Micro-Raman spectroscopic analysis of liquid-liquid phase separation

Suin Choi, So Yeon Chun, Kyungwon Kwak, Minhaeng Cho

Research output: Contribution to journalArticlepeer-review

3 Citations (Scopus)


Liquid-liquid phase separation (LLPS) plays a significant role in various biological processes, including the formation of membraneless organelles and pathological protein aggregation. Although many studies have found various factors that modulate the LLPS process or the liquid-to-solid phase transition (LSPT) using microscopy or fluorescence-based methods, the molecular mechanistic details underlying LLPS and protein aggregation within liquid droplets remain uncharacterized. Therefore, structural information on proteins inside liquid droplets is required to understand the mechanistic link to amyloid formation. In the present study, we monitored droplet formation related to protein fibrillation using micro-Raman spectroscopy in combination with differential interference contrast (DIC) microscopy to study the conformational change in proteins and the hydrogen-bonding (H-bonding) structure of water during LLPS. Interestingly, we found that the O-D stretching band for water (HOD in H2O) inside the droplets exhibited a distinct Raman spectrum from that of the bulk water, suggesting that the time-dependent change in the hydration environment in the protein droplets during the process of LLPS can be studied. These results demonstrate that the superior spatial resolution of micro-Raman spectroscopy offers significant advantages in investigating the molecular mechanisms of LLPS and following LSPT processes.

Original languageEnglish
JournalPhysical Chemistry Chemical Physics
Publication statusAccepted/In press - 2023

Bibliographical note

Publisher Copyright:
© 2023 The Royal Society of Chemistry.

ASJC Scopus subject areas

  • General Physics and Astronomy
  • Physical and Theoretical Chemistry


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