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Quantitative STED microscopy with DNA-fluorophore labels

Kessler, L. F.; Li, Y.; Balakrishnan, A.; Heilemann, M.

2025-12-07 biophysics
10.64898/2025.12.05.692534 bioRxiv
Show abstract

Stimulated emission depletion (STED) microscopy enables super-resolution imaging of complex biological samples in 3D, in large volumes, and live. However, molecular quantification with STED has remained underexplored. Here, we present a straightforward approach for quantitative STED that enables molecule counting. For this purpose, we designed DNA-fluorophore labels that enable signal amplification and allow for reliable intensity-based quantitative imaging. We demonstrate accurate molecule counting on DNA origami. Furthermore, we visualized and quantified EGF receptor monomers and dimers in cells. In summary, we introduce a robust, fast, and easy-to-implement tool for quantitative STED microscopy with single-protein resolution.

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