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A Versatile Tool to Predict and Guide RESOLFT Images Based on Photoswitching, Labelling and Optical Properties
KTH, School of Engineering Sciences (SCI), Applied Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab.ORCID iD: 0000-0001-9302-7576
KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Engineering Sciences (SCI), Applied Physics.ORCID iD: 0000-0002-3488-6845
KTH, School of Engineering Sciences (SCI), Applied Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
KTH, School of Engineering Sciences (SCI), Applied Physics, Biophysics. KTH, Centres, Science for Life Laboratory, SciLifeLab.ORCID iD: 0000-0003-4005-4997
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2026 (English)In: ChemPhysChem, ISSN 1439-4235, E-ISSN 1439-7641, Vol. 27, no 8, article id e202500780Article in journal (Refereed) Published
Abstract [en]

Reversibly switchable fluorescent proteins (RSFPs) transition many times between dark and fluorescent states under minimal light doses. The photoswitching can happen at different speed, contrast and length, and it is often challenging for users to select the optimal imaging scheme to generate images with high contrast and spatial resolution. Here, we experimentally investigate the photophysical properties of different RSFPs under imaging conditions, together with an in silico exploration of their role in nanoscale image formation. We developed open-source software that uses measured parameters such as brightness, switching speed, photoswitching fatigue, labelling densities, noise and illumination type to generate the related RESOLFT (reversible saturable/switchable optical fluorescence transition) super-resolution image. This tool can be used to select optimal imaging schemes for known RSFPs and to guide the rational development of new proteins.

Place, publisher, year, edition, pages
Wiley , 2026. Vol. 27, no 8, article id e202500780
Keywords [en]
fluorescent proteins, imaging simulation, photoswitching, RESOLFT, super-resolution microscopy
National Category
Biophysics Molecular Biology Medical Imaging
Identifiers
URN: urn:nbn:se:kth:diva-380692DOI: 10.1002/cphc.202500780PubMedID: 42014022Scopus ID: 2-s2.0-105036202167OAI: oai:DiVA.org:kth-380692DiVA, id: diva2:2058572
Note

QC 20260508

Available from: 2026-05-08 Created: 2026-05-08 Last updated: 2026-05-08Bibliographically approved

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Boden, AndreasAl-Kadhimi, StaffanTesta, IlariaPennacchietti, Francesca

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Boden, AndreasAl-Kadhimi, StaffanUriati, EleonoraTesta, IlariaPennacchietti, Francesca
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