title: Development of practical red fluorescent probe for cytoplasmic calcium ions with greatly improved cell-membrane permeability.
abstract: Fluorescence imaging of calcium ions (Ca(2+)) has become an essential technique for investigation of signaling pathways involving Ca(2+) as a second messenger. But, Ca(2+) signaling is involved in many biological phenomena, and therefore simultaneous visualization of Ca(2+) and other biomolecules (multicolor imaging) would be particularly informative. For this purpose, we set out to develop a fluorescent probe for Ca(2+) that would operate in a different color region (red) from that of probes for other molecules, many of which show green fluorescence, as exemplified by green fluorescent protein (GFP). We previously developed a red fluorescent probe for monitoring cytoplasmic Ca(2+) concentration, based on our established red fluorophore, TokyoMagenta (TM), but there remained room for improvement, especially as regards efficiency of introduction into cells. We considered that this issue was probably mainly due to limited water solubility of the probe. So, we designed and synthesized a red-fluorescent probe with improved water solubility. We confirmed that this Ca(2+) red-fluorescent probe showed high cell-membrane permeability with bright fluorescence. It was successfully applied to fluorescence imaging of not only live cells, but also brain slices, and should be practically useful for multicolor imaging studies of biological mechanisms.
authors.affiliation: Graduate School of Pharmaceutical Sciences, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
authors.forename: Kazuhisa
authors.initials: K
authors.lastname: Hirabayashi
authors.affiliation: Graduate School of Pharmaceutical Sciences, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan. Electronic address: khanaoka@mol.f.u-tokyo.ac.jp.
authors.forename: Kenjiro
authors.initials: K
authors.lastname: Hanaoka
authors.affiliation: Graduate School of Pharmaceutical Sciences, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
authors.forename: Takahiro
authors.initials: T
authors.lastname: Egawa
authors.affiliation: Graduate School of Pharmaceutical Sciences, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
authors.forename: Chiaki
authors.initials: C
authors.lastname: Kobayashi
authors.affiliation: Graduate School of Pharmaceutical Sciences, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
authors.forename: Shodai
authors.initials: S
authors.lastname: Takahashi
authors.affiliation: Graduate School of Pharmaceutical Sciences, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan; Precursory Research for Embryonic Science and Technology (PRESTO), Japan Science and Technology Agency (JST), 4-1-8 Honcho, Kawaguchi, Saitama 332-0012, Japan.
authors.forename: Toru
authors.initials: T
authors.lastname: Komatsu
authors.affiliation: Graduate School of Pharmaceutical Sciences, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
authors.forename: Tasuku
authors.initials: T
authors.lastname: Ueno
authors.affiliation: Graduate School of Pharmaceutical Sciences, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
authors.forename: Takuya
authors.initials: T
authors.lastname: Terai
authors.affiliation: Graduate School of Pharmaceutical Sciences, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
authors.forename: Yuji
authors.initials: Y
authors.lastname: Ikegaya
authors.affiliation: Drug Discovery Initiative, The University of Tokyo, Tokyo 113-0033, Japan.
authors.forename: Tetsuo
authors.initials: T
authors.lastname: Nagano
authors.affiliation: Graduate School of Pharmaceutical Sciences, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan; Graduate School of Medicine, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan; CREST, AMED, Saitama 332-0012, Japan. Electronic address: uranokun@m.u-tokyo.ac.jp.
authors.forename: Yasuteru
authors.initials: Y
authors.lastname: Urano
mesh_terms: D000818:Animals; D002118:Calcium / Q000737:chemistry* / Q000378:metabolism; D002463:Cell Membrane Permeability; D002478:Cells, Cultured; D003593:Cytoplasm / Q000737:chemistry* / Q000378:metabolism; D005456:Fluorescent Dyes / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D007477:Ions / Q000737:chemistry / Q000378:metabolism; D051379:Mice; D015394:Molecular Structure; D041681:NIH 3T3 Cells; D013050:Spectrometry, Fluorescence; D013056:Spectrophotometry, Ultraviolet
chemical_list: D005456:Fluorescent Dyes; D007477:Ions; D002118:Calcium
keywords: Calcium; Fluorescence; Imaging; Small molecule; Spectroscopy