Cyanine compounds and their application as quenching compounds

• US 9,150,922 B2
• Filed: 12/17/2013
• Issued: 10/06/2015
• Est. Priority Date: 08/11/2003
• Status: Active Grant

• Current Assignee: Life Technologies Corporation (Thermo Fisher Scientific Incorporated)
• Original Assignee: Life Technologies Corporation (Thermo Fisher Scientific Incorporated)
• Inventors: Haugland, Rosaria, Cheung, Ching-Ying, Yue, Stephen
• Primary Examiner(s): KOSACK, JOSEPH R
• Application Number:
  US14/109,475
• Publication Number:
  US 20140295437A1
• Time in Patent Office: 658 Days
• US Class Current: 1/1
• CPC Class Codes: C07D 209/12 Radicals substituted by oxygen atoms
• CPC Class Codes: C07D 209/14 Radicals substituted by nitrogen atoms, not forming part of a nitro radical
• CPC Class Codes: C07D 209/24 with an alkyl or cycloalkyl radical attached to the ring nitrogen atom
• CPC Class Codes: C07D 401/04 directly linked by a ring-member-to-ring-member bond
• CPC Class Codes: C09B 23/12 the polymethine chain being branched "branched" means that the substituent on the polymethine chain forms a new conjugated system, e.g. most trinuclear cyanine dyes
• CPC Class Codes: C12Q 1/6876 Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes

Abstract

The present invention provides methods and non-fluorescent carbocyanine quencher compounds having the general formula:

Wherein the A moiety is a substituted pyridinium, unsubstituted pyridinium, substituted quinolinium, unsubstituted quinolinium, substituted benzazolium, unsubstituted benzazolium, substituted indolinium, or substituted indolinium. The invention further provides luminescent donor molecule-quencher pairs and luminescent donor molecule-quencher-luminescent acceptor molecule conjugates wherein the quencher is a cyanine compound of the present invention. The energy transfer pairs are used to detect an analyte of interest in a sample.

Claims

1. A compound having the formula:

2. The compound according to claim 1, wherein the reactive group, solid support and carrier molecule independently comprise a linker that is a single covalent bond, or a covalent linkage that is linear or branched, cyclic or heterocyclic, saturated or unsaturated, having 1-20 nonhydrogen atoms selected from the group consisting of C, N, P, O and S;

and are composed of any combination of ether, thioether, amine, ester, carboxamide, sulfonamide, hydrazide bonds and aromatic or heteroaromatic bonds.

3. The compound according to claim 1, wherein the reactive group is an acrylamide, an activated ester of a carboxylic acid, a carboxylic ester, an acyl azide, an acyl nitrile, an aldehyde, an alkyl halide, an anhydride, an aniline, an amine, an aryl halide, an azide, an aziridine, a boronate, a diazoalkane, a haloacetamide, a haloalkyl, a halotriazine, a hydrazine, an imido ester, an isocyanate, an isothiocyanate, a maleimide, a phosphoramidite, a reactive platinum complex, a silyl halide, a sulfonyl halide, a thiol or a photoactivatable group.

4. The compound according to claim 1, wherein the reactive group is carboxylic acid, succinimidyl ester of a carboxylic acid, hydrazide, amine or a maleimide.

5. The compound according to claim 1, wherein the carrier molecule is an amino acid, a peptide, a protein, a polysaccharide, a nucleoside, a nucleotide, an oligonucleotide, a nucleic acid polymer, a hapten, a biotin-binding protein, a psoralen, a drug, a hormone, a lipid, a lipid assembly, a synthetic polymer, a polymeric microparticle, a biological cell or a virus.

6. The compound according to claim 1, wherein the carrier molecule is an antibody or fragment thereof, an avidin or streptavidin, a biotin, a blood component protein, a dextran, an enzyme, an enzyme inhibitor, a hormone, an IgG binding protein, a fluorescent protein, a growth factor, a lectin, a lipopolysaccharide, a microorganism, a metal binding protein, a metal chelating moiety, a non-biological microparticle, a peptide toxin, a phosphotidylserine-binding protein, a structural protein, a small-molecule drug, or a tyramide.

7. The compound according to claim 1, wherein the solid support is a microfluidic chip, a silicon chip, a microscope slide, a microplate well, a silica gel, a polymeric membrane, a particle, a derivatized plastic film, a glass bead, cotton, a plastic bead, an alumina gel, a polysaccharide, polyvinylchloride, polypropylene, polyethylene, nylon, latex bead, magnetic bead, paramagnetic bead, or superparamagnetic bead.

8. The compound according to claim 1, wherein the solid support is Sepharose, poly(acrylate), polystyrene, poly(acrylamide), polyol, agarose, agar, cellulose, dextran, starch, FICOLL, heparin, glycogen, amylopectin, mannan, inulin, nitrocellulose, diazocellulose or starch.

9. A composition comprising;

a) a luminescent donor molecule;

b) a luminescent acceptor molecule; and

c) a compound that is non-fluorescent acceptor molecule, wherein the compound has the formula according to claim 1.

11. A method for detecting the presence or absence of an analyte in a sample, wherein the method comprises:

a) contacting the sample with a composition to prepare a labeled sample, wherein the composition comprises a carrier molecule, a luminescent donor molecule, a luminescent acceptor molecule and a compound having the formula according to claim 1;

b) incubating the labeled sample for a sufficient amount of time for the carrier molecule to associate with the analyte to form an incubated sample;

c) illuminating the incubated sample with an appropriate wavelength to form an illuminated sample; and

d) observing the illuminated sample, whereby the presence or absence of the analyte in the sample is detected.

14. A kit for labeling a carrier molecule, wherein the kit comprises:

a non-fluorescent compound, wherein the compound has the formula according to claim 1.

10. A method for absorbing residual energy from an energy donor molecule during energy transfer between a luminescent donor molecule and a luminescent acceptor molecule, wherein the method comprises:

a) covalently bonding a luminescent donor molecule to a luminescent acceptor molecule to prepare an energy transfer pair;

b) covalently bonding a compound to the energy transfer pair to prepare a quencher labeled energy transfer pair, wherein the compound has the formula;

12. A kit for detecting a ligand in a sample, wherein the kit comprises a composition comprising a carrier molecule, a luminescent donor molecule, a luminescent acceptor molecule, and a non-fluorescent acceptor molecule having the formula:

13. The kit according to claim 12, further comprising instructions for detecting the ligand.