Abstract：

Spatially displaced structured beams enable nanometer‑scale localization of single molecules, with MINFLUX setting the photon efficiency benchmark. Three raster‑scanning alternatives—RASTMIN, RASTMAX, and RASTED—can be implemented on standard microscopes with minimal modification, with RASTMIN already demonstrating MINFLUX‑level precision. We compare these methods using simulations and single‑molecule experiments, assessing precision, photon efficiency, robustness, throughput, and implementation. RASTMIN offers highest efficiency but low throughput; RASTMAX is simpler with slightly reduced precision; RASTED requires STED but achieves similar precision with 20–50× higher throughput. These results define optimal use cases and show that nanometer‑scale localization is broadly achievable on standard scanning microscopes.

Biography:

Prof. Stefani holds a Ph.D. from the University of Mainz/Max Planck Institute for Polymer Research, was researcher at the Institute of Photonic Sciences (ICFO, Barcelona), Assistant Prof. at the LMU Munich, Max Planck Partner Group leader with Stefan W. Hell, co-developer of MINFLUX. Currently is Full Prof. at the Physics Dept. of the University of Buenos Aires, Superior Investigator of National Scientific and Technical Research Council and Director of the Center for Bionanoscience Research.