Thesis (Ph. D.)--University of Wisconsin--Madison, 1947. / Typescript. Vita. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references (leaf [39]).

Thesis (PH. D.)--Cornell university. / "Reprinted from the Journal of physical chemistry, vol. 15, p. 845 (1911)" "References": p. 864-865.

Fluorescence techniques have been widely used in chemistry and many areas of biology due to its high sensitivity, simplicity, fast response, and capability of spatial imaging. Currently there are six major classes of small-molecular fluorophores-coumarins, boron dipyrromethene (BODIPY) dyes, fluoresceins, rhodamines, oxazines, and cyanines. This dissertation mainly focuses on modifications of fluoresceins and development of pH probes based on rhodamines. We have modified the xanthene core of fluorescein by replacing the xanthene oxygen with a dimethylsilyl group which shows 90 nm red-shift relative to fluorescein (λabs/λem =582 nm/598 nm). These fluorophores have advantages over their oxygen counterparts because longer-wavelength allows deeper penetration of the light, causing less damage to the cells and less interference from the biological medium. Based on this scaffold, we have tried to develop copper, mercury, and hydrogen peroxide probes by functionalizing the hydroxyl group of the fluorophore with different groups. Although the results weren’t satisfactory, we have gained some new insights about this scaffold. The hydroxyl group of the fluorophore is not very reactive and this scaffold is not stable under strong basic conditions and its electronic configuration is very different with the regular fluorescein. We have synthesized and characterized Si-rhodol fluorophore, the hybrid structure of Si-fluorescein and Si-rhodamine. The preliminary investigation of Si-rhodol, UV-vis and fluorescence, has been performed. It shows that the absorption and fluorescence emission of Si-rhodol are around 30 nm red-shift compared to Si-fluorescein (λabs/λem =617 nm/630 nm). And Si-rhodol is more fluorescent than Si-fluorescein (pKa = 6.8) at pH 7.4 (biological condition) due to its lower pKa (5.3). Despite the excellent photophysical properties of Si-rhodol, there are actually limited examples about its application. Among these examples, none of them gave the detailed information about functionalization of the hydroxyl group of Si-rhodol. We have found a method to functionalize the hydroxyl group of Si-rhodol. We have developed a series of pH probes through modification of rhodamines which possess either an aniline-methyl or cyclic ring moiety. These analogues maintain a coloress, non-fluorescent spirocyclic structure at high pH and the fluorescence goes up while the pH decreases. The substituent effects on anilinomethylrhodamines (AnMR) system follows Hammett’s equation. The effect of cycloalkane ring size on the acid/base properties of the AMR system can be predicted by Baeyer’s ring strain theory. The pKa of these rhodamine analogues can be tuned from around 3 to 8. We have also investigated aminomethylfluoroscein (AMF) system. In this system, we found that once the phenolic hydroxyl group was functionalized at one side, the fluorescence was off at pH>3. In addition, variation of the substituents on aminomethyl moiety didn't affect the pKa of this system significantly. Therefore, we were set out to develop mitochondria-targetable hydrogen peroxide probe. However, in the presence of triphenylphosphine cation, the conversion from the OTf group to the boronate group didn't work.

M.Ing. (Electrical and Electronic Engineering) / Advances in bio-imaging have triggered the development of a highly sophisticated imaging tool known as fluorescence microscopy. Fluorescence microscopy is used in many biological applications to visualize sub-cellular processes and gives the ability to image three-dimensional (3D) structures in living cells. The use of fluorescence microscopy and specific staining methods make biological molecules appear as bright spots in image data. The analysis of fluorescence microscopy images requires the detection and tracking of hundreds spots in image data and is of great importance for biologists to better understand cell functions. However, the analysis of these data is still performed manually in most biological laboratories worldwide. The manual analysis of these data is both time consuming, laborious and susceptible to human errors. Several computer-based algorithms have been proposed for the detection and tracking of spots in microscopy images. Most of these methods were validated on limited image data and relatively few studies have been performed for the comparison of these methods in real applications. This study quantitatively compared the performance of four detection and two tracking methods applied in microscopy images for the analysis of bright spots. The performance of the algorithms was validated on both synthetic and real images. The synthetic images offered a better way of validating algorithms against ground truth reference results. Results indicate that there are major differences in algorithm performance for both detection and tracking. In the detection results the Isotropic Undecimated Wavelet Transform (IUWT) and the Laplacian of Gaussian (LoG) achieved better results than the other methods in comparison when values are considered. The tracking results indicate that the Interacting Multiple Model (IMM) method achieved better results than the Feature Point Tracking (FPT) method when Jaccard Similarity Scores (JSC) are considered.

M.Sc. (Chemistry) / The aim of this investigation was to study, and if possible, to correct curvature of calibration lines when solutions are analysed by X-ray fluorescence spectrometry. The two main metallic elements used for this purpose were copper and cadmium, with a short study of nickel, cobalt and iron. The concentration ranges investigated were from 0,01 % (rn/rn) to 10 % (rn/rn) in the solution sample ...

The absorption and emission of energy is a fundamental property of any atomic system.When the emission occurs in the visible or near visible region the phenomenon is usually referred to as luminescence or fluorescence. This must be distinguished from thermal radiation as it does not follow Kirchoff's law.

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Thesis (M.S.)--University of Montana, 2007. / Title from title screen. Description based on contents viewed Sept. 25, 2007. Includes bibliographical references (p. 44-45).

Calcium (Ca2+), a crucial effector for many biological systems, has been associated with diseases such as cardiovascular disease, Alzheimer’s, Parkinson’s, cancer, and osteoporosis. It is important to develop calcium sensors to measure intracellular Ca2+ dynamics at various biological and pathological states. Our lab has engineered such probes by designing a Ca2+ binding site into fluorescent proteins such as Enhanced Green Fluorescent Protein (EGFP) and mCherry. In this thesis, we aim to improve optical properties and metal binding properties of green EGFP-based sensor CatchER and mCherry based red sensors by site-directed mutagenesis and protein engineering, various spectroscopic methods and cell imaging. The green EGFP-based sensor CatchER, with a Ca2+ binding pocket charge of -5, displays the greatest affinity for Ca2+ and has the greatest fluorescence intensity change with Ca2+ when compared to its variants with a less negative binding pocket charge. In addition, we have also designed several SR/ER targeting CatchER variants using Ryanodine receptor and Calnexin transmembrane domains. These constructs were shown to display a strong presence in the SR/ER lumen and further designed for a new luminal orientation. Further, we have shown that the optical properties of two red calcium sensors can be significantly improved by modifying the local environment of the chromophore.