Physics Commons^™

Imaging The Three-Dimensional Orientation And Rotational Mobility Of Fluorescent Emitters Using The Tri-Spot Point Spread Function, Oumeng Zhang, Jin Lu, Tianben Ding, Matthew D. Lew

Electrical & Systems Engineering Publications and Presentations

Fluorescence photons emitted by single molecules contain rich information regarding their rotational motions, but adapting single-molecule localization microscopy (SMLM) to measure their orientations and rotational mobilities with high precision remains a challenge. Inspired by dipole radiation patterns, we design and implement a Tri-spot point spread function (PSF) that simultaneously measures the three-dimensional orientation and the rotational mobility of dipole-like emitters across a large field of view. We show that the orientation measurements done using the Tri-spot PSF are sufficiently accurate to correct the anisotropy-based localization bias, from 30 nm to 7 nm, in SMLM. We further characterize the emission anisotropy …