# US 10,481,077 B1 · Cell capture system and method of use

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- Number: US 10,481,077 B1
- Document ID: US10481077B1
- Status: Granted
- Kind: B1
- Application number: 16/536,155
- Filed: 2019-08-08
- Published: 2019-12-05
- Granted: 2019-11-19
- Earliest claimed: 2011-08-01
- Assignees: Celsee Diagnostics, Inc.
- Applicants: Celsee Diagnostics, Inc.
- Inventors: Kalyan Handique
- Web page: https://patentgrep.com/patent/US10481077B1

## CPC

- G01N 15/0272: {with screening; with classification by filtering (B01D takes precedence)}
- G01N 15/1484: {microstructural devices}
- B01L 3/021: {Pipettes, i.e. with only one conduit for withdrawing and redistributing liquids}
- B01L 3/502715: {characterised by interfacing components, e.g. fluidic, electrical, optical or mechanical interfaces}
- B01L 3/502746: {characterised by the means for controlling flow resistance, e.g. flow controllers, baffles or throttle valves}
- 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}
- C12M 47/04: {Cell isolation or sorting (purging biological preparations of unwanted cells C12N5/0081, determining the presence or kind of microorganism C12Q1/04)}
- G01N 1/20: for flowing or falling materials ({G01N1/2035} G01N1/12, G01N1/14 take precedence)
- G01N 1/28: Preparing specimens for investigation {including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q}(mounting specimens on microscopic slides G02B21/34; means for supporting the objects or the materials to be analysed in electron microscopes H01J37/20 {; laboratory gas handling apparatus B01L5/00})
- G01N 1/40: Concentrating samples
- G01N 1/405: {by adsorption or absorption}
- G01N 15/1433: using image recognition
- G01N 15/1436: {the optical arrangement forming an integrated apparatus with the sample container, e.g. a flow cell}
- B01L 2200/0652: Sorting or classification of particles or molecules
- B01L 2200/0668: Trapping microscopic beads
- B01L 2300/0636: Integrated biosensor, microarrays
- B01L 2300/0654: Lenses; Optical fibres
- B01L 2300/0672: Integrated piercing tool
- B01L 2300/0816: Cards, e.g. flat sample carriers usually with flow in two horizontal directions
- B01L 2300/0819: Microarrays; Biochips
- B01L 2300/0848: Specific forms of parts of containers
- B01L 2300/0877: Flow chambers
- B01L 2300/168: Specific optical properties, e.g. reflective coatings
- B01L 2400/086: using baffles or other fixed flow obstructions
- G01N 1/4077: {by other techniques involving separation of suspended solids}
- G01N 15/01: specially adapted for biological cells, e.g. blood cells (investigating sedimentation of particle suspensions in blood G01N15/05)
- G01N 15/149: specially adapted for sorting particles, e.g. by their size or optical properties
- G01N 33/48728: {Investigating individual cells, e.g. by patch clamp, voltage clamp (investigating individual particles in general G01N15/10)}
- G01N 2015/1006: {for cytology}
- G01N 2015/1486: {Counting the particles}
- G01N 2015/1497: {Particle shape}
- G01N 2035/00158: {Elements containing microarrays, i.e. "biochip"}
- G06V 20/69: Microscopic objects, e.g. biological cells or cellular parts
- 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)
- B01L: CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
- C12M: APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; {APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS}
- G06V: IMAGE OR VIDEO RECOGNITION OR UNDERSTANDING

## Abstract

A cell capture system including an array, an inlet manifold, and an outlet manifold. The array includes a plurality of parallel pores, each pore including a chamber and a pore channel, an inlet channel fluidly connected to the chambers of the pores; an outlet channel fluidly connected to the pore channels of the pores. The inlet manifold is fluidly connected to the inlet channel, and the outlet channel is fluidly connected to the outlet channel. A cell removal tool is also disclosed, wherein the cell removal tool is configured to remove a captured cell from a pore chamber.

## Claims

### Claim 1

A method comprising:
providing a fluidic network comprising an inlet channel at an upstream end, one or more outlet channels at a downstream end, and a set of structures in fluid communication with the inlet channel and the one or more outlet channels, wherein flow from the inlet channel is configured to reach at least one of the one or more outlet channels only by way of the set of structures;
receiving a fluid sample comprising a set of target cells into the fluidic network;
capturing and partitioning the set of target cells in single-cell format, by way of the set of structures, wherein capturing and partitioning comprises isolating a target cell of the set of target cells in single-cell format with an isolation material within at least a portion of the fluidic network; and
transmitting material associated with the set of target cells from the fluidic network.

### Claim 2 (depends on 1)

The method of claim 1, wherein the set of target cells is co-captured with a set of functionalized microspheres, thereby forming a set of complexes, and wherein capturing and partitioning the set of target cells in single-cell format comprises capturing and partitioning the set of complexes in single-complex format.

### Claim 3 (depends on 1)

The method of claim 1, wherein isolating each of the set of target cells with the isolation material comprises isolating each of the set of target cells with an oil.

### Claim 4 (depends on 1)

The method of claim 1, wherein receiving the fluid sample comprises receiving a diluted fluid sample comprising the set of target cells.

### Claim 5 (depends on 1)

The method of claim 1, wherein transmitting material associated with the set of target cells from the fluidic network comprises establishing communication between the fluidic network and a removal tool, and delivering the material into the removal tool.

### Claim 6 (depends on 5)

The method of claim 5, wherein the removal tool comprises a tip configured to enter the fluidic network, extract the material from the fluidic network, and be reversibly retracted from the fluidic network with the target material.

### Claim 7 (depends on 1)

The method of claim 1, further comprising contacting the target cell with a reagent, wherein the reagent comprises a lysing reagent.

### Claim 8 (depends on 7)

The method of claim 7, further comprising performing a downstream processing operation with cellular lysate of the target cell, wherein the downstream processing operation comprises at least one of: a molecular reaction, an amplification process, immunostaining, single cell proteomic analysis, nucleic acid analysis, and genomic sequencing.

### Claim 9 (depends on 1)

The method of claim 1, wherein capturing and partitioning comprises segregating each of the set of target cells in single-cell format in an arrayed distribution within the fluidic network.

### Claim 10 (depends on 9)

The method of claim 9, wherein providing the fluidic network comprises providing a plurality of pores, each pore in the plurality of pores displaced from at least one other pore in the plurality of pores and comprising: an upstream opening and a downstream opening, having a pore channel width configured to block target particle egress,
wherein the inlet channel is fluidly coupled to the upstream end and at least one of the one or more outlet channels is directly fluidly coupled to the downstream end of each of the plurality of pores.

### Claim 11 (depends on 1)

The method of claim 1, wherein the fluidic network comprises at least one channel with a characteristic dimension of less than 100 micrometers.

### Claim 12 (depends on 1)

The method of claim 1, wherein receiving the fluid sample comprises transmitting the fluid sample into the inlet channel with a pressure difference of less than 10,000 Pascals between the inlet channel and the one or more outlet channels.

### Claim 13

A method comprising:
providing a fluidic network comprising an inlet channel, an outlet channel, and a set of structures in fluid communication with the inlet channel and one or more outlet channels, wherein flow from the inlet channel is configured to reach at least one of the one or more outlet channels only by way of the set of structures;
receiving a fluid sample comprising a set of target cells into the fluidic network;
capturing and partitioning the set of target cells in single-cell format, by way of the set of structures, wherein capturing and partitioning comprises isolating a target cell of the set of target cells in single-cell format with an isolation material within at least a portion of the fluidic network;
generating cellular lysate upon lysing the target cell with a lysis reagent; and
performing a reaction with the cellular lysate.

### Claim 14 (depends on 13)

The method of claim 13, wherein the fluid sample comprises a diluted fluid sample comprising the set of target cells.

### Claim 15 (depends on 14)

The method of claim 14, wherein the isolation material comprises an oil.

### Claim 16 (depends on 15)

The method of claim 15, wherein at least one of the set of target cells is co-captured with a functionalized microsphere, thereby forming a complex, and wherein capturing and partitioning the set of target cells in single-cell format comprises capturing the complex in the oil.

### Claim 17 (depends on 13)

The method of claim 13, further comprising transmitting the target cell from the fluidic network prior to performing the reaction.

### Claim 18 (depends on 17)

The method of claim 17, wherein the reaction comprises a molecular reaction.

### Claim 19 (depends on 17)

The method of claim 17, wherein transmitting the target cell comprises establishing communication between the fluidic network and a removal tool, delivering the material into the removal tool, and delivering the target cell from the removal tool into a container.

### Claim 20 (depends on 13)

The method of claim 13, wherein the microfluidic network comprises at least one channel with a characteristic dimension of less than 100 micrometers.