Optical Imaging of the Nanoscale Structure and Dynamics of Biological Membranes

Biological membranes serve as the fundamental unit of life, allowing the compartmentalization of cellular contents into subunits with specific functions. The bilayer structure, consisting of lipids, proteins, small molecules, and sugars, also serves many other complex functions in addition to maintaining the relative stability of the inner compartments. Signal transduction, regulation of solute exchange, active transport, and energy transduction through ion gradients all take place at biological membranes, primarily with the assistance of membrane proteins. For these functions, membrane structure is often critical. The fluid-mosaic model introduced by Singer and Nicolson in 1972 evokes the dynamic and fluid nature of biological membranes.(1) According to this model, integral and peripheral proteins are oriented in a viscous phospholipid bilayer. Both proteins and lipids can diffuse laterally through the two-dimensional structure. Modern experimental evidence has shown, however, that the structure of the membrane is considerably more complex; various domains in the biological membranes, such as lipid rafts and confinement regions, form a more complicated molecular organization. The proper organization and dynamics of the membrane components are critical for the function of the entire cell. For example, cell signaling is often initiated at biological membranes and requires receptors to diffuse and assemble into complexes and clusters, and the resulting downstream events have consequences throughout the cell. Revealing the molecular level details of these signaling events is the foundation to understanding numerous unsolved questions regarding cellular life

Synthesis of germanium nanocrystals from solid-state disproportionation of a chloride-derived germania glass

Germanium nanocrystals (Ge NCs) have potential to be used in several optoelectronic applications such as photodetectors and light-emitting diodes. Here, we report a solid-state route to synthesizing Ge NCs through thermal disproportionation of a germania (GeOX) glass, which was synthesized by hydrolyzing a GeCl2·dioxane complex. The GeOX glass synthesized in this manner was found to have residual Cl content. The process of nanocrystal nucleation and growth was monitored using powder X-ray diffraction, transmission electron microscopy, X-ray photoelectron spectroscopy and Raman spectroscopy. Compared to existing solid-state routes for synthesizing colloidal Ge NCs, this approach requires fewer steps and is amenable to scaling to large-scale reactions

Directional Raman spectroscopy: A surface-sensitive tool for measuring the chemical, optical, and physical properties of adsorbates on metal surfaces

There is a need for sensitive methods to analyze thin films of polymers, biological cells, dielectric waveguides, and self-assembled monolayers. In this dissertation, we discuss a newly-developed instrument with combined benefits of surface plasmon resonance, plasmon waveguide resonance, and Raman spectroscopy for collecting the chemical information of adsorbates with monolayer sensitivity. Additionally, the instrument is applicable for measuring angle-dependent molecular interactions. Directional-surface-plasmon-coupled Raman spectroscopy (i.e., directional Raman scattering) is a viable non-destructive method equivalent to total internal reflection Raman spectroscopy using a smooth metal film. The excitation of surface plasmons produces directional Raman scattering in the plane of the metal film (in-coupling) and the emission of the scattered light through a Weierstrass prism (out-coupling). A hollow cone of directional scattering at a sharply defined angle results in the surface-plasmon-polariton cone radiating from the Weierstrass prism. The directionality of the signal, as well as the enhanced electric field, produces relatively large Raman signals at a smooth metal interface, without the use of surface-enhanced Raman substrates. The electric field intensity is amplified by 20-fold due to the directional emission of the scattered light and the collection of the entire surface-plasmon-polariton cone. The directional Raman spectrometer has the capability of measuring the full surface-plasmon-polariton cone image, cone intensity, and directional Raman scattering radiating from the cone as a function of the incident angle. On the same instrument, the Kretschmann and reverse-Kretschmann configurations can provide multimodal spectral data (e.g., thickness and refractive indices) collection. The directional Raman spectrometer utilizes translational stages (as opposed to rotational stages, commonly used in surface plasmon resonance sensing). The instrument design provides faster acquisition times and precise control of the light incident on the prism interface with 0.06° angle resolution. We can quantify the surface-plasmon-polariton cone properties and intensity from the digitized surface-plasmon-polariton cone image by extracting the cone diameters from the cone angles. The calculated cone parameters are obtained using three-dimensional finite-difference time-domain simulations of the far-field angular radiation pattern in combination with Fresnel reflectivity calculations. The approach has equivalent sensitivity to alternative methods used to collect surface plasmon resonance and plasmon waveguide resonance data. Further, we can simultaneously measure the adsorption and chemical identification of thin films, waveguides, and self-assembled monolayers. The sensitivity of all the waveguide-coupled surface-plasmon-polariton cone modes is between 0.009 and 0.02° nm-1. The incident angles that produce the surface-plasmon-polariton cones and the surface-plasmon-polariton cone angles are linearly dependent; therefore, it is straightforward to determine the optimum incident angle for collecting directional Raman scattering. According, the acquisition time is reduced for collecting plasmon waveguide resonance data. The thickness and chemical composition for thin films, as well as the structure and orientation of guided modes in waveguides, can be obtained in our multi-detection directional Raman scattering instrument. Directional Raman spectroscopy can be applied to study photovoltaic thin films, polymer brushes, energy harvesting devices, optoelectronics, and sensor readout devices where the chemical composition, orientation, and morphology are essential to their function. This spectroscopic technique will propel new and emerging technologies in which functionalization of a surface is required.</p

Optical Imaging of the Nanoscale Structure and Dynamics of Biological Membranes

Biological membranes serve as the fundamental unit of life, allowing the compartmentalization of cellular contents into subunits with specific functions. The bilayer structure, consisting of lipids, proteins, small molecules, and sugars, also serves many other complex functions in addition to maintaining the relative stability of the inner compartments. Signal transduction, regulation of solute exchange, active transport, and energy transduction through ion gradients all take place at biological membranes, primarily with the assistance of membrane proteins. For these functions, membrane structure is often critical. The fluid-mosaic model introduced by Singer and Nicolson in 1972 evokes the dynamic and fluid nature of biological membranes.(1) According to this model, integral and peripheral proteins are oriented in a viscous phospholipid bilayer. Both proteins and lipids can diffuse laterally through the two-dimensional structure. Modern experimental evidence has shown, however, that the structure of the membrane is considerably more complex; various domains in the biological membranes, such as lipid rafts and confinement regions, form a more complicated molecular organization. The proper organization and dynamics of the membrane components are critical for the function of the entire cell. For example, cell signaling is often initiated at biological membranes and requires receptors to diffuse and assemble into complexes and clusters, and the resulting downstream events have consequences throughout the cell. Revealing the molecular level details of these signaling events is the foundation to understanding numerous unsolved questions regarding cellular life.This document is the unedited Author’s version of a Submitted Work that was subsequently accepted for publication in Analytical Chemistry, copyright © American Chemical Society after peer review. To access the final edited and published work see DOI:10.1021/acs.analchem.8b04755. Posted with permission.</p

Combined Measurement of Directional Raman Scattering and Surface-Plasmon-Polariton Cone from Adsorbates on Smooth Planar Gold Surfaces

Directional-surface-plasmon-coupled Raman scattering (directional RS) has the combined benefits of surface plasmon resonance and Raman spectroscopy, and provides the ability to measure adsorption and monolayer-sensitive chemical information. Directional RS is performed by optically coupling a 50-nm gold film to a Weierstrass prism in the Kretschmann configuration and scanning the angle of the incident laser under total internal reflection. The collected parameters on the prism side of the interface include a full surface-plasmon-polariton cone and the full Raman signal radiating from the cone as a function of incident angle. An instrument for performing directional RS and a quantitative study of the instrumental parameters are herein reported. To test the sensitivity and quantify the instrument parameters, self-assembled monolayers and 10 to 100-nm polymer films are studied. The signals are found to be well-modeled by two calculated angle-dependent parameters: three-dimensional finite-difference time-domain calculations of the electric field generated in the sample layer and projected to the far-field, and Fresnel calculations of the reflected light intensity. This is the first report of the quantitative study of the full surface-plasmon-polariton cone intensity, cone diameter, and directional Raman signal as a function of incident angle. We propose that directional RS is a viable alternative to surface plasmon resonance when added chemical information is beneficial.</p