<p>The visible wavelength range has proven to be a useful spectral window for observing biophotonic events such as absorption in materials (oxy-hemoglobin and deoxy-hemoglobin), light scattering in biological tissue, and biochemical and fluorescence reactions. Diffuse reflectance spectroscopy (DRS) is a technique that utilizes the diffuse reflectance spectra from turbid media (e.g. biological tissue) to quantify the optical properties (e.g. absorption and scattering) of those media. DRS in the visible wavelength range can be utilized to optically differentiate between healthy and cancerous tissue, and thus has applications in intra-operative tumor margin assessment. </p><p>The footprint of conventional DRS systems used for intra-operative tissue margin assessment prohibits their widespread use inside the surgical suite, where space is at a premium. Conventional quantitative DRS imaging systems utilize unwieldy fiber probes, cooled CCD cameras, and imaging spectrographs for imaging tissue margins. These system components not only increase system size, limiting their use inside the surgical suite, but also limit imaging resolution, imaging speed, and increase overall system cost. </p><p>Silicon is an attractive candidate for the development of compact, customized photodetector elements for biophotonic imaging applications such as intra-operative tumor margin assessment using DRS. This thesis deals with the design and development of a customized DRS imaging probe composed of custom silicon imaging arrays for intra-operative breast tumor margin assessment. The first generation of the customized imaging probe consisted of a 4x4 array of annular epitaxial Si pn junction photodiodes (PDs) with a measured responsivity of 0.28 A/W - 0.37 A/W for &#955;= 470 nm - 600 nm, and a measured dark current density of 1.456 nA/cm2 - 4.48 nA/cm2. The imaging array was used to detect diffuse reflectance when placed in direct contact with tissue. A quartz light delivery tube coupled to a xenon lamp was optimized to deliver light to the tissue through the holes of the annular imaging array across a 256 mm2 imaging area. The pixel-to-pixel spacing in the imaging array was 4.5 mm, the highest resolution reported to date for a multi-pixel DRS probe. This resolution was limited by pixel-to-pixel optical crosstalk, which was theoretically calculated and experimentally characterized, to validate the theoretical model for future designs. This first generation probe was successfully tested on diffuse reflectance standards, tissue-mimicking phantoms, animal tissue, and human breast tissue, and yielded an SNR of 30 dB - 55 dB on all measured specimens. </p><p>The next generation of the customized imaging probe consisted of a 4x4 array of annular thin-film Si pn junction PDs heterogeneously bonded to a transparent Pyrex substrate, to enable integration with a guided wave light delivery system. The 4x4 thin-film PD array design and development was prototyped using a 1x2 thin-film PD array heterogeneously bonded to a Pyrex substrate. The responsivity and dark current of the thin-film PDs in the 1x2 array were measured to be 0.19 A/W - 0.34 A/W for &#955;= 470 nm - 600 nm and 0.63 nA/cm2, respectively. The process for the 1x2 thin-film PD array was scaled to fabricate a 4x4 array of thin-film PDs for DRS, and the 4x4 array was optically and electrically characterized. These heterogeneously bonded thin-film single crystal Si PDs have the highest uncooled responsivity to dark current density ratio (greater than 0.30 - 0.54 cm2/nW for &#955;= 470 nm - 600 nm) reported to date, to the best of our knowledge. The 1x2 array of thin-film PDs were also heterogeneously bonded to a flexible substrate without any degradation in PD optical and electrical characteristics, opening the door towards conformal tissue imaging.</p> / Dissertation

<p> The rapid development of the use of lasers in therapeutic and diagnostic medicine in the past few years has generated interest in measuring the optical properties of tissue. In particular, the development of photodynamic therapy (PDT) has necessitated studies of the optical properties of tissue at wavelengths around 630nm, this being the wavelength at which the photosensitizer commonly used in PDT, namely dihematoporphyrin ether (DHE), is normally activated.</p> <p> The control of the volume of tissue from which information about the interaction coefficients of the tissue is obtained is an important problem in diffuse reflectance spectroscopy and other applications of light, because it is critical to understanding which tissue volumes are sampled by the injected photons that eventually are re-emitted. This report describes a simple model that predicts the parameters that control the volume of tissue interrogated by photons during reflectance spectroscopy.</p> <p> In optical fiber based diffuse reflectance spectroscopy, incident radiation is applied at one point on a tissue surface and collected at another point, a radial distance, r, away. Information about the light multiply scattered by the tissue is used to deduce optical scattering and absorption coefficients of the tissue. In this report both steady state and pulse techniques are studied. In the steady state method, the spatial dependence of the backscattered light is the measured quantity, while the pulse technique uses the temporal broadening of a picosecond (ps) pulse to determine the interaction coefficients.</p> <p> The relative contribution of a volume element of tissue to the observed signal depends on its location, the measurement geometry and the optical properties of the tissue. Knowledge of this dependence would allow some control of the volume interrogated by reflectance spectroscopy, and would provide insight into the influence of inhomogeneities.</p> <p> In the work reported here a simple diffusion model of light propagation in tissue based on the Boltzmann radiative transfer equation has been used to derive mathematical expressions for the relative time spent by photons in a given tissue volume element. Using optical interaction coefficients typical of mammalian soft tissues, results are presented for both steady state and pulse irradiation in both semi-infinite and infinite media.</p> <p> The residency time depth profile calculated by this model for index matched and zero fluence boundary conditions has the same shape as that predicted by Weiss (1989), who used a somewhat different model based on a 3-dimensional random walk theory. These profiles are characterized by a build-up region near the surface and exponential fall far away from the surface in the 'diffusion region'. The influence of the absorption coefficient μa and the fiber separation on the residency time as predicted by this model is in good agreement to that predicted by the random-walk theory (i.e the depth-profile of the residency time tends to sharpen as the absorption coefficient increases. This is attributed to the fact that long trajectories are less likely with large absorption probabilities. As well, the greater the fiber separation, the wider and flatter the depth distribution of the residency time. This is because photons that reach the surface at greater r values have, in general, migrated farther from the immediate vicinity of the source and detector and hence have sampled a larger volume of tissue). All the integrations in this report were performed numerically using the IMSL/LIB on the Microvax computer system in the Hamilton Regional Cancer Center. The adequacy of this numerical integration was tested and was found to be good.</p> <p> This model therefore suggests that during diffuse reflectance spectroscopy, the volume sampled by re-emitted photons can be controlled by changing parameters such as the fiber separation (in both steady state and time-resolved techniques) and the detection time (in the time-resolved method).</p> / Thesis / Master of Science (MSc)

This dissertation focuses on the development of computational models and algorithms related to diffuse reflectance spectroscopy. Specifically, this work aims to advance diffuse reflectance spectroscopy to a technique that is capable of measuring depth dependent properties in tissue. First, we introduce the Monte Carlo lookup table (MCLUT) method for extracting optical properties from diffuse reflectance spectra. Next, we extend this method to a two-layer tissue geometry so that it can extract depth dependent properties in tissue. We then develop a computational model that relates photon sampling depth to optical properties and probe geometry. This model can be used to aid in design of application specific diffuse reflectance probes. In order to provide justification for using a two-layer model for extracting tissue properties, we show that the use of a one-layer model can lead to significant errors in the extracted optical properties. Lastly, we use our two-layer MCLUT model and a probe that was designed based on our sampling depth model to extract tissue properties from the skin of 80 subjects at 5 anatomical locations. The results agree with previously published values for skin properties and show that can diffuse reflectance spectroscopy can be used to measured depth dependent properties in tissue. / text

Made available in DSpace on 2014-06-11T19:29:07Z (GMT). No. of bitstreams: 0 Previous issue date: 2011-01-31Bitstream added on 2014-06-13T20:18:54Z : No. of bitstreams: 1 gigante_ac_me_araiq.pdf: 810984 bytes, checksum: d628a07a7c532c812f60d96301f7afff (MD5) / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / O presente trabalho propõe o desenvolvimento de um método limpo para determinação de ureia em amostras comerciais de fertilizantes utilizando a Espectroscopia de Reflectância Difusa combinada com spot test. O método desenvolvido é baseado na reação entre a ureia e o reagente cromogênico p-dimetilaminocinamaldeído (p-DAC) em meio de ácido clorídrico diluído, que resulta em um produto de coloração rósea cujo valor máximo de reflectância ocorre no comprimento de onda de 535 nm. Os parâmetros experimentais foram otimizados através dos planejamentos fatorial e composto central para a obtenção da superfície de resposta, os quais indicaram uma maior sensibilidade para o método quando se utiliza o reagente p-DAC na concentração de 0,196% (m/v) em meio de ácido clorídrico de concentração 0,0549 mol L-1, empregando etanol como solvente para as soluções. O suporte sólido para a reação é um papel de filtro qualitativo onde são colocados 20,0μL de solução de ureia seguidos de secagem com ar frio de um secador de cabelos e 20,0μL de solução de reagente p-DAC 0,196% (m/v) em meio de HCl 0,0549 mol L-1 e secagem ao ar livre, submetendo-se então o spot à leitura reflectométrica em máx. = 535 nm. Uma relação linear (R = 0,996) foi estabelecida na faixa de concentração de ureia compreendida entre 50,0 – 300 mg L-1. Os limites de detecção e de quantificação foram determinados em 5,13 mg L-1 e 17,10 mg L-1, respectivamente. O método limpo desenvolvido foi aplicado na determinação de ureia em amostras comerciais de fertilizantes onde demonstrou apresentar ótima precisão e exatidão, evidenciadas pela boa recuperação (94,2 – 107,4%), além de oferecer vantagens como simplicidade de execução e baixo consumo de reagentes, geração de mínima quantidade de resíduos, rapidez e segurança... / This work proposes the development of a clean method for determination of urea in commercial samples of fertilizers using the Diffuse Reflectance Spectroscopy combined with spot test. The method is based on the reaction between urea and the chromogenic reagent p-dimethylaminocinnamaldehyde (p-DAC) in diluted hydrochloric acid medium, which results in a pink colored product whose maximum value of reflectance occurs at a wavelength of 535 nm. The experimental parameters were optimized through a factorial and a central composite experimental design to obtain the response surface, which indicated a higher sensitivity for the method when using the reagent p-DAC in the concentration of 0.196% (w/v) in a hydrochloric acid 0.0549 mol L-1 medium, using ethanol as a solvent for the solutions. The solid support for the reaction is a qualitative filter paper where they are placed 20.0μL of urea solution followed by drying with cold air from a hair dryer and 20.0μL of reagent solution p-DAC 0.196% (w/v) in the HCl 0.0549 mol L-1 medium and drying in the air, then submitting the spot to the reflectometric reading at max = 535 nm. A linear relationship (R = 0.996) was established in the range of urea concentration between 50.0 to 300 mg L-1. The limits of detection and quantification were determined at 5.13 mg L-1 and 17.10 mg L-1, respectively. The method was applied to the clean determination of urea in commercial samples of fertilizers, which has shown to present great precision and accuracy as evidenced by the good recovery (94.2 to 107.4%) and also offers advantages such as simplicity of implementation and low consumption of reagents, generation of minimum quantity of wastes, rapidity and security, producing reliable results. As shown, the method is designed as environmentally friendly for determination of urea, as it is consistent with the Principles advocated by the Green Chemistry.

No description available.

Tissue optical clearing aims to increase the penetration depth of near-collimated light in biological tissue to enhance optical diagnostic, therapeutic, and cosmetic procedures. Previous studies have shown the effects of chemical optical clearing on tissue optical properties. Drawbacks associated with chemical clearing include the introduction of potentially toxic exogenous chemicals into the tissue, poor site targeting, as well as slow transport of the chemicals through tissue. Thus, alternative clearing methods have been investigated. Mechanical compression is one such alternative tissue optical clearing technique. The mechanisms of action of mechanical compression may be similar to those of chemical clearing, though they have yet to be investigated systematically. This research describes the design and execution of a number of procedures useful for the quantification of the tissue optical clearing effects of localized mechanical compression. The first experimental chapter presents the effects of compression on image resolution and contrast of a target imaged through ex vivo biological tissue. It was found that mechanical optical clearing allowed recovery of smaller targets at higher contrast sensitivity when compared to chemical clearing. Also, thickness-independent tissue clearing effects were observed. In the second experimental chapter, dynamic changes in tissue optical properties, namely scattering and absorption coefficients (?s' and ?a, respectively) were monitored during a controlled compression protocol using different indentation geometries. A reduction in ?s' and ?a was evident for all indentation geometries, with greater changes occurring with smaller surface area. Results indicate that localized mechanical compression may be harnessed as a minimally-invasive tissue optical clearing technique. / Master of Science

Radio frequency (RF) lesioning and deep brain stimulation (DBS) are the two prevailing surgical treatments for movement disorders within the field of stereotactic and functional neurosurgery. For RF-lesioning, a small volume of brain tissue is coagulated and knowledge of the lesion size and growth is of great importance for the safety and outcome of the procedure. This thesis deals with adapting the laser Doppler perfusion monitoring (LDPM) technique for measurements in brain tissue during RF-lesioning. The relation between LDPM signal changes and developed lesion size was investigated. LDPM measurements were evaluated both in vitro (albumin protein solution) and in vivo in the porcine brain during RF-lesioning corresponding to a bilateral thalamotomy in man. The investigated signals from the LDPI measurements can be used for following the lesioning time course and to detect if a lesion was created, both in vitro and in the animal model. For the albumin model, both the total backscattered light intensity and the perfusion signal can be used as markers for estimating the final coagulation size, while in the animal model this conclusion was not statistical verified. Independent on surgical method, RF-lesioning or DBS, intracerebral guidance is an important aspect within stereotactic and functional neurosurgery. To increase the accuracy and precision of reaching the correct target, different methods for intracerebral guidance exist, such as microelectrode recording and impedance methods. In this thesis, the possibility of developing an optical intracerebral guidance method has been investigated. Diffuse reflectance spectroscopy served as technology and all measurements were performed stereotactically in both porcine and human brain. Measurements of white and gray matter showed large differences, with higher reflectivity for white brain matter, both in porcine and in human brain. For the human measurements during DBS-implants, large differences between white matter and functional targets were found. Additionally, differences between native and lesioned porcine brain matter were detected. Both studies support the idea of using diffuse reflectance spectroscopy for developing an intracerebral guidance method.

<p>Diffuse reflectance spectroscopy (DRS) is a simple, yet powerful technique that has the potential to offer practical, non-invasive, and cost effective information for op- tical diagnostics and therapeutics guidance. Any progress towards moving DRS systems from their current laboratory settings to clinical settings, field settings and ambitiously to home settings, is a significant contribution to society in terms of reducing ever growing healthcare expenditures of an aging society. Additionally, im- proving on the existing mathematical models used to analyze DRS signals; in terms of speed, robustness, accuracy, and capability in accounting for larger feature space dimensionality (i.e. extraction of more tissue-relevant information) is equally im- portant for real-time diagnosis in the desired settings and to enable use of DRS in as many biomedical applications (e.g. skin cancer diagnosis, diabetics care, tissue oxygenation monitoring) as possible. Improving the reflectance signal complexity and density through novel DRS instrumentation, would facilitate development of the desired models or put the existing ones built on simulations in practical use; which otherwise could not go beyond being a theoretical demonstration.</p><p>DRS studies tissue morphology and composition through quantification of one or more (ideally all of them) of the tissue- and wavelength-specific optical properties: absorption coefficient (μa), reduced scattering coefficient (μ1s), scattering anisotropy (g), tissue thickness, and scattering phase function details (e.g. higher order moments of the scattering phase function). DRS involves sampling of diffusely reflected photons which experience multiple scattering and absorption as they travel within the tissue, at the tissue surface. Spatially resolved diffuse reflectance spectroscopy (SRDRS) is a subset of general DRS technique, which involves sampling of diffuse reflectance signals at multiple distances to an illumination source. SRDRS provides additional spatial information about the photon path; yielding depth-resolved tissue information critical to layered tissue analysis and early cancer diagnostics. Exist- ing SRDRS systems use fiber optic probes, which are limited in accommodation of large number and high-density collection fibers (i.e. yielding more and dense spa- tially resolved diffuse reflectance (SRDR) measurement data) due to difficulty of fiber multiplexing. The circular shape of the fibers restricts the implementable probe ge- ometries and reduces the fill factor for a given source to detector (i.e. collection fiber) separation (SDS); resulting in reduced light collection efficiency. The finite fiber nu- merical aperture (NA) reduces the light collection efficiency well as; and prevents selective interrogation of superficial tissues where most cancers emerge. Addition- ally, SRDR systems using fiber optic probes for photon collection, require one or more photodetectors (i.e. a cooled CCD); which are often expensive components of the systems.</p><p>This thesis deals with development of an innovative silicon SRDRS probe, which partially addresses the challenge of realizing high measurement density, miniaturized, and inexpensive SRDRS systems. The probe is fabricated by conventional, flexible and inexpensive silicon fabrication technology, which demonstrates the feasibility of developing SRDRS probes in any desired geometry and complexity. Although this approach is simple and straightforward, it has been overlooked by the DRS community due to availability of the conventional fiber optic probe technology. This new probe accommodates large number and high density of detectors; and it is in the form of a concentric semi-annular photodiode (PD) array (CMPA) with a central illumination aperture. This is the first multiple source-detector spacing Si SRDRS probe reported to date, and the most densely packed SRDRS probe reported to date for all types of SRDRS systems. The closely spaced and densely packed detectors enable higher density SRDR measurements compared to fiber-based SRDR probes, and the higher PD NA compared to that of fibers results in a higher SNR increasing light collection efficiency. The higher NA of the PDs and the presence of PDs positioned at very short distances from the illumination aperture center enable superficial tissue analysis as well as depth analysis.</p> / Dissertation

<p>Worldwide, head and neck squamous cell cancers (HNSCC) account for over 375,000 deaths annually. The majority of these cancers arise in the outermost squamous cells which progress through a series of precancerous changes before developing into invasive HNSCC. It is widely accepted that prognosis is strongly correlated to the stage of diagnosis, with early detection more than doubling the patient’s chance of survival. Currently, however, 60% of HNSCCs are diagnosed when they have already progressed to stage 3 or stage 4 disease. The current diagnostic method of visual examination often fails to recognize early indicators of HNSCC, thereby missing an important prevention window.</p><p> </p><p>Determination of cancer from non-malignant tissues is dependent on pathological examination of lesion biopsies. Thus, all patients with any clinically suspicious lesions undergo surgical biopsies. Furthermore, these surgical biopsies carry risks. In addition to the risk of general anesthesia for patients undergoing panedoscopy, some patients have poor healing and develop ulcerations or infections as a result of surgical biopsy at any anatomical site. Additionally, studies have shown that approximately 50% of suspected biopsies are later pathologically confirmed normal. An enormous amount of labor, facility, and monetary resources are expended on non-malignant biopsies and patients who ultimately have no malignancy. It would be of immense overall benefit to clinicians and patients to have a non-invasive and portable technique that could rapidly identify those patients that would benefit from further surgical biopsy from those that only need follow-up clinical observations.</p><p> </p><p>Once carcinoma is confirmed in a patient, treatment currently involves modalities of surgery, radiation, and chemotherapy. Radiotherapy plays a significant role, particularly in the management of localized HNSCC, because it is a non-invasive and function-preserving modality. However, the effectiveness of radiotherapy is limited by hypoxia. Previous studies showed that tumors reoxygenated during radiotherapy treatment may have a better prognosis. Despite decades of work, there is still no reliable, cost-effective way for measuring tumor hypoxia over multiple time points to estimate the prognosis. </p><p>To address these unmet clinical needs, three aims were proposed. The first aim was to improve early detection by identifying biomarkers of early pre-cancer as well as developing an objective algorithm to detect early disease. Neovasculature is an important biomarker for early cancer diagnosis. Even before the development of a clinically detectable lesion, the tumor vasculature undergoes structural and morphological changes in response to oncogenic signaling pathways [8]. Without receiving a sufficient supply of oxygen and nutrients to proliferate, early tumor growth is limited to only 1-2 mm. High-resolution optical imaging is well suited to characterize the earliest neovascularization changes that accompany neoplasia owing to its sensitivity to hemoglobin absorption and resolution to visualize capillary level architecture. Dark field microscopy is a low-cost and robust method to image the neovasculature. We imaged neovascularization in vivo in a spontaneous hamster oral mucosa carcinogen model using a label-free, reflected-light spectral dark field microscope. Hamsters’ cheek pouches were painted with 7, 12-Dimethylbenz[a]anthracene (DMBA) to induce precancerous to cancerous changes, or mineral oil as control. Spectral dark field images were obtained during carcinogenesis and in control oral mucosa, and quantitative vascular features were computed. Vascular tortuosity increased significantly in oral mucosa diagnosed as hyperplasia, dysplasia and squamous cell carcinoma (SCC) compared to normal. Vascular diameter and area fraction decreased significantly in dysplasia and SCC compared to normal. The areas under the receiver operative characteristic (ROC) curves (AUC) computed using a Support Vector Machine (SVM) were 0.95 and 0.84 for identifying SCC or dysplasia, respectively, vs. normal and hyperplasia oral mucosa combined. To improve AUCs for identifying dysplasia, quantitative vascular features were computed again after the vessels were split into large and small vessels based on diameter. The large vessels preserved the same significant trends, while small vessels demonstrated the opposite trends. Significant increases in diameter and decreases in area fraction were observed in SCC and dysplasia. The AUCs were improved to 0.99 and 0.92 for identifying SCC and dysplasia. These results suggest that dark field vascular imaging is a promising tool for pre-cancer detection.</p><p>Optical imaging can also be applied to quantifying other important characteristics of solid tumors in head and neck cancer (HNC), such as hypoxia, abnormal vascularity and cell proliferation. Diffuse reflectance spectroscopy is a simple and robust method to measure tissue oxygenation, vascularity and cell density. It is particularly suitable for applications in the operation room because of its compact design and portability. In addition, a fiber probe-based system is ideal for obtaining measurements at suspicious lesions in the head and neck area during surgery. Thus, my second aim was to reduce the number of unnecessary HNSCC biopsies by developing a robust tool and rapid analysis method appropriate for clinical settings. We propose the use of morphological optical biomarkers for rapid detection of human HNSCC by leveraging the underlying tissue characteristics in the aerodigestive tracts Prior to biopsy, diffuse reflectance spectra were obtained from malignant and contra-lateral non-malignant tissues of 57 patients undergoing panendoscopy. Oxygen saturation (SO2), total hemoglobin concentration ([THb]), and the reduced scattering coefficient were extracted using an inverse Monte Carlo (MC) method previously developed by former student in our lab. Differences in malignant and non-malignant tissues were examined based on two different groupings: by anatomical site and by morphological tissue type. Measurements were acquired from 252 sites, 51 of which were pathologically classified as SCC. Optical biomarkers exhibited statistical differences between malignant and non-malignant samples. Contrast was enhanced when parsing tissues by morphological classification rather than by anatomical subtype for unpaired comparisons. Corresponding linear discriminant models using multiple optical biomarkers showed improved predictive ability when accounting for morphological classification, particularly in node-positive lesions. The false-positive rate was retrospectively found to decrease by 34.2% in morphologically- vs. anatomically-derived predictive models. In glottic tissue, the surgeon exhibited a false-positive rate of 45.7% while the device showed a lower false-positive rate of only 12.4%. Additionally, comparisons of optical parameters were made to further understand the physiology of tumor staging and potential causes of high surgeon false-positive rates. Optical spectroscopy is a user-friendly, non-invasive tool capable of providing quantitative information to discriminate malignant from non-malignant head and neck tissues. Predictive models demonstrated promising results for diagnostics. Furthermore, the strategy described appears to be well suited to reduce the clinical false-positive rate.</p><p>To further improve the speed for extracting the tissue oxygenation and [THb] to reduce the time when patients were under anesthesia, the third aim was to develop a rapid heuristic ratiometric analysis for estimating tissue [THb] and SO2 from measured tissue diffuse reflectance spectra. The analysis was validated in tissue-mimicking phantoms and applied to clinical measurements in head and neck, cervical and breast tissues. The analysis works in two steps. First, a linear equation that translates the ratio of the diffuse reflectance spectra at 584 nm to 545 nm to estimate the tissue [THb] using a Monte carlo (MC)-based lookup table was developed. This equation is independent of tissue scattering and oxygen saturation. Second, SO2 was estimated using non-linear logistic equations that translate the ratio of the diffuse reflectance spectra at 539 nm to 545 nm into the tissue SO2. Correlations coefficients of 0.89 (0.86), 0.77 (0.71) and 0.69 (0.43) were obtained for the tissue hemoglobin concentration (oxygen saturation) values extracted using the full spectral MC and the ratiometric analysis, for clinical measurements in head and neck, breast and cervical tissues, respectively. The ratiometric analysis was more than 4000 times faster than the inverse MC analysis for estimating tissue [THb] and SO2 in simulated phantom experiments. In addition, the discriminatory power of the two analyses was similar. These results show the potential of such empirical tools to rapidly estimate tissue hemoglobin and oxygenation for real-time applications.</p><p>In addition to its use as a diagnostic marker for various cancers, tissue oxygenation is believed to play a role in the success of cancer therapies, particularly radiotherapy. However, since little effort has been made to develop tools to exploit this relationship, the fourth aim was to estimate patient prognosis by measuring tumor hypoxia over multiple time points so physicians are able to develop more informed and effective clinical treatment plan. To test if oxygenation kinetics correlates with the likelihood for local tumor control following fractionated radiotherapy, we again used diffuse reflectance spectroscopy to noninvasively measure tumor vascular oxygenation and [THb] associated with radiotherapy of 5 daily fractions (7.5, 9 or 13.5 Gy/day) in FaDu xenografts. Spectroscopy measurements were obtained immediately before each daily radiation fraction and during the week after radiotherapy. SO2 and [THb] were computed using an inverse MC model. Oxygenation kinetics during and after radiotherapy, but before a change in tumor volume, was associated with local tumor control. Locally controlled tumors exhibited significantly faster increases in oxygenation after radiotherapy (days 12-15) compared with tumors that recurred locally. (2) Within the group of tumors that recurred, faster increases in oxygenation during radiotherapy (days 3-5) were correlated with earlier recurrence times. An AUC of 0.74 was achieved when classifying the local control tumors from all irradiated tumors using the oxygen kinetics with a logistic regression model. (3) The rate of increase in oxygenation was radiation dose dependent. Radiation doses ≤9.5 Gy/day did not initiate an increase in oxygenation whereas 13.5 Gy/day triggered significant increases in oxygenation during and after radiotherapy. Additional confirmation is required in other tumor models, but these results suggest that monitoring tumor oxygenation kinetics could aid in the prediction of local tumor control after radiotherapy.</p><p>Angiogenesis is a highly regulated process to support tissue growth. Neovasculature is designed by nature to grow toward areas lacking nutrition and oxygen. Cancer cells proliferate too quickly to have their nutritional and oxygen needs completely satisfied, which results in an imbalanced state of angiogenesis leading to tortuous blood vessels, hypoxic tissues and radioresistance. We characterized the tumor-induced vascular features with simple, robust and low-cost dark field microscopy and spectroscopy to enable early cancer diagnosis, improvement of surgical biopsy accuracy and better predict the prognosis of radiotherapy for HNC. Our results demonstrated that these noninvasively measured, label-free vascular features are able to detect pre-cancer, reduce unnecessary surgical biopsies and predict prognosis of radiotherapy.</p> / Dissertation

<p>Breast conserving surgery (BCS) is a common treatment option for breast cancer patients. The goal of BCS is to remove the entire tumor from the breast while preserving as much normal tissue as possible for a better cosmetic outcome after surgery. Specifically, the excised specimen must have at least 2 mm of normal tissue surrounding the diseased mass. Unfortunately, a staggering 20-70% of patients undergoing BCS require repeated surgeries due to the incomplete removal of the tumor diagnosed post-operatively. Due to these high re-excision rates as well as limited post-operative histopathological sampling of the tumor specimen, there is an unmet clinical need for margin assessment. Quantitative diffuse reflectance spectral imaging has previously been explored as a promising, method for providing real-time visual maps of tissue composition to help surgeons determine breast tumor margins to ensure the complete removal of the disease during breast conserving surgery. We have leveraged the underlying sources of contrast in breast tissue, specifically total hemoglobin content, beta-carotene content, and tissue scattering, and developed various fiber optics based spectral imaging systems for this clinical application. Combined with a fast inverse Monte Carlo model of reflectance, previous studies have shown that this technology may be able to decrease re-excision rates for BCS. However, these systems, which all consist of a broadband source, fiber optics probes, an imaging spectrograph and a CCD, have severe limitations in system footprint, tumor area coverage, and speed for acquisition and analysis. The fiber based spectral imaging systems are not scalable to smaller designs that cover a large surveillance area at a very fast speed, which ultimately makes them impractical for use in the clinical environment. The objective of this dissertation was to design, develop, test, and show clinical feasibility of a novel wide field spectral imaging system that utilizes the same scientific principles of previously developed fiber optics based imaging systems, but improves upon the technical issues, such as size, complexity, and speed,to meet the demands of the intra-operative setting. </p><p>First, our simple re-design of the system completely eliminated the need for an imaging spectrograph and CCD by replacing them with an array of custom annular photodiodes. The geometry of the photodiodes were designed with the goal of minimizing optical crosstalk, maximizing SNR, and achieving the appropriate tissue sensing depth of up to 2 mm for tumor margin assessment. Without the imaging spectrograph and CCD, the system requires discrete wavelengths of light to launch into the tissue sample. A wavelength selection method that combines an inverse Monte Carlo model and a genetic algorithm was developed in order to optimize the wavelength choices specifically for the underlying breast tissue optical contrast. The final system design consisted of a broadband source with an 8-slot filter wheel containing the optimized set of wavelength choices, an optical light guide and quartz light delivery tube to send the 8 wavelengths of light in free space through the back apertures of each annular photodiode in the imaging array, an 8-channel integrating transimpedance amplifier circuit with a switch box and data acquisition card to collect the reflectance signal, and a laptop computer that controls all the components and analyzes the data.</p><p>This newly designed wide field spectral imaging system was tested in tissue-mimicking liquid phantoms and achieved comparable performance to previous clinically-validated fiber optics based systems in its ability to extract optical properties with high accuracy. The system was also tested in various biological samples, including a murine tumor model, porcine tissue, and human breast tissue, for the direct comparison with its fiber optics based counterparts. The photodiode based imaging system achieved comparable or better SNR, comparable extractions of optical properties extractions for all tissue types, and feasible improvements in speed and coverage for future iterations. We show proof of concept in performing fast, wide field spectral imaging with a simple, inexpensive design. With a reduction in size, cost, number of wavelengths used, and overall complexity, the system described by this dissertation allows for a more seamless scaling to higher pixel number and density in future iterations of the technology, which will help make this a clinically translatable tool for breast tumor margin assessment.</p> / Dissertation