Title: Phototoxicity of Hoechst 33342 in time-lapse fluorescence microscopy
Authors: Purschke, Martin
Rubio, Noemi
Held, Kathryn D
Redmond, Robert W # ×
Issue Date: 2010
Publisher: Royal Society of Chemistry
Series Title: Photochemical & Photobiological Sciences vol:9 issue:12 pages:1634-1639
Abstract: Dyes that bind to DNA, such as Hoechst 33342, are commonly used to visualize chromatin in live cells by fluorescence microscopy. A caveat is that the probes themselves should not perturb cellular responses and under normal conditions the dyes are generally non-toxic. However, researchers are increasingly using computerized time-lapse microscopy (CTLM), where cells stained with fluorescent dyes are often imaged frequently over a period of several days, to follow cellular responses in real time. Little is currently known about possible toxicity of fluorescent DNA dyes under CTLM conditions. In this study we demonstrate that the common live-cell DNA stain Hoechst 33342 can cause apoptosis under CTLM conditions. Although toxicity is evident at long times in the absence of imaging at high dye concentrations, phototoxicity from repeated excitation of the dye in the imaging process is dominant. We show that phototoxicity is a function of the product of light fluence and dye concentration, irrespective of irradiance, frequency and total number of scans. Thus, phototoxicity can be prevented by a combination of dye concentration and imaging procedure that is below this threshold. These quantitative data can be used as a guide to others performing time-lapse microscopy studies with this common live-cell DNA stain and serves as a caution for researchers when using other fluorescent stains under CTLM conditions.
ISSN: 1474-905X
Publication status: published
KU Leuven publication type: IT
Appears in Collections:Non-KU Leuven Association publications
× corresponding author
# (joint) last author

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