Development of a time gated, compressive detection Raman instrument for effective fluorescence rejection and multilayer analysis

In this thesis we have developed and implemented a Raman instrument of a novel design that uses a combination of state-of-the-art technologies to provide a temporal dimension in the measured spectrum with the goal of effective fluorescence suppression. Combining high repetition rate, picosecond laser pulses and high temporal resolution detection systems with MEMs devices we developed a time gated Raman instrument that is capable of effective fluorescence suppression and producing time-gated Raman maps thanks to the improved signal achievable through compressive detection. Time gated Raman spectroscopy was then investigated for pigmented samples of relevance to cultural heritage and to biology. The instrument was able to recover Raman spectral information of highly fluorescent samples where a typical CCD based Raman instrument with CW laser typically fails. Multilayer samples were measured, and 3D mapping performed. By utilizing multiplexing, depth analysis was measured and calculated via photon time of flight. The design of the instrument was optimized to overcome some of the issues that plague current time gated Raman techniques (portability, low fill factors, temporal resolution). The system uses a single element single photon avalanche diode as the detector to provide the best possible temporal resolution and a digital micromirror device (DMD) as the wavelength selective component to provide a high fill factor for maximum throughput. The DMD allows multiple spectral features to be directed to the detector at any one time (multiplexed compressive detection). The increase in signal from multiplexing allows time gated Raman maps to be performed on a time scale comparable to conventional Raman instruments (seconds per pixel). This study demonstrates the feasibility of time gated Raman mapping for a range of applications

Development of a time gated, compressive detection Raman instrument for effective fluorescence rejection and multilayer analysis

In this thesis we have developed and implemented a Raman instrument of a novel design that uses a combination of state-of-the-art technologies to provide a temporal dimension in the measured spectrum with the goal of effective fluorescence suppression. Combining high repetition rate, picosecond laser pulses and high temporal resolution detection systems with MEMs devices we developed a time gated Raman instrument that is capable of effective fluorescence suppression and producing time-gated Raman maps thanks to the improved signal achievable through compressive detection. Time gated Raman spectroscopy was then investigated for pigmented samples of relevance to cultural heritage and to biology. The instrument was able to recover Raman spectral information of highly fluorescent samples where a typical CCD based Raman instrument with CW laser typically fails. Multilayer samples were measured, and 3D mapping performed. By utilizing multiplexing, depth analysis was measured and calculated via photon time of flight. The design of the instrument was optimized to overcome some of the issues that plague current time gated Raman techniques (portability, low fill factors, temporal resolution). The system uses a single element single photon avalanche diode as the detector to provide the best possible temporal resolution and a digital micromirror device (DMD) as the wavelength selective component to provide a high fill factor for maximum throughput. The DMD allows multiple spectral features to be directed to the detector at any one time (multiplexed compressive detection). The increase in signal from multiplexing allows time gated Raman maps to be performed on a time scale comparable to conventional Raman instruments (seconds per pixel). This study demonstrates the feasibility of time gated Raman mapping for a range of applications

Feasibility of integrated high‐wavenumber Raman imaging and fingerprint Raman spectroscopy for fast margin assessment in breast cancer surgery

Intraoperative assessment of surgical margins remains one of the main challenges in cancer surgery. Raman spectroscopy can detect cancer cells with high accuracy, but it is time‐consuming. In this paper, we investigated a selective‐sampling Raman spectroscopy approach, based on high wavenumber (HW) Raman imaging (spectral range 2,500–3,500 cm−1) and fingerprint Raman spectroscopy (spectral range 600–1,800 cm−1), to reduce the overall tissue analysis time while maintaining high diagnostic accuracy. HW Raman mapping was used as a first step to identify the adipose tissue regions based on the C–H stretching bands at 2,700–2,950 cm−1. As residual tumors are typically found in nonadipose tissue, an algorithm was developed to allocate sampling points for fingerprint Raman spectroscopy at locations corresponding to low intensity in the HW‐Raman maps. Preliminary results show that HW‐Raman imaging based on a 671 nm laser is effective and fast for mapping of adipose tissue in breast resections, with typical imaging times of 2 min for tissue areas as large as 2 × 2 cm2 areas. Albeit the remaining high fluorescence background in the fingerprint region prevents the use of single 671‐nm laser, the HW Raman imaging can be still exploited in combination with 785‐nm excitation Raman spectroscopy for identifying residual tumor. Although this study demonstrates the feasibility of this approach, further improvements, such as using single element detectors for HW Raman imaging, are required to increase the analysis speed further towards intraoperative use in the routine clinical setting

Time-gated Raman spectroscopy for biomedical application under ambient or strong background light conditions

Many biomedical applications require measurements of Raman spectra of tissue under ambient lighting conditions. However, the background light often swamps the weaker Raman signal. The use of time-gated (TG) Raman spectroscopy based on a single photon avalanche diode (SPAD) operating in time-correlated single photon counting and near-infrared laser excitation was investigated for acquisition of Raman spectra and spectral images of biological tissue. The results obtained using animal tissue samples (adipose tissue and muscle) show that the time gating modality enables measurement of Raman spectra under background light conditions of similar quality as conventional continuous wave Raman spectroscopy in the absence of background light. Optimal suppression of the background light was observed for time gate widths of 300–1000 ps. The results also showed that TG Raman spectroscopy was able to detect subtle spectral differences required for medical diagnostics, such as differences in Raman spectra of cancer and normal tissue. While the current instrument required scanning of the grating in order to obtain full Raman spectra, leading to impractical times for multi-wavenumber Raman mapping, imaging time could be drastically reduced by spectral multiplexing (compressed detection) using digital micromirror devices or by using SPAD arrays

Fast Raman spectral mapping of highly fluorescing samples by time-gated spectral multiplexed detection

We present a time-gated Raman micro-spectroscopy technique suitable for fast Raman mapping of samples eliciting large laser-induced fluorescence backgrounds. To achieve the required time resolution for effective fluorescence rejection, a picosecond pulsed laser and a single-photon avalanche diode were used. A module consisting of a spectrometer, digital micromirror device, and two prisms was used for high-resolution spectral filtering and multiplexing, which is required for a high chemical specificity and short integration times. With this instrument, we demonstrated time-gated Raman imaging of highly fluorescent samples, achieving acquisition times as short as 3 min for 40×40 pixel resolution images

Sub-Surface Molecular Analysis and Imaging in Turbid Media Using Time-Gated Raman Spectral Multiplexing

Obtaining molecular information deeper within optically turbid samples is valuable in many applications. However, in many cases this is challenging, in particular when the sample elicits strong laser-induced fluorescence emission. Here, we investigated the use of time-gated and micro-spatially offset Raman spectroscopy (micro-SORS) based on spectral multiplexing detection to obtain sub-surface molecular analysis and imaging for both fluorescing and non-fluorescing samples. The multiplexed spectral detection achieved with a digital micromirror device (DMD) allowed fast acquisition of the time-gated signals to enable three-dimensional Raman mapping (raster scanning in the lateral x,y plane and using time-of-flight calibration for the axial z-direction). Sub-millimeter resolution molecular depth mapping was achieved with dwell times on the order of seconds per pixel. To suppress fluorescence backgrounds and enhance Raman bands, time-gated Raman spectroscopy was combined with micro-SORS to recover Raman signals of red pigments placed behind a layer of optically turbid material. Using a defocusing micro-SORS approach, both fluorescence and Raman signals from the surface layers were further suppressed, which enhanced the Raman signals from the deeper sublayers containing the pigment. These results demonstrate that time-gated Raman spectroscopy based on spectral multiplexed detection, and in combination with micro-SORS, is a powerful technique for sub-surface molecular analysis and imaging, which may find practical applications in medical imaging, cultural heritage, forensics, and industry

In-situ polymerisation of fully bioresorbable polycaprolactone/phosphate glass fibre composites: in vitro degradation and mechanical properties

Fully bioresorbable composites have been investigated in order to replace metal implant plates used for hard tissue repair. Retention of the composite mechanical properties within a physiological environment has been shown to be significantly affected due to loss of the integrity of the fibre/matrix interface. This study investigated phosphate based glass fibre (PGF) reinforced polycaprolactone (PCL) composites with 20%, 35% and 50% fibre volume fractions (Vf) manufactured via an in-situ polymerisation (ISP) process and a conventional laminate stacking (LS) followed by compression moulding. Reinforcing efficiency between the LS and ISP manufacturing process was compared, and the ISP composites revealed significant improvements in mechanical properties when compared to LS composites. The degradation profiles and mechanical properties were monitored in phosphate buffered saline (PBS) at 37°C for 28 days. ISP composites revealed significantly less media uptake and mass loss (p<0.001) throughout the degradation period. The initial flexural properties of ISP composites were substantially higher (p<0.0001) than those of the LS composites, which showed that the ISP manufacturing process provided a significantly enhanced reinforcement effect than the LS process. During the degradation study, statistically higher flexural property retention profiles were also seen for the ISP composites compared to LS composites. SEM micrographs of fracture surfaces for the LS composites revealed dry fibre bundles and poor fibre dispersion with polymer rich zones, which indicated poor interfacial bonding, distribution and adhesion. In contrast, evenly distributed fibres without dry fibre bundles or polymer rich zones, were clearly observed for the ISP composite samples, which showed that a superior fibre/matrix interface was achieved with highly improved adhesion

Critical moments? Life transitions and energy biographies

Family and youth research has highlighted the importance of lifecourse transitions, illustrating how they can have a substantial impact on people’s everyday lives and anticipated futures. Given their apparent significance, it is surprising that relatively little attention has been paid to life transitions – particularly unexpected ones – to explore how they can impact upon everyday energy use. This is a central concern of Energy Biographies project. The project’s qualitative longitudinal design makes an original contribution, affording a detailed view of how transitions unfold and their significance for energy demand and environmental action. Central to elucidating these issues is the concept of ‘linked lives’, recognising that people live interdependently. In this paper, we explore the accounts of three participants who experienced one or more life transitions during the course of the project, in order to consider the impacts of these events (both planned and unanticipated) on their everyday energy use and environmental actions as part of their linked lives with others

Evaluating the Effects of SARS-CoV-2 Spike Mutation D614G on Transmissibility and Pathogenicity.

Global dispersal and increasing frequency of the SARS-CoV-2 spike protein variant D614G are suggestive of a selective advantage but may also be due to a random founder effect. We investigate the hypothesis for positive selection of spike D614G in the United Kingdom using more than 25,000 whole genome SARS-CoV-2 sequences. Despite the availability of a large dataset, well represented by both spike 614 variants, not all approaches showed a conclusive signal of positive selection. Population genetic analysis indicates that 614G increases in frequency relative to 614D in a manner consistent with a selective advantage. We do not find any indication that patients infected with the spike 614G variant have higher COVID-19 mortality or clinical severity, but 614G is associated with higher viral load and younger age of patients. Significant differences in growth and size of 614G phylogenetic clusters indicate a need for continued study of this variant

Exponential growth, high prevalence of SARS-CoV-2, and vaccine effectiveness associated with the Delta variant

SARS-CoV-2 infections were rising during early summer 2021 in many countries associated with the Delta variant. We assessed RT-PCR swab-positivity in the REal-time Assessment of Community Transmission-1 (REACT-1) study in England. We observed sustained exponential growth with average doubling time (June-July 2021) of 25 days driven by complete replacement of Alpha variant by Delta, and by high prevalence at younger less-vaccinated ages. Unvaccinated people were three times more likely than double-vaccinated people to test positive. However, after adjusting for age and other variables, vaccine effectiveness for double-vaccinated people was estimated at between ~50% and ~60% during this period in England. Increased social mixing in the presence of Delta had the potential to generate sustained growth in infections, even at high levels of vaccination