title: TO-PRO-3 iodide: a novel HeNe laser-excitable DNA stain as an alternative for propidium iodide in multiparameter flow cytometry.
abstract: A new red emitting fluorophore, TO-PRO-3 iodide (TP3), which is best excited by an HeNe laser (633 nm), has been compared with propidium iodide (PI) for measuring relative DNA content. TP3, which has a peak absorbance at 642 nm and emission at 661 nm, has been tested on peripheral blood lymphocytes (PBL) and keratinocytes in a two-laser system. As an example, we present a three-color flow cytometric application utilizing TP3 in combination with fluorescein-isothiocyanate (FITC) and phycoerythrin (PE) conjugated to monoclonal antibodies in this paper. A subequilibrium concentration of 1 microM TP3, most preferably used in combination with RNase treatment, proved to be a powerful alternative for DNA amount determination. In human- and mouse-Balb/MK-keratinocyte populations with different S-phase fractions, PI and TP3 showed a good correlation. Finally, in the triple labelling experiment we clearly demonstrated that TP3 is readily applied to the analysis of binding of two antibodies and relative DNA content simultaneously.
authors.affiliation: Department of Dermatology, University Hospital Nijmegen, The Netherlands.
authors.forename: C A
authors.initials: CA
authors.lastname: Van Hooijdonk
authors.forename: C P
authors.initials: CP
authors.lastname: Glade
authors.forename: P E
authors.initials: PE
authors.lastname: Van Erp
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D004247:DNA / Q000032:analysis*; D015342:DNA Probes; D005069:Evaluation Studies as Topic; D005434:Flow Cytometry / Q000379:methods*; D016650:Fluorescein-5-isothiocyanate; D005456:Fluorescent Dyes; D006801:Humans; D015603:Keratinocytes / Q000737:chemistry; D007834:Lasers; D008214:Lymphocytes / Q000737:chemistry; D051379:Mice; D010799:Phycoerythrin; D011419:Propidium; D013194:Staining and Labeling
chemical_list: D015342:DNA Probes; D005456:Fluorescent Dyes; D010799:Phycoerythrin; D011419:Propidium; D004247:DNA; D016650:Fluorescein-5-isothiocyanate