A modular scaffold for cellularly-retained fluorogenic probes for sensitive cell-resolved bioactivity imaging

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Contributors

  • Philipp Mauker - , Chair of Organic Chemistry II, TUD Dresden University of Technology, Ludwig Maximilian University of Munich (Author)
  • Lucas Dessen-Weissenhorn - , Ludwig Maximilian University of Munich (Author)
  • Carmen Zecha - , Ludwig Maximilian University of Munich (Author)
  • Nynke A. Vepřek - , Ludwig Maximilian University of Munich (Author)
  • Julia I. Brandmeier - , Ludwig Maximilian University of Munich (Author)
  • Daniela Beckmann - , Hospital of the Ludwig-Maximilians-University (LMU) Munich, Ludwig Maximilian University of Munich (Author)
  • Annabel Kitowski - , Ludwig Maximilian University of Munich (Author)
  • Tobias Kernmayr - , Ludwig Maximilian University of Munich (Author)
  • Julia Thorn-Seshold - , University Medicine (Faculty of Medicine and University Hospital), Chair of Organic Chemistry II, Institute of Clinical Chemistry and Laboratory Medicine, TUD Dresden University of Technology, University Hospital Carl Gustav Carus Dresden (Author)
  • Martin Kerschensteiner - , Hospital of the Ludwig-Maximilians-University (LMU) Munich, Ludwig Maximilian University of Munich, Munich Cluster for Systems Neurology (SyNergy) (Author)
  • Oliver Thorn-Seshold - , Chair of Organic Chemistry II (Author)

Abstract

Here, we develop a general design for high-quality fluorogenic activity probes to quantify biochemical processes within live cells, via the release of a fully cell-retained, bright fluorescent soluble product upon reaction. Live cell probes must be membrane-permeable to access intracellular biochemistry, but often that means their fluorophore products are similarly permeable resulting in rapid signal loss from the activating cell, which limits their cell-by-cell resolution as well as their sensitivity for quantifying low-turnover processes. Current strategies to retain fluorescent products within cells usually disrupt native biology e.g. by non-specific alkylation or solid precipitation. Here, scanning charge- and polarity-based approaches to swap from permeable to cell-retained states, we developed a bright fluorogenic rhodol-based platform, Trappable Green (TraG), balancing all key requirements for signal integration (rapid probe entry, but effective product retention, across many cell lines) and being modular so it can be adapted to quantify many biochemical target types (examples shown here include probes for GSH, TrxR, and H2 O2). The simple and rugged TraG scaffold can now permit straightforward implementation in a range of cell-retained enzyme activity probes, which will enable more accurate cell-resolved imaging as well as higher-sensitivity integration of low-turnover processes, without the drawbacks of alkylation or precipitation-based strategies.

Details

Original languageEnglish
Pages (from-to)22630-22637
Number of pages8
JournalChemical science
Volume16
Issue number47
Publication statusPublished - 21 Dec 2025
Peer-reviewedYes

External IDs

WOS 001603074600001
Scopus 105023953229
ORCID /0000-0003-3981-651X/work/233047354

Keywords

ASJC Scopus subject areas