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Journal of Histochemistry and Cytochemistry, Vol. 45, 49-54, Copyright © 1997 by The Histochemical Society, Inc.

ARTICLE

DNA Staining for Fluorescence and Laser Confocal Microscopy

Takeshi Suzukia, Keiko Fujikuraa, Tetsuya Higashiyamab, and Kuniaki Takataa
a Department of Cell Biology, Institute for Molecular and Cellular Regulation, Gunma University, Gunma, Japan
b Department of Plant Sciences, Graduate School of Science, University of Tokyo, Tokyo, Japan

Correspondence to: Takeshi Suzuki, Dept. of Cell Biology, Inst. for Molecular and Cellular Regulation, Gunma University, Showa-machi 3-39-15, Maebashi, Gunma 371, Japan.

We examined five nucleic acid binding fluorescent dyes, propidium iodide, SYBR Green I, YO-PRO-1, TOTO-3, and TO-PRO-3, for nuclear DNA staining, visualized by fluorescence and laser confocal microscopy. The optimal concentration, co-staining of RNA, and bleaching speeds were examined. SYBR Green I and TO-PRO-3 almost preferentially stained the nuclear DNA, and the other dyes co-stained the cytoplasmic RNA. RNAse treatment completely prevented the cytoplasmic RNA staining. In conventional fluorescence microscopy, these dyes can be used in combination with fluorescence-labeled antibodies. Among the dyes tested, TOTO-3 and TO-PRO-3 stained the DNAs with far-red fluorescence under red excitation. Under Kr/Ar-laser illumination, TOTO-3 and TO-PRO-3 were best suited as the nuclear staining dyes in the specimens immunolabeled with fluorescein and rhodamine (or Texas red). (J Histochem Cytochem 45:49-53, 1997)

Key Words: Laser confocal microscopy, Cell nuclear DNA, Propidium iodide, SYBR green I, YO-PRO-1, TOTO-3, TO-PRO-3


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