Advances in Correlative Single-Molecule Localization Microscopy and Electron Microscopy

During the last few decades, correlative fluorescence light and electron microscopy (FLM-EM) has gained increased interest in the life sciences concomitant with the advent of fluorescence light microscopy. It has become, accompanied by numerous developments in both techniques, an important tool to study bio-cellular structure and function as it combines the specificity of fluorescence labeling with the high structural resolution and cellular context information given by the EM images. Having the recently introduced single-molecule localization microscopy techniques (SMLM) at hand, FLM-EM can now make use of improved fluorescence light microscopy resolution, single-molecule sensitivity and quantification strategies. Here, currently used methods for correlative SMLM and EM including the special requirements in sample preparation and imaging routines are summarized and an outlook on remaining challenges concerning methods and instrumentation is provided.

Single-molecule FRET for Ultrasensitive Detection of Biomolecules

Single-molecule Förster resonance energy transfer (sm- FRET) has been widely employed to detect biomarkers and to probe the structure and dynamics of biomolecules. By monitoring the biological reaction in a spatio-temporal manner, smFRET can reveal the transient intermediates of biological processes that cannot be obtained by conventional ensemble measurements. This review provides an overview of singlemolecule FRET and its applications in ultrasensitive detection of biomolecules, including the major techniques and the molecular probes used for smFRET as well as the biomedical applications of smFRET. Especially, the combination of sm- FRET with new technologies might expand its applications in clinical diagnosis and biomedical research

The Synthesis of a Luminescent Plasmonic Nanomaterial

Gold nanoparticles were synthesised in toluene and stabilised by a fluorescent conjugated polymer (F8BT). The gold/conjugated polymer nanoparticles system was also found to be stable in dichloromethane (DCM), exhibiting both fluorescence (quantum yield = 14%) and a surface plasmon in organic solvents

Pathways to optical STED microscopy

Optical far-field microscopy such as confocal fluorescence microscopy is a very popular technique for investigating the living cell. Unfortunately, its spatial resolution is limited to around 200 nm, impeding the imaging of small molecular assemblies. Recent decades have seen the development of optical nanoscopy, optical far-field microscopy with a spatial resolution down to molecular scales. STED microscopy was the first of such nanoscopy techniques. Despite the fact, that it in principle only requires the addition of a strong STED laser to a conventional microscope, STED nanoscopy was for a long time considered as a very complex technique, impossible to be applicable as a turn-key technique in everyday biological research. However, recent years has seen important improvements of the STED nanoscopy approach which have significantly simplified the setup. These developments mainly followed from optimization of fluorescent labels, laser technology and optical simplifications. As a result, STED microscopy setups have got more compact and have been realized on commercial instruments, allowing access to lessexperienced users in open imaging facilities. Here, we give a brief overview of the recent improvements in STED microscopy that made these important developments possible