Dark quenchers for donor-acceptor energy transfer

• US 8,410,255 B2
• Filed: 08/06/2010
• Issued: 04/02/2013
• Est. Priority Date: 05/09/2000
• Status: Expired due to Fees

• Current Assignee: HSB Corporate Trust Company (UK) Limited
• Original Assignee: Biosearch Technologies Incorporated
• Inventors: Cook, Ronald M., Lyttle, Matt, Dick, Daren
• Primary Examiner(s): Riley, Jezia
• Application Number:
  US12/852,387
• Publication Number:
  US 20110178280A1
• Time in Patent Office: 970 Days
• Field of Search: 534/558, 534/561, 430/141
• US Class Current: 534/558
• CPC Class Codes: C07B 2200/11 Compounds covalently bound to a solid support
• CPC Class Codes: C07C 245/08 with the two nitrogen atoms of azo groups bound to carbon atoms of six-membered aromatic rings, e.g. azobenzene
• CPC Class Codes: C07D 241/46 Phenazines
• CPC Class Codes: C07F 9/091 with hydroxyalkyl compounds with further substituents on alkyl
• CPC Class Codes: C07F 9/2408 of hydroxyalkyl compounds
• CPC Class Codes: C07H 21/00 Compounds containing two or more mononucleotide units having separate phosphate or polyphosphate groups linked by saccharide radicals of nucleoside groups, e.g. nucleic acids
• CPC Class Codes: C09B 31/043 Amino-benzenes
• CPC Class Codes: C09B 56/00 Azo dyes containing other chromophoric systems
• CPC Class Codes: C09B 56/12 Anthraquinone-azo dyes from diazotised aminoanthracene C09B29/0022, azo dyes containing hydroxyl groups acylated with polyfunctional anthraquinone derivatives C09B43/26
• CPC Class Codes: C12Q 1/6818 involving interaction of two or more labels, e.g. resonant energy transfer
• CPC Class Codes: C40B 40/00 Libraries per se, e.g. arrays, mixtures
• CPC Class Codes: G01N 33/542 with steric inhibition or signal modification, e.g. fluorescent quenching
• CPC Class Codes: G01N 33/582 with fluorescent label

Abstract

The present invention provides a family of dark quenchers, termed Black Hole Quenchers (“BHQs”), that are efficient quenchers of excited state energy but which are themselves substantially non-fluorescent. Also provided are methods of using the BHQs, probes incorporating the BHQs and methods of using the probes.

Claims

1. A quencher of excited state energy having a formula which is a member selected from:

2. A quencher of excited state energy, having the formula:

6. The quencher of excited state energy according to claim 2, having a formula which is a member selected from:

8. The quencher of excited state energy according to claim 2, wherein R′

is unsubstituted alkyl and R″

is substituted alkyl.

9. The quencher of excited state energy according to claim 8, wherein said reactive functional group is a member selected from carboxy, N-hydroxysuccinimide ester, N-hydroxybenztriazole ester, acid halide, acyl imidazole, thioester, p-nitrophenyl ester, alkyl ester, alkenyl ester, alkynyl ester, aromatic ester, and —

NH₂.

10. The quencher of excited state energy according to claim 8, wherein X⁵ is

3. A quencher of excited state energy, having the formula:

11. The quencher of excited state energy according to claim 3, wherein R′

is unsubstituted alkyl and R″

is substituted alkyl.

4. A quencher of excited state energy, having the formula:

5. A quencher of excited state energy, having the formula:

7. The quencher of excited state energy according to claim 5, having a formula which is a member selected from:

12. The quencher of excited state energy according to claim 5,

wherein

for X⁶, R′

and R″

are each unsubstituted alkyl.

13. The quencher of excited state energy according to claim 12,

wherein

for X⁵, R′

is unsubstituted alkyl and R″

is substituted alkyl.

14. The quencher of excited state energy according to claim 13, wherein said reactive functional group is a member selected from carboxy, N-hydroxysuccinimide ester, N-hydroxybenztriazole ester, acid halide, acyl imidazole, thioester, p-nitrophenyl ester, alkyl ester, alkenyl ester, alkynyl ester, aromatic ester, and —

NH₂.

15. The quencher of excited state energy according to claim 13, wherein X⁵ is