Raster-scanning Donut simplifies MINFLUX and provides alternative implement on other scanning-based microscopes

Abstract

A donut excitation moves around a single molecule with a zigzag configuration lattice by lattice. Such a method implemented in scanning fluorescence microscopy simplifies the conventional MINFLUX process. Consisting of hollow zero-intensity excitation, single-pixel detection, time-correlated single photon counting, and drift stabilization, the system achieves localization precision and resolution very close to conventional MINFLUX theoretically and experimentally. An averaged high-SNR reference, and pixel-registered intensity from a single molecule is essential to reconstruct localization in maximum likelihood estimation. With performance reaching nearly conventional MINFLUX’s, the proposed raster-scanning MINFLUX can inspire researchers expertized in STED or confocal setup to quickly transform to MINFLUX and develop for further exploring on bio-specimens or optical applications.