Fluorescent rhodol derivatives

Versatile, photostable labels and tracers

James E. Whitaker, Rosaria P. Haugland, Diane Ryan, Peter C. Hewitt, Richard P. Haugland, Franklyn G. Prendergast

Research output: Contribution to journalArticle

92 Citations (Scopus)

Abstract

A series of chemically reactive, fluorescent rhodol derivatives was prepared and evaluated. Reactive functional groups included activated esters, amines, haloacetamides, fixable hydrazide derivatives, acrylamides, and photoaffinity reagents. Depending on the choice of substituents, absorption maxima of the dyes varied from 490 to 550 nm with extinction coefficients that were generally greater than 50,000 m-1 cm-1 in aqueous solution and emission maxima from 520 to 580 nm. Most of the compounds investigated exhibited fluorescence lifetimes between 3 and 4 ns. Individual derivatives were suitable for excitation with the 488 and 514-nm lines of the argon ion laser and the 546-nm line of the mercury arc lamp and were compatible for use with standard fluorescein and rhodamine filter sets. The rhodol dyes were more photostable and less sensitive to pH changes in the physiological range than fluorescein derivatives. Some examples show absorption maxima at or near 514 nm, an excitation wavelength that is useful for multicolor fluorescence microscopy, flow cytometry, and DNA sequencing. Derivatives were also prepared that exhibit absorption and emission maxima similar to those of tetramethylrhodamine (TMR) analogs but with higher quantum yields in aqueous solution. A number of the dyes had higher solubilities in aqueous systems and were less quenched on conjugation to proteins than TMR derivatives. Appropriate substitution results in a wider range of solubilities in hydrophilic or lipophilic solvents than is easily accomplished with fluorescein or TMR derivatives. Conjugates of a number of the rhodol fluorophores were generally more photostable and less pH sensitive than fluorescein conjugates and more fluorescent than TMR conjugates.

Original languageEnglish (US)
Pages (from-to)267-279
Number of pages13
JournalAnalytical Biochemistry
Volume207
Issue number2
DOIs
StatePublished - 1992

Fingerprint

Fluorescein
Labels
Derivatives
Coloring Agents
Solubility
Acrylamides
Rhodamines
Gas Lasers
Mercury
DNA Sequence Analysis
Fluorescence Microscopy
Amines
Flow Cytometry
Esters
Arc lamps
Fluorescence
Mercury vapor lamps
Flow cytometry
Fluorophores
Argon

ASJC Scopus subject areas

  • Biochemistry
  • Biophysics
  • Molecular Biology

Cite this

Whitaker, J. E., Haugland, R. P., Ryan, D., Hewitt, P. C., Haugland, R. P., & Prendergast, F. G. (1992). Fluorescent rhodol derivatives: Versatile, photostable labels and tracers. Analytical Biochemistry, 207(2), 267-279. https://doi.org/10.1016/0003-2697(92)90011-U

Fluorescent rhodol derivatives : Versatile, photostable labels and tracers. / Whitaker, James E.; Haugland, Rosaria P.; Ryan, Diane; Hewitt, Peter C.; Haugland, Richard P.; Prendergast, Franklyn G.

In: Analytical Biochemistry, Vol. 207, No. 2, 1992, p. 267-279.

Research output: Contribution to journalArticle

Whitaker, JE, Haugland, RP, Ryan, D, Hewitt, PC, Haugland, RP & Prendergast, FG 1992, 'Fluorescent rhodol derivatives: Versatile, photostable labels and tracers', Analytical Biochemistry, vol. 207, no. 2, pp. 267-279. https://doi.org/10.1016/0003-2697(92)90011-U
Whitaker JE, Haugland RP, Ryan D, Hewitt PC, Haugland RP, Prendergast FG. Fluorescent rhodol derivatives: Versatile, photostable labels and tracers. Analytical Biochemistry. 1992;207(2):267-279. https://doi.org/10.1016/0003-2697(92)90011-U
Whitaker, James E. ; Haugland, Rosaria P. ; Ryan, Diane ; Hewitt, Peter C. ; Haugland, Richard P. ; Prendergast, Franklyn G. / Fluorescent rhodol derivatives : Versatile, photostable labels and tracers. In: Analytical Biochemistry. 1992 ; Vol. 207, No. 2. pp. 267-279.
@article{c06fe0ca51894f439df726339a2555bc,
title = "Fluorescent rhodol derivatives: Versatile, photostable labels and tracers",
abstract = "A series of chemically reactive, fluorescent rhodol derivatives was prepared and evaluated. Reactive functional groups included activated esters, amines, haloacetamides, fixable hydrazide derivatives, acrylamides, and photoaffinity reagents. Depending on the choice of substituents, absorption maxima of the dyes varied from 490 to 550 nm with extinction coefficients that were generally greater than 50,000 m-1 cm-1 in aqueous solution and emission maxima from 520 to 580 nm. Most of the compounds investigated exhibited fluorescence lifetimes between 3 and 4 ns. Individual derivatives were suitable for excitation with the 488 and 514-nm lines of the argon ion laser and the 546-nm line of the mercury arc lamp and were compatible for use with standard fluorescein and rhodamine filter sets. The rhodol dyes were more photostable and less sensitive to pH changes in the physiological range than fluorescein derivatives. Some examples show absorption maxima at or near 514 nm, an excitation wavelength that is useful for multicolor fluorescence microscopy, flow cytometry, and DNA sequencing. Derivatives were also prepared that exhibit absorption and emission maxima similar to those of tetramethylrhodamine (TMR) analogs but with higher quantum yields in aqueous solution. A number of the dyes had higher solubilities in aqueous systems and were less quenched on conjugation to proteins than TMR derivatives. Appropriate substitution results in a wider range of solubilities in hydrophilic or lipophilic solvents than is easily accomplished with fluorescein or TMR derivatives. Conjugates of a number of the rhodol fluorophores were generally more photostable and less pH sensitive than fluorescein conjugates and more fluorescent than TMR conjugates.",
author = "Whitaker, {James E.} and Haugland, {Rosaria P.} and Diane Ryan and Hewitt, {Peter C.} and Haugland, {Richard P.} and Prendergast, {Franklyn G.}",
year = "1992",
doi = "10.1016/0003-2697(92)90011-U",
language = "English (US)",
volume = "207",
pages = "267--279",
journal = "Analytical Biochemistry",
issn = "0003-2697",
publisher = "Academic Press Inc.",
number = "2",

}

TY - JOUR

T1 - Fluorescent rhodol derivatives

T2 - Versatile, photostable labels and tracers

AU - Whitaker, James E.

AU - Haugland, Rosaria P.

AU - Ryan, Diane

AU - Hewitt, Peter C.

AU - Haugland, Richard P.

AU - Prendergast, Franklyn G.

PY - 1992

Y1 - 1992

N2 - A series of chemically reactive, fluorescent rhodol derivatives was prepared and evaluated. Reactive functional groups included activated esters, amines, haloacetamides, fixable hydrazide derivatives, acrylamides, and photoaffinity reagents. Depending on the choice of substituents, absorption maxima of the dyes varied from 490 to 550 nm with extinction coefficients that were generally greater than 50,000 m-1 cm-1 in aqueous solution and emission maxima from 520 to 580 nm. Most of the compounds investigated exhibited fluorescence lifetimes between 3 and 4 ns. Individual derivatives were suitable for excitation with the 488 and 514-nm lines of the argon ion laser and the 546-nm line of the mercury arc lamp and were compatible for use with standard fluorescein and rhodamine filter sets. The rhodol dyes were more photostable and less sensitive to pH changes in the physiological range than fluorescein derivatives. Some examples show absorption maxima at or near 514 nm, an excitation wavelength that is useful for multicolor fluorescence microscopy, flow cytometry, and DNA sequencing. Derivatives were also prepared that exhibit absorption and emission maxima similar to those of tetramethylrhodamine (TMR) analogs but with higher quantum yields in aqueous solution. A number of the dyes had higher solubilities in aqueous systems and were less quenched on conjugation to proteins than TMR derivatives. Appropriate substitution results in a wider range of solubilities in hydrophilic or lipophilic solvents than is easily accomplished with fluorescein or TMR derivatives. Conjugates of a number of the rhodol fluorophores were generally more photostable and less pH sensitive than fluorescein conjugates and more fluorescent than TMR conjugates.

AB - A series of chemically reactive, fluorescent rhodol derivatives was prepared and evaluated. Reactive functional groups included activated esters, amines, haloacetamides, fixable hydrazide derivatives, acrylamides, and photoaffinity reagents. Depending on the choice of substituents, absorption maxima of the dyes varied from 490 to 550 nm with extinction coefficients that were generally greater than 50,000 m-1 cm-1 in aqueous solution and emission maxima from 520 to 580 nm. Most of the compounds investigated exhibited fluorescence lifetimes between 3 and 4 ns. Individual derivatives were suitable for excitation with the 488 and 514-nm lines of the argon ion laser and the 546-nm line of the mercury arc lamp and were compatible for use with standard fluorescein and rhodamine filter sets. The rhodol dyes were more photostable and less sensitive to pH changes in the physiological range than fluorescein derivatives. Some examples show absorption maxima at or near 514 nm, an excitation wavelength that is useful for multicolor fluorescence microscopy, flow cytometry, and DNA sequencing. Derivatives were also prepared that exhibit absorption and emission maxima similar to those of tetramethylrhodamine (TMR) analogs but with higher quantum yields in aqueous solution. A number of the dyes had higher solubilities in aqueous systems and were less quenched on conjugation to proteins than TMR derivatives. Appropriate substitution results in a wider range of solubilities in hydrophilic or lipophilic solvents than is easily accomplished with fluorescein or TMR derivatives. Conjugates of a number of the rhodol fluorophores were generally more photostable and less pH sensitive than fluorescein conjugates and more fluorescent than TMR conjugates.

UR - http://www.scopus.com/inward/record.url?scp=0026987490&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0026987490&partnerID=8YFLogxK

U2 - 10.1016/0003-2697(92)90011-U

DO - 10.1016/0003-2697(92)90011-U

M3 - Article

VL - 207

SP - 267

EP - 279

JO - Analytical Biochemistry

JF - Analytical Biochemistry

SN - 0003-2697

IS - 2

ER -