portion of the mixture by the percentage of the portion of the imaging chamber of which the images are captured and further dividing by the known volume of sample. ### Claim 63 (depends on 62) The analyzer of claim 62 further including analytical software for calculating the number of the one or more type of biological particles per unit volume in the sample based on a know n dilution ratio resulting from mixing the know n amount of sample with a known amount of diluent and/or stain from the chamber containing diluent and/or stain. ### Claim 64 (depends on 55) The analyzer of claim 55 further including image processing and analytical software for counting all of five different types of white cells in the captured images and calculating a five-part differential of the white cells based on the statistical representation of the captured images and the known volume of sample. ### Claim 65 (depends on 55) The analyzer of claim 55 further including image processing and analytical software for counting all of three different types of white cells in the captured images and calculating a three-part differential of the white cells based on the statistical representation of the captured images in the known volume of sample. ### Claim 66 (depends on 55) The analyzer of claim 55 further including image processing and analytical software for counting at least one of cells, platelets, sperm, bacteria, spores, and inorganic particles. ### Claim 67 (depends on 55) The analyzer of claim 55 further comprising a screen to display the digital images of the particles. ### Claim 68 (depends on 35) The test cartridge of claim 35 further including a mixing chamber that is operatively connected to the imaging chamber and operatively connected to the metering mechanism, and wherein the chamber containing the diluent and/or stain is operatively connected to the metering mechanism, whereby the mixing chamber can receive the sample from the metering mechanism and diluent and/or stain from the metering mechanism, and in which the sample and diluent and/or stain can be mixed to make a substantially uniform mixture of sample and diluent and/or stain.