SPAD imagers for super resolution localization microscopy enable analysis of fast fluorophore blinking

Journal article (2017)

Authors

I.M. Antolović (OLD)Applied Quantum Architectures
S. Burri Swiss Federal Institute of Technology
Claudio Bruschini Swiss Federal Institute of Technology
Ron A. Hoebe Universiteit van Amsterdam
E. Charbon-Iwasaki-Charbon OLD QCD/Charbon Lab , (OLD)Applied Quantum Architectures , Swiss Federal Institute of Technology
Research Group
(OLD)Applied Quantum Architectures
Copyright: © 2017 I.M. Antolović, S. Burri, Claudio Bruschini, Ron A. Hoebe, E. Charbon-Iwasaki-Charbon
DOI: https://doi.org/10.1038/srep44108
More Info
expand_more

Published Date

13-03-2017

Language

English

Research Group

(OLD)Applied Quantum Architectures

Abstract

sCMOS imagers are currently utilized (replacing EMCCD imagers) to increase the acquisition speed in super resolution localization microscopy. Single-photon avalanche diode (SPAD) imagers feature frame rates per bit depth comparable to or higher than sCMOS imagers, while generating microsecond 1-bit-frames without readout noise, thus paving the way to in-depth time-resolved image analysis. High timing resolution can also be exploited to explore fluorescent dye blinking and other photophysical properties, which can be used for dye optimization. We present the methodology for the blinking analysis of fluorescent dyes on experimental data. Furthermore, the recent use of microlenses has enabled a substantial increase of SPAD imager overall sensitivity (12-fold in our case), reaching satisfactory values for sensitivity-critical applications. This has allowed us to record the first super resolution localization microscopy results obtained with a SPAD imager, with a localization uncertainty of 20 nm and a resolution of 80 nm.