Fluorescence lifetime imaging of high-speed particles with single-photon image sensors

Istvan Gyongy; Andrew Green; Sam W. Hutchings; Amy Davies; Neale A. W. Dutton; Rory R. Duncan; Colin Rickman; Robert K. Henderson; Paul A. Dalgarno

doi:10.1117/12.2510773

4 March 2019 Fluorescence lifetime imaging of high-speed particles with single-photon image sensors

Istvan Gyongy, Andrew Green, Sam W. Hutchings, Amy Davies, Neale A. W. Dutton, Rory R. Duncan, Colin Rickman, Robert K. Henderson, Paul A. Dalgarno

Author Affiliations +

Proceedings Volume 10889, High-Speed Biomedical Imaging and Spectroscopy IV; 108890O (2019) https://doi.org/10.1117/12.2510773
Event: SPIE BiOS, 2019, San Francisco, California, United States

Abstract

The capability of Single-Photon Avalanche Diodes (SPADs) to detect photons with picosecond timing precision and shotnoise limited performance has given rise to a range of biological and biomedical applications, from Fluorescence Lifetime Imaging Microscopy (FLIM) to Raman Spectroscopy and Positron Emission Tomography (PET). The use of SPAD sensors has also been successfully demonstrated in Single-Molecule Localisation Microscopy. Traditionally implemented as point detectors, recent advances in SPAD technology, such as compact, binary pixels and back-side illuminated, 3D-stacked architectures, have led to 2-D imaging arrays of increasing resolution and fill factor. Combined with high frame rates (in the kFPS range), and negligible read noise, the sensors offer an exciting prospect for capturing fast temporal dynamics in life science cellular imaging. The work in this paper considers the application of SPAD imaging arrays to widefield fluorescence lifetime imaging of high-speed particles in microscopy. We demonstrate, using a time-gated binary SPAD array, that by tracking particles, and spatially re-assigning the underlying photon counts accordingly, lifetime estimates for fast-moving objects are possible. Moreover, we give the first demonstration of FLIM using a SPAD imaging array with on-chip histogramming of photon arrival time, with potential frame rates of several 100FPS. Both FLIM techniques are illustrated using experimental results based on fluorescent microspheres undergoing Brownian motion. The results pave the way towards applications in live-cell microscopy, such as the monitoring of the fluorescence lifetime of highly mobile cell structures, with a view, for example, to study molecular interactions using Förster Resonance Energy Transfer (FRET) measurements.

Conference Presentation

Citation Download Citation

Istvan Gyongy, Andrew Green, Sam W. Hutchings, Amy Davies, Neale A. W. Dutton, Rory R. Duncan, Colin Rickman, Robert K. Henderson, and Paul A. Dalgarno "Fluorescence lifetime imaging of high-speed particles with single-photon image sensors", Proc. SPIE 10889, High-Speed Biomedical Imaging and Spectroscopy IV, 108890O (4 March 2019); https://doi.org/10.1117/12.2510773

ACCESS THE FULL ARTICLE

INSTITUTIONAL
Select your institution to access the SPIE Digital Library.

SELECT YOUR INSTITUTION

PERSONAL
Sign in with your SPIE account to access your personal subscriptions or to use specific features such as save to my library, sign up for alerts, save searches, etc.

PERSONAL SIGN IN

No SPIE Account? Create one

PURCHASE THIS CONTENT

SUBSCRIBE TO DIGITAL LIBRARY

50 downloads per 1-year subscription

Members: $195

Non-members: $335 ADD TO CART

25 downloads per 1 - year subscription

Members: $145

Non-members: $250 ADD TO CART

PURCHASE SINGLE ARTICLE

Includes PDF, HTML & Video, when available