US 5,310,687 AGranted

Luminescent metal chelate labels and means for detection

AssigneeIgen, Inc.

InventorsAllen J. Bard, George M. Whitesides

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Abstract

A chemical moiety is disclosed which comprises a chemical, biochemical, or biological substance attached to one or more electrochemiluminescent organometallic compounds. In a preferred embodiment of the invention the substance is attached to one or more ruthenium-containing or osmium-containing luminescent organometallic compounds. Methods are disclosed for detecting low concentrations of the chemical moiety using chemiluminescent, electrochemiluminescent, and photoluminescent means. Compounds are disclosed which are useful for labeling substances of interest with ruthenium-containing and osmium-containing labels or other electrochemiluminescent labels. These labeled substances are useful in methods provided for detecting and quantifying analytes of interest in binding assays and competitive binding assays. The labeled substances are of particular use in homogeneous binding assays. These methods form the bases for systems designed to enable the rapid, efficient, and sensitive determination of a broad array of chemical, biochemical, and biological materials of interest.

Claims18

Dependent claims

A method of determining the presence of a chemical moiety, which comprises

(a) forming a reagent mixture containing the chemical moiety, or the chemical moiety and an agent which upon exposure of the reagent mixture to electrochemical energy forms either a reductant or an oxidant;
said chemical moiety having the formula ##STR4## wherein:
M is ruthenium or osmium;
P, L.sup.1, and L.sup.2, each is a bidentate aromatic heterocyclic nitrogen-containing ligand of M selected from the group consisting of bipyridyl, substituted bipyridyl, bipyrazyl, substituted bipyrazyl, phenanthrolyl and substituted phenanthrolyl, wherein each of said substituted groups is substituted by an alkyl, aryl, aralkyl, carboxylate, carboxaldehyde, carboxamide, cyano, amino, hydroxycarbonyl, hydroxyamino, aminocarbonyl, amidine, guanidinium, ureide, sulfur-containing group, phosphorus-containing group, or the carboxylate ester of N-hydroxysuccinimide, each of said ligands being the same or not the same as each other ligand;
B is a biological substance, a synthetic substance which is capable of competing with a biological substance in a competitive binding reaction with a complementary material, or a non-biological polymer;
m is 1;
each of n and o is 1;
t is an integer equal to or greater than 1;
and
u is 1;
said link being covalent bonding linking B with at least one of P, L.sup.1 or L.sup.2 ;
P, L.sup.1, L.sup.2 and B being of such composition and number that the chemical moiety is capable of being induced to electrochemiluminesce;
(b) exposing the reagent mixture to electrochemical energy the potential of which oscillates between a potential sufficiently positive to oxidize the chemical moiety and a potential sufficiently negative to reduce the chemical moiety, or to electrochemical energy such that said chemical moiety is oxidized and the agent forms a reductant, or such that said chemical moiety is reduced and the agent forms an oxidant, thereby to induce the chemical moiety to electrochemiluminesce;
and
(c) detecting emitted luminescence thereby to determine the presence of the chemical moiety.

12 more elements

A method of determining the presence of an analyte of interest which binds to a chemical moiety, which comprises

(a) forming a reagent mixture comprising the chemical moiety, or comprising the chemical moiety and an agent which upon exposure of the reagent mixture to electrochemical energy forms either a reductant or an oxidant, and the analyte of interest;
such that the chemical moiety and the analyte specifically bind to one another, said chemical moiety having the formula ##STR5## wherein:
M is ruthenium or osmium;
P, L.sup.1, and L.sup.2, each is a bidentate aromatic heterocyclic nitrogen-containing ligand of M selected from the group consisting of bipyridyl, substituted bipyridyl, bipyrazyl, substituted bipyrazyl, phenanthrolyl and substituted phenanthrolyl, wherein each of said substituted groups is substituted by an alkyl, aryl, aralkyl, carboxylate, carboxaldehyde, carboxamide, cyano, amino, hydroxycarbonyl, hydroxyamino, aminocarbonyl, amidine, guanidinium, ureide, sulfur-containing group, phosphorus-containing group, or the carboxylate ester of N-hydroxysuccinimide, each of said ligands being the same or not the same as each other ligand;
B is a biological substance, or a synthetic substance which is capable of competing with a biological substance in a competitive binding reaction with a complementary material, said biological substance or synthetic substance being capable of specifically binding to the analyte of interest;
m is 1;
each of n and o is 1;
t is an integer equal to or greater than 1;
and
u is 1;
said link being covalent bonding linking B with at least one of P, L.sup.1 or L.sup.2 ;
or
P, L.sup.1, L.sup.2 and B being of such composition and number that the chemical moiety is capable of being induced to electrochemiluminesce;
(b) exposing the reagent mixture to electrochemical energy the potential of which oscillates between a potential sufficiently positive to oxidize the chemical moiety and a potential sufficiently negative to reduce the chemical moiety, or to electrochemical energy such that said chemical moiety is oxidized and the agent forms a reductant, or such that said chemical moiety is reduced and the agent forms an oxidant, thereby to induce the chemical moiety to electrochemiluminesce;
and
(c) detecting emitting luminescence thereby to determine the presence of the analyte of interest.

13 more elements

A competitive binding method of determining the presence of an analyte of interest wherein the analyte and a chemical moiety bind competitively to a complementary material, which comprises

(a) forming a reagent mixture comprising the analyte of interest, the complementary material and the chemical moiety, or the chemical moiety and an agent which upon exposure of the reagent mixture to electrochemical energy forms either a reductant or an oxidant, such that the chemical moiety and the analyte of interest bind competitively to the complementary material;
said chemical moiety having the formula ##STR6## wherein:
M is ruthenium or osmium;
P, L.sup.1, and L.sup.2, each is a bidentate aromatic heterocyclic nitrogen-containing ligand of M selected from the group consisting of bipyridyl, substituted bypyridyl, bipyrazyl, substituted bipyrazyl, phenantholyl and substituted phenanthrolyl, wherein each of said substituted groups is substituted by an alkyl, aryl, aralkyl, carboxylate, carboxaldehyde, carboxamide, cyano, amino, hydroxycarbonyl, hydroxyamino, aminocarbonyl, amide, guanidinium, ureide, sulfur-containing group, phosphorus-containing group, or the carboxylate ester of N-hydroxysuccinimide, each of said ligands being the same or not the same as each other ligand;
B is a biological substance, or a synthetic substance which is capable of competing with a biological substance in a competitive binding reaction with said complementary material, said biological substance or synthetic substance being capable of binding to the complementary material;
m is 1;
each of n an o is 1;
t is an integer equal to or greater than 1;
and
u is 1;
said link being covalent bonding linking B with at least one of P, L.sup.1 or L.sup.2 ;
or
P, L.sup.1 L.sup.2 and B being of such composition and number that the chemical moiety is capable of being induced to electrochemiluminesce;
(b) exposing the reagent mixture to electrochemical energy the potential of which oscillates between a potential sufficiently positive to oxidize the chemical moiety and a potential sufficiently negative to reduce the chemical moiety, or to electrochemical energy such that said chemical moiety is oxidized and the agent forms a reductant, or such that said chemical moiety is reduced and the agent forms an oxidant, thereby to induce the chemical moiety to electrochemiluminesce;
and
(c) detecting emitted luminescence thereby to determine the presence of the analyte of interest.

13 more elements

A system for determining the presence of a chemical moiety, which system comprises

(a) a reagent mixture comprising the chemical moiety, or the chemical moiety and an agent which upon exposure of the reagent mixture to electrochemical energy forms either a reductant or an oxidant, said chemical moiety having the formula ##STR7## wherein:
M is ruthenium or osmium;
P, L.sup.1, and L.sup.2, each is a bidentate aromatic heterocyclic nitrogen-containing ligand of M selected from the group consisting of bipyridyl, substituted bipyridyl, bipyrazyl, substituted bipyrazyl, phenanthrolyl and substituted phenanthrolyl, wherein each of said substituted groups is substituted by an alkyl, aryl, aralkyl, carboxylate, carboxaldehyde, carboxamide, cyano, amino, hydroxycarbonyl, hydroxyamino, aminocarbonyl, amidine, guanidinium, ureide, sulfur-containing group, phosphorus-containing group, or the carboxylate ester of N-hydroxysuccinimide, each of said ligands being the same or not the same as each other ligand;
B is a biological substance, a synthetic substance which is capable of competing with a biological substance in a competitive binding reaction with a complementary material, or a non-biological polymer;
m is 1;
each of n is o is 1;
t is an integer equal to or greater than 1;
and
u is 1;
said link being covalent bonding linking B with at least one of P, L.sup.1 or L.sup.2 ;
or
P, L.sup.1, L.sup.2 and B being of such composition and number that the chemical moiety is capable of being induced to electrochemiluminesce;
(b) means for exposing the reagent mixture to electrochemical energy the potential of which oscillates between a potential sufficiently positive to oxidize the chemical moiety and a potential sufficiently negative to reduce the chemical moiety, or to electrochemical energy such that said chemical moiety is oxidized and the agent forms a reductant, or such that said chemical moiety is reduced and the agent forms an oxidant, thereby to induce the chemical moiety to electrochemiluminesce;
and
(c) means for detecting emitted luminescence thereby to determine the presence of the chemical moiety.

12 more elements

A system for determining the presence of an analyte of interest which binds to a chemical moiety, which system comprises

(a) a reagent mixture comprising the chemical moiety, or comprising the chemical moiety and an agent which upon exposure of the reagent mixture to electrochemical energy forms either a reductant or an oxidant, and the analyte of interest;
said chemical moiety having the formula ##STR8## wherein:
m is ruthenium or osmium;
P, L.sup.1, and L.sup.2, each is a bidentate aromatic heterocyclic nitrogen-containing ligand of M selected from the group consisting of bipyridyl, substituted bipyridyl, bipyrazyl, substituted bipyrazyl, phenanthrolyl and substituted phenanthrolyl, wherein each of said substituted groups is substituted by an alkyl, aryl, aralkyl, carboxylate, carboxaldehyde, carboxamide, cyano, amino, hydroxycarbonyl, hydroxyamino, aminocarbonyl, amidine, guanidinium, ureide, sulfur-containing group, phosphorus-containing group, or the carboxylate ester of N-hydroxysuccinimide, each of said ligands being the same or not the same as each other ligand;
B is a biological substance, or a synthetic substance which is capable of competing with a biological substance in a competitive binding reaction with a complementary material, or synthetic substance being capable of binding to the analyte of interest;
m is 1;
each of n is o is 1;
t is an integer equal to 1;
and
u is 1;
said link being covalent bonding linking B with one of P, L.sup.1 or L.sup.2 ;
or
P, L.sup.1, L.sup.2 and B being of such composition and number that the chemical moiety is capable of being induced to electrochemiluminesce;
(b) means for contacting the chemical moiety with the analyte of interest to form a reagent mixture;
(c) means for exposing the reagent mixture to electrochemical energy the potential of which oscillates between a potential sufficiently positive to oxidize the chemical moiety and a potential sufficiently negative to reduce the chemical moiety, or to electrochemical energy such that said chemical moiety is oxidized and the agent forms a reductant, or such that said chemical moiety is reduced and the agent forms an oxidant, thereby to induce the chemical moiety to electrochemiluminesce;
and
(d) means for detecting emitted luminescence thereby to determine the presence of the chemical moiety.

14 more elements

Dependencies are inferred from the claim text.

Facts

Application: 07/789,418Filed 4 Nov 1991 · continuation
Priority: 31 Oct 1984
Granted: 10 May 1994
Term ends: Not estimatedThe source records don’t support an estimate.

Classification11 codes

C07F 15/0053
Ruthenium compounds: without a metal-carbon linkage
C07F 15/0026
Osmium compounds: without a metal-carbon linkage
G01N 33/533
Production of labelled immunochemicals: with fluorescent label
Y10S 436/80
Chemistry: analytical and immunological testing: fluorescent dyes, e.g. rhodamine

Family

Serial666,987Parent of this continuation-in-part · filed 31 Oct 1984
Serial789,113Parent of this continuation-in-part · filed 24 Oct 1985
Serial858,353Parent of this continuation · filed 30 Apr 1986
US 5,235,808 AParent of this continuation-in-part
US 5,310,687 AThis patent

From the US filing history. Foreign family members aren’t listed.