An ensemble and single-molecule fluorescence microscopy investigation of phase-separated monolayer films stained with Nile Red.
Phase-separated Langmuir-Blodgett monolayer films prepared from mixtures of arachidic acid (C19H39COOH) and perfluorotetradecanoic acid (C13F27COOH) were stained via spin-casting with the polarity sensitive phenoxazine dye Nile Red, and characterized using a combination of ensemble and single-molecule fluorescence microscopy measurements. Ensemble fluorescence microscopy and spectromicroscopy showed that Nile Red preferentially associated with the hydrogenated domains of the phase-separated films, and was strongly fluorescent in these areas of the film. These measurements, in conjunction with single-molecule fluorescence imaging experiments, also indicated that a small sub-population of dye molecules localizes on the perfluorinated regions of the sample, but that this sub-population is spectroscopically indistinguishable from that associated with the hydrogenated domains. The relative importance of selective dye adsorption and local polarity sensitivity of Nile Red for staining applications in phase-separated LB films as well as in cellular environments is discussed in context of the experimental results.
Department of Chemistry, University of Saskatchewan 110 Science Place, Saskatoon, SK S7N 5C9, Canada.
Yin
Y
Lu
Robyn
R
Porterfield
Terri
T
Thunder
Matthew F
MF
Paige
D004537:Eicosanoic Acids / Q000737:chemistry; D005466:Fluorocarbons / Q000737:chemistry; D018625:Microscopy, Atomic Force; D008856:Microscopy, Fluorescence / Q000379:methods*; D019814:Myristic Acid / Q000737:chemistry; D010078:Oxazines / Q000737:chemistry*; D044367:Phase Transition; D013050:Spectrometry, Fluorescence; D013194:Staining and Labeling
D004537:Eicosanoic Acids; D005466:Fluorocarbons; D010078:Oxazines; D019814:Myristic Acid; C073608:perfluorooctane; C044808:nile red; C094477:arachidic acid