Cooperative Blinking from Dye Ensemble Activated by Energy Transfer for Super-resolution Cellular Imaging

Area of Science:

• Nanotechnology
• Biophotonics
• Materials Science

Background:

• Photoblinking is a key characteristic of single fluorophores, crucial for advanced imaging techniques.
• Achieving controlled and enhanced photoblinking in ensemble systems has been a significant challenge.
• Existing fluorescent probes often lack the desired on/off ratios and cooperative blinking for super-resolution applications.

Purpose of the Study:

• To develop a strategy for achieving pronounced, high on/off ratio, and cooperative photoblinking in multifluorophore systems.
• To create novel fluorescent probes based on dye-doped semiconducting polymer dots (Pdots) for enhanced super-resolution imaging.
• To demonstrate the utility of these blinking Pdots in subcellular structure labeling and resolution enhancement.

Main Methods:

• Doping organic dye molecules into semiconducting polymer dots (Pdots) to create donor-acceptor multifluorophore systems.
• Investigating energy transfer mechanisms between Pdots and dye acceptors to induce cooperative photoblinking.
• Utilizing hole polaron quenching of semiconducting polymers to generate the dark state in the blinking process.
• Applying the blinking Pdots for subcellular structure labeling in high-order super-resolution optical fluctuation nanoscopy.

Main Results:

• Demonstrated robust photoblinking with a high on/off ratio in dye-doped Pdots, unlike pristine Pdots or dyes in inert matrices.
• Established that energy transfer from Pdots to dye acceptors drives a cooperative blinking process.
• Observed that the bright state arises from the dye ensemble, while the dark state is due to polymer quenching by hole polarons.
• Achieved approximately 3.6-fold enhancement in imaging resolution using blinking Pdots in super-resolution nanoscopy compared to conventional microscopy.

Conclusions:

• Successfully transformed a nonquantized ensemble into a single-emitter-like optical source with controllable photoblinking.
• Developed an effective approach for generating superior photoblinking fluorescent probes.
• Showcased the potential of these probes for significantly advancing super-resolution imaging capabilities, particularly in subcellular structure analysis.