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Updated: Feb 18, 2026

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Imaging Ca2+ with a Fluorescent Rhodol.

Alisha A Contractor1, Evan W Miller1

  • 1Department of Chemistry, ‡Department of Molecular and Cell Biology, and §Helen Wills Neuroscience Institute, University of California , Berkeley, California 94720, United States.

Biochemistry
|November 29, 2017
PubMed
Summary

Researchers developed a new calcium (Ca2+) sensor, rhodol Ca2+ sensor 1 (RCS-1), for improved cellular imaging. This novel fluorophore offers enhanced two-photon absorption, enabling better visualization of Ca2+ dynamics in neurons.

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Area of Science:

  • Biochemistry and Biophysics
  • Cellular Signaling
  • Fluorescent Probes and Imaging

Background:

  • Calcium ions (Ca2+) are critical mediators of cellular signaling pathways.
  • Synthetic Ca2+-sensitive fluorophores are essential tools for studying Ca2+ dynamics.
  • Rhodol-based fluorophores represent an underexplored class for Ca2+ sensing applications.

Purpose of the Study:

  • To design, synthesize, and evaluate a novel rhodol-based Ca2+ sensor.
  • To assess the sensor's performance, including Ca2+ affinity and photophysical properties.
  • To demonstrate the sensor's utility in live-cell imaging, particularly in neurons.

Main Methods:

  • Chemical synthesis of a chlorinated pyrrolidine-based rhodol derivative (RCS-1).
  • Characterization of Ca2+ binding affinity (Kd = 240 nM) and dynamic range (10-fold response).

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  • Measurement of two-photon absorption cross-section (σTPA = 76 GM at 840 nm).
  • Live-cell imaging of cytosolic Ca2+ fluctuations in cultured neurons using one- and two-photon microscopy.

    • Main Results:

    • RCS-1 exhibits a high Ca2+ binding affinity suitable for physiological Ca2+ levels.
    • RCS-1 demonstrates a significant 10-fold fluorescence turn-on response to Ca2+.
    • RCS-1 possesses a superior two-photon absorption cross-section compared to fluorescein-based sensors.
    • The acetoxy-methyl ester form of RCS-1 effectively stains live cell cytosol for imaging.

    Conclusions:

    • Rhodol-based scaffolds are highly promising for developing advanced Ca2+ sensors.
    • RCS-1 provides a valuable new tool for studying Ca2+ dynamics with enhanced two-photon imaging capabilities.
    • This study highlights the potential of rhodol fluorophores for neurobiological research involving Ca2+ signaling.