We report super-resolution fluorescence imaging of live cells with high spatiotemporal resolution using stochastic optical reconstruction microscopy (STORM). By labeling proteins either directly or via SNAP tags with photoswitchable dyes, we obtained two-dimensional (2D) and 3D super-resolution images of living cells, using clathrin-coated pits and the transferrin cargo as model systems. Bright, fast-switching probes enabled us to achieve 2D imaging at spatial resolutions of 1/425 nm and temporal resolutions as fast as 0.5 s. We also demonstrated live-cell 3D super-resolution imaging. We obtained 3D spatial resolution of 1/430 nm in the lateral direction and 1/450 nm in the axial direction at time resolutions as fast as 1-2 s with several independent snapshots. Using photoswitchable dyes with distinct emission wavelengths, we also demonstrated two-color 3D super-resolution imaging in live cells. These imaging capabilities open a new window for characterizing cellular structures in living cells at the ultrastructural level.
Bibliographical noteFunding Information:
We thank M. Davidson (Florida State University) and L. Looger (Janelia Farm) for Eos fluorescent protein plasmids. This work is supported in part by the US National Institutes of Health (to X.Z.) and a Collaborative Innovation Award (43667) from Howard Hughes Medical Institute. S.-H.S. is in part supported by the Mary Fieser fellowship. X.Z. receives support from the Howard Hughes Medical Institute.
ASJC Scopus subject areas
- Molecular Biology
- Cell Biology