2. Academic Validation
  3. Detection of LacZ-Positive Cells in Living Tissue with Single-Cell Resolution

Detection of LacZ-Positive Cells in Living Tissue with Single-Cell Resolution

  • Angew Chem Int Ed Engl. 2016 Aug 8;55(33):9620-4. doi: 10.1002/anie.201603328.
Tomohiro Doura 1,  Mako Kamiya 2 3,  Fumiaki Obata 4,  Yoshifumi Yamaguchi 4 5,  Takeshi Y Hiyama 6 7,  Takashi Matsuda 7,  Akiyoshi Fukamizu 8 9,  Masaharu Noda 6 7,  Masayuki Miura 4 10,  Yasuteru Urano 11 12 13
Affiliations

Affiliations

  • 1 Graduate School of Medicine, The University of Tokyo, 7-3-1, Hongo, Bunkyo-ku, Tokyo, 113-0033, Japan.
  • 2 Graduate School of Medicine, The University of Tokyo, 7-3-1, Hongo, Bunkyo-ku, Tokyo, 113-0033, Japan. mkamiya@m.u-tokyo.ac.jp.
  • 3 PRESTO, Japan, Science and Technology Agency, 4-1-8 Honcho, Kawaguchi, Saitama, 332-0012, Japan. mkamiya@m.u-tokyo.ac.jp.
  • 4 Graduate School of Pharmaceutical Sciences, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo, 113-0033, Japan.
  • 5 PRESTO, Japan, Science and Technology Agency, 4-1-8 Honcho, Kawaguchi, Saitama, 332-0012, Japan.
  • 6 Division of Molecular Neurobiology, National Institute for Basic Biology, 5-1 Higashiyama, Myodaiji-cho, Okazaki, Aichi, 444-8787, Japan.
  • 7 School of Life Science, The Graduate University for Advanced Studies, 5-1 Higashiyama, Myodaiji-cho, Okazaki, Aichi, 444-8787, Japan.
  • 8 Life Science Center, Tsukuba Advanced Research Alliance, Tsukuba, Ibaraki, 305-8577, Japan.
  • 9 Graduate School of Life and Environmental Sciences, University of Tsukuba, Tsukuba, Ibaraki, 305-8577, Japan.
  • 10 CREST, Japan, Agency for Medical Research and Development, 1-7-1 Otemachi, Chiyoda-ku, Tokyo, 100-0004, Japan.
  • 11 Graduate School of Medicine, The University of Tokyo, 7-3-1, Hongo, Bunkyo-ku, Tokyo, 113-0033, Japan. uranokun@m.u-tokyo.ac.jp.
  • 12 Graduate School of Pharmaceutical Sciences, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo, 113-0033, Japan. uranokun@m.u-tokyo.ac.jp.
  • 13 CREST, Japan, Agency for Medical Research and Development, 1-7-1 Otemachi, Chiyoda-ku, Tokyo, 100-0004, Japan. uranokun@m.u-tokyo.ac.jp.
PMID: 27400827 DOI: 10.1002/anie.201603328
Abstract

The LacZ gene, which encodes Escherichia coli β-galactosidase, is widely used as a marker for cells with targeted gene expression or disruption. However, it has been difficult to detect lacZ-positive cells in living organisms or tissues at single-cell resolution, limiting the utility of existing lacZ reporters. Herein we present a newly developed fluorogenic β-galactosidase substrate suitable for labeling live cells in culture, as well as in living tissues. This precisely functionalized fluorescent probe exhibited dramatic activation of fluorescence upon reaction with the Enzyme, remained inside cells by anchoring itself to intracellular proteins, and provided single-cell resolution. Neurons labeled with this probe preserved spontaneous firing, which was enhanced by application of ligands of receptors expressed in the cells, suggesting that this probe would be applicable to investigate functions of targeted cells in living tissues and organisms.

Keywords

fluorescent probes; lacZ; quinone methide intermediates; single-cell resolution; β-galactosidase.

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