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  4. Single-Photon, Time-Gated, Phasor-Based Fluorescence Lifetime Imaging through Highly Scattering Medium
 
research article

Single-Photon, Time-Gated, Phasor-Based Fluorescence Lifetime Imaging through Highly Scattering Medium

Ankri, Rinat
•
Basu, Arkaprabha
•
Ulku, Arin Can  
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January 1, 2020
Acs Photonics

Fluorescence lifetime imaging (FLI) is increasingly recognized as a powerful tool for biochemical and cellular investigations, including in vivo applications. Fluorescence lifetime is an intrinsic characteristic of any fluorescent dye which, to a large extent, does not depend on excitation intensity and signal level. In particular, it allows distinguishing dyes with similar emission spectra, offering additional multiplexing capabilities. However, in vivo FLI in the visible range is complicated by the contamination by (i) tissue autofluorescence, which decreases contrast, and by (ii) light scattering and absorption in tissues, which significantly reduce fluorescence intensity and modify the temporal profile of the signal. Here, we demonstrate how these issues can be accounted for and overcome, using a new time-gated single-photon avalanche diode array camera, SwissSPAD2, combined with phasor analysis to provide a simple and fast visual method for lifetime imaging. In particular, we show how phasor dispersion increases with increasing scattering and/or decreasing fluorescence intensity. Next, we show that as long as the fluorescence signal of interest is larger than the phantom autofluorescence, the presence of a distinct lifetime can be clearly identified with appropriate background correction. We use these results to demonstrate the detection of A459 cells expressing the fluorescent protein mCyRFP1 through highly scattering and autofluorescent phantom layers. These results showcase the possibility to perform FLI in challenging conditions, using standard, bright, visible fluorophore or fluorescence proteins.

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Type
research article
DOI
10.1021/acsphotonics.9b00874
Web of Science ID

WOS:000508475800008

Author(s)
Ankri, Rinat
Basu, Arkaprabha
Ulku, Arin Can  
Bruschini, Claudio  
Charbon, Edoardo  
Weiss, Shimon
Michalet, Xavier
Date Issued

2020-01-01

Publisher

AMER CHEMICAL SOC

Published in
Acs Photonics
Volume

7

Issue

1

Start page

68

End page

79

Subjects

Nanoscience & Nanotechnology

•

Materials Science, Multidisciplinary

•

Optics

•

Physics, Applied

•

Physics, Condensed Matter

•

Science & Technology - Other Topics

•

Materials Science

•

Physics

•

fluorescence lifetime imaging

•

phasor lifetime analysis

•

time-gated camera

•

singlephoton detection

•

scattering medium

•

optical-properties

•

intracellular delivery

•

proteins

•

tomography

•

microscopy

•

performance

•

tissues

•

cells

Editorial or Peer reviewed

REVIEWED

Written at

EPFL

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Available on Infoscience
March 3, 2020
Use this identifier to reference this record
https://infoscience.epfl.ch/handle/20.500.14299/166731
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