Energy transfer hybridization assay composition

• US 6,239,271 B1
• Filed: 08/31/1999
• Issued: 05/29/2001
• Est. Priority Date: 02/24/1989
• Status: Expired due to Term

• Current Assignee: Enzo Diagnostics, Inc.
• Original Assignee: Enzo Diagnostics, Inc.
• Inventors: Rabbani, Elazar, Hurley, Ian
• Primary Examiner(s): Horlick, Kenneth R.
• Application Number:
  US09/386,695
• Time in Patent Office: 637 Days
• Field of Search: 536/24.3, 435/6, 435/810, 436/94
• US Class Current: 536/24.3
• 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: C07H 23/00 Compounds containing boron, silicon, or a metal, e.g. chelates, vitamin B12 esters with inorganic acids C07H11/00; metal salts, see parent compounds
• CPC Class Codes: C12Q 1/6818 involving interaction of two or more labels, e.g. resonant energy transfer
• CPC Class Codes: C12Q 1/6825 Nucleic acid detection involving sensors
• CPC Class Codes: C12Q 2563/137 Metal/ion, e.g. metal label
• CPC Class Codes: C12Q 2563/173 staining/intercalating agent, e.g. ethidium bromide

Abstract

Disclosed is a nucleic acid hybridization assay composition for detecting the presence or absence of a target oligo- or polynucleotide in a sample. The composition comprises: a solid matrix having at least one surface which is substituted with a first intercalator capable of binding dsDNA, dsRNA, or DNA-RNA hybrids; a second intercalator, which may or may not comprise at least one fluorophore, said intercalator or said fluorophore, each acting as either an energy donor or an energy acceptor; and an oligo- or polynucleotide probe which is specifically hybridizable with the target oligo- or polynucleotide and has directly or indirectly bound thereto, at least one lanthanide metal chelate or at least one fluorophore, each acting as either an energy donor or an energy acceptor. Also disclosed are a method and kit for its use.

Claims

1. A nucleic acid hybridization assay composition for detecting the presence or absence of a target oligo- or polynucleotide in a sample, which composition comprises:

a solid matrix having at least one surface to which is fixed or immobilized a first intercalator capable of binding dsDNA, dsRNA or DNA-RNA hybrids;

a second intercalator, which may or may not comprise at least one fluorophore, said second intercalator or said fluorophore, each acting as either an energy donor or an energy acceptor; and

an oligo- or polynucleotide probe which is specifically hybridizable with the target oligo- or polynucleotide and has directly or indirectly bound thereto as least one lanthanide metal or at least one fluorophore, each acting as either an energy donor or an energy acceptor.

2. The composition of claim 1, wherein said solid matrix is selected from the group consisting of a polymeric siliceous material and a polysaccharide material.

3. The composition of claim 2, wherein said polymeric siliceous material comprises glass.

4. The composition of claim 3, wherein said glass surface has been treated or derivatized or activated to enhance fixation or immobilization.

5. The composition of claim 1, wherein said first intercalator is bound to said surface directly or through a linkage group.

6. The composition of claim 5, wherein said solid matrix comprises activated glass and said linkage group comprises an amino linkage group on said activated glass.

7. The composition of claim 6, wherein said linkage group terminates in a substituent capable of reacting with amino groups and said linkage group does not substantially interfere with other components in said composition.

8. The composition of claim 1, wherein said first intercalator is selected from the group consisting of phenanthridines and psoralenamines.

9. The composition of claim 8, wherein said phenanthridine is selected from the group consisting of ethidium bromide and 6-(4′

-carboxyphenyl)-3,8-diamine-5-methyl phenanthridine chloride.

10. The composition of claim 8, wherein said psoralenamine is selected from the group consisting of 8-[[[diethylamino)methyl]propyl]oxy]psoralen and 5-(N-piperadinyl)-8-methoxypsoralen derivatized with linkers that contain secondary or tertiary amino groups or quaternary ammonium salts.

11. The composition of claim 1, wherein said second intercalator is selected from the group consisting of acridines and coumarins.

12. The composition of claim 11, wherein said acridine comprises 9-aminoacridine.

13. The composition of claim 11, wherein said coumarin is selected from the group consisting of 7-hydroxy-4-methylcoumarin, 7-amino-4-methylcoumarin and 4-methyl-7-sulphatomethoxycoumarin.

14. The composition of claim 1, wherein said first intercalator comprises 6-(4′

-carboxyphenyl)-3,8-diamine-5-methyl phenanthridine chloride and said second intercalator comprises 9-aminoacridine.

15. The composition of claim 1, wherein said fluorophore in the second intercalator is selected from the group consisting of a chelated lanthanide metal, a naphthalene sulfonamide and a pyrene compound.

16. The composition of claim 1, wherein said first intercalator comprises 6-(4′

-carboxyphenyl)-3,8-diamine-5-methyl phenanthridine chloride, said second intercalator comprises 9-aminoacridine and said lanthanide metal comprises europium.

17. The composition of claim 1, wherein upon hybridization of said oligo- or polynucleotide to a complementary oligo- or polynucleotide of interest, said energy donor and said energy acceptor are within proximate distance of each other such that energy from said energy donor is absorbed by said energy acceptor.

18. The composition of claim 17, wherein said proximate distance is equal to or less than Furster'"'"'s radius.

19. The composition of claim 18, wherein said proximate distance is 30 Angstroms or less.

20. The composition of claim 1, wherein said lanthanide metal is selected from the group consisting of europium and terbium.

21. The composition of claim 1, wherein said lanthanide metal is chelated.

22. The composition of claim 21, wherein said chelator is selected from the group consisting of diethylenetriamine pentaacetic acid (DTPA) and transdiaminocyclohexane tetraacetic acid (DCTA).