Modified carbocyanine dyes and their conjugates
AssigneeInvitrogen Corporation
InventorsWai-Yee LEUNG, Ching-Ying CHEUNG, Stephen YUE
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Abstract
Chemically reactive carbocyanine dyes incorporating an indolium ring moiety that is substituted at the 3-position by a reactive group or by a conjugated substance, and their uses, are described. Conjugation through this position results in spectral properties that are uniformly superior to those of conjugates of spectrally similar dyes wherein attachment is at a different position. The invention includes derivative compounds having one or more benzo nitrogens.
Claims47
1-17. (canceled)
A compound of Formula:
- wherein,
- δ is 0 or 1;
- n is 1, 2 or 3;
- W represents the atoms necessary to form one fused aromatic ring having 6 atoms, which atoms are selected from —CH and —CR1′;
- each R1′ is sulfo;
- R2 is C1-C6 alkyl, which is optionally substituted by sulfo;
- R3 is C1-C6 alkyl;
- R4 is C1-C6 alkyl;
- R6 is H or sulfo;
- R7 is H or sulfo;
- R8 is H or sulfo;
- R9 is H or sulfo;
- R12 is -L-Rx or -L-Sc, or a C1-C6 alkyl, which is optionally substituted one or more times by F, Cl, Br, I, hydroxy, carboxy, sulfo, or amino;
- R13 is -L-Rx or -L-Sc;
- R14 is C1-C6 alkyl;
- Rx is the reactive group;
- Sc is a conjugated substance;
- and
- L is a covalent linkage.
A compound of Formula:
- wherein,
- R2 is C1-C6 alkyl, which is substituted by sulfo;
- R3 is C1-C6 alkyl;
- R4 is C1-C6 alkyl;
- R12 is C1-C6 alkyl;
- R13 is -L-Rx or -L-Sc;
- R14 is C1-C6 alkyl;
- δ is 0 or 1;
- and
- n is 1, 2, or 3.
A carbocyanine dye according to the formula:
- A-BRIDGE-B
- wherein A and B are indolium rings, wherein the substituent on the 3-carbon of one of the indolium rings contains a chemically reactive group or a conjugated substance;
- and
- BRIDGE is a polymethine linker.
A method of staining a biological sample, comprising:
- combining a dye solution comprising a compound having the formula:
- wherein,
- δ is 0 or 1;
- n is 1, 2 or 3;
- W represents the atoms necessary to form one fused aromatic ring having 6 atoms, which atoms are selected from —CH and —CR1′;
- each R1′ is sulfo;
- R2 is C1-C6 alkyl, which is substituted by sulfo;
- R3 is C1-C6 alkyl;
- R4 is C1-C6 alkyl;
- R6 is H or sulfo;
- R7 is H or sulfo;
- R8 is H or sulfo;
- R9 is H or sulfo;
- R12 is -L-Rx or -L-Sc, or a C1-C6 alkyl, which is optionally substituted one or more times by F, Cl, Br, I, hydroxy, carboxy, sulfo, or amino;
- R13 is -L-Rx or -L-Sc;
- R14 is C1-C6 alkyl;
- Rx is the reactive group;
- Sc is a conjugated substance;
- and
- L is a covalent linkage;
- with a biological sample in a concentration sufficient to yield a detectable optical response under desired conditions.
A method of staining a biological sample, comprising:
- combining a dye solution comprising a compound having the formula:
- A-BRIDGE-B
- wherein A and B are indolium rings, wherein the substituent on the 3-carbon of one of the indolium rings contains a chemically reactive group or a conjugated substance;
- and
- BRIDGE is a polymethine linker;
- with a biological sample in a concentration sufficient to yield a detectable optical response under desired conditions.
A method for forming a dye-conjugate of a protein, peptide or a nucleic acid polymer, said method comprising:
- combining a dye solution comprising a compound of the formula
- wherein,
- δ is 0 or 1;
- n is 1, 2 or 3;
- W represents the atoms necessary to form one fused aromatic ring having 6 atoms, which atoms are selected from —CH and —CR1′;
- each R1′ is sulfo;
- R2 is C1-C6 alkyl, which is substituted by sulfo;
- R3 is C1-C6 alkyl;
- R4 is C1-C6 alkyl;
- R6 is H or sulfo;
- R7 is H or sulfo;
- R8 is H or sulfo;
- R9 is H or sulfo;
- R12 is -L-Rx or -L-Sc, or a C1-C6 alkyl, which is optionally substituted one or more times by F, Cl, Br, I, hydroxy, carboxy, sulfo, or amino;
- R13 is -L-Rx or -L-Sc;
- R14 is C1-C6 alkyl;
- Rx is the reactive group;
- Sc is a conjugated substance;
- and
- L is a covalent linkage;
- with a sample comprising a protein, peptide or a nucleic acid polymer whereby a dye conjugate is formed.
A method for forming a dye-conjugate of a protein, peptide or a nucleic acid polymer, said method comprising:
- combining a dye solution comprising a compound of the formula
- A-BRIDGE-B
- wherein A and B are indolium rings, wherein the substituent on the 3-carbon of one of the indolium rings contains a chemically reactive group or a conjugated substance;
- and
- BRIDGE is a polymethine linker;
- with a sample comprising a protein, peptide or a nucleic acid polymer whereby a dye conjugate is formed.
A kit for fluorescent labeling of a biological sample, comprising:
- a dye solution comprising a compound having the formula:
- wherein,
- δ is 0 or 1;
- n is 1, 2 or 3;
- W represents the atoms necessary to form one fused aromatic ring having 6 atoms, which atoms are selected from —CH and —CR1′;
- each R1′ is sulfo;
- R2 is C1-C6 alkyl, which is substituted by sulfo;
- R3 is C1-C6 alkyl;
- R4 is C1-C6 alkyl;
- R6 is H or sulfo;
- R7 is H or sulfo;
- R8 is H or sulfo;
- R9 is H or sulfo;
- R12 is -L-Rx or -L-Sc, or a C1-C6 alkyl, which is optionally substituted one or more times by F, Cl, Br, I, hydroxy, carboxy, sulfo, or amino;
- R13 is -L-Rx or -L-Sc;
- R14 is C1-C6 alkyl;
- Rx is the reactive group;
- Sc is a conjugated substance;
- and
- L is a covalent linkage;
- and
- a buffer.
A kit for fluorescent labeling of a biological sample, comprising:
- a dye solution comprising a compound having the formula:
- A-BRIDGE-B
- wherein A and B are indolium rings, wherein the substituent on the 3-carbon of one of the indolium rings contains a chemically reactive group or a conjugated substance;
- and
- BRIDGE is a polymethine linker;
- and
- a buffer.
A carbocyanine dye comprising at least one substituted indolium ring system wherein a substituent on the 3-carbon of the indolium ring contains a chemically reactive group or a conjugated substance.
The carbocyanine dye of claim 62, comprising two substituted indolium ring systems wherein a substituent on the 3-carbon of one of the indolium ring contains a chemically reactive group or a conjugated substance.
A compound comprising 3-(5-carboxypentyl)-2,3-dimethyl-5-sulfoindolium or a salt thereof.
Dependencies are inferred from the claim text.