title: Whole-object fluorescence lifetime setup for efficient non-imaging quantitative intracellular fluorophore measurements.
abstract: In the present study we introduce a Whole-Object Fluorescence Life Time (wo-FLT) measurement approach for ease and a relatively inexpensive method of tracing alterations in intracellular fluorophore distribution and in the physical-chemical features of the microenvironments hosting the fluorophore. Two common fluorophores, Rhodamine 123 and Acridine Orange, were used to stain U937 cells which were incubated, with and without either Carbonyl cyanide 3-chlorphenylhydrazon or the apoptosis inducer H(2)O(2). The wo-FLT, which is a non-imaging quantitative measurement, was able to detect several fluorescence decay components and corresponding weights in a single cell resolution. Following cell treatment, both decay time and weight were altered. Results suggest that the prominent factor responsible for these alterations and in some cases to a shift in emission spectrum as well, is the intracellular fluorophore local concentration. In this study it was demonstrated that the proposed wo-FLT method is superior to color fluorescence based imaging in cases where the emission spectrum of a fluorophore remains unchanged during the investigated process. The proposed wo-FLT approach may be of particular importance when direct imaging is impossible.
authors.affiliation: The Biophysical Interdisciplinary Jerome Schottenstein Center for the Research and the Technology of the Cellome, Physics Department, Bar-Ilan University, Ramat-Gan, Israel.
authors.forename: Yaniv
authors.initials: Y
authors.lastname: Namer
authors.forename: Lior
authors.initials: L
authors.lastname: Turgeman
authors.forename: Mordechai
authors.initials: M
authors.lastname: Deutsch
authors.forename: Dror
authors.initials: D
authors.lastname: Fixler
mesh_terms: D000165:Acridine Orange / Q000737:chemistry*; D017209:Apoptosis / Q000187:drug effects; D002258:Carbonyl Cyanide m-Chlorophenyl Hydrazone / Q000494:pharmacology; D005453:Fluorescence; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology; D020112:Rhodamine 123 / Q000737:chemistry*; D013050:Spectrometry, Fluorescence; D013194:Staining and Labeling; D020298:U937 Cells
chemical_list: D020112:Rhodamine 123; D002258:Carbonyl Cyanide m-Chlorophenyl Hydrazone; D006861:Hydrogen Peroxide; D000165:Acridine Orange