Electrochemiluminescent label for DNA probe assays

• US 5,597,910 A
• Filed: 06/07/1995
• Issued: 01/28/1997
• Est. Priority Date: 12/11/1991
• Status: Expired due to Term

• Current Assignee: BioVeris Corporation (Roche Holding AG)
• Original Assignee: IGEN, Inc. (Teligent Incorporated)
• Inventors: Gudibande, Satyanarayana R., Kenten, John H.
• Primary Examiner(s): Jones, W. Gary
• Assistant Examiner(s): Sisson, Bradley L.
• Application Number:
  US08/479,817
• Time in Patent Office: 601 Days
• Field of Search: 435/6, 435/91.1, 435/91.2, 536/24.3, 536/24.31, 536/24.32, 536/25.32, 536/26.41, 542/2, 542/26, 436/800, 436/801, 250/361 C, 252/301.16, 252/301.4
• 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: C12Q 1/6816 characterised by the detection means C12Q1/6804 takes precedence
• CPC Class Codes: C12Q 1/6876 Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
• CPC Class Codes: C12Q 1/703 Viruses associated with AIDS
• CPC Class Codes: C12Q 1/708 for papilloma
• CPC Class Codes: C12Q 2563/103 luminescence
• CPC Class Codes: G01N 27/44726 using specific dyes, markers or binding molecules
• CPC Class Codes: Y10S 428/917 Electroluminescent
• CPC Class Codes: Y10S 436/80 Fluorescent dyes, e.g. rhodamine
• CPC Class Codes: Y10S 436/801 Electron dense compounds, e.g. ferritin
• CPC Class Codes: Y10S 436/805 Optical property
• CPC Class Codes: Y10S 436/905 Photochemical activation of reactions
• CPC Class Codes: Y10T 436/143333 Saccharide [e.g., DNA, etc.]

Abstract

This invention relates to a new electrochemiluminescent (ECL) label for oligonucleotides using phosphoramidite chemistry.

Claims

1. A compound of the formula:

##STR23## wherein M is selected from the group consisting of Ru, Os and Re;

u.t=v, u is 1 or 2 and t is 1 or 2;

L₁, L₂, and L₃ are the same or not all the same and each is ##STR24## wherein R₁, R₂, R₃, R₄, R₅ and R₆ are the same or not all the same and each is H, alkyl of from 1-5 carbon atoms, ##STR25## and at least one of R₁, R₂, R₃, R₄, R₅ and R₆ is either ##STR26## n is an integer in the range from 1-20;

X is selected from the group consisting of O, S, SO₂, COO, and CONH;

a and b are --N(CH--(CH₃)₂)₂, --NH--CH--(CH₃)₂, O--(CH₂)₂ --CN, O--CH₃ or morpholino, provided that a and b are not the same; and

Z is a counterion, provided that said counterion is an anion associated with the synthesis or purification of the compound; and

further provided that ligands L₁, L₂ and L₃ are selected such that the compound is capable of electrochemiluminescence.

2. A compound as recited in claim 1 wherein L₁, L₂ and L₃ are all substituted or unsubstituted bipyridyl and n is from 4 to 7.

3. A compound as recited in claim 1 wherein the ratio of the relative ECL capability of the compound to the relative ECL capability of its bipyridyl analog is greater than or equal to 0.10.

4. A compound of the formula:

##STR27## wherein M is selected from the group consisting of Ru, Os and Re;

L₁, L₂ and L₃ are the same or different and each is ##STR28## wherein R₁, R₂, R₃, R₄, R₅ and R₆ are the same or different and each is H or an alkyl group of 1-5 carbon atoms provided, however, that one of R₁ -R₆ is a linker of the formula ##STR29## wherein n is an integer from 1-20;

X is selected from the group consisting of O, S, SO₂, COO and CONH;

y is an integer of from 1-20;

z is 0 or 1; and

B is a biomolecule or a synthetic analog thereof, and the phosphodiester group is bound to said biomolecule;

said compound being capable of electrochemiluminescence.

5. A compound as recited in claim 4 wherein the ratio of the relative ECL capability of the compound to the relative ECL capability of its bipyridyl analog is greater than or equal to 0.10.

6. A complex of the formula ##STR30## wherein M is ruthenium, osmium or rhenium;

R₁, R₂, R₃, R₄ and R₅ are the same or different and each is H or alkyl of 1-4 carbon atoms;

n is an integer from 1 to 20;

X is selected from the group consisting of O, S, SO₂, COO and CONH;

y is an integer of from 1-20;

z is 0 or 1;

m is an integer of from 1-1000; and

NAB is a nucleic acid base which may be modified or unmodified;

said complex being able to electrochemiluminesce.

7. A complex as recited in claim 6 wherein said nucleic acid is a primer for a polymerase amplification reaction.

8. A complex as recited in claim 6 wherein said nucleic acid is a hybridization probe.

9. A complex as recited in claim 6 wherein the ratio of the relative ECL capability of the complex to the relative ECL capability of its bipyridyl analog is greater than or equal to 0.25.