Abstract
.Significance: Fluorescence lifetime imaging microscopy (FLIM) measures the decay rate of fluorophores, thus providing insights into molecular interactions. FLIM is a powerful molecular imaging technique that is widely used in biology and medicine.Aim: This perspective highlights some of the major advances in FLIM instrumentation, analysis, and biological and clinical applications that we have found impactful over the last year.Approach: Innovations in FLIM instrumentation resulted in faster acquisition speeds, rapid imaging over large fields of view, and integration with complementary modalities such as single-molecule microscopy or light-sheet microscopy. There were significant developments in FLIM analysis with machine learning approaches to enhance processing speeds, fit-free techniques to analyze images without a priori knowledge, and open-source analysis resources. The advantages and limitations of these recent instrumentation and analysis techniques are summarized. Finally, applications of FLIM in the last year include label-free imaging in biology, ophthalmology, and intraoperative imaging, FLIM of new fluorescent probes, and lifetime-based Förster resonance energy transfer measurements.Conclusions: A large number of high-quality publications over the last year signifies the growing interest in FLIM and ensures continued technological improvements and expanding applications in biomedical research.
Highlights
The fluorescence lifetime is the average time that a fluorophore remains in an excited state before returning to the ground state and emitting a photon
Datta et al.: Recent innovations in fluorescence lifetime imaging microscopy for biology and medicine applications to live cell and tissue imaging.[1]. The goal of this perspective paper is to provide updates of innovations in Fluorescence lifetime imaging microscopy (FLIM) which we, as experts in the field, found impactful within the last year. These new developments have been categorized into advances in instrumentation, new approaches to FLIM data analysis, and recent applications of FLIM in biology and medicine
A real-time digital frequency domain FLIM system using field programmable gate arrays (FPGA) at 250 MS∕s was implemented by Serafino et al.[7] that is capable of simultaneous three-channel detection using avalanche photodiodes (APD)
Summary
FLIM is a widely used tool for biomedical imaging and has advanced the field of microscopy in the past few decades In this perspective, we provided updates of innovations in FLIM within the last year that we as experts in the field found compelling. We provided updates of innovations in FLIM within the last year that we as experts in the field found compelling These new developments have been categorized into advances in instrumentation, new approaches to analyze FLIM data, and recent applications of FLIM in biology and medicine. There is an emphasis on open source FLIM analysis tools that can be integrated into other data processing pipelines These computational tools unravel spatial and molecular features of cellular physiology that are not apparent with qualitative observation of FLIM images. The numerous high-quality publications this year indicate that FLIM technologies, analysis, and applications will continue to develop, gain popularity, and impact biomedical research
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