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<p>Mechanical alterations of protein condensates are increasingly recognized in the etiology of several neurodegenerative diseases, yet their characterization remains technically challenging. Although Brillouin microscopy could offer a promising solution, its use is hindered by instrumental instabilities demanding frequent adjustments and manual calibrations with reference materials. Here, we present an enhanced Brillouin Microscope that incorporates an electro-optic modulator, serving simultaneously as frequency reference, spectrometer calibrator, and temporal stabilizer. This integration enables robust, real-time spectral stability over multiple days in a fully automated workflow. Using this system, we quantify Brillouin shifts of several protein condensates in living cells and validate our findings with FRAP. The correlation between techniques reveals a fractal internal architecture of the condensates, providing important insights into their physical nature while probing the mechanical behavior of entire compartments containing multiple protein species. Our method offers a unique framework for distinguishing physiological from pathological condensates, paving the way for long-term, user-independent, high-precision mechanical measurements in living cells.</p>

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Stabilized real-time Brillouin microscopy reveals fractal organization of protein condensates in living cells

  • 期刊论文

  • 出版模式：

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  • 发表日期：

    2026年2月5日

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  • Claudia Testi,
  • Emanuele Pontecorvo,
  • Chiara Bartoli,
  • Chiara Marzaro,
  • Fabrizio Gala,
  • Li Zhang,
  • Giulia Zanini,
  • Noemi D’Abbondanza,
  • Maria Giovanna Garone,
  • Valeria de Turris,
  • Andrea Giuliani,
  • Gaia Di Timoteo,
  • Irene Bozzoni,
  • Alessandro Rosa,
  • Giancarlo Ruocco

摘要

Mechanical alterations of protein condensates are increasingly recognized in the etiology of several neurodegenerative diseases, yet their characterization remains technically challenging. Although Brillouin microscopy could offer a promising solution, its use is hindered by instrumental instabilities demanding frequent adjustments and manual calibrations with reference materials. Here, we present an enhanced Brillouin Microscope that incorporates an electro-optic modulator, serving simultaneously as frequency reference, spectrometer calibrator, and temporal stabilizer. This integration enables robust, real-time spectral stability over multiple days in a fully automated workflow. Using this system, we quantify Brillouin shifts of several protein condensates in living cells and validate our findings with FRAP. The correlation between techniques reveals a fractal internal architecture of the condensates, providing important insights into their physical nature while probing the mechanical behavior of entire compartments containing multiple protein species. Our method offers a unique framework for distinguishing physiological from pathological condensates, paving the way for long-term, user-independent, high-precision mechanical measurements in living cells.