Single-Molecule Metal-Induced Energy Transfer: From Basics to Applications
by Narain Karedla
Date of Examination:2016-06-02
Date of issue:2016-12-16
Advisor:Prof. Dr. Jörg Enderlein
Referee:Prof. Dr. Andreas Janshoff
Referee:Prof. Dr. Sarah Köster
Referee:Prof. Dr. Holger Stark
Referee:Dr. Gopalakrishnan Balasubramanian
Referee:Dr. Andreas Neef
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Description:Doctoral Dissertation: Single-Molecule Metal Induced Energy Transfer
Abstract
English
Single-molecule detection and spectroscopy have revolutionized the field of fluorescence microscopy. Due to their enormous potential in studying physics, chemistry and biology at molecular level, the number of single-molecule based techniques and methods has grown exponentially in the last two decades. A recent addition to the pool of existing single-molecule based techniques are superresolution imaging methods, which are used for resolving structures far below the diffraction limit of an optical microscope. However, a major limitation faced by most of the methods developed so far is the resolution along the axial direction, which is still an order of magnitude worse than the maximum lateral resolution achievable. In this thesis, we present a new concept for measuring distances of single molecules from a metal surface with nanometer accuracy using the energy transfer from the excited molecules to the surface plasmons of a metal film, which we term single-molecule Metal-Induced Energy Transfer (smMIET). We perform the first proof of principle experiments on single dye molecules and demonstrate an axial localization with nanometer accuracy. Here, we build the theoretical outline for the description of smMIET, and throw light on the potential for its application in structural biology. Apart from this, in this thesis, we present the first experimental approach to determine simultaneously the three-dimensional excitation and emission dipole geometry of individual emitters. Here, we use defocused imaging in conjugation with radially polarized excitation scanning to characterize the emission and excitation transition probabilities. We demonstrate this approach on two commercially available dye molecules and obtain the distributions of the angle between their excitation and emission transition dipoles. This experimental tool can be used for elucidating more complex excitation/ emission geometries, such as those found in fluorescent nano-crystals (quantum dots) and also for verifying the quantum chemical calculations that are used for predicting the structure and geometry of the molecular orbitals involved in an electronic transition.
Keywords: Superresolution microscopy; Axial resolution; Förster Resonance Energy Transfer; Electrodynamics; Transition dipole; Pattern matching; Fluorescence Lifetime Imaging Microscopy; Defocused Imaging; Single-molecule imaging; Single-molecule localization microscopy