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Updated: May 2, 2026

Time-Resolved Fluorescence Anisotropy from Single Molecules for Characterizing Local Flexibility in Biomolecules
Published on: April 25, 2025
Single-molecule FRET of protein structure and dynamics - a primer
Abstract:
Single-molecule spectroscopy has developed into a widely used method for probing the structure, dynamics, and mechanisms of biomolecular systems, especially in combination with Förster resonance energy transfer (FRET). In this introductory tutorial, essential concepts and methods will be outlined, from the FRET process and the basic considerations for sample preparation and instrumentation to some key elements of data analysis and photon statistics. Different approaches for obtaining dynamic information over a wide range of timescales will be explained and illustrated with examples, including the quantitative analysis of FRET efficiency histograms, correlation spectroscopy, fluorescence trajectories, and microfluidic mixing.
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