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Topics in living cell miultiphoton laser scanning microscopy (MPLSM) image analysisZhang, Weimin 30 October 2006 (has links)
Multiphoton laser scanning microscopy (MPLSM) is an advanced fluorescence
imaging technology which can produce a less noisy microscope image and minimize the
damage in living tissue. The MPLSM image in this research is the dehydroergosterol
(DHE, a fluorescent sterol which closely mimics those of cholesterol in lipoproteins and
membranes) on living cell's plasma membrane area. The objective is to use a statistical
image analysis method to describe how cholesterol is distributed on a living cell's
membrane. Statistical image analysis methods applied in this research include image
segmentation/classification and spatial analysis. In image segmentation analysis, we
design a supervised learning method by using smoothing technique with rank statistics.
This approach is especially useful in a situation where we have only very limited
information of classes we want to segment. We also apply unsupervised leaning methods
on the image data. In image data spatial analysis, we explore the spatial correlation of
segmented data by a Monte Carlo test. Our research shows that the distributions of DHE
exhibit a spatially aggregated pattern. We fit two aggregated point pattern models, an
area-interaction process model and a Poisson cluster process model, to the data. For the area interaction process model, we design algorithms for maximum pseudo-likelihood
estimator and Monte Carlo maximum likelihood estimator under lattice data setting. For
the Poisson Cluster process parameter estimation, the method for implicit statistical
model parameter estimate is used. A group of simulation studies shows that the Monte
Carlo maximum estimation method produces consistent parameter estimates. The
goodness-of-fit tests show that we cannot reject both models. We propose to use the area
interaction process model in further research.
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Nanoscale engineering of semiconductor heterostructures for quadratic nonlinear optics and multiphoton imagingZieliński, Marcin 09 February 2011 (has links) (PDF)
Nonlinear coherent scattering phenomena from single nanoparticles have been recently proposed as alternative processes for fluorescence in multiphoton microscopy staining. Commonly applied nanoscale materials, however, have reached a certain limit in size dependent detection efficiency of weak nonlinear optical signals. None of the recent efforts in detection of second-harmonic generation (SHG), the lowest order nonlinear process, have been able to cross a ~40 nm size barrier for nanoparticles (NPs), thus remaining at the level of "large" nanoscatterers, even when resorting to the most sensitive detection techniques such as single-photon counting technology. As we realize now, this size limitation can be significantly lowered when replacing dielectric insulators or wide gap semiconductors by direct-gap semiconducting quantum dots (QDs). Herein, a new type of highly nonlinear nanoprobes is engineered in order to surpass above mentioned size barrier at the single nanoparticle scale. We consider two-photon resonant excitation in individual zinc-blende CdTe QDs of about 12.5 nm diameter, which provide efficient coherent SHG radiation, as high as 105 Hz, furthermore exhibiting remarkable sensitivity to spatial orientation of their octupolar crystalline lattice. Moreover, quantum confinement effects have been found to strongly contribute to the second-order nonlinear optical susceptibility χ(2) features. Quantitative characterization of the χ(2) of QDs by way of their spectral dispersion and size dependence is therefore undertaken by single particle spectroscopy and ensemble Hyper-Rayleigh Scattering (HRS) studies. We prove that under appropriate conditions, χ(2) of quantum confined semiconducting structures can significantly exceed that of bulk. Furthermore, a novel type of semiconducting hybrid rod-on-dot (RD) QDs is developed by building up on crystalline moieties of different symmetries, in order to increase their effective quadratic nonlinearity while maintaining their size close to a strong quantum confinement regime. The new complex hybrid χ(2) tensor is analyzed by interfering the susceptibilities from each component, considering different shape and point group symmetries associated to octupolar and dipolar crystalline structures. Significant SHG enhancement is consequently observed, exceeding that of mono-compound QDs, due to a coupling between two nonlinear materials and slower decoherence, which we attribute to the induced spatial charge separation upon photoexcitation.
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Resonance Enhanced Multiphoton Ionization Studies of Dichlorotoluenses, Dichloroanilines, and Dichlorophenolsde Laat, Richard 09 January 2013 (has links)
A new instrument using a time-of-flight (TOF) mass filter (MF) for resonance enhanced multiphoton ionization (REMPI) studies of gas phase molecules was developed. This instrument was designed to make use of the selectivity of the REMPI process and the sensitivity of a TOF-MF with a microchannel plate detector. A pulsed valve inlet system was tested to determine its effectiveness in “cooling” molecules but it was not used for the bulk of the studies performed.
The instrument was tested using molecular and atomic bromine, atomic carbon, and atomic iodine. The atomic bromine, carbon, and iodine, were generated by the photodissociation of molecular bromine, hydrocarbons, and methyl iodide respectively. Nitrogen gas in air was used to test the pulsed valve system.
The instrument was then used to conduct REMPI studies of five dichlorotoluene (DCT) isomers (2,4-DCT; 2,5-DCT; 3,4-DCT; 2,6-DCT; and 2,3-DCT). REMPI studies of six dichloroaniline (DCA) isomers, including 2,5-DCA; 3,4-DCA; 3,5-DCA; 2,6-DCA; 2,4-DCA; and 2,3-DCA were conducted. Six isomers of dichlorophenol (DCP) were studied (2,5-DCP; 3,4-DCP; 3,5-DCP; 2,3-DCP; 2,4-DCP; and 2,6-DCP). It was determined that the 2,3-DCP; 2,4-DCP; and 2,6-DCP isomers photodissociated to form CCl, which itself could be observed through a REMPI process.
The results from the REMPI studies of the dichloroaromatics and data from previous ultraviolet, infrared, and RAMAN studies of these molecules was used in order to assign the observed peaks.
The observed 0,0 π→π* transition energies of the dichloroaromatics studied were used along with ultraviolet 0,0 π→π* transition energies from previous works in order to discuss substituent effects. A qualitative method of predicting the relative location of 0,0 π→π* transition energies of dichloroaromatics was developed.
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Rôle des états de Rydberg dans la dynamique de photoionisation et de formation de paires d'ions (NO+,O-) de la molécule NO2 : photoémission induite par rayonnement synchrotron et impulsions lasers femtosecondesMarggi Poullain, Sonia 14 January 2014 (has links) (PDF)
L'étude comparée des réactions de formation de paires d'ions et de simple photoionisation de la molécule NO2 induites par rayonnement synchrotron (RS) d'une part et par impulsions laser femtosecondes (fs) d'autre part, démontre le rôle remarquable de l'excitation résonante d'états de Rydberg dans la dynamique électronique et nucléaire induite. Trois réactions principales, la photoionisation non dissociative (NO2+ (X 1Σ+g) + e), la photoionisation dissociative (NO+ (X 1Σ+) + O(3P) + e) et la formation de paires d'ions, (NO+ (X 1Σ+) + O- (2P)), ont été caractérisées en utilisant la méthode des corrélations vectorielles ou spectroscopie en coïncidence des impulsions du photoélectron et des photoions, auprès des sources RS (SOLEIL, DESIRS) et lasers fs (CEA, Saclay), respectivement. Le diagramme de corrélation des énergies cinétiques électron-ion, première observable issue de ces mesures, met en évidence un partage de l'énergie en excès entre noyaux et électrons qui dépend fortement du mode d'excitation photonique. Les déviations significatives observées par rapport aux profils d'ionisation de type Franck Condon sont attribuées à des couplages vibroniques entre états excités NO2*, tels que ceux induits par une intersection conique. Les chemins réactionnels identifiés confirment le rôle de l'excitation des séries de Rydberg [R*(6a1)-1] et [R*(4b2)-1] intervenant comme états intermédiaires dans l'excitation multiphotonique ou dans le continuum d'ionisation exploré. Une étude complémentaire par spectroscopie à haute résolution des états [R*(6a1)-1] a été mise en œuvre (UBC, Vancouver).Pour une réaction de photoionisation dissociative (PID), l'observable la plus complète est la distribution angulaire des photoélectrons dans le référentiel lié à la vitesse de recul de l'ion fragment (RFPAD) déduite de la mesure de la corrélation vectorielle (Vi, Ve, P). Afin d'accéder aux éléments de matrice dipolaire décrivant la photoionisation de l'état électronique considéré, le formalisme développé en collaboration avec R. R. Lucchese (Texas A&M) décrivant la photoémission dans le référentiel moléculaire pour la simple PID d'une molécule linéaire par excitation à un photon, a été étendu à l'étude des réactions de PID par excitation multiphotonique d'une molécule polyatomique, telle que la molécule NO2 de symétrie C2v. L'analyse multivariée de la RFPAD multiphotonique proposée constitue une stratégie fructueuse en vue d'extraire l'information optimale sur la dynamique complexe de photoionisation et de réaliser une comparaison détaillée entre les résultats expérimentaux et les calculs de photoionisation des états excités de la molécule.
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Coherent Anti-Stokes Raman Scattering Miniaturized MicroscopeSmith, Brett 04 July 2013 (has links)
Microscopy techniques have been developed and refined over multiple decades, but innovation around single photon modalities has slowed. The advancement of the utility of information acquired, and minimum resolution available is seemingly reaching an asymptote. The fusion of light microscopy and well-studied nonlinear processes has broken through this barrier and enabled the collection of vast amounts of additional information beyond the topographical information relayed by traditional microscopes. Through nonlinear imaging modalities, chemical information can also be extracted from tissue. Nonlinear microscopy also can beat the resolution limit caused by diffraction, and offers up three-dimensional capabilities. The power of nonlinear imaging has been demonstrated by countless research groups, solidifying it as a major player in biomedical imaging.
The value of a nonlinear imaging system could be enhanced if a reduction in size would permit the insertion into bodily cavities, as has been demonstrated by linear imaging endoscopes. The miniaturization of single photon imaging devices has led to significant advancements in diagnostics and treatment in the medical field. Much more information can be extracted from a patient if the tissue can be imaged in vivo, a capability that traditional, bulky, table top microscopes cannot offer. The development of new technologies in optics has enabled the miniaturization of many critical components of standard microscopes. It is possible to combine nonlinear techniques with these miniaturized elements into a portable, hand held microscope that can be applied to various facets of the biomedical field.
The research demonstrated in this thesis is based on the selection, testing and assembly of several miniaturized optical components for use as a nonlinear imaging device. This thesis is the first demonstration of a fibre delivered, microelectromechanical systems mirror with miniaturized optics housed in a portable, hand held package. Specifically, it is designed for coherent anti-Stokes Raman scattering, second harmonic generation, and two-photon excitation fluorescence imaging. Depending on the modality being exploited, different chemical information can be extracted from the sample being imaged. This miniaturized microscope can be applied to diagnostics and treatments of spinal cord diseases and injuries, atherosclerosis research, cancer tumour identification and a plethora of other biomedical applications. The device that will be revealed in the upcoming text is validated by demonstrating all designed-for nonlinear modalities, and later will be used to perform serialized imaging of myelin of a single specimen over time.
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Investigating multiphoton phenomena using nonlinear dynamicsHuang, Shu 20 March 2008 (has links)
Many seemingly simple systems can display extraordinarily complex dynamics which has been studied and uncovered through nonlinear dynamical theory. The leitmotif of this thesis is changing phase-space structures and their (linear or nonlinear) stabilities by adding control functions
(which act on the system as external perturbations) to the relevant Hamiltonians. These phase-space structures may be periodic orbits, invariant tori or their stable and unstable manifolds. One-electron systems and diatomic molecules are fundamental and important staging ground for new discoveries in nonlinear dynamics. In past years, increasing emphasis and effort has been put on the control or manipulation of these systems. Recent developments of nonlinear dynamical tools can
provide efficient ways of doing so. In the first
subtopic of the thesis, we are adding a control function to restore tori at prescribed locations in phase space. In the remainder of the
thesis, a control function with parameters is used to change the linear stability of the periodic orbits which govern the processes in question.
In this thesis, we report our theoretical analyses on multiphoton ionization of Rydberg atoms exposed to strong microwave fields and
the dissociation of diatomic molecules exposed to bichromatic lasers using nonlinear dynamical tools. This thesis is composed of three subtopics. In the first subtopic, we employ
local control theory to reduce the stochastic ionization of hydrogen atom in a strong microwave field by adding a relatively small control term to the original Hamiltonian. In the second subtopic, we perform periodic orbit analysis to investigate multiphoton ionization driven by a
bichromatic microwave field. Our results show quantitative and qualitative agreement with previous studies, and hence identify the mechanism through which short periodic orbits organize the dynamics in multiphoton ionization. In addition, we achieve substantial time
savings with this approach. In the third subtopic we extend our periodic orbit analysis to the dissociation of diatomic molecules driven
by a bichromatic laser. In this problem, our results based on periodic orbit analysis again show good agreement with previous work, and hence promise more potential applications of this
approach in molecular physics.
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Low-energy electron induced processes in hydrocarbon films adsorbed on silicon surfacesShepperd, Kristin 06 July 2009 (has links)
The deposition of hydrocarbons on silicon substrates is a topic of wide interest. This is generally related to the technological importance of silicon carbide (SiC) and a growing interest in graphene and graphitic materials. Methods for producing these materials predominantly involve high processing temperatures. In the case of SiC, these high processing temperatures often result in the formation of surface defects, which compromise the electronic properties of the material. In an effort to grow SiC films at low temperatures, a technique known as electron-beam chemical vapor deposition (EBCVD) has been developed. Most electron beam deposition techniques employ a focused beam of high-energy (20-30 keV) electrons to form nanometer-sized solid deposits on a surface. However, in an effort to deposit macroscale films, a broad beam of low-energy electrons was used.
In addition to investigating the applications of low-energy electrons in semiconductor film growth, the fundamental chemical and physical processes induced by the bombardment of adsorbate-covered surfaces with low-energy electrons were examined. Specifically, the electron-stimulated desorption of various adsorbate-substrate systems such as acetylene adsorbed on silicon, graphene oxide on silicon, and ultrathin graphite films on silicon carbide have been investigated. The yields of cation and neutral desorbates as a function incident electron energy were measured, appearance thresholds were determined and mechanisms of desorption were proposed.
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Bladder microstructural and biomechanical modelling : in vivo, in vitro and in silicoHornsby, Jack January 2016 (has links)
Lower urinary tract disorders are significant prognostic indicators of institutionalisation and lower quality of life in the elderly and their incidence increases with age. Urodynamics, the gold standard in diagnosis, replicates symptoms to assess functionality through controlled filling and voiding of the bladder but its interpretation is subjective and may be inconclusive; often requiring further testing or leading to inappropriate treatment. Normal filling and voiding biomechanics of the bladder relate directly to the structural composition of the bladder wall. Alterations to tissue composition in aging and pathology have significant impacts on biomechanics but are yet to be fully described. The aim of this thesis was to gain insight into the individual microstructural components of the bladder wall and how they relate to the gross mechanical response. Additionally, representation of these observations in a mathematical model that can be used to improve our understanding of urodynamic data. This aim was achieved through a combination of in situ mechanical testing and the development of a microstructural constitutive model, which was then included within an overall micturition framework to simulate filling and voiding functions, and evaluated with clinical data. Coupled systems of multiphoton microscopy and uniaxial, biaxial and inflation testing were used to correlate extra cellular matrix interactions with the mechanical response of young and aged murine bladder. Wall-layer specific collagen fibre orientation, dispersion and recruitment were quantified and implemented into a novel microstructural constitutive model. The bladder was modelled as a nonlinear elastic, constrainedmixture planar membrane with contribution from smooth muscle and collagen fibres in the detrusor. Collagen recruitment in the detrusor was observed to occur at a finite stretch; correlated with a steep increase in stiffness of the tissue, while collagen of the lamina propria plays a capacitance role. Collagen recruitment was modelled using a triangular probability density function; quantified from sequential microscopy images and fitted to mechanical data. Increased collagen area fraction and changes in dominant fibre orientation were attributed to reduced compliance in aged bladder. This behaviour was captured by the model. The microstructural model was modified to an isotropic thin-walled spherical membrane for the filling phase of a micturition model framework, consisting of a bladder outlet relation and urethral resistance relation. A contractile smooth muscle element was included in the active response. In the first steps towards clinical application the model was applied to male and female 'normal' patient urodynamic data to observe quality of model fit and estimate baseline parameter values. The model simulated key filling and voiding features seen in normal male and female clinical data. Mechanobiological modelling combined with clinically relevant micturition modelling has the potential to quantify bladder dysfunction. Moreover, improved understanding of how the microstructure influences macroscopic mechanics will yield improved understanding of how changes to the bladder impair its functionality. We predict that modelling will become a clinically relevant tool in urodynamics; leading to new options for diagnosis and management of patients with bladder dysfunction.
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Produção de nanopartículas de Au induzida por pulsos laser de femtossegundos formatados / Gold nanoparticles production induced by shaped femtosecond laser pulsesPaulo Henrique Dias Ferreira 27 October 2011 (has links)
Neste trabalho investigamos a dinâmica de formação de nanopartículas de Au por pulsos de femtossegundos formatados (800 nm, 30 fs, 1 kHz e 2 mJ), induzida pela ionização da molécula de quitosana. Inicialmente desenvolvemos um sistema de formatação de pulsos ultracurtos que faz uso de um modulador espacial de luz, constituído por um arranjo linear de cristais líquidos, com o qual somos capazes de impor distintas modulações de fase ao pulso laser. Para monitorar o processo de produção de nanopartículas, montamos um sistema de excitação (pulsos de femtossegundos) e prova (luz branca), o qual permite a observação em tempo real do aparecimento da banda de plásmon e, consequentemente, da dinâmica de formação das nanopartículas. Resultados obtidos para pulsos não formatados (limitados por Transformada de Fourier) demonstraram que a formação de nanopartículas deve-se à ionização não linear da quitosana, a qual está relacionada à oxidação do grupo hidroxila para o grupo carbonila. Medidas de microscopia eletrônica de transmissão forneceram os tamanhos (entre 20 e 100 nm) e formatos (esferas, pirâmides, hexágonos, bastões, etc) das nanopartículas geradas. Ainda, nossos resultados revelaram que esta ionização é iniciada por absorção multifotônica, mais especificamente por absorção de 4 fótons. Utilizando pulsos formatados com fase espectrais constante, degrau e cossenoidal com diferentes frequências, investigamos a influência destes na formação de nanopartículas. Concluímos que os pulsos mais longos são mais favoráveis ao processo de ionização, e consequente redução dos íons de Au para a formação de nanopartículas metálicas. Este comportamento se deve, provavelmente, à redistribuição da energia absorvida para os modos vibracionais, o que é mais provável para pulsos mais longos. Assim, o método apresentado pode abrir novas maneiras para a formação de nanopartículas de metálicas, as quais podem ser mais exploradas dos pontos de vista aplicado e fundamental. / In this work we have studied the synthesis of Au nanoparticles using shaped ultrashort pulses (800 nm, 30 fs, 1 kHz and 2 mJ), induced by the ionization of the chitosan. Initially we developed a pulse shaping setup that uses a spatial light modulator (liquid crystals array), with which we are able to impose distinct phase mask to the laser pulse. In order to monitor the nanoparticles production process, we used a pump-probe system, consisting of femtosecond pulses (pump) and white light (probe), which allows the observation of the plasmon band enhancement and hence the nanoparticles formation dynamics. The results obtained by Fourier Transform limited pulses have shown that the nanoparticles formation is due to the nonlinear ionization of chitosan, which is related to hydroxyl group oxidation to the carbonyl group. Transmission electron microscopy measurements provided the sizes (20-100 nm) and shapes (spheres, pyramids, hexagons, rods, etc.) of the produced nanoparticles. Moreover, our results revealed that ionization is initiated by multiphoton absorption, more specifically by four photons absorption. Using pulses shaped with constant, step and cossenoidal (with different frequencies) spectral phase masks, we investigated their influence in the nanoparticles formation. We conclude that longer pulses are more favorable to the ionization process and, consequently, to the gold ions reduction for the synthesis of the metallic nanoparticles. This behavior is probably due to the redistribution of the absorbed energy to the vibrational modes, which is more likely for longer pulses. Therefore, the approach presented here can open new ways to produce metallic nanoparticles, which can be further explored from applied and fundamental points of view.
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Photonic approach for the study of dental hard tissues and carious lesion detection / Approche photonique pour l’étude des tissus durs dentaires et la détection des lésions carieusesSlimani, Amel 23 November 2017 (has links)
Les propriétés photoniques des tissus durs dentaires nous ont permis d’étudier l’email et la dentine a un niveau moléculaire (in vitro) en utilisant des techniques de microscopie optique non linéaires. La microscopie confocale Raman est technique d’imagine de haute résolution permettant d’analyse d’échantillon sans préparation spécifique ni marquage. Cette méthode nous a permis de reconstituer une cartographie de la réticulation du collagène et de la cristallinité au niveau de la jonction émail-dentine et cela avec une résolution spatiale non atteinte jusque-là. Cette analyse chimique de la jonction émail-dentine a permis de redéfinir la largeur de cette zone de transition. Cette largeur est nettement supérieure à celles proposées par les études précédentes. Par ailleurs, l’étude portant sur les changements de fluorescence intrinsèque entre les tissues dentaires sains et cariés suggèrent l’implication de la protoporphyrin IX et de la pentosidine dans l’expression de la fluorescence rouge des tissus cariés. La microscopie multiphotonique quant à elle nous a permis de détecter la lésion carieuse et de suivre son développement en utilisant la génération de seconde harmonique (SHG) et la fluorescence par excitation à deux photons (2PEF). Nos études ont démontré la validité du ratio SHG/2PEF comme paramètre fiable pour la détection de la lésion carieuse. Les études proposées par cette thèse montrent le potentiel des propriétés photoniques de l’émail et de la dentine en utilisant les microscopies Raman et multiphotoniques dans l’étude de ces tissus au niveau moléculaire. Cela offre de nouvelles perspectives en recherche et en applications cliniques. / Photonic properties of dental hard tissues allowed us to proceed to in vitro analysis of enamel and dentin on a molecular level. Confocal Raman microscopy has been used to produce a mapping of collagen cross-link and crystallinity of human dentin–enamel junction (DEJ) with a spatial resolution not achieved up to now. The method is a non-invasive, label-free and a high spatial resolution imaging technique. This chemical analysis of DEJ led us to redefine a wider width of this transition zone and advance our understanding of dental histology. A study on the intrinsic fluorescence changes of sound and carious tissues using conventional fluorescence microscopy suggests the involvement of protoporphyrin IX and pentosidine in the fluorescence red-shift observed in carious tissues. Multiphoton microscopy allowed to detect nonlinear optical signal changes during caries process using second harmonic generation (SHG) and two-photon excitation fluorescence (2PEF). Our studies led us to propose the ratio SHG/2PEF as valuable parameter to monitor caries lesion. Collectively, advances described in this thesis show the potential of photonic properties of enamel and dentin using Raman and multiphoton microcopies for molecular investigations on sound as much as on carious tissues. It opens new perspective in dental research and clinical applications.
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