Recommendations for empowering early career researchers to improve research culture and practice.

Early career researchers (ECRs) are important stakeholders leading efforts to catalyze systemic change in research culture and practice. Here, we summarize the outputs from a virtual unconventional conference (unconference), which brought together 54 invited experts from 20 countries with extensive experience in ECR initiatives designed to improve the culture and practice of science. Together, we drafted 2 sets of recommendations for (1) ECRs directly involved in initiatives or activities to change research culture and practice; and (2) stakeholders who wish to support ECRs in these efforts. Importantly, these points apply to ECRs working to promote change on a systemic level, not only those improving aspects of their own work. In both sets of recommendations, we underline the importance of incentivizing and providing time and resources for systems-level science improvement activities, including ECRs in organizational decision-making processes, and working to dismantle structural barriers to participation for marginalized groups. We further highlight obstacles that ECRs face when working to promote reform, as well as proposed solutions and examples of current best practices. The abstract and recommendations for stakeholders are available in Dutch, German, Greek (abstract only), Italian, Japanese, Polish, Portuguese, Spanish, and Serbian.

Challenges on optical printing of colloidal nanoparticles.

While colloidal chemistry provides ways to obtain a great variety of nanoparticles with different shapes, sizes, material compositions, and surface functions, their controlled deposition and combination on arbitrary positions of substrates remain a considerable challenge. Over the last ten years, optical printing arose as a versatile method to achieve this purpose for different kinds of nanoparticles. In this article, we review the state of the art of optical printing of single nanoparticles and discuss its strengths, limitations, and future perspectives by focusing on four main challenges: printing accuracy, resolution, selectivity, and nanoparticle photostability.

Thermometries for Single Nanoparticles Heated with Light.

The development of efficient nanoscale photon absorbers, such as plasmonic or high-index dielectric nanostructures, allows the remotely controlled release of heat on the nanoscale using light. These photothermal nanomaterials have found applications in various research and technological fields, ranging from materials science to biology. However, measuring the nanoscale thermal fields remains an open challenge, hindering full comprehension and control of nanoscale photothermal phenomena. Here, we review and discuss existent thermometries suitable for single nanoparticles heated under illumination. These methods are classified in four categories according to the region where they assess temperature: (1) the average temperature within a diffraction-limited volume, (2) the average temperature at the immediate vicinity of the nanoparticle surface, (3) the temperature of the nanoparticle itself, and (4) a map of the temperature around the nanoparticle with nanoscale spatial resolution. In the latter, because it is the most challenging and informative type of method, we also envisage new combinations of technologies that could be helpful in retrieving nanoscale temperature maps. Finally, we analyze and provide examples of strategies to validate the results obtained using different thermometry methods.

Super-Resolved FRET Imaging by Confocal Fluorescence-Lifetime Single-Molecule Localization Microscopy.

Fluorescence Resonance Energy Transfer (FRET)-based approaches are unique tools for sensing the immediate surroundings and interactions of (bio)molecules. FRET imaging and Fluorescence Lifetime Imaging Microscopy (FLIM) enable the visualization of the spatial distribution of molecular interactions and functional states. However, conventional FLIM and FRET imaging provide average information over an ensemble of molecules within a diffraction-limited volume, which limits the spatial information, accuracy, and dynamic range of the observed signals. Here, an approach to obtain super-resolved FRET imaging based on single-molecule localization microscopy using an early prototype of a commercial time-resolved confocal microscope is demonstrated. DNA Points Accumulation for Imaging in Nanoscale Topography with fluorogenic probes provides a suitable combination of background reduction and binding kinetics compatible with the scanning speed of usual confocal microscopes. A single laser is used to excite the donor, a broad detection band is employed to retrieve both donor and acceptor emission, and FRET events are detected from lifetime information.

Super-resolution FRET measurements.

Super-resolution fluorescence microscopy and Förster Resonance Energy Transfer (FRET) form a well-established family of techniques that has provided unique tools to study the dynamic architecture and functionality of biological systems, as well as to investigate nanomaterials. In the last years, the integration of super-resolution methods with FRET measurements has generated advances in two fronts. On the one hand, FRET-based probes have enhanced super-resolution imaging. On the other, the development of super-resolved FRET imaging methods has allowed the visualization of molecular interaction patterns with higher spatial resolution, less averaging and higher dynamic range. Here, we review these advances and discuss future perspectives, including the possible integration of FRET with next generation super-resolution techniques capable of reaching true molecular-scale spatial resolution.