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Protein Dynamics in Living Cells

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Utilizing pHluorin-tagged Receptors to Monitor Subcellular Localization and Trafficking

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Published on: March 16, 2017

Chemically Tuned, Reversible Fluorogenic Electrophile for Live Cell Nanoscopy.

Richard Lincoln¹, Wenzhou Zhang¹, Terri C Lovell¹

¹Department of Chemistry and Quebec Center for Advanced Materials (QCAM/CQMF), McGill University, 801 Sherbrooke Street West, Montreal H3A 0B8, Quebec, Canada.

|January 5, 2022

Summary

This summary is machine-generated.

We developed a novel fluorogenic probe, cyanoAcroB, for live-cell super-resolution imaging. This probe utilizes reversible chemical reactions to enable long-term imaging of cellular dynamics with nanoscale precision.

Keywords:

electrophilic stress fluorogenic BODIPY probes nucleophiles reversible fluorescent probes single-molecule localization microscopy

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Internalization and Observation of Fluorescent Biomolecules in Living Microorganisms via Electroporation

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Published on: February 8, 2015

Area of Science:

Chemical Biology
Biophysics
Cell Biology

Background:

Live-cell imaging requires probes that are non-phototoxic and photostable.
Super-resolution microscopy demands probes with controllable fluorescence emission.
Existing probes often suffer from photobleaching and limited temporal resolution.

Purpose of the Study:

To develop a chemically tuned fluorogenic probe for live-cell super-resolution imaging.
To exploit stochastic reversible alkylation for controlled fluorescence.
To visualize organelle dynamics with high spatial and temporal resolution.

Main Methods:

Design and synthesis of a novel fluorogenic probe (cyanoAcroB) combining a BODIPY fluorophore and a cyanoacrylate warhead.
Application of single-molecule localization microscopy (SMLM) and super-resolution radial fluctuation (SRRF) imaging.
Live-cell imaging experiments to observe organelle dynamics.

Main Results:

The cyanoAcroB probe exhibits intermittent fluorescence through a reversible thiolate Michael addition and retro-Michael reaction.
Long-term super-resolution imaging of live cells was achieved without significant background increase.
Electrophile alkylation was found to be prevalent in mitochondria and endoplasmic reticulum.
2D dynamics of these organelles were resolved with nanometer spatial and sub-second temporal resolution.

Conclusions:

The reversible fluorogenic probe cyanoAcroB enables robust live-cell super-resolution imaging.
This approach overcomes limitations of fluorescent product accumulation and probe depletion.
The study highlights the potential of bioinspired reversible probes for dynamic cellular process visualization.