# US 11,796,548 B2 · Continuous stable cell line for identification of infectious African swine fever virus in clinical samples

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- Number: US 11,796,548 B2
- Document ID: US11796548B2
- Status: Granted
- Kind: B2
- Application number: 17/236,284
- Filed: 2021-04-21
- Published: 2021-10-28
- Granted: 2023-10-24
- Earliest claimed: 2020-04-22
- Assignees: The United States of America as represented by the Secretary of Agriculture
- Applicants: The United States of America, as represented by the Secretary of Agriculture
- Inventors: Douglas P. Gladue; Ayushi Rai; Manuel V. Borca
- Web page: https://patentgrep.com/patent/US11796548B2

## CPC

- G01N 33/56983: {Viruses}
- G01N 33/80: involving blood groups or blood types {or red blood cells (white blood cells G01N33/56972)}
- C12N 7/00: Viruses; Bacteriophages; Compositions thereof; Preparation or purification thereof (preparing medicinal viral antigen or antibody compositions, e.g. virus vaccines, A61K39/00)
- C12N 2710/12031: Uses of virus other than therapeutic or vaccine, e.g. disinfectant
- C12N 2710/12051: Methods of production or purification of viral material
- G01N 2333/01: DNA viruses
- G01N 2800/26: Infectious diseases, e.g. generalised sepsis
- 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)
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)

## Abstract

Methods are provided herein utilizing a stable cell line capable of efficient infection by African swine fever virus (ASFV) and also provides for the detection of the presence of virus in samples applied to the cells. Detection of the virus by means such as red blood cell rosetting is a surprising result given that the cell line is derived from African green monkeys. This cell line provides a marked improvement over the currently available testing strategies.

## Claims

### Claim 1

A method of detecting African Swine Fever Virus (ASFV) in a sample, comprising the steps of:
a. contacting the sample with a culture of Ma-104 cells under conditions that allow for infection of the cells by ASFV;
b. incubating the Ma-104 cell and sample mixture under conditions that allow for ASFV detection; and
c. analyzing the mixture from step b for the presence or absence of replicating ASFV.

### Claim 2 (depends on 1)

The method of claim 1, wherein the ASFV comprises a strain of ASFV that is not attenuated for growth in Ma-104 cells.

### Claim 3 (depends on 1)

The method of claim 1, wherein the ASFV comprises a strain of ASFV that has not been passaged in cell culture prior to contacting with Ma-104 cells.

### Claim 4 (depends on 1)

The method of claim 1, wherein the ASFV comprises the Georgia/2007 strain.

### Claim 5 (depends on 1)

The method of claim 1, wherein the sample comprises a sample from a swine.

### Claim 6 (depends on 1)

The method of claim 1, wherein the analyzing step comprises adding red blood cells to the mixture from step b, and examining the Ma-104 cells for the presence or absence of hemadsorbed red blood cells on the Ma-104 cells.

### Claim 7 (depends on 6)

The method of claim 6, wherein the presence or absence of hemadsorbed red blood cells is determined within 24 hours after the red blood cells are added to the Ma-104 cell and sample mixture.

### Claim 8

A composition comprising Ma-104 cells, African Swine Fever Virus (ASFV), and red blood cells.

### Claim 9 (depends on 8)

The composition of claim 8, wherein the ASFV comprises the Georgia/2007 strain.

### Claim 10 (depends on 1)

The composition of claim 1, wherein the Ma-104 cells are genetically unmodified.

### Claim 11 (depends on 8)

The composition of claim 8, wherein the Ma-104 cells are genetically unmodified.