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Updated: Jun 17, 2025

Dual-Color Fluorescence Cross-Correlation Spectroscopy to Study Protein-Protein Interaction and Protein Dynamics in Live Cells

Dual-Color Fluorescence Cross-Correlation Spectroscopy to Study Protein-Protein Interaction and Protein Dynamics in Live Cells

Published on: December 11, 2021

Room-Temperature Solution Fluorescence Excitation Correlation Spectroscopy.

Yusuke Yoneda^1,2, Hikaru Kuramochi^1,2

¹Research Center of Integrative Molecular Systems (CIMoS), Institute for Molecular Science, National Institutes of Natural Sciences, 38 Nishigo-Naka, Myodaiji, Okazaki 444-8585, Japan.

The Journal of Physical Chemistry Letters

|August 12, 2024

Summary

This study introduces fluorescence excitation correlation spectroscopy to observe rapid molecular spectral changes in solutions at room temperature. This new method achieves microsecond resolution, enabling the study of dynamic molecular processes.

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High Precision FRET at Single-molecule Level for Biomolecule Structure Determination

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Area of Science:

Physical Chemistry
Spectroscopy
Single-Molecule Biophysics

Background:

Single-molecule fluorescence spectroscopy is vital for studying molecular properties.
Investigating fast dynamics of freely diffusing molecules in solution at room temperature is challenging.

Purpose of the Study:

To develop a method for studying fast fluctuation dynamics of excitation spectra in room-temperature solutions at the single-molecule level.
To enable the investigation of spontaneous spectral changes with microsecond time resolution.

Main Methods:

Utilized fluorescence excitation correlation spectroscopy.
Employed Fourier transform spectroscopy with broadband femtosecond pulses.
Applied time-correlated single-photon counting for single-molecule measurements.

Main Results:

Achieved single-molecule fluorescence excitation spectroscopy of room-temperature solutions.
Obtained excitation wavelength-resolved fluorescence autocorrelation functions in the microsecond to millisecond range.
Demonstrated the method's potential for studying time-dependent excitation spectra changes.

Conclusions:

The developed method allows for the study of fast, spontaneous excitation spectrum fluctuations in statistically equilibrated systems.
Further development can enable the study of spectral exchange with unprecedented time resolution.
This technique offers new insights into molecular dynamics in solution.