The promise and peril of comparing fluorescence lifetime in biology revealed by simulations
Signaling dynamics are crucial in biological systems, and biosensor-based real-time imaging has revolutionized their analysis. Fluorescence lifetime imaging microscopy (FLIM) excels over the widely used fluorescence intensity imaging by allowing the measurement of absolute signal levels independent...
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| Format: | Article |
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eLife Sciences Publications Ltd
2025-08-01
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| Series: | eLife |
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| Online Access: | https://elifesciences.org/articles/101559 |
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| author | Pingchuan Ma Peter Chen Scott Sternson Yao Chen |
| author_facet | Pingchuan Ma Peter Chen Scott Sternson Yao Chen |
| author_sort | Pingchuan Ma |
| collection | DOAJ |
| description | Signaling dynamics are crucial in biological systems, and biosensor-based real-time imaging has revolutionized their analysis. Fluorescence lifetime imaging microscopy (FLIM) excels over the widely used fluorescence intensity imaging by allowing the measurement of absolute signal levels independent of sensor concentration. This capability enables the comparison of signaling dynamics across different animals, body regions, and timeframes. However, FLIM’s advantage can be compromised by factors like autofluorescence in biological experiments. To address this, we introduce FLiSimBA, a flexible computational framework for realistic Fluorescence Lifetime Simulation for Biological Applications. Through simulations, we analyze the signal-to-noise ratios of fluorescence lifetime data, determining measurement uncertainty and providing necessary error bars for lifetime measurements. Furthermore, we challenge the belief that fluorescence lifetime is unaffected by sensor expression and establish quantitative limits to this insensitivity in biological applications. Additionally, we propose innovations, notably multiplexed dynamic imaging that combines fluorescence intensity and lifetime measurements. This innovation can transform the number of signals that can be simultaneously monitored, thereby enabling a systems approach in studying signaling dynamics. Thus, by incorporating different factors into our simulation framework, we uncover surprises, identify limitations, and propose advancements for fluorescence lifetime imaging in biology. This quantitative framework supports rigorous experimental design, facilitates accurate data interpretation, and paves the way for technological advancements in fluorescence lifetime imaging. |
| format | Article |
| id | doaj-art-eb7536a536bc4d6b8a73bbfbdbc285af |
| institution | Kabale University |
| issn | 2050-084X |
| language | English |
| publishDate | 2025-08-01 |
| publisher | eLife Sciences Publications Ltd |
| record_format | Article |
| series | eLife |
| spelling | doaj-art-eb7536a536bc4d6b8a73bbfbdbc285af2025-08-20T11:17:24ZengeLife Sciences Publications LtdeLife2050-084X2025-08-011310.7554/eLife.101559The promise and peril of comparing fluorescence lifetime in biology revealed by simulationsPingchuan Ma0https://orcid.org/0000-0001-9964-2852Peter Chen1https://orcid.org/0009-0008-8531-5509Scott Sternson2https://orcid.org/0000-0002-0835-444XYao Chen3https://orcid.org/0000-0003-1509-6634Department of Neuroscience, Washington University in St. Louis, St. Louis, United States; Ph.D. Program in Neuroscience, Washington University in St. Louis, St. Louis, United StatesDepartment of Neuroscience, Washington University in St. Louis, St. Louis, United States; Master’s Program in Biomedical Engineering, Washington University in St. Louis, St. Louis, United StatesDepartment of Neuroscience, Howard Hughes Medical Institute, University of California, San Diego, San Diego, United StatesDepartment of Neuroscience, Washington University in St. Louis, St. Louis, United StatesSignaling dynamics are crucial in biological systems, and biosensor-based real-time imaging has revolutionized their analysis. Fluorescence lifetime imaging microscopy (FLIM) excels over the widely used fluorescence intensity imaging by allowing the measurement of absolute signal levels independent of sensor concentration. This capability enables the comparison of signaling dynamics across different animals, body regions, and timeframes. However, FLIM’s advantage can be compromised by factors like autofluorescence in biological experiments. To address this, we introduce FLiSimBA, a flexible computational framework for realistic Fluorescence Lifetime Simulation for Biological Applications. Through simulations, we analyze the signal-to-noise ratios of fluorescence lifetime data, determining measurement uncertainty and providing necessary error bars for lifetime measurements. Furthermore, we challenge the belief that fluorescence lifetime is unaffected by sensor expression and establish quantitative limits to this insensitivity in biological applications. Additionally, we propose innovations, notably multiplexed dynamic imaging that combines fluorescence intensity and lifetime measurements. This innovation can transform the number of signals that can be simultaneously monitored, thereby enabling a systems approach in studying signaling dynamics. Thus, by incorporating different factors into our simulation framework, we uncover surprises, identify limitations, and propose advancements for fluorescence lifetime imaging in biology. This quantitative framework supports rigorous experimental design, facilitates accurate data interpretation, and paves the way for technological advancements in fluorescence lifetime imaging.https://elifesciences.org/articles/101559fluorescence lifetimesimulationbiosensorFLiSimBAsensor expressionmultiplexed dynamic imaging |
| spellingShingle | Pingchuan Ma Peter Chen Scott Sternson Yao Chen The promise and peril of comparing fluorescence lifetime in biology revealed by simulations eLife fluorescence lifetime simulation biosensor FLiSimBA sensor expression multiplexed dynamic imaging |
| title | The promise and peril of comparing fluorescence lifetime in biology revealed by simulations |
| title_full | The promise and peril of comparing fluorescence lifetime in biology revealed by simulations |
| title_fullStr | The promise and peril of comparing fluorescence lifetime in biology revealed by simulations |
| title_full_unstemmed | The promise and peril of comparing fluorescence lifetime in biology revealed by simulations |
| title_short | The promise and peril of comparing fluorescence lifetime in biology revealed by simulations |
| title_sort | promise and peril of comparing fluorescence lifetime in biology revealed by simulations |
| topic | fluorescence lifetime simulation biosensor FLiSimBA sensor expression multiplexed dynamic imaging |
| url | https://elifesciences.org/articles/101559 |
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