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Title: Single-molecule orientation localization microscopy for resolving structural heterogeneities between amyloid fibrils
Simultaneous measurements of single-molecule positions and orientations provide critical insight into a variety of biological and chemical processes. Various engineered point spread functions (PSFs) have been introduced for measuring the orientation and rotational diffusion of dipole-like emitters, but the widely used Cramér-Rao bound (CRB) only evaluates performance for one specific orientation at a time. Here, we report a performance metric, termed variance upper bound (VUB), that yields a global maximum CRB for all possible molecular orientations, thereby enabling the measurement performance of any PSF to be computed efficiently ( ∼<#comment/> 1000 ×<#comment/> faster than calculating average CRB). Our VUB reveals that the simple polarized standard PSF provides robust and precise orientation measurements if emitters are near a refractive index interface. Using this PSF, we measure the orientations and positions of Nile red (NR) molecules transiently bound to amyloid aggregates. Our super-resolved images reveal the main binding mode of NR on amyloid fiber surfaces, as well as structural heterogeneities along amyloid fibrillar networks, that cannot be resolved by single-molecule localization alone.  more » « less
Award ID(s):
1653777
PAR ID:
10158810
Author(s) / Creator(s):
; ; ; ;
Publisher / Repository:
Optical Society of America
Date Published:
Journal Name:
Optica
Volume:
7
Issue:
6
ISSN:
2334-2536
Format(s):
Medium: X Size: Article No. 602
Size(s):
Article No. 602
Sponsoring Org:
National Science Foundation
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