# US 12,377,414 B2 · Microfluidic device and method for detecting an infected cell in a fluid sample > Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt - Number: US 12,377,414 B2 - Document ID: US12377414B2 - Status: Granted - Kind: B2 - Application number: 17/674,087 - Filed: 2022-02-17 - Published: 2022-08-18 - Granted: 2025-08-05 - Earliest claimed: 2021-02-18 - Assignees: City University of Hong Kong; The Hong Kong Polytechnic University - Applicants: City University of Hong Kong; The Hong Kong Polytechnic University - Inventors: Bee Luan Khoo; Song Lin Chua; Junchen Liao - Web page: https://patentgrep.com/patent/US12377414B2 ## CPC - B01L 3/502715: {characterised by interfacing components, e.g. fluidic, electrical, optical or mechanical interfaces} - G01N 15/0255: {with mechanical, e.g. inertial, classification, and investigation of sorted collections (with centrifuges G01N15/042)} - B01L 3/502753: {characterised by bulk separation arrangements on lab-on-a-chip devices, e.g. for filtration or centrifugation} - B01L 3/502761: {specially adapted for handling suspended solids or molecules independently from the bulk fluid flow, e.g. for trapping or sorting beads or physically stretching molecules} - G01N 1/30: Staining; Impregnating {; Fixation; Dehydration; Multistep processes for preparing samples of tissue, cell or nucleic acid material and the like for analysis} - G01N 15/1433: using image recognition - G01N 21/6428: {Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes" (in vivo A61B5/00; immunoassay G01N33/53)} - G01N 21/6458: {Fluorescence microscopy (fluorescence microscopes per se G02B21/0076 and G02B21/16)} - G01N 33/569: for microorganisms, e.g. protozoa, bacteria, viruses - B01L 3/502707: {characterised by the manufacture of the container or its components} - B01L 2200/0636: Focussing flows, e.g. to laminate flows - B01L 2200/0652: Sorting or classification of particles or molecules - B01L 2300/0864: comprising only one inlet and multiple receiving wells, e.g. for separation, splitting - B01L 2300/088: Channel loops - B01L 2400/0463: Hydrodynamic forces, venturi nozzles - G01N 15/01: specially adapted for biological cells, e.g. blood cells (investigating sedimentation of particle suspensions in blood G01N15/05) - G01N 15/075: by optical means - G01N 15/149: specially adapted for sorting particles, e.g. by their size or optical properties - G01N 2015/016: {White blood cells} - G01N 2015/0288: {Sorting the particles} - G01N 2015/0294: {Particle shape} - G01N 2015/1006: {for cytology} - G01N 2015/1486: {Counting the particles} - G01N 2015/1493: {Particle size} - G01N 2015/1495: {Deformation of particles} - G01N 2015/1497: {Particle shape} - G01N 2021/6439: {with indicators, stains, dyes, tags, labels, marks} - 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 A microfluidic device containing an inlet, a microchannel in fluid communication with the inlet, and a plurality of outlets in fluid communication with the microchannel. The microchannel contains a loop; or from about 1 loop to about 50 loops; or from about 2 loops to about 25 loops; or from about 5 loops to about 15 loops. A method for detecting an infected cell may employ the microfluidic device. ## Claims ### Claim 1 A method for detecting an immune cell comprising an internalized microbe in a fluid sample comprising the steps of: A. providing a microfluidic device comprising: i. an inlet to receive a fluid sample with an immune cell with an internalized microbe and an immune cell without an internalized microbe; ii. a curvilinear microchannel in fluid communication with the inlet, the curvilinear microchannel is configured in a loop concentrically surrounding the inlet, wherein at least a curve portion of the loop has a rectangular cross-section along a substantial portion of the length of the curved portion enabling the detection of the immune cell with an internalized microbe from the immune cell without an internalized microbe in the fluid sample through their differences in cell deformability, wherein the microchannel has an inner wall; and iii. a plurality of outlets in fluid connection with the curvilinear microchannel, wherein at least two of the outlets closest to the inner wall of the microchannel receive the immune cell with an internalized microbe; each of the outlets receive the immune cell without an internalized microbe, wherein the plurality of outlets is from about 2 outlets to about 20 outlets; B. introducing the fluid sample into the inlet at a predetermined flow rate of from about 1.3 ml/min to about 2.1 ml/min; or from about 1.5 ml/min to about 1.9 ml/min; or about 1.7 ml/min; C. individually collecting the output from the plurality of outlets; and D. quantifying a proportion of the immune cells with an internalized microbe in each of the outputs, thereby enabling determination of infection stage of the fluid sample; wherein the quantifying step comprises an optical imaging step. ### Claim 2 (depends on 1) The method according to claim 1, wherein the inlet consists essentially of one inlet and the plurality of outlets comprise from about 2 outlets to about 15 outlets; or from about 3 outlets to about 10 outlets; or wherein the plurality of outlets consists of 5 outlets. ### Claim 3 (depends on 1) The method according to claim 1, wherein the curvilinear microchannel branches to form the plurality of outlets. ### Claim 4 (depends on 3) The method according to claim 3, wherein the distance between the adjacent outlets is from about 100 μm to about 500 μm; or from about 150 μm to about 450 μm; or from about 200 μm to about 400 μm; or from about 250 μm to about 350 μm; or about 250 μm. ### Claim 5 (depends on 1) The method according to claim 1, wherein the curvilinear microchannel has a width of from about 200 μm to about 800 μm; or from about 300 μm to 700 μm; or from about 400 μm to about 650 μm; or from about 450 μm to about 550 μm. ### Claim 6 (depends on 1) The method according to claim 1, wherein the curvilinear microchannel has a height of from about 50 μm to about 350 μm; or from about 80 μm to about 325 μm; or from about 100 μm to about 310 μm; or from about 130 μm to about 275 μm; or from about 150 μm to about 250 μm, as measured horizontally between the inner and outer walls of the channel. ### Claim 7 (depends on 1) The method according to claim 1, wherein at least a portion of the microfluidic device produced by a method comprising lithography, 3D printing, and a combination thereof. ### Claim 8 (depends on 1) The method according to claim 1, wherein the individually collecting step comprises collecting the output from each outlet to comprise a plurality of outputs, and wherein the quantifying step comprises quantifying the proportion of the immune cell with an internalized microbe in each of the plurality of outputs. ### Claim 9 (depends on 1) The method according to claim 1, wherein the fluid sample comprises a body fluid selected from the group consisting essentially of blood, urine, sputum, lymph fluid, spinal fluid, semen, a tissue extract, and a combination thereof. ### Claim 10 (depends on 1) The method according to claim 1, wherein the immune cell comprises a myeloid cell, a lymphoid cell, and a combination thereof; or a phagocyte, a T cell, a B cell, a natural killer cell, and a combination thereof; or a monocyte, a neutrophil, and a combination thereof; or a monocyte; or a neutrophil. ### Claim 11 (depends on 10) The method according to claim 10, wherein the immune cell binds to or contains at least one microbe; or a pathogen, a parasite and a combination thereof; or wherein the microbe is selected from the group consisting essentially of a bacterium, a fungi, a virus, a parasite, an amoeba, a protozoa, a virus, and a combination thereof; or wherein the microbe is a Gram-negative bacteria. ### Claim 12 (depends on 1) The method according to claim 1, wherein the fluid sample is a blood sample and wherein the method further comprises the step of pre-treating the blood sample to lyse the red blood cells prior to introducing the fluid sample into the inlet. ### Claim 13 (depends on 1) The method according to claim 1, wherein the fluid sample comprises an immune cell at a concentration of from about 0 cells/ml to about 10×106 cells/ml; or from about 1×103 cells/ml to about 1×106 cells/ml. ### Claim 14 (depends on 1) The method according to claim 1, wherein the microbe is selected from the group consisting of bacterium, a fungi, a virus, a parasite, an amoeba, a protozoa, a yeast, and a combination thereof.