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En Viticulture de Précision, l’imagerie multi-spectrale est principalement utilisée pour des dispositifs de télédétection. Ce manuscrit s’intéresse à son utilisation en proxidétection, pour la caractérisation du feuillage. Il présente un dispositif expérimental terrestre mobile composé d’un GPS, d’une caméra multi-spectrale acquérant des images visible et proche infrarouge, et d’un Greenseeker RT-100 mesurant l’indice Normalized Difference Vegetation Index (NDVI). Ce système observe le feuillage de la vigne dans le plan de palissage, en lumière naturelle. La parcelle étudiée comporte trois cépages (Pinot Noir, Chardonnay et Meunier) plantés en carré latin. En 2013, six jeux de données ont été acquis à différents stades phénologiques.Pour accéder aux propriétés spectrales de la végétation, il est nécessaire de calibrer les images en réflectance. Cela requiert l’utilisation d’une mire de MacBeth comme référence radiométrique. Lorsque la mire est cachée par les feuilles, les paramètres de calibration sont estimés par une interpolation linéaire en fonction des images les plus proches sur lesquelles la mire est visible. La cohérence de la méthode d’estimation employée est vérifiée par une validation croisée (LOOCV).La comparaison du NDVI fournie par le Greenseeker avec celui déterminé via les images corrigées permet de valider les données générées par le dispositif. La polyvalence du système est évaluée via les images où plusieurs indices de végétation sont déterminés. Ils permettant des suivis de croissance de la végétation originaux offrant des potentialités de phénotypage ou une caractérisation de l’état sanitaire de la végétation illustrant la polyvalence et le gain en précision de cette technique. / Mutispectral imaging systems are widely used in remote sensing for Precision Viticulture. In this work, this technique was applied in the proximal sensing context to characterize vine foliage. A mobile terrestrial experimental system is presented, composed of a GPS receiver, a multi-spectral camera acquiring visible and near infrared images, and a Greenseeker RT-100 which measures the Normalized Difference Vegetative Index (NDVI). This optical system observes vine foliage in the trellis plan, in natural sunlight. The experimental field is planted with Chardonnay, Pinot Noir and Meunier cultivars in a latin squared pattern. In 2013, six datasets were acquired at various phenological stages.Spectral properties of the vegetation are accessible on images when they are calibrated in reflectance. This step requires the use of a MacBeth colorchart as a radiometric reference. When the chart is hidden by leaves, the calibration parameters are estimated by simple linear interpolation using the results from resembling images, which have a visible chart. The performance of this method is verified with a cross-validation technique (LOOCV).To validate the data provided by the experimental system, the NDVI given by the Greenseeker was compared to those computed from the calibrated images. The assessment of the versatility of the system is done with the images where several indices were determined. It allows an innovative follow-up of the vegetative growth, and offering phenotyping applications. Moreover, the characterization of the sanitary state of the foliage prove that this technique is versatile and accurate.

This thesis reports the work done towards design and fabrication of a versatile and low cost, frequency domain DOT (Diffuse Optical Tomography) Imager. A design which uses only a single fiber for the source and a single fiber bundle for the detector is reported. From near the source, to diametrically opposite to the source, the detected intensity of scattered light varies by three to four orders in magnitude, depending on the tissue/phantom absorption and scattering properties. The photo multiplier tube’s (PMT’s) gain is controlled to operate it in the linear range, thus increasing the dynamic range of detection. Increasing the dynamic range by multi channel data acquisition is also presented. Arresting the oscillations of a stepper using a negative torque braking method is also adopted in this application for increasing the speed of data acquisition. The finite element method (FEM) for obtaining photon density solution to the transport equation and the model based iterative image reconstruction (MPBIIR) algorithm are developed for verifying the experimental prototype. Simulation studies presented towards the end of this thesis work provide insight into the nature of measurements. The optical absorption reconstructed images from the simulation, verified the validity of implementation of the reconstruction method for further reconstructions from data gathered from the developed imager. A single iteration of MOBIIR to segment the region of interest (ROI) using an homogeneous measurement estimate is presented. Using the single iteration MOBIIR to obtain a relatively more accurate starting value for the optical absorption coefficient, and the reconstruction results for data obtained from tissue mimicking solid epoxy-resin phantom with a single in-homogeneity inclusion is also presented to demonstrate the imager prototype.

Medical devices are used widely at every stage of disease diagnosis and treatment. To eradicate certain infectious diseases, the development of highly sensitive diagnostic tools and techniques is essential. The work reported in this thesis presents a novel approach, which can be used for the diagnosis of various diseases in the field of clinical cytology. The central theme of this approach was to develop a simple, holistic and completely automated system for point-of-care (POC) diagnostics. This is realized through the Development of an Absorption Flow-Cytometer with Synergistic Integration of Microfluidic, Optics and simple Electronics. Quantitative diagnosis of malaria has been taken as test case for the characterization and validation of the developed technology. Malaria is a life-threatening disease widely prevalent in developing countries. Approximately half the world population undergoes a test of malaria and it kills close to half a million people every year. Early detection and treatment will reduce the number of fatalities and also decrease its transmission rate. In the recent past, several diagnostic tools have been developed to detect malaria but there are varied demands on diagnostic instruments in healthcare settings and endemic contexts. The objective of this thesis is to develop an instrument capable of identifying malaria-infected red blood cells (i-RBCs) from a given few micro-liters of whole blood. The optical absorption properties of blood cells were measured at a single-cell level to diagnose malaria. The proof-of-concept for the instrument was established in four stages, after which a prototype was also developed and validated. In the first stage, a system capable of simultaneously imaging cells and also measuring their optical absorbance properties was developed. The developed system was employed to characterize absorption properties of red blood cells (malaria-infected and healthy ones) on blood-smear. A custom-made bright-field transmission microscope in combination with a pair of laser diode and photo-detector was used to simultaneously image and measure transmittance of infected and uninfected RBCs. In the second stage, the technique was extended to enable high-throughput measurements with the use of microfluidic sample handling and synchronous data acquisition. Using this technique, the optical absorbance and morphology of infected and healthy RBCs have been characterized in statistically significant numbers. The correlation between cell morphology (from images) and single-cell optical absorbance level helped to establish the thresholds for differentiating healthy and infected cells. In the third stage, a portable prototype capable of assessing optical absorbance levels of single cells was fabricated. The developed prototype is capable of assessing cells at throughputs of about 1800 cells/ second. It was initially validated with sample suspensions containing infected and healthy RBCs obtained from malaria cultures. For the device to be usable at the field-level, it has to function in the presence of all other cellular components of whole blood. The optical absorbance of other cellular components of blood like white blood cells and platelets, were characterized. The device was finally tested with blood samples spiked with malaria-infected RBCs validating the overall proof-of-concept and the developed prototype. The deployment of such cost-effective, automated POC system would enable malaria diagnosis at remote locations and play a crucial role in the ongoing efforts to eradicate malaria. In future, the presented technology can be extended to develop POC diagnostic tool for other diseases as well. As it enables quantitative estimation of malaria, the present optical absorption flow analyzer would also find application in disease prognosis monitoring, anti-malarial drug development and other studies requiring measurements on a single-cell basis. The hyper-imaging system can be used to characterize and validate the threshold information, and can be incorporated in the prototype. Thus, it is a continuous process to characterization and implementation in the prototype. The optofluidic absorption flow analyzer will help enable affordable clinical diagnostic testing in resource limited settings. This approach will be extended to diagnose other diseases, using differences in optical absorption as criteria for differentiating healthy and infected cells.

Aujourd’hui, les difficultés pour établir des liens entre caractéristiques des nanomatériaux et réponses biologiques sont principalement issues du manque de contrôle de la synthèse des nanomatériaux, ne permettant pas de faire varier leurs paramètres physico-chimiques clés une à une.Pour identifier certains mécanismes gouvernant la toxicité des nanomatériaux nous avons utilisé un nanotube inorganique modèle dont la synthèse est bien contrôlée : les Ge-imogolites. Les effets de la longueur, du nombre de parois, de la cristallinité et de la composition chimique des Ge-imogolites ont été étudiés sur une bactérie des sols: Pseudomonas brassicacearum. Il a été identifié que la présence de sites réactifs (en bordure de tubes) induit une toxicité due à une interaction forte des nanotubes avec les cellules bactériennes, ainsi que la génération d’espèces réactives de l’oxygène. Ajouter des sites réactifs via la présence de défauts structuraux augmente la dégradation des tubes ainsi que la rétention d’éléments nutritifs essentiels, ce qui augmente leur toxicité. Enfin, l’ajout de fer dans leur structure transforme les Ge-imogolites en source de fer, qui sont dégradées et deviennent promoteurs de croissance. Dans tous ces cas, les interactions entre nanomatériaux et cellules ont été identifiées comme cruciales pour comprendre et prévenir les effets des nanomatériaux. Ce travail de thèse a également permis de mettre en avant la capacité de nouveaux outils pour le suivi de l’internalisation de nanomatériaux dans les organismes. / Only a few studies of (eco)toxicology linked the physico-chemical properties of nanoparticles to the toxicity mechanisms or the stress they induce. Moreover, no clear conclusions can be drawn at present because of the variability of nanoparticles used in studies. The present study used the inorganic Ge-imogolite nanotubes as a model compound. The toxic effects of length, number of walls, structural defects, and chemical composition were assessed towards the soil bacteria Pseudomonas brassicacearum. Several mechanisms modulating the toxicity of Ge-imogolite were then identified. Indeed, reactive sites at the tube ends induce a slight toxicity via a strong cell interaction and the generation of reactive oxygen species. Creating vacant sites on the surface of Ge-imogolite (ant thus increasing the number of reactive sites), appears to cause a deficiency of nutrients in the culture media correlated with a higher degradation of the tubes, leading to a high bacterial growth decrease. Finally, structural iron incorporation into Ge-imogolite transforms them into an iron source, being degraded and becoming growth promoters. In this work, the new tools capacities for the study of nanomaterials/cells interaction have been studied.

Le mélanome est un cancer rare de la peau qui se développe à partir des cellules responsables de la pigmentation : les mélanocytes. Depuis quelques années, nous observons une augmentation significative du nombre de personnes atteintes par cette maladie, de mauvais pronostic et très agressive (132 000 nouveaux cas chaque année dans le monde). Le taux de mortalité de ce cancer est très élevé en raison de la rapidité de propagation des cellules cancéreuse dans d’autres régions du corps. En France, on remarque un nombre relativement élevé de cas détectés, plus particulièrement en Bretagne. La tumeur se présente comme une tâche dont l’apparence est très proche du grain de beauté. Si elle est détectée suffisamment tôt, un prélèvement suffit à la guérison et les risques de récidives sont très faibles. En revanche, une fois que les métastases se propagent, les chances de survie à long terme sont très faibles. Malgré les récentes avancées en thérapie ciblée, les traitements du mélanome métastatique restent encore limités. En partenariat avec le groupe Malakoff Médéric, nous développons actuellement un système optique pour la détection précoce du mélanome cutané. Celui-ci est basé sur les propriétés de polarisation de la lumière. La thèse présente la conception du système, de l’étude de faisabilité jusqu’à l’étape finale d’étalonnage. Nous proposons également différents tests sur des échantillons de simulation. Ces derniers nous permettent de démontrer la corrélation qu’il existe entre les effets de polarisation et les modifications biologiques en cours lors du développement du mélanome. Cette étude préliminaire nous prépare aux expérimentations sur de vrais échantillons. / Melanoma is a rare cancer that develops from the pigmentation cells of the skin. Recently, we notice a significant increase in the number of people affected by this aggressive disease with a poor prognosis (132 000 new cases each year worldwide). The mortality rate of this cancer is very high, which is due to the rapid spread of cancerous cells to other parts of the body. In France, there is a relatively high number of cases detected, especially in Britanny. The tumor is a spot which looks like a mole. If detected early, a levy is sufficient to healing and the risk of recurrence is very low. However, once metastasis spread, the long-term prognosis is very low. Despite recent advances in targeted therapy, the treatments of metastatic melanoma are still limited. In partnership with the Malakoff Mederic group, we are currently developing an optical system for early detection of cutaneous melanoma. It is based on the polarization properties of light. The PhD focuses on the design of the system, from the feasibility study to the final calibration. We also present various tests on samples of simulation. These allow us to demonstrate the correlation between the effects of polarization and biological changes during the development of melanoma. This preliminary study prepares us to experiments on real samples.

Diffuse Optical Tomography is a diagnostic imaging modality where optical parameters such as absorption coefficient, scattering coefficient and refractive index distributions are recovered to form the internal tissue metabolic image. Near-infrared (NIR) light has the potential to be used as a noninvasive means of diagnostic imaging within the human breast. Due to the diffusive nature of light in tissue, computational model-based methods are required for functional imaging. The main goal is to recover the spatial variation of optical properties which shed light on the different metabolic states of tissue and tissue like media. This thesis addresses the issue of quantitative recovery of optical properties of tissue-mimicking phantom and pork tissue using diffuse optical tomography (DOT). The main contribution of the present work is the development of robust, efficient and fast optical property reconstruction algorithms for a direct 3-D DOT imaging system. There are both theoretical and experimental contributions towards the development of an imaging system and procedures to minimize accurate data collection time, overall system automation as well as development of computational algorithms. In nurturing the idea of imaging using NIR light into a fully developed direct 3-D imaging system, challenges from the theoretical and computational aspects have to be met. The recovery of the optical property distribution in the interior of the object from the often noisy boundary measurements on light, is an ill-posed ( and nonlinear) problem. This is particularly true, when one is interested in a direct 3-D image reconstruction instead of the often employed stacking of 2-D cross-sections obtained from solving a set of 2-D DOT problems. In order to render the DOT, a useful diagnostic imaging tool and a robust reconstruction procedure giving accurate and reliable parameter recovery in the scenario, where the number of unknowns far outnumbers the number of independent data sets that can be gathered (for example, the direct 3-D recovery mentioned earlier) is essential. Here, the inversion problem is often solved through iterative methods based on nonlinear optimization for the minimization of a data-model misfit function. An interesting development in this direction has been the development of Broyden’ s and adjoint Broyden’ s methods that avoids direct Jacobian computation in each iteration thereby making the full 3-D a reality. Conventional model based iterative image reconstruction (MoBIIR) algorithm uses Newton’ s and it’s variant methods, where it required repeated evaluation of whole Jacobian, which consumes bulk time in reconstruction process. The explicit secant and adjoint information based fast 2-D/3-D image reconstruction algorithms without repeated evaluation of the Jacobian is proposed in diffuse optical tomography, where the computational time has been decreased many folds by updating the Jacobian successively through low rank update. An alternative route to the iterative solution is attempted by introducing an artificial dynamics in the system and treating the steady-state response of the artificially evolving dynamical system as a solution. The objective is to consider a novel family of pseudo-dynamical 2-D and 3-D systems whose numerical integration in time provides an asymptotic solution to the inverse problem at hand. We convert Gauss-Newton’ s equation for updates into a pseudo-dynamical (PD) form by explicitly adding a time derivative term. As the pseudo-time integration schemes do not need such explicit matrix inversion and depending on the pseudo-time step size, provides for a layer of regularization that in turn helps in superior quality of 2-D and 3-D image reconstruction. A cost effective frequency domain Matlab based 2-D/3-D automated imaging system is designed and built. The complete instrumentation (including PC-based control software) has been developed using a single modulated laser source (wavelength 830nm) and a photo-multiplier tube (PMT). The source and detector fiber change their positions dynamically allowing us to gather data at multiple source and detector locations. The fiber positions are adjusted on the phantom surface automatically for scanning variable size phantoms. A heterodyning scheme was used for reading out the measurement using a lock-in-amplifier. The Matlab program carries out sequence of actions such as instrument control, data acquisition, data organization, data calibration and reconstruction of image. The Gauss-Newton’ s, Broyden’ s, adjoint Broyden’ s and pseudo-time integration algorithms are evaluated using the simulation data as well as data from the experimental DOT system. Validation of the system and the reconstruction algorithms were carried out on a real tissue, a pork tissue with an embedded fat inhomogeneity. The results were found to match the known parameters closely.

The thesis deals with physical principles of ultrasound with following specialization on ophtalmic ultrasound system Nidek Echoscan 4000 and its properties. And also it deals with description of investigative techniques in ophthalmology (A-mode, B-mode, biometry and pachymetry). Next, they was suggested procedures of measurement and that is spatial resolution (axial and lateral spatial resolution) and propagation speed of ultrasound depending on the temperature. Then these properties was tested on laboratory phantoms of eye – ETETECH LTD. and Multipurpose Ultrasound Phantom but on created phantoms of eye too, which will be made from agarose gel with build-in materail (fishing line, plastic film, plastic bag and rubber). At the end, measured values was processed in the generated program.

<p>Cadmium (Cd) is a highly mobile and toxic heavy metal that negatively affects plants, soil biota, animals and humans, even in very low concentrations. Currently, Cd contamination of cocoa produced in Latin American countries is a significant problem, as concentrations can exceed acceptable levels set by the European Union (0.5 mg/kg), sometimes by more than 10 times allowable levels. In South America, <i>Theobroma cacao</i> is an essential component of the basic market basket. This crop contributes to the Latin-American trade balance, as these countries export cacao and chocolate-based products to major consumer countries such as the United States and Europe. Some soil amendments can alter the bioavailability and uptake of Cd into edible plant tissues, though cacao plants can accumulate Cd without displaying any visible symptoms of phytotoxicity, which makes it difficult to determine if potential remediation strategies are successful. Currently, the only effective way to quantify Cd accumulation in plant tissues is via destructive post-harvest practices that are time-consuming and expensive. New hyperspectral imaging (HSI) technologies developed for use in high-throughput plant phenotyping are powerful tools for monitoring environmental stress and predicting the nutritional status in plants. Consequently, the experiments described in this thesis were conducted to determine if HSI technologies could be adapted for monitoring plant stress caused by Cd, and estimating its concentration in vegetative plant tissues. Two leafy green crops were used in these experiments, basil (<i>Ocimum basilicum L.</i> var. Genovese) and kale (<i>Brassica oleracea L</i>. var. Lacinato), because they are fast growing, and therefore, could serve as indicator crops on cacao farms. In addition, we expected these two leafy green crops would differ in their morphological responses to Cd stress. Specifically, we predicted that stress responses would be visible in basil, but not kale, which is known to be a hyperaccumulator. The plants were subject to four levels of soil Cd (0, 5, 10 and 15 ppm), and half of the pots were amended with biochar at a rate of 3% (v/v), as this amendment has previously been demonstrated to improve plant health and reduce Cd uptake. The experiments were conducted at Purdue’s new Controlled Environment Phenotyping Center (CEPF). The plants were imaged weekly and manual measurements of plant growth and development were collected at the same times, and concentrations of Cd as well as many other elements were determined after harvest. Fourteen vegetation indices generated using HSI images collected from the side and top view of plants were evaluated for their potential to identify subtle signs of plant stress due soil Cd and the biochar amendment. In addition, three mathematical models were evaluated for their potential to estimate Cd concentrations in the plant biomass and determine if they exceed safe standards (0.28 mg/kg) set by the Food and Agriculture Organization (FAO) for leafy greens. Results of these studies confirm that like many plants, these leafy green crops can accumulate Cd levels that are well above safety thresholds for human health, but exhibit few visible symptoms of stress. The normalized difference vegetation index (NDVI) and the chlorophyll index at the red edge (CI_RE) were the best indices for detecting Cd stress in these crops, and the plant senescence and reflectance index (PSRI) and anthocyanin reflectance index (ARI) were the best at detecting subtle changes in plant physiology due to the biochar amendment. The heavy metal stress index (HMSSI), developed exclusively for detecting heavy metal stress, was only able to detect Cd stress in basil when images were taken from the top view. Results of the mathematical models indicated that principal components analysis (PCA) and partial least squares (PLS) models overfit despite efforts to transform the data, indicating that they are not capable of predicting Cd concentrations in these crops at these levels. However, the artificial neural networks (ANN) was able to predict whether leafy greens had levels of Cd that were above or below critical thresholds suggested by the FAO, indicating that HSI could be further developed to predict Cd concentrations in plant tissues. Further research conducted in the field and in the presence of other environmental stress factors are needed to confirm the utility of these tools, and determine whether they can be adapted to monitor Cd uptake in cacao plants.</p>

<p dir="ltr">Cancer biological mechanisms are a vastly researched area in the field, yet they are not well understood in the various contexts in which cancer is found. Cancerous tumors often exist in harsh, stressful environments for normal cells, but cancer cells can thrive in these conditions. The tumor microenvironment (TME) typically has low oxygen levels (hypoxia), high acidity, and low nutrition. Exposure to the TME leads to many metabolic changes in the cells, enabling cancer to continue proliferating and migrating. However, these metabolic changes are not well understood, especially at the single-cell level. The ability to monitor cells in real time to determine the physical characteristics they undergo is critical to understanding the impact of these metabolic changes. Conventional methods focus on determining the genomic and proteomic changes in large numbers of cells, which may be overlooked if the changes are homogeneous across samples. In this work, we demonstrate the power of using multiple imaging techniques in combination with biochemical methods to visualize metabolic changes and determine the causes in various cancer cells under extreme hypoxia conditions.</p><p dir="ltr">The changes in the microtubule network that occur under hypoxia at the single-cell level are not widely researched. The use of confocal fluorescence microscopy can determine microtubule polymerization in conjunction with eGFP-transfected EB3, a protein that assists in microtubule polymerization. We have determined that hypoxic HeLa cells produce finger-like protrusions when exposed to hypoxia that help with cell migration and, ultimately, cancer cell metastasis. The formation of these protrusions is facilitated by localized mitochondria activities in the protrusions.</p><p dir="ltr">The metabolic changes in lipid droplets (LDs) under hypoxia at the single-cell level remain an elusive topic. The use of stimulated Raman spectroscopy (SRS) and coherent anti-Stokes Raman scattering (CARS) can determine the quantity and spatial-temporal distribution of LDs in cancer cells. We have found that LDs redistribute to the endoplasmic reticulum (ER) and increase in intensity in hypoxic MIA PaCa-2 and A549 cells. Time-lapse CARS microscopy revealed a release-accumulate process of these LDs on ER in hypoxia. We also studied the impact of carbon sources on LD formation and found that MIA PaCa2 cells prefer direct lipid uptake while glucose is also essential to reduce lipotoxicity. The use of hyperspectral stimulated Raman scattering (hSRS) also reveals that the content of the LDs changes to include less cholesteryl ester and a decrease in lipid saturation level.</p><p dir="ltr">Collectively, these findings shed new light on the understanding of cytoskeleton dynamics and lipid metabolism in hypoxic conditions. The discoveries made within this research would lead to better treatment strategies for effective treatment of hypoxia-resistant cancer cells.</p>