Development of widefield time-resolved fluorescence anisotropy imaging (TR-FAIM) using a single photon avalanche diode (SPAD) array

Louis Obeid Mogridge; Jakub Nedbal; Shirwa Awale; Jess Tallis; Emily J. Johnston; Francesco Mattioli Della Rocca; Susan J. Rosser; Robert K. Henderson; Klaus Suhling

doi:10.1117/12.3001710

Development of widefield time-resolved fluorescence anisotropy imaging (TR-FAIM) using a single photon avalanche diode (SPAD) array

Louis Obeid Mogridge^*, Jakub Nedbal, Shirwa Awale, Jess Tallis, Emily J. Johnston, Francesco Mattioli Della Rocca, Susan J. Rosser, Robert K. Henderson, Klaus Suhling

^*Corresponding author for this work

Research output: Chapter in Book/Report/Conference proceeding › Conference contribution

Abstract

The time-resolved fluorescence anisotropy of a biological sample can reveal rotational dynamics and structure of a fluorescent probe’s local environment. Here, methods are presented towards developing widefield time-resolved fluorescence anisotropy imaging (TR-FAIM) using a single photon avalanche diode (SPAD) array (QuantICAM). Such detector allows for simultaneous time-correlated single photon counting (TCSPC) measurements in each pixel with single-photon sensitivity and picosecond time resolution. Our method integrates the SPAD array with a widefield microscope, and automated rotating polarizers in the excitation and emission pathways to demonstrate TR-FAIM. We have shown the robustness of the method through spectroscopic measurements of the fluorophore PM546, and we have demonstrated the usefulness of simultaneous FLIM and TR-FAIM for studying properties of plasma membranes in live yeast cells.

Original language	English
Title of host publication	Proceedings Volume 12847
Subtitle of host publication	Multiphoton Microscopy in the Biomedical Sciences XXIV
Editors	Ammasi Periasamy, Peter T. So
Place of Publication	Bellingham
Publisher	SPIE
Chapter	0J
Number of pages	19
Volume	12847
ISBN (Electronic)	9781510669543
ISBN (Print)	9781510669536
DOIs	https://doi.org/10.1117/12.3001710
Publication status	Published - 12 Mar 2024
Event	SPIE BiOS, 2024, San Francisco, California, United States - The Moscone Center, 747 Howard St, San Francisco, CA 94103, San Francisco, United States Duration: 28 Jan 2024 → 30 Jan 2024 https://spie.org/conferences-and-exhibitions/photonics-west/exhibitions/bios-expo?SSO=1

Conference

Conference	SPIE BiOS, 2024, San Francisco, California, United States
Country/Territory	United States
City	San Francisco
Period	28/01/24 → 30/01/24
Internet address	https://spie.org/conferences-and-exhibitions/photonics-west/exhibitions/bios-expo?SSO=1

Keywords / Materials (for Non-textual outputs)

Time-resolved fluorescence anisotropy imaging (TR-FAIM
single-photon avalanche diode (SPAD) array
time-correlated single photon counting (TCSPC)
, fluorescence lifetime imaging microscopy (FLIM)
live-cell imaging
microviscosity
intracellular diffusion

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10.1117/12.3001710

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Mogridge, L. O., Nedbal, J., Awale, S., Tallis, J., Johnston, E. J., Rocca, F. M. D., Rosser, S. J., Henderson, R. K., & Suhling, K. (2024). Development of widefield time-resolved fluorescence anisotropy imaging (TR-FAIM) using a single photon avalanche diode (SPAD) array. In A. Periasamy, & P. T. So (Eds.), Proceedings Volume 12847: Multiphoton Microscopy in the Biomedical Sciences XXIV (Vol. 12847). Article 128470J SPIE. https://doi.org/10.1117/12.3001710

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title = "Development of widefield time-resolved fluorescence anisotropy imaging (TR-FAIM) using a single photon avalanche diode (SPAD) array",

abstract = "The time-resolved fluorescence anisotropy of a biological sample can reveal rotational dynamics and structure of a fluorescent probe{\textquoteright}s local environment. Here, methods are presented towards developing widefield time-resolved fluorescence anisotropy imaging (TR-FAIM) using a single photon avalanche diode (SPAD) array (QuantICAM). Such detector allows for simultaneous time-correlated single photon counting (TCSPC) measurements in each pixel with single-photon sensitivity and picosecond time resolution. Our method integrates the SPAD array with a widefield microscope, and automated rotating polarizers in the excitation and emission pathways to demonstrate TR-FAIM. We have shown the robustness of the method through spectroscopic measurements of the fluorophore PM546, and we have demonstrated the usefulness of simultaneous FLIM and TR-FAIM for studying properties of plasma membranes in live yeast cells.",

keywords = "Time-resolved fluorescence anisotropy imaging (TR-FAIM, single-photon avalanche diode (SPAD) array, time-correlated single photon counting (TCSPC), , fluorescence lifetime imaging microscopy (FLIM), live-cell imaging, microviscosity, intracellular diffusion",

author = "Mogridge, {Louis Obeid} and Jakub Nedbal and Shirwa Awale and Jess Tallis and Johnston, {Emily J.} and Rocca, {Francesco Mattioli Della} and Rosser, {Susan J.} and Henderson, {Robert K.} and Klaus Suhling",

year = "2024",

month = mar,

day = "12",

doi = "10.1117/12.3001710",

language = "English",

isbn = "9781510669536",

volume = "12847",

editor = "Ammasi Periasamy and So, {Peter T.}",

booktitle = "Proceedings Volume 12847",

publisher = "SPIE",

address = "United States",

note = "SPIE BiOS, 2024, San Francisco, California, United States ; Conference date: 28-01-2024 Through 30-01-2024",

url = "https://spie.org/conferences-and-exhibitions/photonics-west/exhibitions/bios-expo?SSO=1",

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Mogridge, LO, Nedbal, J, Awale, S, Tallis, J, Johnston, EJ, Rocca, FMD, Rosser, SJ , Henderson, RK & Suhling, K 2024, Development of widefield time-resolved fluorescence anisotropy imaging (TR-FAIM) using a single photon avalanche diode (SPAD) array. in A Periasamy & PT So (eds), Proceedings Volume 12847: Multiphoton Microscopy in the Biomedical Sciences XXIV. vol. 12847, 128470J, SPIE, Bellingham, SPIE BiOS, 2024, San Francisco, California, United States, San Francisco, United States, 28/01/24. https://doi.org/10.1117/12.3001710

Development of widefield time-resolved fluorescence anisotropy imaging (TR-FAIM) using a single photon avalanche diode (SPAD) array. / Mogridge, Louis Obeid; Nedbal, Jakub; Awale, Shirwa et al.
Proceedings Volume 12847: Multiphoton Microscopy in the Biomedical Sciences XXIV. ed. / Ammasi Periasamy; Peter T. So. Vol. 12847 Bellingham: SPIE, 2024. 128470J.

Research output: Chapter in Book/Report/Conference proceeding › Conference contribution

TY - GEN

T1 - Development of widefield time-resolved fluorescence anisotropy imaging (TR-FAIM) using a single photon avalanche diode (SPAD) array

AU - Mogridge, Louis Obeid

AU - Nedbal, Jakub

AU - Awale, Shirwa

AU - Tallis, Jess

AU - Johnston, Emily J.

AU - Rocca, Francesco Mattioli Della

AU - Rosser, Susan J.

AU - Henderson, Robert K.

AU - Suhling, Klaus

PY - 2024/3/12

Y1 - 2024/3/12

N2 - The time-resolved fluorescence anisotropy of a biological sample can reveal rotational dynamics and structure of a fluorescent probe’s local environment. Here, methods are presented towards developing widefield time-resolved fluorescence anisotropy imaging (TR-FAIM) using a single photon avalanche diode (SPAD) array (QuantICAM). Such detector allows for simultaneous time-correlated single photon counting (TCSPC) measurements in each pixel with single-photon sensitivity and picosecond time resolution. Our method integrates the SPAD array with a widefield microscope, and automated rotating polarizers in the excitation and emission pathways to demonstrate TR-FAIM. We have shown the robustness of the method through spectroscopic measurements of the fluorophore PM546, and we have demonstrated the usefulness of simultaneous FLIM and TR-FAIM for studying properties of plasma membranes in live yeast cells.

AB - The time-resolved fluorescence anisotropy of a biological sample can reveal rotational dynamics and structure of a fluorescent probe’s local environment. Here, methods are presented towards developing widefield time-resolved fluorescence anisotropy imaging (TR-FAIM) using a single photon avalanche diode (SPAD) array (QuantICAM). Such detector allows for simultaneous time-correlated single photon counting (TCSPC) measurements in each pixel with single-photon sensitivity and picosecond time resolution. Our method integrates the SPAD array with a widefield microscope, and automated rotating polarizers in the excitation and emission pathways to demonstrate TR-FAIM. We have shown the robustness of the method through spectroscopic measurements of the fluorophore PM546, and we have demonstrated the usefulness of simultaneous FLIM and TR-FAIM for studying properties of plasma membranes in live yeast cells.

KW - Time-resolved fluorescence anisotropy imaging (TR-FAIM

KW - single-photon avalanche diode (SPAD) array

KW - time-correlated single photon counting (TCSPC)

KW - , fluorescence lifetime imaging microscopy (FLIM)

KW - live-cell imaging

KW - microviscosity

KW - intracellular diffusion

U2 - 10.1117/12.3001710

DO - 10.1117/12.3001710

M3 - Conference contribution

SN - 9781510669536

VL - 12847

BT - Proceedings Volume 12847

A2 - Periasamy, Ammasi

A2 - So, Peter T.

PB - SPIE

CY - Bellingham

T2 - SPIE BiOS, 2024, San Francisco, California, United States

Y2 - 28 January 2024 through 30 January 2024

ER -

Mogridge LO, Nedbal J, Awale S, Tallis J, Johnston EJ, Rocca FMD et al. Development of widefield time-resolved fluorescence anisotropy imaging (TR-FAIM) using a single photon avalanche diode (SPAD) array. In Periasamy A, So PT, editors, Proceedings Volume 12847: Multiphoton Microscopy in the Biomedical Sciences XXIV. Vol. 12847. Bellingham: SPIE. 2024. 128470J doi: 10.1117/12.3001710

Development of widefield time-resolved fluorescence anisotropy imaging (TR-FAIM) using a single photon avalanche diode (SPAD) array

Abstract

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Keywords / Materials (for Non-textual outputs)

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