# US 2019/0376977 A1 · Fluorescent methods and materials for directed biomarker signal amplification

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- Number: US 2019/0376977 A1
- Document ID: US20190376977A1
- Status: Published application
- Kind: A1
- Application number: 16/528,469
- Filed: 2019-07-31
- Published: 2019-12-12
- Earliest claimed: 2006-10-06
- Assignees: Sirigen, Inc.
- Applicants: Sirigen Inc.
- Inventors: Brent S. Gaylord; Janice W. Hong; Tsu-Ju Fu; ChengJun Sun; Russell A. Baldocchi
- Web page: https://patentgrep.com/patent/US20190376977A1

## CPC

- G01N 33/582: {with fluorescent label}
- C08G 61/02: Macromolecular compounds containing only carbon atoms in the main chain of the macromolecule, e.g. polyxylylenes
- C08G 61/12: Macromolecular compounds containing atoms other than carbon in the main chain of the macromolecule
- G01N 33/542: with steric inhibition or signal modification, e.g. fluorescent quenching
- C08G 2261/3142: fluorene-based, e.g. fluorene, indenofluorene, or spirobifluorene
- C08G 2261/3221: containing one or more nitrogen atoms as the only heteroatom, e.g. pyrrole, pyridine or triazole
- C08G 2261/3241: containing one or more nitrogen atoms as the only heteroatom, e.g. carbazole
- C08G 2261/3246: containing nitrogen and sulfur as heteroatoms
- C08G 2261/522: fluorescent
- C08G 2261/78: Complexation
- C08G 2261/94: Applications in sensors, e.g. biosensors
- H10K 85/115: {Polyfluorene; Derivatives thereof}
- Y10T 436/13: Tracers or tags
- 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)
- C08G: MACROMOLECULAR COMPOUNDS OBTAINED OTHERWISE THAN BY REACTIONS ONLY INVOLVING UNSATURATED CARBON-TO-CARBON BONDS
- H10K: ORGANIC ELECTRIC SOLID-STATE DEVICES
- Y10T: TECHNICAL SUBJECTS COVERED BY FORMER US CLASSIFICATION

## Abstract

Methods and compositions are provided that include a multichromophore and/or multichromophore complex for identifying a target biomolecule. A sensor biomolecule, for example, an antibody can be covalently linked to the multichromophore. Additionally, a signaling chromophore can be covalently linked to the multichromophore. The arrangement is such that the signaling chromophore is capable of receiving energy from the multichromophore upon excitation of the multichromophore. Since the sensor biomolecule is capable of interacting with the target biomolecule, the multichromophore and/or multichromophore complex can provide enhanced detection signals for a target biomolecule.

## Claims

### Claim at position 1

-71. (canceled)

### Claim 72

A multichromophore-biomolecule conjugate comprising:
a sensor biomolecule covalently linked to a light harvesting water soluble multichromophore via a bioconjugation site;
wherein the light harvesting water soluble multichromophore is a conjugated polymer having a backbone of π-conjugated repeat units and comprises the structure:
{{[CP1]-a[CP2]b}m[CP1]-a[CP3]c}n
wherein:
CP1 is present and is a polycyclic repeat unit;
CP2 is an aromatic repeat unit;
CP3 is an aromatic repeat unit comprising a bioconjugation functional group capable of covalently linking to a biomolecule or dye;
m is 1 to 10,0000;
n is independently 0 to 10,000 and; and
a, b and c are independently 0 to 250 wherein a+b+c>1.

### Claim 73 (depends on 72)

The multichromophore-biomolecule conjugate according to claim 72, wherein CP1 comprises 1 to 5 aromatic rings.

### Claim 74 (depends on 72)

The multichromophore-biomolecule conjugate according to claim 72, wherein CP1 comprises up to 5 fused and/or bridged rings.

### Claim 75 (depends on 72)

The multichromophore-biomolecule conjugate according to claim 72, wherein CP1 comprises a biphenyl.

### Claim 76 (depends on 75)

The multichromophore-biomolecule conjugate according to claim 75, wherein the biphenyl is bridged.

### Claim 77 (depends on 72)

The multichromophore-biomolecule conjugate according to claim 72, wherein the multichromophore is substituted with a plurality of charge neutral water-soluble groups.

### Claim 78 (depends on 72)

The multichromophore-biomolecule conjugate according to claim 72, wherein CP1 and CP2 are each independently substituted with one or more water-soluble groups.

### Claim 79 (depends on 72)

The multichromophore-biomolecule conjugate according to claim 72, wherein the water-soluble groups comprise one or more groups selected from ethylene glycol oligomer, ethylene glycol polymer, ω-ammonium alkoxy salt and ω-sulfonate alkoxy salt.

### Claim 80 (depends on 72)

The multichromophore-biomolecule conjugate according to claim 72, wherein the sensor biomolecule is a protein, an antibody, a nucleic acid, an affinity ligand, a sugar, a lipid or a peptide.

### Claim 81 (depends on 80)

The multichromophore-biomolecule conjugate according to claim 80, wherein the sensor biomolecule is a protein, an antibody or a nucleic acid.

### Claim 82 (depends on 81)

The multichromophore-biomolecule conjugate according to claim 81, wherein the sensor biomolecule is an antibody.

### Claim 83 (depends on 71)

The multichromophore-biomolecule conjugate according to claim 71, wherein the bioconjugation site results from a linking chemistry selected from maleimide, thiol, succimidylester (NHS ester), amine, azide chemistry, carboxy/EDC (1-Ethyl-3-[3-dimethylaminopropyl]carbodiimide hydrochloride, sulfo-SMCC (sulfosuccinimidyl4-[N-maleimidomethyl]cyclohexane-1-carboxylate), amine/BMPH (N-[β-Maleimidopropionic acid]hydrazide.TFA) and sulfo-SBED sulfosuccinimidyl[2-6-(biotinamido)-2-(p-azidobenzamido)-hexanoamido]-ethyl-1,3′-dithiopropionate.

### Claim 84 (depends on 83)

The multichromophore-biomolecule conjugate according to claim 83, wherein the light harvesting multichromophore further comprises the structure:
wherein: L is a linker to a bioconjugation site A covalently linked to the sensor biomolecule; and *=site for covalent attachment to unsaturated backbone of the multichromophore.

### Claim 85

A light harvesting water soluble multichromophore comprising a conjugated polymer having a backbone of π-conjugated repeat units and a bioconjugation functional group capable of covalently linking to a sensor biomolecule, wherein the multichromophore has the structure:
{{[CP1]-a[CP2]b}m[CP1]-a[CP3]c}n
wherein:
CP1 is present and is a polycyclic repeat unit;
CP2 is an aromatic repeat unit;
CP3 is an aromatic repeat unit comprising a bioconjugation functional group capable of covalently linking to a biomolecule or dye;
m is 1 to 10,0000;
n is independently 0 to 10,000 and; and
a, b and c are independently 0 to 250 wherein a+b+c>1.

### Claim 86 (depends on 85)

The light harvesting water soluble multichromophore according to claim 85, wherein CP1 comprises 1 to 5 aromatic rings.

### Claim 87 (depends on 85)

The light harvesting water soluble multichromophore according to claim 85, wherein CP1 comprises up to 5 fused and/or bridged rings.

### Claim 88 (depends on 85)

The light harvesting water soluble multichromophore according to claim 85, wherein CP1 comprises a biphenyl.

### Claim 89 (depends on 88)

The light harvesting water soluble multichromophore according to claim 88, wherein the biphenyl is bridged.

### Claim 90 (depends on 85)

The light harvesting water soluble multichromophore according to claim 85, wherein the multichromophore is substituted with a plurality of charge neutral water-soluble groups.

### Claim 91 (depends on 90)

The light harvesting water soluble multichromophore according to claim 90, wherein CP1 and CP2 are each independently substituted with one or more water-soluble groups.

### Claim 92 (depends on 91)

The light harvesting water soluble multichromophore according to claim 91, wherein the water-soluble groups comprise one or more groups selected from ethylene glycol oligomer, ethylene glycol polymer, ω-ammonium alkoxy salt and ω-sulfonate alkoxy salt.

### Claim 93 (depends on 85)

The light harvesting water soluble multichromophore according to claim 85, wherein the bioconjugation functional group is selected from maleimide, thiol, succimidyl ester, amine, azide and carboxy/EDC (1-Ethyl-3-[3-dimethylaminopropyl]carbodiimide hydrochloride).