Long emission wavelength chemiluminescent compounds and their use in test assays

Claim 2

A method as claimed in claim 1 wherein each of the chemiluminescent compounds is conjugated to a binding partner which is specific for and binds to a substance to form a reaction product to be detected and/or quantitated in the test sample.

Claim 3

A method as claimed in claim 1 or claim 2 wherein the emission spectra of the chemiluminescent compounds or chemiluminescent conjugates are discernable in that the emission maxima thereof are spaced apart a sufficient distance to discern the signals of each conjugate, preferably the emission maxima are spaced greater than about 60 nanometers apart.

Claim 8

A method as claimed in claim 7 wherein the assays comprise: immunoassays, homogeneous or heterogeneous, hybridization assays and amplification assays.

Claim 11

A method as claimed in claim 10 wherein the internal reference is a target sequence that either is or is not amplified.

Claim 12

A method as claimed in claim 10 or claim 11 wherein it further comprises performing at least one binding reaction, a hybridization reaction, specific to a target sequence of the test sample and/or specific to the internal reference.

Claim 14

A chemiluminescent compound characterised in that, upon chemical treatment, it emits a blue-green, green, yellow, orange and red-orange light.

Claim 15

A compound as claimed in claim 14 wherein it has an emission spectra peak or maximum greater than about 515 nm.

Claim 16

A compound as claimed in claim 14 or claim 15 wherein it includes an aromatic ring fused acridinium compound or a benzacridinium ring system or an N-alkylated benzacridinium ring system.

Claim 18

A method as claimed in claim 17 wherein at least one chemiluminescent compound includes a benzacridinium ring system or an acridinium ring system.

Claim 19

A chemiluminescent label assay method for detection and/or quantitating at least two substances in a test sample characterised in that it comprises: (a) providing a first chemiluminescent reagent to the test sample, said first chemiluminescent reagent including an aromatic ring fused acridinium compound or a benzacridinium ring system, preferably and N-alkylated benzacridinium ring system, and being specific to one substance in the test sample; (b) providing a second chemiluminescent reagent to the test sample, the second chemiluminescent reagent including an acridinium ring system and being specific to another substance in the test sample; and (c) activating the chemiluminescent reagents to provide discernable emission signals for detecting and/or quantitating the substances.

Claim 20

A method as claimed in claim 19 wherein it further comprises a binding step and wherein each chemiluminescent reagent further includes a conjugated molecule specific to a substance in the test sample to enable a binding reaction to occur so that a reaction product is formed.

Claim 21

A test kit for detecting and/or quantitating two or more substances in a test sample characterised in that it comprises at least two difference chemiluminescent compounds, each compound being conjugated to a binding partner which is specific for at least one substance in a test sample.

Claim 22

A test kit for performing a simultaneous assay on two or more substances in a test sample characterised in that it comprises: (a) a first chemiluminescent reagent including an aromatic ring fused acridinium system or a benzacridinium ring system, preferably an N-alkylated benzacridinium ring system; and (b) a second chemiluminescent reagent including an acridinium ring system.

Claim 23

A test kit for performing a simultaneous assay on two or more substances in a test sample characterised in that it comprises at least two different chemiluminescent reagents, each reagent having a discernable emission spectra for detecting and/or quantitating the substances.

Claim 24

A kit as claimed in claim 23 wherein at least one of the chemiluminescent reagents includes a N-alkylated benzacridinium ring system and/or at least one of the chemiluminescent reagents includes an acridinium ring system.

Claim 25

A luminescent compound characterised in that it corresponds to the following general formula: wherein W represents carbon; or C₇, W, C₉ or C₁₀ may be replaced with -N=; or W may be omitted and C₇ connected to C₉, and C₇, C₉ or C₁₀ may be replaced with -O-, -S-, -NH- or -NR-; Y and Y' independently represent alkyl which may contain up to 20 carbon atoms, optionally halogenated, or a polysubstituted aryl moiety corresponding to the following general formula: R₁ represents alkyl, alkenyl, alkynyl or aralkyl which may contain up to 20 heteroatoms; R₂, R₃, R₅, R₇, R₉ and R₁₀ independently represent hydrogen, optionally substituted aryl (ArR or Ar), halide, amino, hydroxyl, nitro, sulfonate, -R, -CN, -COOH, -SCN, -OR, -SR, -SSR, -C(O)R, -C(O)OR, -C(O)NHR, or -NHC(O)R; R₂ represents one or more substituents at C₁₋₄ or a fused aromatic ring with or without heteroatoms; R₃ represents one or more substituents at C₇, W, C₉ or C₁₀; A- represents a counter-ion preferably CH₃SO₄-, FSO₃-, CF₃SO₄-, C₄F₉SO₃-, CH₃C₆H₄SO₃- and halide; X represents a heteroatom, preferably nitrogen, oxygen or sulfur, such that, when X represents oxygen or sulfur, Z is omitted, and, when X represents nitrogen, Z represents -SO₂-Y'; R₄ and R₈ independently represent alkyl, alkenyl, alkynyl, alkoxyl, alkylthiol, or amido; R₆ represents -R₁₁-R₁₂, wherein R₁₁, if present, represents alkyl, optionally substituted aryl or aralkyl which may contain up to 20 heteroatoms; and R₁₂ represents a leaving group or an electrophilic functional group attached with a leaving group or -Q-R-Nu, -Q-R(I) n Nu, -Q-Nu, -R-Nu or -Nu; wherein n represents an integer of at least 1; Nu represents a nucleophilic group; Q represents a functional linkage; I represents an ionic or ionizable group; and R represents alkyl(ese), alkenyl(ese), alkynyl(ene), aryl(ene) or aralkyl(ene) which may contain up to 20 heteroatoms; R₅ and R₆, and R₆ and R₇ being interchangeable.

Claim 26

A compound as claimed in claim 25 wherein it is conjugated with a binding partner for assay use.

Claim 27

An intermediate characterised in that it is selected from; (a) benz[b]acridine-12-carboxylic acid hydrochloride; (b) 3-EtO-benz[b]acridinine-12-carboxylic acid; (c) 3-hydroxy-benz[b]acridine-12-carboxylic acid; (d) 2-MeO-benz[b]acridine-12-carboxylic acid; (e) 2-hydroxy-benz[b]acridine-12-carboxylic acid; (f) 3-MeO-acridine-9-carboxylic acid hydrochloride.

Claim 28

An intermediate characterised in that it corresponds to the following general formula: where W represents carbon; or C₇, W, C₉ or C₁₀ may be replaced with -N=, or W may be omitted and C₇ connected to C₉, and C₇, C₉ or C₁₀ may be replaced with -O-, -S-, -NH- or -NR-; R₂, R₃, R₉ and R₁₀ independently represent hydrogen, optionally substituted aryl (ArR or Ar), halide, amino, hydroxyl, nitro, sulfonate, -R, -CN, -COOH, -SCN, -OR, -SR, -SSR, -C(O)R, -C(O)OR, -C(O)NHR, or -NHC(O)R; R₂ represents one or more substituents at C₁₋₄ or a fused aromatic ring with or without heteroatoms; R₃ represents one or more substituents at C₇, W, C₉ or C₁₀; R represents alkyl(ene), alkenyl(ene), alkynyl(ene), aryl(ene) or aralkyl(ene) which may contain up to 20 heteroatoms.

Claim 29

An intermediate characterised in that it corresponds to the following general formula: where W represents carbon; or C₇, W, C₉ or C₁₀ may be replaced with -N=; or W may be omitted and C₇ connected to C₉ and C₇, C₉ or C₁₀ may be replaced with -O-, -S-, -NH- or -NR-; Y and Y' independently represent alkyl which may contain up to 20 carbon atoms, optionally halogenated, or a polysubstituted aryl moiety corresponding to the following general formula: R₂, R₃, R₉ and R₁₀ independently represent hydrogen, optionally substituted aryl (ArR or Ar), halide, amino, hydroxyl, nitro, sulfonate, -R, -CN, -COOH, -SCN, -OR, -SR, -SSR, -C(O)R, -C(O)OR, -C(O)NHR, or NHC(O)R; R₂ represents one or more substituents at C₁₋₄ or a fused aromatic ring with or without heteroatoms; R₃ represents one or more substituents at C₇, W, C₉ or C₁₀; X represents a heteroatom, preferably nitrogen, oxygen or sulfur, such that, when X represents oxygen or sulfur, Z is omitted, and, when X represents nitrogen, Z represents -SO₂-Y'; R₄ and R₈ independently represent alkyl, alkenyl, alkynyl, alkoxyl, alkylthiol or amido; R₆ represents -R₁₁-R₁₂, wherein R₁₁, if present, represents alkyl, optionally substituted aryl or aralkyl which may contain up to 20 heteroatoms; and R₁₂ represents a leaving group or an electrophilic functional group attached with a leaving group or -Q-R-Nu, -Q-R(I) n Nu, -Q-Nu, -R-Nu or Nu; wherein n represents an integer of at least 1; Nu represents a nucleophilic group; Q represents a functional linkage; I represents an ionic or ionizable group; and R represents alkyl(ese), alkenyl(ene), alkynyl(ene), aryl(ene) or aralkyl(ene) which may contain up to 20 heteroatoms; R₅ and R₆, and R₆ and R₇ being interchangeable.

Claim 30

A compound as claimed in claim 25 wherein the isomeric, furano, thiopheno, pyrro and pyrido moiety rings include substituent(s) comprising R₃.