Rotational mobility of single molecules affects localization accuracy in super-resolution fluorescence microscopy.

The asymmetric nature of single-molecule (SM) dipole emission patterns limits the accuracy of position determination in localization-based super-resolution fluorescence microscopy. The degree of mislocalization depends highly on the rotational mobility of SMs; only for SMs rotating within a cone half angle α > 60° can mislocalization errors be bounded to ≤10 nm. Simulations demonstrate how low or high rotational mobility can cause resolution degradation or distortion in super-resolution reconstructions.

[1]  W. Vaughan,et al.  Dielectric relaxation by restricted rotational diffusion , 1978 .

[2]  Yale E. Goldman,et al.  Rotational motions of macro-molecules by single-molecule fluorescence microscopy. , 2005 .

[3]  N. Maiti,et al.  Fluorescence Dynamics of Dye Probes in Micelles , 1997 .

[4]  Shu Jia,et al.  Ultra-bright Photoactivatable Fluorophores Created by Reductive Caging , 2012, Nature Methods.

[5]  Matthew D Lew,et al.  Simultaneous, accurate measurement of the 3D position and orientation of single molecules , 2012, Proceedings of the National Academy of Sciences.

[6]  S. Achilefu,et al.  Fluorescence lifetime measurements and biological imaging. , 2010, Chemical reviews.

[7]  Y. Goldman,et al.  Measurement of single macromolecule orientation by total internal reflection fluorescence polarization microscopy. , 2005, Biophysical journal.

[8]  K. Kinosita,et al.  A theory of fluorescence polarization decay in membranes. , 1977, Biophysical journal.

[9]  T. Laurence,et al.  Polarization Spectroscopy of Single Fluorescent Molecules , 1999 .

[10]  W. Webb,et al.  Fluorescence correlation spectroscopy: diagnostics for sparse molecules. , 1997, Proceedings of the National Academy of Sciences of the United States of America.

[11]  W. E. Moerner,et al.  Microscopy beyond the diffraction limit using actively controlled single molecules , 2012, Journal of microscopy.

[12]  Mark Bates,et al.  Evaluation of fluorophores for optimal performance in localization-based super-resolution imaging , 2011, Nature Methods.

[13]  V. Subramaniam,et al.  One- and two-photon excited fluorescence lifetimes and anisotropy decays of green fluorescent proteins. , 2000, Biophysical journal.

[14]  A. Szabó,et al.  Effect of librational motion on fluorescence depolarization and nuclear magnetic resonance relaxation in macromolecules and membranes. , 1980, Biophysical journal.

[15]  Paul R Selvin,et al.  Polarization effect on position accuracy of fluorophore localization. , 2006, Optics express.

[16]  Robert M. Dickson,et al.  Imaging Three-Dimensional Single Molecule Orientations , 1999 .

[17]  Thorsten Staudt,et al.  Molecular orientation affects localization accuracy in superresolution far-field fluorescence microscopy. , 2011, Nano letters.

[18]  S. Brasselet,et al.  Quantitative imaging of molecular order in lipid membranes using two-photon fluorescence polarimetry , 2009, CLEO/QELS: 2010 Laser Science to Photonic Applications.

[19]  S. Stallinga,et al.  Accuracy of the gaussian point spread function model in 2D localization microscopy. , 2010, Optics express.

[20]  A. Verkman,et al.  Photobleaching recovery and anisotropy decay of green fluorescent protein GFP-S65T in solution and cells: cytoplasmic viscosity probed by green fluorescent protein translational and rotational diffusion. , 1997, Biophysical journal.

[21]  Michael J Rust,et al.  Sub-diffraction-limit imaging by stochastic optical reconstruction microscopy (STORM) , 2006, Nature Methods.

[22]  Rahul Roy,et al.  A practical guide to single-molecule FRET , 2008, Nature Methods.

[23]  Michael D. Mason,et al.  Ultra-high resolution imaging by fluorescence photoactivation localization microscopy. , 2006, Biophysical journal.

[24]  J. Lippincott-Schwartz,et al.  Imaging Intracellular Fluorescent Proteins at Nanometer Resolution , 2006, Science.

[25]  Shimon Weiss,et al.  TEMPORAL FLUCTUATIONS OF FLUORESCENCE RESONANCE ENERGY TRANSFER BETWEEN TWO DYES CONJUGATED TO A SINGLE PROTEIN , 1999 .

[26]  S. Hell.  Microscopy and its focal switch , 2008, Nature Methods.