Recent innovations in fluorescence lifetime imaging microscopy for biology and medicine

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 Forster 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. (C) The Authors. Published by SPIE under a Creative Commons Attribution 4.0 Unported License.

Automated summary

Fluorescence lifetime imaging microscopy (FLIM) is a powerful molecular imaging technique that measures the decay rate of fluorophores to provide insights into molecular interactions. Major advances in FLIM instrumentation, analysis, and biological and clinical applications over the last year have contributed to the growing interest in FLIM and continuous technological improvements in biomedical research.