# US 12,600,935 B2 · Device for forming and counting spheroids and method for producing same, and spheroid culturing method and counting method using same

> Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt

- Number: US 12,600,935 B2
- Document ID: US12600935B2
- Status: Granted
- Kind: B2
- Application number: 16/760,583
- Filed: 2018-10-25
- Published: 2020-08-20
- Granted: 2026-04-14
- Earliest claimed: 2017-11-03
- Assignees: Korea Research Institute of Chemical Technology; The Industry & Academic Cooperation in Chungnam National University (IAC)
- Applicants: Korea Research Institute of Chemical Technology; The Industry & Academic Cooperation In Chungnam National University (Iac)
- Inventors: Sun Woong Kang; Kang Moo Huh; Sung Hwan Moon; Da Eun Kim; Hye Min Oh; Hye Eun Shim
- Web page: https://patentgrep.com/patent/US12600935B2

## CPC

- C12M 41/36: {of biomass, e.g. colony counters or by turbidity measurements (electrooptical investigation of individual particles G01N15/14, flow cytometers G01N15/1404)}
- C12M 23/20: {Material Coatings (immunocoatings C12M25/00)}
- C12M 23/22: {Transparent or translucent parts (glassware for laboratory use B01L3/00)}
- C12M 25/02: {Membranes; Filters (filters or filtration in general B01D24/00-B01D41/00)}
- C12Q 1/06: Quantitative determination
- 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}
- C12Q: MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS (immunoassay G01N33/53); COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES

## Abstract

A device for forming and counting spheroids according to one embodiment of the present invention may comprise a substrate, and a mesh member which is configured to be able to be positioned on the surface of the substrate and has a plurality of protruding members. The plurality of protruding members intersects one another in the form of a mesh so as to form a plurality of pores.

## Claims

### Claim 1

A device for forming and counting spheroids, comprising:
a flat substrate; and
a flexible mesh member, configured with a plurality of protruding members intersecting parallel to each other in a mesh configuration to form a plurality of pores,
wherein the substrate and the mesh member are each coated with hexanoyl glycol chitosan;
wherein each pore is configured to have a flat bottom surface and a suitable size to accommodate one spheroid of uniform size;
wherein the mesh member is fixed by an assembly member positioned at the side of the substrate to place the mesh member in a position above the surface of the substrate and to provide a gap between the mesh member and the surface of the substrate; and
wherein the gap between the mesh member and the surface of the substrate is smaller than the size of the spheroid.

### Claim 2 (depends on 1)

The device for forming and counting spheroids according to claim 1, wherein the pores have a width of 50 to 500 μm.

### Claim 3 (depends on 1)

The device for forming and counting spheroids according to claim 1, wherein the substrate is a slide glass.

### Claim 4 (depends on 1)

The device for forming and counting spheroids according to claim 1, wherein the protruding members comprise:
a plurality of first protruding members extending in a first direction and spaced apart from one another; and
a plurality of second protruding members extending in a second direction intersecting with the first direction and spaced apart from one another.

### Claim 5 (depends on 1)

A method for manufacturing the device for forming and counting spheroids according to claim 1, comprising:
a solution preparation step of preparing a hexanoyl glycol chitosan solution;
a substrate coating step of coating the substrate by spraying the prepared solution onto the substrate, followed by drying the substrate;
a mesh member coating step of coating the mesh member by spraying the prepared solution onto the mesh member, followed by drying the mesh member, and
a step of assembling the coated substrate and the mesh member using the assembly member.

### Claim 6 (depends on 5)

The method for manufacturing the device for forming and counting spheroids according to claim 5, wherein in the mesh member coating step, the mesh member is sprayed with the prepared solution while the mesh member is in a state of being erected and fixed.

### Claim 7 (depends on 5)

The method for manufacturing the device for forming and counting spheroids according to claim 5, wherein the hexanoyl glycol chitosan solution has a concentration of 0.01 to 1 wt %.

### Claim 8 (depends on 1)

A method for culturing spheroids using the device for forming and counting spheroids according to claim 1 comprising:
seeding a plurality of cells in the device;
and
obtaining spheroids in each pore of the device after several days.

### Claim 9 (depends on 1)

A method for counting spheroids using the device for forming and counting spheroids according to claim 1 comprising:
seeding spheroids in the device,
positioning the spheroids into each pore of the device by shaking the device in both directions parallel to one face of the device;
placing the device on a stage of a microscope; and
counting the spheroids in a field of view by moving a position on an image in the microscope.