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. 2021 Oct 1;60(44):23835–23841. doi: 10.1002/anie.202110327

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© 2021 The Authors. Angewandte Chemie International Edition published by Wiley-VCH GmbH

This is an open access article under the terms of the http://creativecommons.org/licenses/by-nc-nd/4.0/ License, which permits use and distribution in any medium, provided the original work is properly cited, the use is non‐commercial and no modifications or adaptations are made.

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MINFLUX measurements of ^StrepEnc_SM{HaloTag‐AF647}. A) Off‐switching count trace with visible off‐switching steps for a single ^StrepEnc_SM{HaloTag‐AF647} construct. B) Estimated number of fluorophores per ^StrepEnc_SM{HaloTag‐AF647} construct for all measurements (histogram, n=125) and median of the distribution (dashed line). C) 3D MINFLUX localizations of the single ^StrepEnc_SM{HaloTag‐AF647} construct. Each scatter dot represents an individual localization estimate for the position of a fluorescent AF647 molecule. The color‐coding describes when the localization was registered after starting the acquisition. Individual fluorescent molecules can deliver tens of localizations. We centered the localization with the origin by subtracting the mean position from all localizations. A sphere of 14 nm diameter (gray) encloses 94 % of localizations.