Fluorescence lifetime imaging (FLIM): simulation and error analysis

Masilamani Elangovan; Peter Kionga-Kamau; Ammasi Periasamy

doi:10.1117/12.384230

27 April 2000 Fluorescence lifetime imaging (FLIM): simulation and error analysis

Masilamani Elangovan, Peter Kionga-Kamau, Ammasi Periasamy

Proceedings Volume 3921, Optical Diagnostics of Living Cells III; (2000) https://doi.org/10.1117/12.384230
Event: BiOS 2000 The International Symposium on Biomedical Optics, 2000, San Jose, CA, United States

Abstract

Real-time monitoring of reorganization and molecular interactions of live cellular components on a precise spatial and temporal scale, requires a high-speed and high- sensitivity imaging system. Theoretically, excited state fluorescence lifetime imaging techniques provide high temporal resolution for dynamics studies of biological samples. In order to provide a comprehensive foundation for the development of the technique, we simulated images using the decay and normal equations for a single component with different windowing schemes. In this model we varied several parameters involved in the simulation to produce images under different theoretical conditions. To obtain a realistic result, we considered equations for noise due to readout, dark current, photon shot noises, and other factors. In addition, the simulant was validated using sample decay images with known lifetimes, which were obtained with current fluorescent decay systems. The software developed through this research is intended to create a compete tutorial describing the theory and procedures of lifetime image acquisition. This theoretical simulation is verified with experiments using known fluorophore lifetimes.

Citation Download Citation

Masilamani Elangovan, Peter Kionga-Kamau, and Ammasi Periasamy "Fluorescence lifetime imaging (FLIM): simulation and error analysis", Proc. SPIE 3921, Optical Diagnostics of Living Cells III, (27 April 2000); https://doi.org/10.1117/12.384230

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KEYWORDS

Luminescence

Fluorescence lifetime imaging

Error analysis

Image processing

Software development

Imaging systems

Microscopy

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