Sustained 3D isotropic imaging of subcellular dynamics using adaptive VCD light-field microscopy 2.0

Long-term and high-spatiotemporal-resolution 3D imaging of living cells remains an unmet challenge for super-resolution microscopy, owing to the noticeable phototoxicity and limited scanning speed. While emerging light-field microscopy can mitigate this issue through three-dimensionally capturing biological dynamics with merely single snapshot, it suffers from suboptimal resolution insufficient for resolving subcellular structures. Here we propose an adaptive light-field microscopy (VCD-LFM 2.0) with new optics-aware and adaptive-tuning deep learning strategies (VCD 2.0) capable for highly-generalizable light-field reconstruction of diverse subcellular dynamics. VCD-LFM 2.0 accomplishes isotropic spatial resolution down to 120 nm without compromising superior temporal resolution and low phototoxicity. With the development of a compact light-field add-on and an open-source reconstruction software, we realize rapid (up to 333 volumes per second), sustained (up to 60 hours) 3D super-resolution imaging of diverse intracellular dynamics simply on a retrofitted inverted microscope. With VCD-LFM 2.0 approach, we quantitatively analyze both the instantaneous processes of lysosome-mitochondrial interaction and long-term evolution of mitochondrial across two complete cell cycles.