Quantitative multi-parameter analysis of single molecule dynamics by PIE FastFLIM microscopy

Yuansheng Sun; Ulas Coskun; Phoebe S. Tsoi; Josephine C. Ferreon; Allan Chris Ferreon; Beniamino Barbieri; Shih-Chu Jeff Liao

doi:10.1117/12.2255961

21 February 2017 Quantitative multi-parameter analysis of single molecule dynamics by PIE FastFLIM microscopy

Yuansheng Sun, Ulas Coskun, Phoebe S. Tsoi, Josephine C. Ferreon, Allan Chris Ferreon, Beniamino Barbieri, Shih-Chu Jeff Liao

Author Affiliations +

Proceedings Volume 10069, Multiphoton Microscopy in the Biomedical Sciences XVII; 100691X (2017) https://doi.org/10.1117/12.2255961
Event: SPIE BiOS, 2017, San Francisco, California, United States

Abstract

PIE FastFLIM microscopy allows the quantitative multi-parameter measurement of single molecule protein folding and dynamics. Using donor-acceptor FRET pair-labeled proteins, we detect changes in protein conformation and dynamics by monitoring FRET efficiency, stoichiometry and lifetime. Together with anisotropy decay information, we acquire rotational relaxation times for single molecules. By applying antibunching, FLCS and burst analysis, multi-parameters (such as copy numbers in protein complexes), diffusion coefficient and molecular brightness can be fitted for deeper understanding of the conformational dynamic behavior of single protein molecules. In this paper, we’ll focus on the multiparameters of FRET efficiency, stoichiometry and lifetime.

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Yuansheng Sun, Ulas Coskun, Phoebe S. Tsoi, Josephine C. Ferreon, Allan Chris Ferreon, Beniamino Barbieri, and Shih-Chu Jeff Liao "Quantitative multi-parameter analysis of single molecule dynamics by PIE FastFLIM microscopy", Proc. SPIE 10069, Multiphoton Microscopy in the Biomedical Sciences XVII, 100691X (21 February 2017); https://doi.org/10.1117/12.2255961

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