Resolving single-molecule assembled patterns with superresolution blink-microscopy.
In this paper we experimentally combine a recently developed AFM-based molecule-by-molecule assembly (single-molecule cut-and-paste, SMCP) with subdiffraction resolution fluorescence imaging. Using "Blink-Microscopy", which exploits the fluctuating emission of single molecules for the reconstruction of superresolution images, we resolved SMCP assembled structures with features below the diffraction limit. Artificial line patterns then served as calibration structures to characterize parameters, such as the labeling density, that can influence resolution of Blink-Microscopy besides the localization precision of a single molecule. Finally, we experimentally utilized the adjustability of blink parameters to demonstrate the general connection of photophysical parameters with spatial resolution and acquisition time in superresolution microscopy.
Cordes
Thorben
T
Applied Physics-Biophysics and Center for NanoScience, Ludwig-Maximilians-Universität, Amalienstrasse 54, D-80799 München, Germany.
Strackharn
Mathias
M
Stahl
Stefan W
SW
Summerer
Wolfram
W
Steinhauer
Christian
C
Forthmann
Carsten
C
Puchner
Elias M
EM
Vogelsang
Jan
J
Gaub
Hermann E
HE
Tinnefeld
Philip
P
D000465:Algorithms; D002138:Calibration; D004247:DNA / Q000737:chemistry; D005456:Fluorescent Dyes / Q000494:pharmacology; D007091:Image Processing, Computer-Assisted / Q000379:methods; D018625:Microscopy, Atomic Force / Q000379:methods; D008856:Microscopy, Fluorescence / Q000379:methods; D015394:Molecular Structure; D053761:Nanocomposites / Q000737:chemistry*; D049329:Nanostructures / Q000737:chemistry; D036103:Nanotechnology / Q000379:methods*
D005456:Fluorescent Dyes; D004247:DNA