Single-molecule spectroscopy has become a powerful method for using organic fluorescent molecules in numerous applications. Along with sensing applications in biology and solid-state physics or a... Show moreSingle-molecule spectroscopy has become a powerful method for using organic fluorescent molecules in numerous applications. Along with sensing applications in biology and solid-state physics or a variety of applications in quantum information technology, molecules offer interesting possibilities for fundamental research. One of the very interesting areas is the study of charge transport and electric field sensing at the nanoscale. Developing molecular nanosensors for electric fields can not only help to fundamentally explore the motion of charges in conductors and semiconductors but can also lead to very sensitive and accurate instruments for quasi-static charge tracing or even single-electron charge detection. Such research could eventually lead to the construction of precise electric field sensors that can act as an interface between the quantum state of an electron and the outside word. We developed fluorescence molecular systems and electronic circuits with the aim of electric-field sensing and optical detection of one single electron. Show less
In human cells, a meter-long DNA is condensed inside a micrometer-sized cell nucleus. Simultaneously, the genetic code must remain accessible for its replication and transcription to functional... Show moreIn human cells, a meter-long DNA is condensed inside a micrometer-sized cell nucleus. Simultaneously, the genetic code must remain accessible for its replication and transcription to functional proteins. Such plasticity of the genome is maintained by dynamic folding and unfolding of DNA-protein spools called nucleosomes. It is unclear, however, how this process is controlled when multiple nucleosomes stack on top of each other and form compact chromatin fibers. This is particularly important since nucleosomes are rarely present in isolation inside a densely packed cell nucleus. Therefore, the aim of this thesis was to increase the understanding of the chromatin fiber structure and its dynamics. Knowing these details would provide many new insights into the mechanisms of gene expression (epigenetic regulation) which, upon malfunction, may cause severe diseases. The presented work consists of an experimental approach involving the application of single-molecule force spectroscopy, and makes use of theoretical modelling based on statistical mechanics. By using magnetic tweezers, we stretched and twisted individual chromatin fibers reconstituted in vitro in order to unfold its nucleosomes. These studies show that folding of nucleosomes into chromatin fibers opens up a plethora of regulatory pathways for controlling the level of DNA organization in cells. Show less
Single-molecule fluorescence was invented in the 1990s and has quickly developed into an indispensable technique in the biomedical sciences and condensed-matter research. It has revolutionized... Show moreSingle-molecule fluorescence was invented in the 1990s and has quickly developed into an indispensable technique in the biomedical sciences and condensed-matter research. It has revolutionized the fields of molecular biology, imaging (super-resolution), and catalysis, to name a few. In this thesis, we will apply fluorescence enhancement by single gold nanorods to extend single-molecule studies to chromophores with low fluorescence quantum yields and to high concentrations of probe molecules. Following single-molecule trajectories, we will explore variations in the electron-transfer rates of the metalloprotein azurin both from molecule to molecule and for the same molecule as a function of time. Evidence for conformational substates will be discussed based on dynamic heterogeneity. Show less
In this thesis, we perform fundamental spectroscopic studies of organic fluorescent chromophores (dyes) for temperatures from 1-10 Kelvins. We use confocal and wide-field fluorescence microscopy... Show moreIn this thesis, we perform fundamental spectroscopic studies of organic fluorescent chromophores (dyes) for temperatures from 1-10 Kelvins. We use confocal and wide-field fluorescence microscopy techniques. We analyzed the spectroscopic properties of well-known dyes, such as perylene, terrylene and dibenzoterrylene embedded in different solids. By doing this systematically, we correlate the presence of methyl groups in the host-crystal (Chapter 2) with the strong spectral diffusion of the resonant line, which lead to spectral broadening and frequency instabilities of the guest DBT. In contrast, the absence or substitution of methyl groups by chloride showed any of these effects. We found that the fluorescent properties of perylene in ortho-dichlorobenzene and terrylene in para-dichlorobenzene solids were those expected for the single quantum system. No reports of lifetime-limited excitation lines for perylene were ever reported till now. Even further, we report the coupling of an acoustic wave created by a macroscopic object to the optical response of a single molecule. The shot-noise limited sensitivity from the response of the single molecule to the displacement of the fork's prongs was found to be in the sub-nanometer regime. Perylene in o-DCB will be used to perform the single molecule all-optical transistor proposed in the introduction. Show less
The research in this thesis comprises two separate topics: single-molecule spectroscopy and resonant Raman spectroscopy. The first part concerns single-molecule (SM) spectroscopy on polyethylene ... Show moreThe research in this thesis comprises two separate topics: single-molecule spectroscopy and resonant Raman spectroscopy. The first part concerns single-molecule (SM) spectroscopy on polyethylene (PE) films. Ultra thin (200 nm) films of pure high density PE were produced by spincoating. By determining the position (accuracy 10 nm) and in-plane orientation (acc. 5 deg.) of single 2.3,8.9-dibenzanthanthrene (DBATT) guest molecules, by means of SM microscopy and spectroscopy respectively, we demonstrated that these thin PE films have a shish-kebab morphology, instead of the spherulitic morphology common in thicker PE films. We have also investigated the alignment process of individual quest chromophores by stretching thicker (50 um), low density PE films. Using SM spectroscopy, we have shown that individual guest chromophores are not better aligned along the stretch direction, as draw ratio increases. Instead alignment occurs suddenly, due to the destruction and (oriented) reformation of local crystalline regions and subsequent adsorption of chromophores. Each chromophore's orientation is determined by specific interactions with the oriented PE crystal surface. The second part of this thesis concerns a quantum-chemical analysis of the resonant Raman spectrum of the carotenoid spheroidene reconstituted in the photosynthetic reaction center (RC) of Rhodobacter sphaeroides. Our analysis demonstrates that spheroidene can adopt at least two cis forms in the RC. One of these has been conclusively shown to be the 15,15'-cis structure. Show less
