Abstract
Conventional laser scanning microscopy for multiple fluorescent stains can be a useful tool if the problems of autofluorescence and cross-talk are eliminated. The technique of spectral imaging was employed to unmix five different fluorophores - ranging in emission from 435 to 665 nm - applied to a Pseudomonas aeruginosa biofilm with overlapping spectra and which was not possible using traditional channel mode operation. Using lambda scanning and linear unmixing, the five fluorophores could be distinguished with regions of differentiation apparent.
Copyright © 2012 Elsevier B.V. All rights reserved.
Publication types
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Research Support, N.I.H., Extramural
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Research Support, U.S. Gov't, Non-P.H.S.
MeSH terms
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Benzenesulfonates / chemistry
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Biofilms*
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Calibration
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Concanavalin A / chemistry
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Fluorescein-5-isothiocyanate / chemistry
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Fluorescent Dyes / chemistry
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Microscopy, Confocal / methods*
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Organic Chemicals / chemistry
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Organometallic Compounds / chemistry
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Pseudomonas aeruginosa / cytology
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Pseudomonas aeruginosa / physiology*
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Rhodamines / chemistry
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Staining and Labeling
Substances
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Benzenesulfonates
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Fluorescent Dyes
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Organic Chemicals
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Organometallic Compounds
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Rhodamines
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SYTO 9
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Sypro Ruby
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Concanavalin A
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tetramethylrhodamine
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C.I. Fluorescent Brightening Agent 28
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Fluorescein-5-isothiocyanate