Quantifying Lifetime Uncertainty in Gated-ICCD Fluorescence Lifetime Imaging
Year: 2024
Type: preprint
Abstract: Fluorescence lifetime imaging (FLI) is a versatile technique for quantifying molecular processes in biomedical applications. Wide-field FLI analyzes fluorescence temporal decays computationally, often using intensified charge-coupled device (ICCD) cameras for both microscopic (FLIM) and macroscopic (MFLI) imaging. While uncertainty in lifetime parameter estimation has been extensively studied for photon-counting detectors, ICCDs pose unique challenges due to their nonlinear noise characteristics. Factors such as exposure time, mi- crochannel plate (MCP) gain, and illumination power significantly influence recorded fluorescence intensities. These variations complicate accurate photon count es- timation and, consequently, the determination of un- certainty in decay parameters. To address these chal- lenges, we propose a heuristic method for estimating un- certainties in ICCD-based fluorescence lifetimes. This method directly correlates uncertainty with recorded intensity and camera settings. It replaces the conven- tional shot-noise-limited uncertainty model used for photon-counting detectors with a modified expression, supported by a lookup table (LUT) that accounts for spe- cific experimental setups and camera configurations. We validated this approach through extensive studies with phantoms and demonstrated its in vivo application by imaging HER2+ breast tumor xenografts in nude mice. The imaging utilized Trastuzumab labeled with Alexa Fluor 700 and IRDye 800 2DG, highlighting the method’s reliability and effectiveness in preclinical biomedical imaging scenarios.
Language: English
Subfield: Biophysics
Domain: Life Sciences
Open Access status: green
OpenAlex ID: W4405826535