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. Author manuscript; available in PMC: 2021 Aug 14.
Published in final edited form as: Curr Opin Cell Biol. 2020 Dec 9;70:18–26. doi: 10.1016/j.ceb.2020.11.002

Figure 4. Microscopy overview and particle tracking considerations.

Figure 4

(a) Overview of trade-offs in fluorescence microscopy. High laser power provides a high signal, high spatial resolution, and low localization error but leads to phototoxicity and bleaching. Fast scan speed/exposure time yields high temporal resolution and limits ‘motion blurring’ but at the cost of lower signal and lower spatial resolution. Simultaneous use of high laser power and fast scan speed/exposure time can yield both high spatial and high temporal resolution but at the cost of high phototoxicity and bleaching. Time-lapse imaging makes it possible to follow the same cell and loci over time, detect dynamics, and record trajectories. However, longer time series lead to higher photobleaching. Optimal SRLCI imaging requires careful parameter optimization according to these considerations. (b) Workflow diagram for image analysis, trajectory generation, and trajectory analysis. SRLCI, super-resolution live-cell imaging.