# US 2014/0193805 A1 · HPV E6, E7 MRNA Assay and Methods of Use Thereof

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- Number: US 2014/0193805 A1
- Document ID: US20140193805A1
- Status: Published application
- Kind: A1
- Application number: 14/149,747
- Filed: 2014-01-07
- Published: 2014-07-10
- Earliest claimed: 2005-02-02
- Assignees: IncellDx, Inc.
- Applicants: IncellDx, Inc.
- Inventors: Bruce K. Patterson
- Web page: https://patentgrep.com/patent/US20140193805A1

## CPC

- C12Q 1/708: {for papilloma}
- C12Q 1/6841: In situ hybridisation
- G16Z 99/00: Subject matter not provided for in other main groups of this subclass
- C12Q 2600/158: Expression markers
- 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
- G16Z: INFORMATION AND COMMUNICATION TECHNOLOGY [ICT] SPECIALLY ADAPTED FOR SPECIFIC APPLICATION FIELDS, NOT OTHERWISE PROVIDED FOR

## Abstract

Provided is an HPV E6, E7 mRNA assay, referenced herein as the “In Cell HPV Assay,” that is capable of sensitive and specific detection of normal cervical cells undergoing malignant transformation as well as abnormal cervical cells with pre-malignant or malignant lesions. The In Cell HPV Assay identifies HPV E6, E7 mRNA via in situ hybridization with oligonucleotides specific for HPV E6, E7 mRNA and quantitates the HPV E6, E7 mRNA via flow cytometry. The In Cell HPV Assay can be carried out in less than three hours directly from liquid-based cervical (“LBC”) cytology specimens. The In Cell HPV Assay provides an efficient and highly sensitive alternative to the Pap smear for determining abnormal cervical cytology.

## Claims

### Claim at position 1

1-28. (canceled)

### Claim 29

A set of reagents comprising:
HPV E6, E7 mRNA specific oligonucleotides; and
a permeabilization reagent.

### Claim 30 (depends on 29)

The set of reagents according to claim 29, further comprising a first pre-hybridization wash fluid.

### Claim 31 (depends on 30)

The set of reagents according to claim 30, further comprising a second pre-hybridization wash fluid.

### Claim 32 (depends on 29)

The set of reagents according to claim 29, further comprises a hybridization mix.

### Claim 33 (depends on 29)

The set of reagents according to claim 29, wherein the HPV specific E6, E7 mRNA oligonucleotides are labeled.

### Claim 34 (depends on 33)

The set of reagents according to claim 33, wherein the HPV specific E6, E7 mRNA oligonucleotides are labeled with fluorescein.

### Claim 35 (depends on 34)

The set of reagents according to claim 34, wherein the HPV specific E6, E7 mRNA oligonucleotides are 3′ and 5′ labeled with fluorescein.

### Claim 36 (depends on 29)

The set of reagents according to claim 29, wherein the permeablization reagent is a fixation reagent.

### Claim 37 (depends on 36)

The set of reagents according to claim 36, wherein the set further comprises a first post-hybridization stringency wash.

### Claim 38 (depends on 37)

The set of reagents according to claim 37, wherein the set further comprises a second post-hybridization stringency wash.

### Claim 39 (depends on 29)

The set of reagents according to claim 29, further comprising a first antibody.

### Claim 40 (depends on 39)

The set of reagents according to claim 39, further comprising a second antibody.

### Claim 41 (depends on 40)

The set of reagents according to claim 40, wherein the first and second antibodies are labeled.

### Claim 42 (depends on 41)

The set of reagents according to claim 41, wherein the first and second antibodies are fluorescently labeled.

### Claim 43 (depends on 41)

The set of reagents according to claim 41, wherein the first and second antibodies comprise anti-CD16 and anti-CAM5.2.

### Claim 44

A set of reagents comprising:
HPV E6, E7 mRNA specific oligonucleotides;
a fixation/permeabilization reagent;
first and second pre-hybridization wash fluids;
a hybridization mix; and
first and second post-hybridization wash fluids.

### Claim 45 (depends on 44)

The set of reagents according to claim 44, wherein the HPV specific E6, E7 mRNA oligonucleotides are labeled.

### Claim 46 (depends on 45)

The set of reagents according to claim 45, wherein the HPV specific E6, E7 mRNA oligonucleotides are labeled with fluorescein.

### Claim 47 (depends on 46)

The set of reagents according to claim 46, wherein the HPV specific E6, E7 mRNA oligonucleotides are 3′ and 5′ labeled with fluorescein.

### Claim 48 (depends on 44)

The set of reagents according to claim 44, further comprising a first antibody.

### Claim 49 (depends on 48)

The set of reagents according to claim 48, further comprising a second antibody.