Spectral imaging in preclinical research and clinical pathology.

Spectral imaging methods are attracting increased interest from researchers and practitioners in basic science, pre-clinical and clinical arenas. A combination of better labeling reagents and better optics creates opportunities to detect and measure multiple parameters at the molecular and cellular level. These tools can provide valuable insights into the basic mechanisms of life, and yield diagnostic and prognostic information for clinical applications. There are many multispectral technologies available, each with its own advantages and limitations. This chapter will present an overview of the rationale for spectral imaging, and discuss the hardware, software and sample labeling strategies that can optimize its usefulness in clinical settings

Multimodal Multispectral Optical Endoscopic Imaging for Biomedical Applications

Optical imaging is an emerging field of clinical diagnostics that can address the growing medical need for early cancer detection and diagnosis. Various human cancers are amenable to better prognosis and patient survival if found and treated during early disease onset. Besides providing wide-field, macroscopic diagnostic information similar to existing clinical imaging techniques, optical imaging modalities have the added advantage of microscopic, high resolution cellular-level imaging from in vivo tissues in real time. This comprehensive imaging approach to cancer detection and the possibility of performing an ‘optical biopsy’ without tissue removal has led to growing interest in the field with numerous techniques under investigation. Three optical techniques are discussed in this thesis, namely multispectral fluorescence imaging (MFI), hyperspectral reflectance imaging (HRI) and fluorescence confocal endomicroscopy (FCE). MFI and HRI are novel endoscopic imaging-based extensions of single point detection techniques, such as laser induced fluorescence spectroscopy and diffuse reflectance spectroscopy. This results in the acquisition of spectral data in an intuitive imaging format that allows for quantitative evaluation of tissue disease states. We demonstrate MFI and HRI on fluorophores, tissue phantoms and ex vivo tissues and present the results as an RGB colour image for more intuitive assessment. This follows dimensionality reduction of the acquired spectral data with a fixed-reference isomap diagnostic algorithm to extract only the most meaningful data parameters. FCE is a probe-based point imaging technique offering confocal detection in vivo with almost histology-grade images. We perform FCE imaging on chemotherapy-treated in vitro human ovarian cancer cells, ex vivo human cancer tissues and photosensitiser-treated in vivo murine tumours to show the enhanced detection capabilities of the technique. Finally, the three modalities are applied in combination to demonstrate an optical viewfinder approach as a possible minimally-invasive imaging method for early cancer detection and diagnosis

Dynamic Hyperspectral and Polarized Endoscopic Imaging

The health of rich, developed nations has seen drastic improvement in the last two centuries. For it to continue improving at a similar rate new or improved diagnostic and treatment technologies are required, especially for those diseases such as cancer which are forecast to constitute the majority of disease burden in the future. Optical techniques such as microscopy have long played their part in the diagnostic process. However there are several new biophotonic modalities that aim to exploit various interactions between light and tissue to provide enhanced diagnostic information. Many of these show promise in a laboratory setting but few have progressed to a clinical setting. We have designed and constructed a flexible, multi-modal, multi-spectral laparoscopic imaging system that could be used to demonstrate several different techniques in a clinical setting. The core of this system is a dynamic hyperspectral illumination system based around a supercontinuum laser and Digital Micromirror Device that can provide specified excitation light in the visible and near infra-red ranges. This is a powerful tool for spectroscopic techniques as it is not limited to interrogating a fixed range of wavelengths and can switch between excitation bands instantaneously. The excitation spectra can be customised to match particular fluorophores or absorption features, introducing new possibilities for spectral imaging. A standard 10 mm diameter rigid endoscope was incorporated into the system to reduce cost and demonstrate compatibility with existing equipment. The polarization properties of two commercial endoscopes were characterised and found to be unsuited to current polarization imaging techniques as birefringent materials used in their construction introduce complex, spatially dependent transformations of the polarization state. Preliminary exemplar data from phantoms and ex vivo tissue was collected and the feasibility and accuracy of different analysis techniques demonstrated including multiple class classification algorithms. Finally, a novel visualisation method was implemented in order to display the complex hyperspectral data sets in a meaningful and intuitive way to the user