Advancing Single-Molecule Imaging

In this talk, I will demonstrate how advancements in imaging methods can open new avenues for research at solid-liquid interfaces and also applications in single molecule biophysics. In the first part, I will discuss our efforts to explore nanophotonics at the single-molecule level, with a focus on the optical dynamics and conformations of individual emitters at the solid-liquid interface and in confined environments.
In our recent work, we employed fluorescence microscopy to monitor electrochemical reactions at the single-molecule scale, achieving a wide field of view and high temporal resolution. Additionally, we introduced an advanced bifocal polarization single-molecule localization microscopy (pSMLM) to enable real-time, multi-dimensional observations of quantum emissions formed by organic molecules and h-BN native defects. Our findings reveal a strong correlation between the orientation of quantum emitters and the symmetry of the h-BN lattice.
I will also discuss the use of SPAD cameras for single-particle tracking applications, as well as in high-throughput single-molecule fluorescence lifetime imaging (smFLIM). Finally, we highlight the compatibility of these approaches with the SPiFFI (Single-Frame Super-Resolution Imaging) method developed in our laboratory, which enables ~80 nm spatial resolution from a single acquisition frame using a simple optical implementation.
1. Ronceray, Nathan, Yi You, Evgenii Glushkov, Martina Lihter, Benjamin Rehl, Tzu-Heng Chen, Gwang-Hyeon Nam et al. "Liquid-activated quantum emission from pristine hexagonal boron nitride for nanofluidic sensing." Nature Materials 22, no. 10 (2023): 1236-1242.
2. Mayner, Eveline, Nathan Ronceray, Martina Lihter, Tzu-Heng Chen, Kenji Watanabe, Takashi Taniguchi, and Aleksandra Radenovic. "Monitoring electrochemical dynamics through single-molecule imaging of hBN surface emitters in organic solvents." ACS nano 18 (40), 27401-27410 
3. Guo, Wei, Tzu-Heng Chen, Nathan Ronceray, Eveline Mayner, Kenji Watanabe, Takashi Taniguchi, and Aleksandra Radenovic. "Dipole orientation reveals single-molecule interactions and dynamics on 2D crystals." arXiv preprint arXiv:2408.01207 (2024).
4. Guo, Wei, Lely Feletti, and Aleksandra Radenovic. "Spatial Polarization-Induced Fluorescence Fluctuation Imaging (SPIFFI) Enables Single-shot Super-Resolution and Multidimensional Imaging." bioRxiv (2025): 2025-12.