Three-dimensional structural dynamics of myosin V by single-molecule fluorescence polarization

Nature ◽  
2003 ◽  
Vol 422 (6930) ◽  
pp. 399-404 ◽  
Author(s):  
Joseph N. Forkey ◽  
Margot E. Quinlan ◽  
M. Alexander Shaw ◽  
John E. T. Corrie ◽  
Yale E. Goldman
Keyword(s):  
Structural Dynamics ◽  
Single Molecule ◽  
Fluorescence Polarization ◽  
Three Dimensional ◽  
Myosin V ◽  
Single Molecule Fluorescence

scholarly journals Systematic Assessment of Burst Impurity in Confocal-Based Single-Molecule Fluorescence Detection Using Brownian Motion Simulations

Molecules ◽  
2019 ◽  
Vol 24 (14) ◽  
pp. 2557 ◽  
Author(s):  
Dolev Hagai ◽  
Eitan Lerner
Keyword(s):  
Brownian Motion ◽  
Fluorescence Detection ◽  
Single Molecule ◽  
Three Dimensional ◽  
Resonance Energy Transfer ◽  
Resonance Energy ◽  
Single Molecule Fluorescence ◽  
Experimental Conditions ◽  
Systematic Assessment ◽  
Burst Analysis

Single-molecule fluorescence detection (SMFD) experiments are useful in distinguishing sub-populations of molecular species when measuring heterogeneous samples. One experimental platform for SMFD is based on a confocal microscope, where molecules randomly traverse an effective detection volume. The non-uniformity of the excitation profile and the random nature of Brownian motion, produce fluctuating fluorescence signals. For these signals to be distinguished from the background, burst analysis is frequently used. Yet, the relation between the results of burst analyses and the underlying information of the diffusing molecules is still obscure and requires systematic assessment. In this work we performed three-dimensional Brownian motion simulations of SMFD, and tested the positions at which molecules emitted photons that passed the burst analysis criteria for different values of burst analysis parameters. The results of this work verify which of the burst analysis parameters and experimental conditions influence both the position of molecules in space when fluorescence is detected and taken into account, and whether these bursts of photons arise purely from single molecules, or not entirely. Finally, we show, as an example, the effect of bursts that are not purely from a single molecule on the accuracy in single-molecule Förster resonance energy transfer measurements.

scholarly journals Correction to Monitoring Nanoscale Deformations in a Drawn Polymer Melt with Single-Molecule Fluorescence Polarization Microscopy

ACS Nano ◽  
2016 ◽  
Vol 10 (3) ◽  
pp. 3873-3873
Author(s):  
Stefan Krause ◽  
Martin Neumann ◽  
Melanie Fröbe ◽  
Robert Magerle ◽  
Christian von Borczyskowski
Keyword(s):  
Single Molecule ◽  
Fluorescence Polarization ◽  
Polymer Melt ◽  
Polarization Microscopy ◽  
Single Molecule Fluorescence ◽  
Fluorescence Polarization Microscopy

scholarly journals Systematic Assessment of Freely-Diffusing Single-Molecule Fluorescence Detection using Brownian Motion Simulations

2019 ◽  
Author(s):  
Dolev Hagai ◽  
Eitan Lerner
Keyword(s):  
Brownian Motion ◽  
Fluorescence Detection ◽  
Single Molecule ◽  
Spatial Information ◽  
Three Dimensional ◽  
Single Molecule Fluorescence ◽  
Experimental Conditions ◽  
Systematic Assessment ◽  
Detection Volume ◽  
Burst Analysis

Single-molecule fluorescence detection (SMFD) experiments are useful in distinguishing between sub-populations of molecular species in measurements of heterogeneous samples. One of the experimental platforms for SMFD is based on a confocal microscope setup, where molecules in the solution randomly traverse an effective detection volume, formed by a tightly focused laser beam. The non-uniformity of the excitation profile and the random nature of Brownian motion, produce fluctuating fluorescence signals. For these signals to be distinguished from the background, single-molecule fluorescence burst analysis is frequently used. Yet, the relation between the results of burst analyses and the underlying spatial information of the diffusing molecules is still obscure and requires systematic assessment. In this work we performed three-dimensional Brownian motion simulations of SMFD, and tested the positions from which the molecules emitted photons that were detected and passed the burst analysis criteria for different values of burst analysis parameters. The results of this work verify which of the burst analysis parameters and experimental conditions influence both the position of molecules in space when fluorescence is detected and taken into account, and whether these bursts of photons arise purely from single molecules, or not entirely.

Monitoring Nanoscale Deformations in a Drawn Polymer Melt with Single-Molecule Fluorescence Polarization Microscopy

ACS Nano ◽  
2016 ◽  
Vol 10 (2) ◽  
pp. 1908-1917 ◽  
Author(s):  
Stefan Krause ◽  
Martin Neumann ◽  
Melanie Fröbe ◽  
Robert Magerle ◽  
Christian von Borczyskowski
Keyword(s):  
Single Molecule ◽  
Fluorescence Polarization ◽  
Polymer Melt ◽  
Polarization Microscopy ◽  
Single Molecule Fluorescence ◽  
Fluorescence Polarization Microscopy
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