Principles of fluorescence correlation spectroscopy applied to studies of biomolecular liquid–liquid phase separation

Zhulou Wang; Huizhi Zhang; Lin Jian; Bo Ding; Keying Huang; Wolun Zhang; Qian Xiao; Shaohui Huang

doi:10.52601/bpr.2022.210047

Volume 8 Issue 2

Apr. 2022

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Zhulou Wang, Huizhi Zhang, Lin Jian, Bo Ding, Keying Huang, Wolun Zhang, Qian Xiao, Shaohui Huang. Principles of fluorescence correlation spectroscopy applied to studies of biomolecular liquid–liquid phase separation[J]. Biophysics Reports, 2022, 8(2): 100-118. doi: 10.52601/bpr.2022.210047

Citation:

Zhulou Wang, Huizhi Zhang, Lin Jian, Bo Ding, Keying Huang, Wolun Zhang, Qian Xiao, Shaohui Huang. Principles of fluorescence correlation spectroscopy applied to studies of biomolecular liquid–liquid phase separation[J]. Biophysics Reports, 2022, 8(2): 100-118. doi: 10.52601/bpr.2022.210047

Citation:

Zhulou Wang, Huizhi Zhang, Lin Jian, Bo Ding, Keying Huang, Wolun Zhang, Qian Xiao, Shaohui Huang. Principles of fluorescence correlation spectroscopy applied to studies of biomolecular liquid–liquid phase separation[J]. Biophysics Reports, 2022, 8(2): 100-118. doi: 10.52601/bpr.2022.210047

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Principles of fluorescence correlation spectroscopy applied to studies of biomolecular liquid–liquid phase separation

doi: 10.52601/bpr.2022.210047

1.
Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China
2.
School of Life Sciences, University of Chinese Academy of Sciences, Beijing 101499, China
3.
LightEdge Technologies Limited, Zhongshan, Guangdong 528403, China

Funds: We thank research funding from National Natural Science Foundation of China (21874147).

More Information

Corresponding author: shaohuih@ibp.ac.cn (S. Huang)
Received Date: 28 December 2021
Accepted Date: 10 February 2022

Available Online: 25 March 2022

Publish Date: 22 April 2022

Abstract

Abstract

Fluorescence correlation spectroscopy (FCS) investigates the temporal relationship of fluctuating fluorescence signals reflecting underlying molecular processes occurring in a solution sample or a single live cell. This review article introduces the principles of two basic and most used FCS techniques: fluorescence auto-correlation spectroscopy (FACS) and fluorescence cross-correlation spectroscopy (FCCS). Combined, FACS and FCCS techniques can quantitatively analyze multiple properties of molecule or nanoparticle samples, including molar concentration, diffusion coefficient and hydrodynamic radius, homo- or hetero-interaction, fluorescence brightness, etc. Not surprisingly, FCS techniques have long been used to investigate molecular mechanisms of biomolecular phase separation, first in the lipid bilayer and more recently in cell cytosol and nucleoplasm. The latter applications are especially exciting since a whole new class of membraneless cellular organelles have been discovered, which are proposed to be results of biomolecule liquid-liquid phase separation (LLPS). LLPS research can benefit significantly from the multifunctionality and single-molecule sensitivity of a variety of FCS techniques, particularly for live-cell studies. This review illustrates how FACS and FCCS techniques can be used to investigate multiple aspects of the molecular mechanisms of LLPS, and summarizes FCS applications to LLPS research in vivo and in vitro.
- Liquid–liquid phase separation,
- Fluorescence correlation spectroscopy,
- Diffusion coefficient,
- Hydrodynamic radius,
- Binding affinity

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References(76)

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