# US 12,215,307 B2 · Culture device and methods for enumerating mold colonies

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- Number: US 12,215,307 B2
- Document ID: US12215307B2
- Status: Granted
- Kind: B2
- Application number: 17/304,185
- Filed: 2021-06-16
- Published: 2021-10-14
- Granted: 2025-02-04
- Earliest claimed: 2013-05-06
- Applicants: Neogen Food Safety US Holdco Corporation
- Inventors: Kurt J. Halverson
- Web page: https://patentgrep.com/patent/US12215307B2

## 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 23/04: {Flat or tray type, drawers (C12M23/10, C12M23/12, C12M23/16 take precedence)}
- C12M 23/20: {Material Coatings (immunocoatings C12M25/00)}
- C12M 23/22: {Transparent or translucent parts (glassware for laboratory use B01L3/00)}
- C12M 23/24: {Gas permeable parts}
- C12M 23/28: {disposable or single use}
- C12M 23/48: {Holding appliances; Racks; Supports (holding devices for laboratory apparatus B01L9/00)}
- C12M 25/02: {Membranes; Filters (filters or filtration in general B01D24/00-B01D41/00)}
- C12M 25/06: {Plates; Walls; Drawers; Multilayer plates}
- C12Q 1/045: {Culture media therefor}
- 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 thin film culture device for enumerating mold colonies is provided. The device comprises water-resistant first and second substrates with a growth region disposed therebetween, a dry, cold water-soluble gelling agent disposed in the growth region, and an effective amount of a calcium-chelating compound disposed in the growth region. The effective amount of calcium-chelating compound is capable of reducing a rate of lateral enlargement of the colony-forming unit growing in the culture device relative to the rate of lateral enlargement of a colony of the same mold species growing in an otherwise identical culture device that does not contain the effective amount disposed in the growth region, wherein reducing the rate of lateral enlargement of the colony-forming unit does not substantially delay detection of the colony. A corresponding method is also provided.

## Claims

### Claim 1

A thin-film culture device, comprising:
a water-resistant first substrate;
a water-resistant second substrate;
a growth region disposed between the first and second substrates,
wherein the growth region comprises:
(a) an effective amount of calcium-chelating compound,
(b) a nutrient medium capable of supporting growth of a mold microorganism,
(c) a dry coating comprising ethylenediamine tetraacetic acid, and
(d) a dry, cold-water-soluble gelling agent;
wherein, when the growth region is hydrated with a predetermined volume of aqueous liquid and inoculated with a colony-forming unit of a mold species, the effective amount of calcium-chelating compound is capable of reducing a rate of lateral enlargement of the colony-forming unit growing in the culture device relative to the rate of lateral enlargement of a colony of the same mold species growing in an otherwise identical culture device that does not contain the effective amount disposed in the growth region;
wherein reducing the rate of lateral enlargement of the colony-forming unit does not substantially delay detection of the colony.

### Claim 2 (depends on 1)

The thin-film culture device of claim 1, wherein the ethylenediamine tetraacetic acid comprises disodium ethylenediamine tetraacetic acid dihydrate.

### Claim 3 (depends on 1)

The thin-film culture device of claim 1, further comprising a calcium salt disposed in the growth region.

### Claim 4 (depends on 3)

The thin-film culture device of claim 3, wherein the calcium salt is disposed in the dry coating.

### Claim 5 (depends on 1)

The thin-film culture device of claim 1, wherein the dry, cold-water soluble gelling agent is disposed in the first dry coating.

### Claim 6 (depends on 1)

The thin-film culture device of claim 1, wherein the ethylenediamine tetraacetic acid is the calcium-chelating compound.

### Claim 7 (depends on 1)

The thin-film culture device of claim 1, further comprising an indicator reagent.

### Claim 8 (depends on 1)

The thin film culture device of claim 1, further comprising an air-permeable membrane adhered to the first substrate, wherein the air-permeable membrane is substantially coextensive with the growth region of the culture device.

### Claim 9 (depends on 1)

The thin-film culture device of claim 1, further comprising a predefined volume of an aqueous liquid.

### Claim 10

A kit comprising a thin-film culture device and a container, wherein the thin-film culture device comprises:
a water-resistant first substrate;
a water-resistant second substrate;
a growth region disposed between the first and second substrates, wherein the growth region comprises:
a nutrient medium capable of supporting growth of a mold microorganism, and
a dry, cold-water-soluble gelling agent; and
a predetermined quantity of ethylenediamine tetraacetic acid.

### Claim 11 (depends on 10)

The kit of claim 10, wherein the ethylenediamine tetraacetic acid is disodium ethylenediamine tetraacetic acid dihydrate.

### Claim 12 (depends on 10)

The kit of claim 10, further comprising a reagent selected from the group consisting of a nutrient, a detection reagent, a selective agent, a buffering agent, a dye, and a mixture comprising any two or more of the foregoing reagents.