# US 10,246,738 B2 · DNA-antigen exchange and amplification

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- Number: US 10,246,738 B2
- Document ID: US10246738B2
- Status: Granted
- Kind: B2
- Application number: 15/941,770
- Filed: 2018-03-30
- Published: 2018-10-04
- Granted: 2019-04-02
- Earliest claimed: 2017-03-31
- Assignees: Ultivue, Inc.
- Applicants: Ultivue, Inc.
- Inventors: Stephanie Rae Hennek; Mael Manesse
- Web page: https://patentgrep.com/patent/US10246738B2

## CPC

- C12Q 1/682: Signal amplification
- C12Q 1/6804: Nucleic acid analysis using immunogens (immunoassay G01N33/53)
- C12Q 1/6844: Nucleic acid amplification reactions
- G01N 33/5308: {for analytes not provided for elsewhere, e.g. nucleic acids, uric acid, worms, mites}
- G01N 33/582: {with fluorescent label}
- G01N 2458/10: Oligonucleotides as tagging agents for labelling antibodies
- 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
- 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)

## Abstract

Methods for imaging are described, including, but not limited to a method comprising: (1) contacting a sample being tested for the presence of one or more targets with one or more target-specific binding partners, wherein each target-specific binding partner is linked to a docking strand, and wherein target-specific binding partners of different specificity are linked to different docking strands, (2) optionally removing unbound target-specific binding partners, (3) contacting the sample with antigen-bound imager strands and antigen-specific binding partners linked (directly or indirectly) to optical labels, wherein the antigen-bound imager strands have complementarity to a docking strand, directly or indirectly, and wherein each antigen-specific binding partner is linked to one or more optical labels, and wherein antigen-specific binding partners of different specificity are linked to distinct optical labels, (4) optionally removing unbound antigen-bound imager strands and/or antigen-specific binding partners, (5) imaging the sample to detect bound labeled antigen-specific binding partners, (6) optionally removing/extinguishing signal from the optical labels, and (7) optionally repeating steps (1)-(7), or any subset thereof.

## Claims

### Claim 1

A method comprising:
(1) contacting a sample being tested for the presence of one or more targets with one or more target-specific binding partners, wherein each target-specific binding partner is linked to a first docking strand, and wherein target-specific binding partners of different specificity are linked to different docking strands,
(2) optionally removing unbound target-specific binding partners,
(3) contacting the sample with antigen-bound imager strands and antigen-specific binding partners,
wherein the antigen-bound imager strands have complementarity to the first docking strand, directly or indirectly, and
wherein each antigen-specific binding partner is linked to at least one second docking strand, and wherein the antigen specific binding partner binds to the antigen of the antigen-bound imager strand;
(4) optionally removing unbound antigen-bound imager strands and/or antigen specific binding partners,
(5) increasing the number of second docking strands by a DNA amplification reaction and labeling of the at least one second docking strand using an optical label,
(6) imaging the sample to detect bound labeled antigen-specific binding partners,
(7) optionally extinguishing the signal from the optical label, and
(8) optionally repeating steps (1)-(7), or any subset thereof.

### Claim 2 (depends on 1)

The method of claim 1, wherein the first docking strand and the at least one second docking strand have a different sequence.

### Claim 3 (depends on 1)

The method of claim 1, wherein the first docking strand and the at least one second docking strand have the same sequence.

### Claim 4 (depends on 1)

The method of claim 1, wherein the at least one second docking strand is directly labeled using labeled nucleotides in the DNA amplification reaction.

### Claim 5 (depends on 4)

The method of claim 4, wherein the labeled nucleotides are fluorescently-labeled nucleotides.

### Claim 6 (depends on 1)

The method of claim 1, wherein the at least one second docking strand is labeled using a labeled nucleic acid that binds thereto.

### Claim 7 (depends on 1)

The method of claim 1, wherein the DNA amplification reaction comprises rolling circle amplification.

### Claim 8 (depends on 1)

The method of claim 1, wherein the DNA amplification reaction comprises hybridization chain reaction.

### Claim 9 (depends on 1)

The method of claim 1, wherein the DNA amplification reaction comprises hairpin-based concatemerization (including dendrimerization) reaction.