NIR fluorescence lifetime macroscopic imaging with a time-gated SPAD camera

X. Michalet; A. Ulku; J. T. Smith; C. Bruschini; S. Weiss; E. Charbon; X. Intes

doi:10.1117/12.2607833

3 March 2022 NIR fluorescence lifetime macroscopic imaging with a time-gated SPAD camera

X. Michalet, A. Ulku, J. T. Smith, C. Bruschini, S. Weiss, E. Charbon, X. Intes

Proceedings Volume 11965, Multiphoton Microscopy in the Biomedical Sciences XXII; 1196507 (2022) https://doi.org/10.1117/12.2607833
Event: SPIE BiOS, 2022, San Francisco, California, United States

Abstract

The performance of SwissSPAD2 (SS2), a large scale, widefield time-gated CMOS SPAD imager developed for fluorescence lifetime imaging, has recently been described in the context of visible range and fluorescence lifetime imaging microscopy (FLIM) of dyes with lifetimes in the 2.5 – 4 ns range. Here, we explore its capabilities in the NIR regime relevant for small animal imaging, where its sensitivity is lower and typical NIR fluorescent dye lifetimes are much shorter (1 ns or less). We carry out this study in a simple macroscopic imaging setup based on a compact NIR picosecond pulsed laser, an engineered diffuser-based illumination optics, and NIR optimized imaging lens suitable for well-plate or small animal imaging. Because laser repetition rates can vary between models, but the synchronization signal frequency accepted by SS2 is fixed to 20 MHz, we first checked that a simple frequency-division scheme enables data recording for different laser repetition rates. Next, we acquired data using different time gate widths, including gates with duration longer than the laser period, and analyzed the resulting data using both standard nonlinear least-square fit (NLSF) and phasor analysis. We show that the fixed synchronization rate and large gate widths characterizing SS2 (10 ns and over) are not an obstacle to accurately extracting lifetime in the 1 ns range and to distinguishing between close lifetimes. In summary, SS2 and similar very large gated SPAD imagers appear as a versatile alternative to other widefield time-resolved detectors for NIR fluorescence lifetime imaging, including preclinical molecular applications.

Conference Presentation

Citation Download Citation

X. Michalet, A. Ulku, J. T. Smith, C. Bruschini, S. Weiss, E. Charbon, and X. Intes "NIR fluorescence lifetime macroscopic imaging with a time-gated SPAD camera", Proc. SPIE 11965, Multiphoton Microscopy in the Biomedical Sciences XXII, 1196507 (3 March 2022); https://doi.org/10.1117/12.2607833

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