Water soluble fluorescent or colored dyes and methods for their use
Compounds useful as fluorescent or colored dyes are disclosed. The compounds have the following structure (I): including stereoisomers, salts and tautomers thereof, wherein R 1 , R 2 , R 3 , L 1 , L 2 , L 3 , L 4 , L 5 , L 6 , M 1 , M 2 , A, q, w and n are as defined herein. Methods associated with preparation and use of such compounds are also provided.
1. A compound having the following structure (IIa):
wherein:
M 1 is, at each occurrence, independently a moiety comprising two or more double bonds and at least one degree of conjugation, and at least one occurrence of M 1 is a moiety comprising three or more rings selected from aryl or heteroaryl, wherein one or more occurrence of M 1 is the same and one or more occurrence of M 1 is different;
L 1 , L 7 and L 3 are, at each occurrence, independently linkers comprising atoms selected from C, O, S, N and P;
R 1 is, at each occurrence, independently H, alkyl or alkoxy;
R 2 is a linker comprising a covalent bond to a biomolecule;
R 3 is H, OH, SH, —NH 2, alkyl, alkylether, hydroxylalkyl, aminoalkyl, hydroxylalkylether, sulfhydrylalkyl, sulfyhdrylalkylether, cyanoalkyl, —Oaralkyl, phosphate, thiophosphate, alkylphospho, alkylthiophospho, —Oalkylphospho, —Oalkylthiophospho, alkyletherphospho, alkyletherthiophospho, —Oalkyletherphospho, —Oalkyletherthiophospho phosphoalkyl, phosphoalkylether, thiophosphoalkyl, thiophosphoalkylether, —Ophosphoalkyl, O—phosphoalkylether, —Othiophosphoalkyl or —Othiophosphoalkylether;
R 4 is, at each occurrence, independently O − , S − , OZ, SZ or N(R 6 ) 2, where Z is a cation and each R 6 is independently H or alkyl;
R 5 is, at each occurrence, independently oxo, thioxo or absent; and
n is an integer from 2 to 20.
2. The compound of claim 1 , wherein the compound has the following structure (IIb):
wherein:
R 7 , R 8 , R 9 , R 10 , R 11 and R 12 are, at each occurrence, independently H or alkyl; and
x 1 , y 1 and z 1 are, at each occurrence, independently an integer from 0 to 5.
3. The compound of claim 2 , wherein the compound has one of the following structures (IIc), (IId), (IIe) or (IIf):
4. The compound of claims 1 , wherein R 3 is OH.
5. The compound of claim 1 , wherein R 3 is phosphate, thiophosphate, phospho, thiophospho, Oalkylphospho, Oalkylthiophospho, Oalkyletherphospho, Oalkyletherthiophospho, Ophosphoalkyl, Ophosphoalkylether, Othiophosphoalkyl or Othiophosphoalkylether optionally substituted with a substituent selected from —OH, NH2, and —SH.
6. The compound of claim 5 , wherein R 3 has one of the following structures:
wherein:
R 3a is —OH, —SH, —NH 2, phosphate or thiophosphate;
R 4a and R 4b are independently O − , S − , OZ or SZ, where Z is a cation;
R 5a and R 5b are independently oxo, or thioxo; and
b and c are each independently integers from 1 to 10.
7. The compound of claim 1 , wherein R 4a and R 4b are each O − and R 5a and R 5b are each oxo.
8. The compound of claim 1 , wherein n is an integer from 2 to 15.
9. The compound of claim 1 , wherein n is an integer from 2 to 10.
10. The compound of claim 1 , wherein at least one M 1 is a dimethylaminostilbene, quinacridone, fluorophenyl-dimethyl-BODIPY, his-fluorophenyl-BODIPY, acridine, terrylene, sexiphenyl, porphyrin, benzopyrene, (fluorophenyl-dimethyl-difluorobora-diaza-indacene) phenyl, (bis-fluorophenyl-difluorobora-diaza-indacene) phenyl, quaterphenyl, bi-benzothiazole, ter-benzothiazole, bi-naphthyl, bi-anthracyl, squaraine, squarylium, 9, 10-ethynylanthracene or ter-naphthyl moiety.
11. The compound of claim 1 , wherein at least one M 1 is p-terphenyl, perylene, azobenzene, phenazine, phenanthroline, acridine, thioxanthrene, chrysene, rubrene, coronene, cyanine, perylene imide, or perylene amide or derivative thereof.
12. The compound of claim 1 , wherein at least one M 1 is a coumarin dye, resorufin dye, dipyrrometheneboron difluoride dye, ruthenium bipyridyl dye, energy transfer dye, thiazole orange dye, polymethine or N-aryl-1,8-naphthalimide dye.
13. The compound of claim 1 , wherein each M 1 is pyrene, perylene, perylene monoimide or 6-FAM or derivative thereof.
14. The compound of claim 1 , wherein M 1 has one of the following structures:
15. The compound of claim 1 , wherein the biomolecule is a nucleic acid, amino acid or a polymer thereof.
16. The compound of claim 15 , wherein the biomolecule is an enzyme, receptor, receptor ligand, antibody, glycoprotein, aptamer or prion.
17. A method of staining a sample, comprising adding to said sample the compound of claim 1 in an amount sufficient to produce an optical response when said sample is illuminated at an appropriate wavelength.
18. The method of claim 17 , wherein said optical response is a fluorescent response.
19. The method of claim 17 , wherein said sample comprises cells.
20. The method of claim 19 , further comprising observing said cells by flow cytometry.