Texas red, a hydrophilic, red-emitting flourophore for use with flourescein in dual parameter flow microfluorometric and fluorescence microscopic studies

Titus, Julie A.; Haugland, Richard P.; Sharrow, Susan O.; Segal, David M.

doi:10.1016/0022-1759(82)90225-3

Cited by 267 publications

(140 citation statements)

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“…Another method for reducing autofluorescence is to use a single laser and measure two different fluorescence emission regions: one region covering the optimal emission of the fluorescent label and the other at a longer wavelength region, outside the optimal emission region of the fluorescent label (1,ll). While this method works well with fluorescein, the efficacy decreases when fluorochromes that emit in the red spectral region (6,12,19) are used because of decrease in response of PMTs and increase in noise at the longer wavelengths. An exception to this is R-phycoerythrin (19), which when excited at 488-nm wavelength, emits in the yellow (575 nm).…”

Section: Discussionmentioning

confidence: 99%

“…While this method works well with fluorescein, the efficacy decreases when fluorochromes that emit in the red spectral region (6,12,19) are used because of decrease in response of PMTs and increase in noise at the longer wavelengths. An exception to this is R-phycoerythrin (19), which when excited at 488-nm wavelength, emits in the yellow (575 nm). The green autofluorescence (515 to 545 nm) could then be subtracted from the yellow R-phycoerythrinlabeled antibody plus autofluorescence components.…”

Section: Discussionmentioning

confidence: 99%

“…In addition to using the 413-nm wavelength suboptimally to excite the FITC-labeled antibody, the uv (350 nm) lines from a n argon laser were also tested for this purpose. Antibody labeled with Texas red (19) was also used to demonstrate the methodology. The primary excitation wavelength for Texas red was 568 nm (0.4 W) ( …”

Section: -mentioning

confidence: 99%

“…Other fluorochromes such as the rhodamines (6,19) and the phycoerythrins and phycocyanins (12) can be used. We presently do not know the absolute limit of detection in FITC equivalent molecules per cell, but we were able to detect macrophages labeled at very dilute MAC 1 antibody concentrations.…”

Section: Single-lasermentioning

confidence: 99%

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Dual‐laser, differential fluorescence correction method for reducing cellular background autofluorescence

Steinkamp

Stewart

1986

Cytometry

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A method has been developed for reducing the intrinsic autofluorescence background component in cells labeled with fluorescent antibodies, thus permitting low levels of antibody-binding on highly autofluorescent cells to be quantified. The method is based on the broad autofluorescent excitation spectra compared to the well-defined spectra of the fluorescent label. Two laser wavelengths were used, one optimally to excite the fluorescent label plus autofluorescence and the second to excite only the autofluorescence. Two fluorescence measurements were made in the same wavelength region and the signals were subtracted on a cell-by-cell basis using a difference amplifier to zero the autofluorescence and amplify the signal from the fluorescent label. Test results on unlabeled autofluorescent macrophages showed that the autofluorescence component was reduced by balancing the signal inputs to the difference amplifier. When labeled macrophages were analyzed, the autofluorescence was reduced and the fluorescent-labeled antibody-binding component was amplified, The method was also able to resolve labeled lymphocytes from unlabeled autofluorescent macrophages.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: -mentioning

confidence: 99%

Section: Single-lasermentioning

confidence: 99%

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Dual‐laser, differential fluorescence correction method for reducing cellular background autofluorescence

Steinkamp

Stewart

1986

Cytometry

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“…Texas Red fulfills these needs in combinatin with FITC (20) and is used by other investigators as well (8).…”

Section: Discussionmentioning

confidence: 99%

Inexpensive upgrading of a FACS I and isolation of rare somatic variants by double‐fluorescence sorting

Weichel¹,

Liesegang²,

Gehrke³

et al. 1985

Cytometry

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We have modified a FACS I by addition of a tibodies labelled with FITC or Texas Red, we tunable dye laser and an optical system for have isolated from a murine T-cell lymphoma fluorescence detection that allows physically line variant subclones expressing structurally independent measurement of green and red altered histocompatibility class I (H-2Kk) immunofluorescence. These modifications are molecules. inexpensive and should be applicable to most single-laser systems. By using the modified machine and double-fluorescence activated Key words: Dual immunofluorescence, cell selection with H-2Kk-specific monoclonal an-sorting, Texas Red, H-2 variantsThe experiments reported in this paper were aimed at isolating structural variants from the mouse lymphoma line LDHB that express altered instead of wild type H2Kk molecules on the cell surface. These variants are interesting for the investigation of the structural basis of class I antigen function in immune recognition. Furthermore, they can be used to approach the question of which types of somatic mutation occur in the polymorphic family of class I genes.In our previous work, we had achieved this aim by inhibiting the binding of one fluorescent monoclonal anti-Kk antibody to the cells by an excess of a second, unlabelled one and selecting for rare fluorescent cells in the fluorescence activated cell sorter (FACS) (10). This approach was restricted to the selection of structural variants that had lost the expression of one of two Kk determinants, which had to be sufficiently close to each other (topographically) so that the binding of an antibody to one of them would inhibit the binding of a second antibody to the other. In the present work this limitation was overcome by staining wild type cells with two monoclonal anti-Kk antibodies that are labelled with green and red fluorescent tags, respectively, and enriching the cells that carry one label only. For this purpose, we have upgraded a single-laser FACS I instrument by addition of a dye laser that supplies the excitation light for red-fluorescent dyes, such as Texas Red or propidium iodide, that can be used for dual parameter sorting with fluorescein as a green fluorescent label. In contrast to other approaches to double (or triple) fluorescence (161, the dye laser is pumped by the same argon laser that provides the 488 nm light for fluorescein excitation. MATERIALS AND METHODS Cell SorterThe modifications described here were added to a FACS I instrument (Becton Dickinson, Mountain View, CA; serial no. 04). A 5-watt argon ion laser (SpectraPhysics Model 164-08, Darmstadt, FRG) operated at 4 Watts in an all-line mode to give approximately 800 milliwatts at 488 nm and 1.7 watt at 514 nm served as a source for 488 nm excitation of fluorescein and for pumping a Model 375 dye laser (Spectra-Physics, Darmstadt, FRG). The latter was operated with Rhodamin 6G Cambda Physik, Gottingen, FRG) in a setup as described by . Reflectors, dichroic mirrors, and interference and coloured-glass filters used in the system (Fig. 1)

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Advances in the microscopy of osteoarthritis

Gardner

Salter

Oates

1997

Microsc. Res. Tech.

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Texas red, a hydrophilic, red-emitting flourophore for use with flourescein in dual parameter flow microfluorometric and fluorescence microscopic studies

Cited by 267 publications

References 4 publications

Dual‐laser, differential fluorescence correction method for reducing cellular background autofluorescence

Dual‐laser, differential fluorescence correction method for reducing cellular background autofluorescence

Inexpensive upgrading of a FACS I and isolation of rare somatic variants by double‐fluorescence sorting

Advances in the microscopy of osteoarthritis

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