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US 8,574,881 B2Granted
Compositions useful for detection or quantification of desirable target molecules, novel dyes, composite dyes, and oligonucleotides or polynucleotides comprising such dyes
AssigneeEnzo Life Sciences, Inc.
InventorsPraveen Pande, Yuejun Xiang, Elazar Rabbani
Life of this patent
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Abstract
The present invention provides dyes, reactive dyes and labeled reagents that may be used in the detection or quantification of desirable target molecules, such as proteins and nucleic acids. Dyes are provided that may be used free in solution where the binding of the dye to the target molecule provides signal generation. Dyes are also provided that comprise reactive groups that may be used to attach the dyes to probes that will bind to desirable target molecules. The novel dyes of the present invention have been modified by the addition of charged and polar groups to provide beneficial properties.
Claims33
A dye comprising the formula:
- wherein n can be 1, 2 or 3;
- wherein at least one of R1, R2, R3, R4, R5, R6, R7, R8, R9, R21 or R22 comprises Q wherein Q comprises a phosphate (PO4=), a phosphate monoester (PO3−ER13), a phosphate diester (PO2ER13ER14), a phosphonate (PO3═), a phosphonate monoester (PO2−ER13), a phosphonate diester (POER13ER14), a thiophosphate (PSO3=), a thiophosphate monoester (PSO2−ER13), a thiophosphate diester (PSOER13ER14), a thiophosphonate (PSO2=), a thiophosphonate monoester (PSO−ER13), a thiophosphonate diester (PSER13ER14), a phosphonamide (PONR13R14NR19R20), its thioanalogue (PSNR13R14NR19R20), a phosphoramide (PONR13R14NR15NR19R20), its thioanalogue (PSNR13R14NR15NR19R20), or a phosphoramidite (PO2R19NR13R14), or its thioanalogue (POSR19NR13R14), where E can be independently O or S;
- wherein Q is attached directly, or indirectly through a linker arm comprising carbon, sulfur, oxygen, nitrogen, or any combinations thereof and wherein said linker arm may be saturated or unsaturated, linear or branched, substituted or unsubstituted or any combinations thereof;
- wherein R13, R14, R15, R19 and R20 can be hydrogen, a halogen, an amino group, an alkyl group wherein said alkyl group is saturated or unsaturated, linear or branched, substituted or unsubstituted, an alkoxy group wherein said alkoxy group is saturated or unsaturated, branched or linear, substituted or unsubstituted, or when taken together R13 and R14 form a five- or six-membered ring;
- wherein R16, R17 and the remaining R1, R2, R3, R4, R5, R6, R7, R8, R9, R21 or R22 are independently hydrogen, Z, an alkyl group wherein said alkyl group is saturated or unsaturated, linear or branched, substituted or unsubstituted, an alkoxy group that is saturated or unsaturated, branched or linear, substituted or unsubstituted, or when taken together, R1 and R2, R2 and R3, R3 and R4, R4 and R5, R5 and R16, R16 and R17, R17 and R9, R9 and R8, R8 and R21, R21 and R22, R22 and R7, and R7 and R6 may form a five- or six-membered ring;
- wherein Z comprises a carboxyl group (CO2−), a carbonate ester (COER13), a sulfonate (SO3−), a sulfonate ester (SO2ER13), a sulfoxide (SOR13), a sulfone (SO2CR13R14R15), a sulfonamide (SO2NR3R14), a phosphate (PO4=), a phosphate monoester (PO3−ER13), a phosphate diester (PO2ER13ER14), a phosphonate (PO3=), a phosphonate monoester (PO2−ER13), a phosphonate diester (POER13ER14), a thiophosphate (PSO3=), a thiophosphate monoester (PSO2−ER13), a thiophosphate diester (PSOER13ER14), a thiophosphonate (PSO2=), a thiophosphonate monoester (PSO−ER13), a thiophosphonate diester (PSER13ER14), a phosphonamide (PONR13R14NR19R20), its thioanalogue (PSNR13R14NR19R20), a phosphoramide (PONR13R14NR15NR19R20), its thioanalogue (PSNR13R14NR15NR19R20), a phosphoramidite (PO2R19NR13R14), or its thioanalogue (POSR19NR13R14), where E can be independently O or S;
- wherein Z is attached directly, or indirectly through a linker arm comprising carbon, sulfur, oxygen, nitrogen, or any combinations thereof, and wherein said linker arm may be saturated or unsaturated, linear or branched, substituted or unsubstituted or any combinations thereof;
- wherein any of R1, R2, R3, R4, R5, R6, R7, R8, R9, R21 or R22 may further comprise a heteroatom containing side chain wherein said side chain is joined to the R group by a linkage which comprises an ether linkage (—OR25), a thioether linkage (—SR25), or an amine linkage (—NR25R26 or —N+R25R26R27), and wherein R25, R26 and R27 independently comprise hydrogen, Z, an alkyl group wherein said alkyl group is saturated or unsaturated, linear or branched, substituted or unsubstituted, an alkoxy group that is saturated or unsaturated, branched or linear, substituted or unsubstituted, or when taken together, R25 and R26, and R26 and R27 independently comprise a five- or six-membered ring, and wherein any of R25, R26 or R27 may further comprise said heteroatom containing side chain, and
- Wherein a least one R1, R2, R3, R4, R5, R7, R8, R9, R16, R17, R21, or R22 comprises a reactive group selected from an isocyanate, isothiocyanate, monochlorotriazine, dichlorotriazine, 4,6-dichloro-1,3,5-triazines, mono- or di-halogen substituted pyridine, mono- or di-halogen substituted diazine, maleimide, haloacetamide, aziridine, sulfonyl halide, acid halide, hydroxysuccinimide ester, hydroxysulfosuccinimide ester, imido ester, hydrazine, azidonitrophenol, azide, 3-(2-pyridyl dithio)-proprionamide, glyoxal, aldehyde, reactive thiol, amine, hydroxyl, terminal alkene, terminal alkyne, platinum coordinate group and alkylating agent.
6 more elements
A dye comprising the structure:
- wherein at least one of R1, R2, R3, R6, R7, R8, R9, R21 or R22 comprises Q wherein Q comprises a phosphate (PO4=), a phosphate monoester (PO3−ER13), a phosphate diester (PO2ER13ER14), a phosphonate (PO3=), a phosphonate monoester (PO2−ER13), a phosphonate diester (POER13ER14), a thiophosphate (PSO3=), a thiophosphate monoester (PSO2−ER13), a thiophosphate diester (PSOER13ER14), a thiophosphonate (PSO2=), a thiophosphonate monoester (PSO−ER13), a thiophosphonate diester (PSER13ER14), a phosphonamide (PONR13R14NR19R20), its thioanalogue (PSNR13R14NR19R20), a phosphoramide (PONR13R14NR5NR19R20), its thioanalogue (PSNR13R14NR15NR19R20), a phosphoramidite (PO2R19NR13R14), or its thioanalogue (POSR19NR13R14), where E can be independently O or S;
- wherein Q is attached directly, or indirectly through a linker arm comprising carbon, sulfur, oxygen, nitrogen, or any combinations thereof and wherein said linker arm may be saturated or unsaturated, linear or branched, substituted or unsubstituted or any combinations thereof;
- wherein R13, R14, R15, R19 and R20 is hydrogen, a halogen, an amino group, an alkyl group wherein said alkyl group is saturated or unsaturated linear or branched, substituted or unsubstituted, an alkoxy group wherein said alkoxy group is saturated or unsaturated, branched or linear, substituted or unsubstituted, or when taken together R13 and R14 form a five- or six-membered ring;
- wherein R16, R17 and the remaining R1, R2, R3, R6, R7, R8, R9, R21 or R22 are independently hydrogen, Z, an alkyl group wherein said alkyl group is saturated or unsaturated, linear or branched, substituted or unsubstituted, an alkoxy group that is saturated or unsaturated, branched or linear, substituted or unsubstituted, or when taken together R16 and R17, R17 and R9, R9 and R8, R8 and R21, R21 and R22, R22 and R7, and R7 and R6 may form a five- or six-membered ring;
- wherein Z comprises a carboxyl group (CO2−), a carbonate ester (COER13), a sulfonate (SO3−), a sulfonate ester (SO2ER13), a sulfoxide (SOR13), a sulfone (SO2CR13R14R15), a sulfonamide (SO2NR13R14), a phosphate (PO4=), a phosphate monoester (PO3−ER13), a phosphate diester (PO2ER13ER14), a phosphonate (PO3=), a phosphonate monoester (PO2−ER13), a phosphonate diester (POER13ER14, a thiophosphate (PSO3=), a thiophosphate monoester (PSO2−ER13), a thiophosphate diester (PSOER13ER14), a thiophosphonate (PSO2=), a thiophosphonate monoester (PSO−ER13), a thiophosphonate diester (PSER13ER14), a phosphonamide (PONR13R14NR19R20), its thioanalogue (PSNR13R14NR19R20), a phosphoramide (PONR13R14NR15NR19R20), its thioanalogue (PSNR13R14NR15NR19R20), a phosphoramidite (PO2R19NR13R14), or its thioanalogue (POSR19NR13R14), where E can be independently O or S;
- wherein Z is attached directly, or indirectly through a linker arm comprising carbon sulfur, oxygen, nitrogen, or any combinations thereof and wherein said linker arm may be saturated or unsaturated linear or branched, substituted or unsubstituted or any combinations thereof;
- wherein any of R1, R2, R3, R6, R7, R8, R9, R21 or R22 may further comprise a heteroatom containing side chain wherein said side chain is joined to the R group by a linkage which comprises an ether linkage (—OR25), a thioether linkage (—SR25), or an amine linkage (—NR25R26 or —N+NR25R26R27), and wherein R25, R26 and R27 independently comprise hydrogen, Z, an alkyl group wherein said alkyl group is saturated or unsaturated, linear or branched, substituted or unsubstituted, an alkoxy group that is saturated or unsaturated, branched or linear substituted or unsubstituted, or when taken together, R25 and R26, and R26 and R27 independently comprise a five- or six membered ring, and wherein any of R25, R26 or R27 may further comprise said heteroatom containing side chain;
- wherein R41, R42, R43 and R44 can independently be hydrogen, an alkyl group wherein said alkyl group is saturated or unsaturated, linear or branched, substituted or unsubstituted, an alkoxy group that is saturated or unsaturated, branched or linear, substituted or unsubstituted, or when taken together, R3 and R41, R41 and R42, R42 and R43, R43 and R44, and R44 and R16 may form a five- or six-membered ring
- and wherein any of R41, R42, R43 or R44 may further comprise a heteroatom containing side chain wherein said side chain is joined to the R group by a linkage which comprises an ether linkage (—OR25), a thioether linkage (—SR25), or an amine linkage (—NR25R26 or —NR25R26R27), and wherein R25, R26 and R27 independently comprise hydrogen, Z, an alkyl group wherein said alkyl group is saturated or unsaturated, linear or branched, substituted or unsubstituted, an alkoxy group that is saturated or unsaturated, branched or linear, substituted or unsubstituted, or when taken together, R25 and R26, and R26 and R27 independently comprise a five- or six-membered ring, and wherein any of R25, R26 or R27 may further comprise said heteroatom containing side chain.
6 more elements
Dependencies are inferred from the claim text.