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Evaluation of acridine orange, LysoTracker Red, and quinacrine as fluorescent probes for long‐term tracking of acidic vesicles
Author(s) -
PierzyńskaMach Agnieszka,
Janowski Paweł A.,
Dobrucki Jurek W.
Publication year - 2014
Publication title -
cytometry part a
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.316
H-Index - 90
eISSN - 1552-4930
pISSN - 1552-4922
DOI - 10.1002/cyto.a.22495
Subject(s) - acridine orange , vesicle , fluorescence , phototoxicity , photobleaching , biophysics , chemistry , endocytosis , fluorescence microscope , biochemistry , biology , optics , apoptosis , physics , membrane , cell , in vitro
Abstract Acidic vesicles can be imaged and tracked in live cells after staining with several low molecular weight fluorescent probes, or with fluorescently labeled proteins. Three fluorescent dyes, acridine orange, LysoTracker Red DND‐99, and quinacrine, were evaluated as acidic vesicle tracers for confocal fluorescence imaging and quantitative analysis. The stability of fluorescent signals, achievable image contrast, and phototoxicity were taken into consideration. The three tested tracers exhibit different advantages and pose different problems in imaging experiments. Acridine orange makes it possible to distinguish acidic vesicles with different internal pH but is fairly phototoxic and can cause spectacular bursts of the dye‐loaded vesicles. LysoTracker Red is less phototoxic but its rapid photobleaching limits the range of useful applications considerably. We demonstrate that quinacrine is most suitable for long‐term imaging when a high number of frames is required. This capacity made it possible to trace acidic vesicles for several hours, during a process of drug‐induced apoptosis. An ability to record the behavior of acidic vesicles over such long periods opens a possibility to study processes like autophagy or long‐term effects of drugs on endocytosis and exocytosis. © 2014 International Society for Advancement of Cytometry