During the project period, we achieved several significant milestones that greatly impacted our research. Firstly, we successfully established five fully functional experimental setups, either constructed by our team members, including PhD students, postdocs, and the principal investigator, or acquired from Bruker/JPK. Secondly, we assembled a talented team through the hiring process. The project's aim was to develop new tools to reveal the interfacial dynamics of water, ions, and molecules in nanofluidics platforms. We identified a significant knowledge gap due to the lack of appropriate tools to study these dynamics. To address this, we constructed dedicated experimental setups that combine nanoscale conductance measurements and electrochemistry with optical imaging at the nanoscale. These setups include super-resolution localization microscopy, which enhances spatial, spectral, and temporal resolution, as well as setups that allow us to infer the orientation of molecules (3D polarization setup) and perform wide-field lifetime measurements.
To improve the temporal resolution of single-molecule imaging and tracking, we explored the use of SPAD arrays for fluorescence microscopy, focusing on challenging single-molecule imaging. SPAD cameras enable high-speed imaging of fast-moving emitters, which we applied to single proteins on lipid membranes and 2D crystals of hexagonal boron nitride. Single-molecule imaging was achievable in both cases, with photon arrival sub-information at 10-100 µs resolution, enriching trajectory analysis possibilities. Additionally, we investigated the time-resolved capabilities of SPAD cameras for lifetime measurements of fluorescent emitters on a wide-field basis. This resulted in a new high-throughput method, normally achieved by scanning one molecule at a time on a confocal microscope. Our method could enhance techniques like STORM and PAINT used in single-molecule localization microscopy by adding lifetime information, potentially leading to improved imaging of multiple targets simultaneously and better detection of environmental changes at the super-resolution level.
Conventional SMLM only contains information about the position of the emitter, but features such as its wavelength, polarization, and lifetime are also essential for a complete depiction of the profile. The process of stimulated emission is a consequence of the uneven distribution of charge within a molecule. As a result, the polarization of the emitted fluorescent photon depends on the molecule's orientation. Within this project, we constructed a 3D polarization SMLM imaging platform that allows us to investigate the three-dimensional orientations of these molecules.
1. Nathan Ronceray, et.al Nature Materials 22, no. 10 (2023): 1236-1242.
The paper has been disseminated at the following conferences by Nathan Ronceray (NR) and Aleksandra Radenovic (AR): SMLMS 2023, Vienna (NR); SMLMS 2022, Paris (NR); Heraeus-Seminar on "Defects in Two-dimensional Materials, Bad Honnef (AR); Cambridge 2D TMD 2023, Cambridge (AR); BoronNitrideWorkshop 2023, Montpellier (AR, NR);
2. 2. Emmerich, Theo et al. Nature Electronics (2024): 1-8.
The paper has been disseminated at the following conferences by AR, NR, and Theo Emmerich (TE) Nanofluidics conference, Lenzerheide (AR, NR); Nanofluidics 23 in physics and biology, Lyon (AR); Dubrovnik - Solid State to BioPhysics X, Cavtat, Dubrovnik (AR, NR, TE).
