Multimodal two-photon imaging using a second harmonic generation-specific dye

Mutsuo Nuriya, Shun Fukushima, Atsuya Momotake, Takanori Shinotsuka, Masato Yasui, Tatsuo Arai

Research output: Contribution to journalArticlepeer-review

66 Citations (Scopus)


Second harmonic generation (SHG) imaging can be used to visualize unique biological phenomena, but currently available dyes limit its application owing to the strong fluorescent signals that they generate together with SHG. Here we report the first non-fluorescent and membrane potential-sensitive SHG-active organic dye Ap3. Ap3 is photostable and generates SH signals at the plasma membrane with virtually no fluorescent signals, in sharp contrast to the previously used fluorescent dye FM4-64. When tested in neurons, Ap3-SHG shows linear membrane potential sensitivity and fast responses to action potentials, and also shows significantly reduced photodamage compared with FM4-64. The SHG-specific nature of Ap3 allows simultaneous and completely independent imaging of SHG signals and fluorescent signals from various reporter molecules, including markers of cellular organelles and intracellular calcium. Therefore, this SHG-specific dye enables true multimodal two-photon imaging in biological samples.

Original languageEnglish
Article number11557
JournalNature communications
Publication statusPublished - 2016 May 9

ASJC Scopus subject areas

  • General Chemistry
  • General Biochemistry,Genetics and Molecular Biology
  • General Physics and Astronomy


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