Imaging Dynamic Subcellular Organization at High Spatiotemporal Resolution.

The spatiotemporal organization of intracellular compartments is fundamental to cellular function and to the understanding of the processes underpinning health and disease. Fluorescence microscopy offers a powerful means to observe organelle morphology and dynamics with high specificity. However, no single technique can capture the wide range of relevant spatiotemporal scales due to inherent trade-offs in resolution, speed, field of view, signal-to-noise ratio, and sample viability. In this review, we describe recent developments across high-resolution fluorescence microscopy techniques and associated computational methods, critically evaluating how these advances address key limitations. Through biological examples of organelle dynamics at different scales, we illustrate the impact of these technologies on our understanding of cellular organization and function. Finally, we discuss the current challenges and outline future directions for imaging-based research, highlighting the potential for further innovations to deepen insights into dynamic subcellular processes.