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Tagging live cells that express specific peptidase activity with solid-state fluorescence.

Maxime Prost1, Laurence Canaple, Jacques Samarut

  • 1Laboratoire de Chimie, University of Lyon-UCB Lyon 1, CNRS, ENS de Lyon, 46 allée d'Italie, 69364 Lyon (France).

Chembiochem : a European Journal of Chemical Biology
|June 20, 2014
PubMed
Summary

A novel solid-state fluorophore probe detects peptidase activity in living cells. Its bright, rapid signal offers superior imaging and localization for biomedical applications.

Keywords:
cell taggingcleavage reactionsfluorescence spectroscopyfluorogenic probesspacers

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Area of Science:

  • Biochemistry
  • Cell Biology
  • Chemical Biology

Background:

  • Peptidase activity is crucial in biological processes.
  • Existing probes often suffer from signal diffusion and low signal-to-background ratios.
  • Need for stable, high-resolution probes for live-cell analysis.

Purpose of the Study:

  • To develop a novel three-component probe for detecting peptidase activity in living cells.
  • To achieve high signal-to-background ratios for microscopic imaging.
  • To enable precise localization of enzymatic activity.

Main Methods:

  • Utilized a solid-state fluorophore with an off-on detection mode.
  • Incorporated a tertiary carbamate linkage for probe stability.
  • Varied the degree of chlorination to optimize response time.
  • Applied the probe for live-cell imaging, gel analysis, and agar-based assays.

Main Results:

  • The probe demonstrated an exceptional signal-to-background ratio due to its precipitate formation and bright fluorescence.
  • Probe stability was enhanced by the tertiary carbamate linkage.
  • Chlorination level controlled response time, enabling suitability for live-cell analysis.
  • Achieved highly resolved localization of peptidase activity, outperforming probes releasing soluble fluorophores.

Conclusions:

  • The developed probe offers a robust and sensitive method for detecting peptidase activity.
  • Its solid-state nature and precipitate formation overcome limitations of soluble probes.
  • The probe shows significant potential for applications in flow cytometry and colony assays.