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Protein-based, live-cell PAINT microscopy with fluorogenic exchangeable HaloTag ligands
Preprint   Open access

Protein-based, live-cell PAINT microscopy with fluorogenic exchangeable HaloTag ligands

Tong Zhan, Nels C Gerstner, Julia G Martin, Jack T McCann, Marissa C Lynch, Hsin-Jung Wu, Ke Xu and Evan W Miller
bioRxiv
Cold Spring Harbor Laboratory
07/17/2026
DOI: 10.64898/2026.07.16.738981
PMID: 42523431
url
https://doi.org/10.64898/2026.07.16.738981View
Preprint (Author's original) This preprint has not been evaluated by subject experts through peer review. Preprints may undergo extensive changes and/or become peer-reviewed journal articles. Open Access

Abstract

We report a generalizable method for single-molecule localization super-resolution microscopy in living cells. Live-cell protein-based PAINT (points accumulation for imaging in nanoscale topography) is achieved by pairing exceptionally fluorogenic bis-trifluoromethyl rhodamine (BF) dyes with reversible HaloTag ligands. This far-red small molecule update to protein-based PAINT is readily incorporated into existing super-resolution microscopy workflows: pairing with photoactivatable fluorescent proteins further enables simultaneous two-color live-cell super-resolution microscopy and single-molecule diffusivity mapping (SM M).

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