Danush Taban, Marvin Jungblut, Made Budiarta, Dominic A. Helmerich, Cerridwen Kiesel, Sarah E. Plutkis, Luke D. Lavis, Donatus Krah, Ali H. Shaib, Sören Doose, Philip Kollmannsberger, Silvio O. Rizzoli, Gerti Beliu, Markus Sauer
Expansion microscopy (ExM) enables nanoscale fluorescence imaging on standard microscopes, but its combination with single-molecule localization microscopy (SMLM) remains challenging due to the incompatibility of expanded hydrogels with photoswitching buffers. Here, we introduce a single-step expansion microscopy approach that enables SMLM using spontaneously blinking dyes in 6-14× expanded samples, without re-embedding or buffer exchange. Using this approach, we achieve nanometer-scale spatial resolution by resolving the organization of the nuclear pore complex and the molecular structure of recombinant homotrimeric proliferating cell nuclear antigen.