DNA staining for fluorescence and laser confocal microscopy.
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).
Suzuki
T
T
Department of Cell Biology, Gunma University, Japan.
Fujikura
K
K
Higashiyama
T
T
Takata
K
K
D000818:Animals; D052160:Benzothiazoles; D001583:Benzoxazoles; D002232:Carbocyanines; D002467:Cell Nucleus / Q000737:chemistry*; D004247:DNA / Q000032:analysis*; D003959:Diamines; D005455:Fluorescent Antibody Technique; D005456:Fluorescent Dyes; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence; D009930:Organic Chemicals; D011419:Propidium; D011804:Quinolines; D011806:Quinolinium Compounds; D051381:Rats; D017207:Rats, Sprague-Dawley; D012260:Ribonucleases / Q000378:metabolism; D013844:Thiazoles
D052160:Benzothiazoles; D001583:Benzoxazoles; D002232:Carbocyanines; D003959:Diamines; D005456:Fluorescent Dyes; D009930:Organic Chemicals; D011804:Quinolines; D011806:Quinolinium Compounds; C098830:TO-PRO-3; D013844:Thiazoles; C075295:1,1'-(4,4,7,7-tetramethyl-4,7-diazaundecamethylene)bis-4-(3-methyl-2,3-dihydro(benzo-1,3-thiazole)-2-methylidene)quinolinium; C089813:YO-PRO 1; C098022:SYBR Green I; D011419:Propidium; D004247:DNA; D012260:Ribonucleases