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Published on: June 8, 2020
Spontaneously blinking spiroamide rhodamines for live SMLM imaging of the plasma membrane
Sonia Pfister1, Sophie Walter1, Aurélie Perrier2
1Chemistry of Photoresponsive Systems, Laboratoire de Chémo-Biologie Synthétique et Thérapeutique (CBST) UMR 7199, CNRS, Université de Strasbourg, F-67400 Illkirch, France. mayeul.collot@unistra.fr.
Abstract:
We have developed spontaneously blinking fluorescent probes based on the reversible spirolactamization of rhodamine, to efficiently image the plasma membrane (PM) of live cells with enhanced resolution using SMLM. This study demonstrates that the blinking efficiency of spiroamide PM probes is not solely governed by their pKa; the presence of a charged polar group on the amide should also be taken into account.
Insights
Researchers created new fluorescent probes for live-cell imaging of the plasma membrane (PM). Blinking efficiency depends on probe structure, not just acidity, enabling enhanced resolution with super-resolution microscopy.
Area of Science:
- Chemical biology
- Cell biology
- Microscopy
Background:
- Accurate imaging of the plasma membrane (PM) in live cells is crucial for understanding cellular processes.
- Existing fluorescent probes often face limitations in resolution and signal stability for super-resolution microscopy.
- Rhodamine-based probes offer potential but require optimization for specific cellular targets like the PM.
Purpose of the Study:
- To develop novel, spontaneously blinking fluorescent probes for high-resolution live-cell plasma membrane imaging.
- To investigate the factors influencing the blinking efficiency of these novel probes for super-resolution microscopy (SMLM).
- To enhance the resolution of plasma membrane imaging using these optimized probes.
Main Methods:
- Synthesis of novel rhodamine-based fluorescent probes utilizing reversible spirolactamization.
- Application of the probes for live-cell imaging of the plasma membrane.
- Utilized super-resolution microscopy techniques (SMLM) to assess imaging resolution and probe performance.
- Analyzed probe blinking efficiency in relation to chemical structure, including pKa and the presence of charged polar groups.
Main Results:
- Successfully developed spontaneously blinking fluorescent probes capable of imaging the live-cell plasma membrane.
- Achieved enhanced resolution in plasma membrane imaging using SMLM with the developed probes.
- Demonstrated that probe blinking efficiency is influenced by both the pKa and the presence of a charged polar group on the amide moiety, not solely by pKa.
Conclusions:
- Novel spontaneously blinking fluorescent probes based on rhodamine spirolactamization enable efficient and high-resolution live-cell plasma membrane imaging.
- The design of effective probes requires consideration of the interplay between pKa and charged polar groups for optimal blinking efficiency.
- These findings advance the development of advanced fluorescent tools for super-resolution microscopy applications in cell biology.
