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Journal of Lipid Research, Vol 26, 781-789, Copyright © 1985 by Lipid Research, Inc.
P Greenspan and SD Fowler
We found that the dye nile red, 9-diethylamino-5H-
benzo[alpha]phenoxazine-5-one, can be applied as a fluorescent vital stain
for the detection of intracellular lipid droplets by fluorescence
microscopy and flow cytofluorometry (J. Cell. Biol. 1985. 100: 965- 973).
To understand the selectivity of the staining, we examined the fluorescence
properties of nile red in the presence of organic solvents and model lipid
systems. Nile red was found to be both very soluble and strongly
fluorescent in organic solvents. The excitation and emission spectra of
nile red shifted to shorter wavelengths with decreasing solvent polarity.
However, the fluorescence of nile red was quenched in aqueous medium. Nile
red was observed to fluoresce intensely in the presence of aqueous
suspensions of phosphatidylcholine vesicles (excitation maximum: 549 nm;
emission maximum: 628 nm). When neutral lipids such as triacylglycerols or
cholesteryl esters were incorporated with phosphatidylcholine to form
microemulsions, nile red fluorescence emission maxima shifted to shorter
wavelengths. Serum lipoproteins also induced nile red fluorescence and
produced spectral blue shifts. Nile red fluorescence was not observed in
the presence of either immunoglobulin G or gelatin. These results
demonstrate that nile red fluorescence accompanied by a spectral blue shift
reflects the presence of nile red in a hydrophobic lipid environment and
account for the selective detection of neutral lipid by the dye. Nile red
thus serves as an excellent fluorescent lipid probe.
ARTICLES
Spectrofluorometric studies of the lipid probe, nile red
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