Picosecond wide-field time-correlated single photon counting fluorescence microscopy with a delay line anode detector

Liisa Maija Hirvonen; Wolfgang Becker; James Milnes; Thomas Conneely; Stefan Smietana; Alix Marie Le Marois; Ottmar Jagutzki; Klaus Suhling

doi:10.1063/1.4961054

Picosecond wide-field time-correlated single photon counting fluorescence microscopy with a delay line anode detector

Liisa Maija Hirvonen, Wolfgang Becker, James Milnes, Thomas Conneely, Stefan Smietana, Alix Marie Le Marois, Ottmar Jagutzki, Klaus Suhling

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Abstract

We perform wide-field time-correlated single photon counting-based fluorescence lifetime imaging (FLIM) with a crossed delay line anode image intensifier, where the pulse propagation time yields the photon position. This microchannel plate-based detector was read out with conventional fast timing electronics and mounted on a fluorescence microscope with total internal reflection (TIR) illumination. The picosecond time resolution of this detection system combines low illumination intensity of microwatts with wide-field data collection. This is ideal for fluorescence lifetime imaging of cell membranes using TIR. We show that fluorescence lifetime images of living HeLa cells stained with membrane dye di-4-ANEPPDHQ exhibit a reduced lifetime near the coverslip in TIR compared to epifluorescence FLIM.

Original language	English
Article number	071101
Number of pages	4
Journal	APPLIED PHYSICS LETTERS
Volume	109
Issue number	7
DOIs	https://doi.org/10.1063/1.4961054
Publication status	Published - 15 Aug 2016

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Picosecond wide-field time_HIRVONEN_Accepted 3Aug2016_GOLD VoR
© 2016 Author(s). All article content, except where otherwise noted, is licensed under a Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).
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title = "Picosecond wide-field time-correlated single photon counting fluorescence microscopy with a delay line anode detector",

abstract = "We perform wide-field time-correlated single photon counting-based fluorescence lifetime imaging (FLIM) with a crossed delay line anode image intensifier, where the pulse propagation time yields the photon position. This microchannel plate-based detector was read out with conventional fast timing electronics and mounted on a fluorescence microscope with total internal reflection (TIR) illumination. The picosecond time resolution of this detection system combines low illumination intensity of microwatts with wide-field data collection. This is ideal for fluorescence lifetime imaging of cell membranes using TIR. We show that fluorescence lifetime images of living HeLa cells stained with membrane dye di-4-ANEPPDHQ exhibit a reduced lifetime near the coverslip in TIR compared to epifluorescence FLIM.",

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AU - Hirvonen, Liisa Maija

AU - Becker, Wolfgang

AU - Milnes, James

AU - Conneely, Thomas

AU - Smietana, Stefan

AU - Le Marois, Alix Marie

AU - Jagutzki, Ottmar

AU - Suhling, Klaus

PY - 2016/8/15

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AB - We perform wide-field time-correlated single photon counting-based fluorescence lifetime imaging (FLIM) with a crossed delay line anode image intensifier, where the pulse propagation time yields the photon position. This microchannel plate-based detector was read out with conventional fast timing electronics and mounted on a fluorescence microscope with total internal reflection (TIR) illumination. The picosecond time resolution of this detection system combines low illumination intensity of microwatts with wide-field data collection. This is ideal for fluorescence lifetime imaging of cell membranes using TIR. We show that fluorescence lifetime images of living HeLa cells stained with membrane dye di-4-ANEPPDHQ exhibit a reduced lifetime near the coverslip in TIR compared to epifluorescence FLIM.

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DO - 10.1063/1.4961054

M3 - Article

SN - 0003-6951

VL - 109

JO - APPLIED PHYSICS LETTERS

JF - APPLIED PHYSICS LETTERS

IS - 7

M1 - 071101

ER -

Picosecond wide-field time-correlated single photon counting fluorescence microscopy with a delay line anode detector

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