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

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

Antolović, I.M. (TU Delft (OLD)Applied Quantum Architectures)
Burri, S. (Swiss Federal Institute of Technology)
Bruschini, Claudio (Swiss Federal Institute of Technology)
Hoebe, Ron A. (Universiteit van Amsterdam)
Charbon-Iwasaki-Charbon, E. (TU Delft (OLD)Applied Quantum Architectures; TU Delft OLD QCD/Charbon Lab; Swiss Federal Institute of Technology)

2017-03-13

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.

Electrical and electronic engineering
Imaging and sensing
Super-resolution microscopy

http://resolver.tudelft.nl/uuid:0df3296c-9a60-4243-b677-30507c5b38ea

https://doi.org/10.1038/srep44108

2045-2322

Scientific Reports, 7, 1-11

Institutional Repository

journal article

© 2017 I.M. Antolović, S. Burri, Claudio Bruschini, Ron A. Hoebe, E. Charbon-Iwasaki-Charbon