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FRET measurements can provide dynamic spatial information on length scales smaller than the diffraction limit of light. Several methods exist to measure FRET between fluorophores, including Fluorescence Lifetime Imaging Microscopy (FLIM),…

Quantitative Methods · Quantitative Biology 2017-01-09 Bryan Kaye , Peter J. Foster , Tae Yeon Yoo , Daniel J. Needleman

FRET-based approaches are a unique tool for sensing the immediate surroundings and interactions of (bio)molecules. FRET imaging and FLIM (Fluorescence Lifetime Imaging Microscopy) enable the visualization of the spatial distribution of…

We report the development and detailed calibration of a multiphoton fluorescence lifetime imaging system (FLIM) using a streak camera. The present system is versatile with high spatial (0.2 micron) and temporal (50 psec) resolution and…

Quantitative Methods · Quantitative Biology 2009-11-10 R. V. Krishnan , H. Saitoh , H. Terada , V. E. Centonze , B. Herman

Photoluminesce (PL) spectroscopy offers excellent methods for mapping the PL decay on the nanosecond time scale. However, capturing maps of emission dynamics on the microsecond time scale can be highly time-consuming. We present a new…

Optics · Physics 2021-02-24 Jiri Junek , Karel Zidek

We present a fast and accurate analytical method for fluorescence lifetime imaging microscopy (FLIM) using the extreme learning machine (ELM). We used extensive metrics to evaluate ELM and existing algorithms. First, we compared these…

Biological Physics · Physics 2022-03-28 Zhenya Zang , Dong Xiao , Quan Wang , Zinuo Li , Wujun Xie , Yu Chen , David Day Uei Li

Fluorescence lifetime imaging microscopy (FLIM) is a powerful technique in biomedical research that uses the fluorophore decay rate to provide additional contrast in fluorescence microscopy. However, at present, the calculation, analysis,…

Image and Video Processing · Electrical Eng. & Systems 2021-06-09 Varun Mannam , Yide Zhang , Xiaotong Yuan , Cara Ravasio , Scott S. Howard

Fluorescence lifetime imaging microscopy (FLIM) is a well-established technique with numerous imaging applications. Yet, one of the limitations of FLIM is that it provides information about the emitting state only. Here, we present an…

Biological Physics · Physics 2023-10-27 Pavel Malý , Dita Strachotová , Aleš Holoubek , Petr Heřman

We report the cell biological applications of a recently developed multiphoton fluorescence lifetime imaging microscopy system using a streak camera (StreakFLIM). The system was calibrated with standard fluorophore specimens and was shown…

Cell Behavior · Quantitative Biology 2009-11-10 R. V. Krishnan , Eva Biener , Jian-Hua Zhang , Robert Heckel , Brian Herman

Fluorescence lifetime imaging microscopy (FLIM) is a powerful quantitative technique that provides metabolic and molecular contrast, offering strong translational potential for label-free, real-time diagnostics. However, its clinical…

Computer Vision and Pattern Recognition · Computer Science 2025-12-19 Paloma Casteleiro Costa , Parnian Ghapandar Kashani , Xuhui Liu , Alexander Chen , Ary Portes , Julien Bec , Laura Marcu , Aydogan Ozcan

We present a Fisher-information (FI) framework for photon-efficient fluorescence lifetime imaging microscopy (FLIM) that treats temporal sampling as a controllable design variable under a fixed photon (dose) budget. Starting from a Poisson…

Instrumentation and Detectors · Physics 2026-01-05 J. Sumaya-Martinez , E. Torres-Garcia

In fluorescence microscopy, spectral unmixing aims to recover individual fluorophore concentrations from spectral images that capture mixed fluorophore emissions. Since classical methods operate pixel-wise and rely on least-squares fitting,…

Computer Vision and Pattern Recognition · Computer Science 2026-03-26 Federico Carrara , Talley Lambert , Mehdi Seifi , Florian Jug

Conformational dynamics of biomolecules are of fundamental importance for their function. Single-molecule F\"orster Resonance Energy Transfer (smFRET) is a powerful approach to inform on the structure and the dynamics of labeled molecules.…

Fluorescence lifetime imaging microscopy (FLIM) is a powerful tool for quantitative fluorescence imaging because fluorescence lifetime is independent of concentration of fluorescent molecules or excitation/detection efficiency and is robust…

We present a statistics-aware compression strategy that processes photon timestamps directly from time-correlated single-photon counting (TCSPC) modules for time-domain fluorescence lifetime imaging (FLIM). Rather than storing or…

Image and Video Processing · Electrical Eng. & Systems 2026-05-08 Zhenya Zang , Istvan Gyongy , Mike Davies

Fluorescence lifetime imaging microscopy (FLIM) provides detailed information about molecular interactions and biological processes. A major bottleneck for FLIM is image resolution at high acquisition speeds, due to the engineering and…

Image and Video Processing · Electrical Eng. & Systems 2024-04-23 Valentin Kapitány , Areeba Fatima , Vytautas Zickus , Jamie Whitelaw , Ewan McGhee , Robert Insall , Laura Machesky , Daniele Faccio

Fluorescence lifetime imaging microscopy (FLIM) is an important technique to understand the chemical micro-environment in cells and tissues since it provides additional contrast compared to conventional fluorescence imaging. When two…

Image and Video Processing · Electrical Eng. & Systems 2022-01-19 Varun Mannam , Xiaotong Yuan , Scott Howard

Fluorescent imaging plays a critical role in a myriad of scientific endeavors, particularly in the biological sciences. Three-dimensional imaging of fluorescent intensity often requires serial data acquisition, that is voxel-by-voxel…

Optics · Physics 2015-10-27 Jeffrey J. Field , David G. Winters , Randy A. Bartels

Fluorescence Lifetime Imaging Microscopy (FLIM) using multiphoton excitation techniques is now finding an important place in quantitative imaging of protein-protein interactions and intracellular physiology. We review here the recent…

Quantitative Methods · Quantitative Biology 2009-11-10 R. V. Krishnan , A. Masuda , V. E. Centoze , B. Herman

RAndom Temporal Signals (RATS) method has proven to be a useful and versatile method for measuring photoluminescence (PL) dynamics and fluorescence lifetime imaging (FLIM). Here, we present two fundamental development steps in the method.…

Optics · Physics 2023-02-15 Jiri Junek , Karel Zidek

We demonstrate an electro-optic wide-field method to enable fluorescence lifetime microscopy (FLIM) with high throughput and single-molecule sensitivity. Resonantly driven Pockels cells are used to efficiently gate images at 39 MHz,…

Optics · Physics 2021-09-23 Adam J. Bowman , Mark A. Kasevich
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