Thesis (Ph.D.)--Boston University / The large electromagnetic field enhancement provided by nanostructured noble metal surfaces forms the foundation for a series of enabling optical analytical techniques, such as surface enhanced Raman spectroscopy (SERS), surface enhanced IR absorption spectroscopy (SEIRA), surface enhanced fluorescent microscopy (SEF), to name only a few. Critical sensing applications have, however, other substrate requirements than mere peak signal enhancement. The substrate needs to be reliable, provide reproducible signal enhancements, and be amenable to a combination with microfluidic chips or other integrated sensor platforms. These needs motivate the development of engineerable SERS substrate "chips" with defined near- and far-field responses. In this dissertation, two types of rationally designed SERS substrates - nanoparticle cluster arrays (NCAs) and SERS stamp - will be introduced and characterized. NCAs were fabricated through a newly developed template guided self-assembly fabrication approach, in which chemically synthesized nanoparticles are integrated into predefined patterns using a hybrid top-down/bottom-up approach. Since this method relies on chemically defined building blocks, it can overcome the resolution limit of conventional lithographical methods and facilitates higher structural complexity. NCAs sustain near-field interactions within individual clusters as well as between entire neighboring clusters and create a multi-scale cascaded E-field enhancement throughout the entire array. SERS stamps were generated using an oblique angle metal deposition on a lithographically defined piston. When mounted on a nanopositioning stage, the SERS stamps were enabled to contact biological surfaces with pristine nanostructured metal surfaces for a label-free spectroscopic characterization. The developed engineered substrates were applied and tested in critical sensing applications, including the ultratrace detection of explosive vapors, the rapid discrimination of bacterial pathogens, and the label-free monitoring of the enzymatic degradation of pericellular matrices of cancer cells.

No description available.

The incidence of gastrointestinal (GI) cancers has been steadily increasing in the UK since the mid 1970’s. These include cancers of the colon and oesophagus. Colon cancers have a high incidence rate, being the fourth most common cancer in the UK for both men and women. Oesophageal cancers in comparison are much rarer, however they have a poor survival rate primarily due to a late diagnosis. The key to improving survival for these cancers and many others is to detect and remove the disease at the early stages, to prevent the cancer from advancing. At present the ‘gold standard’ for diagnosis is a biopsy followed by histopathology. This technique is invasive, time consuming and highly subjective. It is therefore important to look towards non-invasive methods for early and rapid diagnosis. Optical techniques have begun to show such promise. By probing the interactions of tissues with light, diagnostic information is able to be obtained non-invasively. Techniques such as Raman spectroscopy utilise inherent molecular vibrations to extract biochemical information from tissues. Raman spectroscopy, however, is currently fundamentally limited by long acquisition times, due to the inherently weak signals produced. Using coherent Raman techniques such as coherent anti-Stokes Raman scattering (CARS) and stimulated Raman scattering (SRS), the molecular vibrations are coherently driven to provide an enhancement in signal. This thesis explored spectral signatures from snap frozen oesophageal sections in the fingerprint (450 cm-1 to 1850 cm-1) and high wavenumber (2800 cm-1 to 3050 cm-1) regions using spontaneous Raman and compared with spectra from hyperspectral SRS. The diagnostic potential for each technique was assessed for four major pathology groups, normal, Barrett’s oesophagus, dysplasia and adenocarcinoma. Samples were classified using a principal component fed linear discriminant analysis (PCA-LDA) approach with a leave-one-out cross validation. Comparisons were made to haematoxylin and eosin (H&E) stained sections. Raman in the fingerprint region was found to be the most promising for diagnosis. There were minimal changes in the high wavenumber region between pathology groups which was also reflected in the SRS spectra and proved to be insufficient for classification. Further comparisons were made between spontaneous and coherent Raman techniques using frozen colon sections. The morphological and structural information available was explored using a k-means cluster analysis. Both spontaneous and coherent Raman were able to distinguish important structural features in the colon, such as the epithelial cells that form the colonic glands and surrounding connective tissue. Both are important visual markers for cancer diagnosis in the current approach. SRS demonstrated higher spatial resolution and faster acquisition times in comparison to spontaneous Raman. This work has discussed the many advantages of using coherent Raman techniques for tissue applications, but has also highlighted some of the limitations for spectral measurements, arising from the complexity of the system.

Non-linear optical processes such as coherent anti-Stokes Raman scattering and sum frequency generation offer a view into the chemical and biological interactions of molecules that is distinctly different from linear techniques like near infra-red and fluorescence. This insight comes at a cost: non-linear techniques are more sensitive to external perturbations of the system, increasing the noise and decreasing the repeatability of the data. We work here on both aspects of these non-linear techniques, taking advantage of their power to offer new imaging techniques as well as working to quantify and reduce the non-resonant noise inherent to the system. In pursuit of the first part, we look at formalin fixed paraffin embedded tissue samples. This is the most common form of tissue storage in the world. However, the paraffin renders them unavailable for spectroscopic study. We introduce a new technique, combination coherent anti-Stokes Raman scattering microscopy and sum frequency generation microscopy, to avoid the issue of paraffin signal contamination. This high resolution, widefield technique allows for the separate identification of paraffin and the tissue embedded within it. We show in this work the capability of this technique to enable high throughput automated detection of osteoporosis in mice. In pursuit of the second part, we demonstrate experimentally for the first time, deferred build up in coherent anti-Stokes Raman scattering. We show that coherent anti-Stokes Raman scattering signal is maximized when the probe pulse is delayed by an amount dependent on the probe width and the material itself. Non-resonant contamination, however, is maximized when the probe delay is zero, meaning that it is possible to decrease the non-resonant noise while increasing the desired signal. We also show that the dephasing time is inversely dependent on the probe width, so narrower probe pulses allow for further delayed probe pulses, which in turn decrease non-resonant noise more. We demonstrate this technique by looking at the effects of hydrogen bonding in pyridine-water complexes.

Redox regulation and homeostasis are critically important in the regulation of cell function; however, there are significant challenges in quantitatively measuring and monitoring intracellular redox potentials. The work in this thesis details a novel approach to intracellular redox monitoring. The approach is based on the use of nanosensors, which comprise molecules capable of sensing the local redox potential, assembled on gold nanoshells. Since the Raman spectra of the sensor molecules change depending on their oxidation state, and since the nanoshells allow a large enhancement of the Raman scattering, intracellular potential can be calculated by simple optical measurements. A full description of the design, fabrication and characterisation (spectroscopic and electrochemical) of the nanosensors is provided within. The ability to deliver nanosensors into cells in a controllable fashion was confirmed using electron microscopy. Results from a range of assays are also presented which reveal that introduction of nanosensors does not result in any cytotoxicity. Sensor utility in monitoring redox potentials as cells responded to physiological and superphysiological oxidative and reductive stimuli was investigated. Importantly, the capability of the nanosensors in monitoring intracellular potentials in a reversible, non-invasive manner, and over a previously unattainable potential range, is demonstrated.

This thesis explores the generation and the coherence properties of Raman frequency combs that are initiated from vacuum fluctuations using hydrogen-filled hollow-core photonic crystal fibre (HC-PCF). The motivation is to explore a novel route for generating attosecond pulses and waveform synthesis. To this end, work has been undertaken in the design and fabrication of HC-PCF, in the generation of Raman comb with a compact set-up and finally in an experimental demonstration of the mutual coherence between the comb spectral components. Here, the well-established photonic bandgap (PBG) HC-PCF is further developed. Surface mode spectral positions are controlled by chemical etching technique, and a single piece of fibre with two robust bandgaps is fabricated. Furthermore, the second established class of HC-PCF; namely large-pitch Kagome-lattice HC-PCF, has experienced challenging developments. This led to the fabrication of a hypocycloid-core seven-cell Kagome HC-PCF with comparable attenuation value to that of PBG HC-PCF while offering much larger bandwidth. Using the fabricated HC-PCF, different Raman frequency comb systems are developed. In addition to the previously-generated multi-octave Raman frequency comb from a large 1064 nm Nd:YAG Q-switch laser, several more compact version of Raman comb sources have been developed, including one whose lines lay in the visible and UV for applications in forensics and biomedicine. The Raman frequency comb generated inside a length of hydrogen-filled HC-PCF is further investigated by studying the coherence of the Raman lines. Despite of vacuum-fluctuation-initiation, it is demonstrated that the comb has self- and mutualcoherence properties within each single shot, bringing thus the possibility of generating attosecond pulses with non-classical properties.

Cellular redox potential is incredibly important for the control and regulation of a vast number of processes occurring in cells. Disruption of the fine redox balance within cells is has been associated with disease. Of particular interest to my research is the redox gradient that develops in cancer tumours, in which the internal regions are further from vascular blood supply and therefore become starved of oxygen and hypoxic. This makes treatment of these areas a lot more challenging, as radiotherapy approaches rely on the presence of oxygen and, with a poor vascular blood supply, drugs delivered through the blood stream will have poor access to these regions. Currently, there is limited knowledge regarding the quantitative nature of this redox gradient in cancerous tumours. To aid the development of drugs and therapies to overcome this problem, a system that enables quantitative mapping of redox potential through a tumour would be a vital tool. In this work redox sensitive molecules attached to gold nanoparticles (NPs) are delivered to cells and give signals using surface enhanced Raman scattering (SERS). Redox potential changes are monitored quantitatively by ratiometric changes in signal intensity of selected signals in the SER spectra acquired. Multicellular tumour spheroids (MTS) are used as a three dimensional (3D) in vitro tumour model, in which the 3D architecture and gradients observed in tumours in vivo develop. As redox potential is pH dependent and pH is another important physiological characteristic in its own right, a SERS pH sensor was developed and ultimately a system that multiplexes intracellular pH and redox measurement by SERS. Initially, simultaneous redox potential and pH measurements were performed in monolayer culture before extending this to MTS. Photothermal optical coherence tomography (OCT) was used to investigate overall 3D NP distribution in the MTS models. It was possible to control NP delivery to MTS to localise NPs to various regions. Redox potential and pH could then be measured using a fibre optic Raman probe, and spatial response to drug treatment monitored. Intracellular NP localisation was investigated using transmission electron microscopy (TEM), scanning electron microscopy (SEM), helium ion microscopy (HIM) and confocal fluorescence microscopy (CFM) and attempts were made to control NP delivery to particular intracellular compartments.

QC 20100628

The armchair variety of single-wall carbon nanotubes (SWCNTs) is the only nanotube species that behaves as a metal with no electronic band gap and massless carriers, making them ideally suited to probe fundamental questions of many-body physics of one-dimensional conductors as well as to serve in applications such as high-current power transmission cables. However, current methods of nanotube synthesis produce bulk material comprising of a mixture of nanotube lengths, diameters, wrapping angles, and electronic types due to the inability to control the growth process at the nanometer level. As a result, measurements of as-grown SWCNTs produce a superposition of electrical and optical responses from multiple SWCNT species. This thesis demonstrates production of aqueous suspensions composed almost entirely of armchair SWCNTs using a post-synthesis separation method employing density gradient ultracentrifugation (DGU) to separate different SWCNT types based on their mass density and surfactant-specific interactions. Resonant Raman spectroscopy determines the relative abundances of each nanotube species, before and after DGU, by measuring the integrated intensity of the radial breathing mode, the diameter-dependent radial vibration of the SWCNT perpendicular to its main axis, and quantifies the degree of enrichment of bulk nanotube samples to exclusively armchair tubes. Raman spectroscopy of armchair-enriched samples of the G-band mode, which is composed of longitudinal (G-) and circumferential (G+) vibrations oscillating parallel and perpendicular to the tube axis, shows that the G- peak, long-held to be an indicator for the presence of metallic SWCNTs, appears only when electronic resonance with narrow-gap semiconducting SWCNTs occurs and shows only the G+ component in spectra containing only armchair species. Finally, by combining optical absorption measurements with nanotube composition as determined earlier via Raman scattering, peak fitting of absorption spectra indicates that interband transitions of armchair SWCNTs are strongly excitonic as shown by the highly symmetric peak lineshapes, a property normally attributed to semiconductors. Such lineshapes allow classification of armchair SWCNTs as a unique hybrid class of optical nanomaterial. Combining absorption and Raman scattering measurements establishes a distinct optical signature that describes the fundamental optical processes within armchair SWCNTs and lays the foundation for future studies of many-body photophysics and electrical applications.

Surface-enhanced Raman scattering (SERS) effect on Ag films with different morphology is studied. We varied the deposition rates and also proposed a new method to control the nano-gaps on the silver island film. By bending the glass substrates during film deposition, we can control the gap width on the fractal Ag film. The measured SERS intensity is related to the metal film morphology and we found that the gap width is the dominant factor to analyze the SERS signal. The 3-layer metal-insulator-metal structure is simulated and the E-field intensity with different gaps fits to our measurement results.