Thesis (M.S.E.C.E.)--University of Delaware, 2007. / Principal faculty advisors: Fouad Kiamilev and Robert F. Rogers, Dept. of Electrical and Computer Engineering. Includes bibliographical references.

Thesis (MSc)--University of Stellenbosch, 2006. / Bibliography. Also available via the Internet.

Thesis (M.Phil.)--Hong Kong University of Science and Technology, 2007. / Includes bibliographical references (leaves 110-116). Also available in electronic version.

No description available.

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Th.--Chim. phys.--Nancy--I.N.P.L., 1985.

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The genome is more than a linear sequence of bases; its spatial organisation is a key part of its function. In humans, three billion base pairs, or approximately two metres of DNA are packaged into a nucleus a few micrometres in diameter. The genome must also be organised so that it can be replicated and partitioned into daughter cells, and so that regulatory elements are positioned to affect their targets. Until recently, little was known about the organisation of the genome at the scale of single genes. The packaging of DNA onto nucleosomes, and the segregation of chromosomes into chromosome territories was well understood, but the development of chromatin conformation capture (3C) techniques has enabled the first thorough study of intermediate scales. These methods provide information about the distances between pairs of genomic loci, which gives indirect information about their positions. By applying these techniques to single cells, it has become possible to calculate a structure from the observed distance restraints. Through the prior constraints placed on the model, such as the existence of a continuous backbone, these structures provide additional information about the conformation of DNA. To overcome the limitations of 3C, it is useful to integrate additional sources of information. I present several methods for the validation and improvement of Hi-C structures by adding data from microscopy, and for characterising dyes used in single-molecule light microscopy. It is found that single-cell Hi-C structures agree with fluorescence microscopy when observing the distance of genes from the edge of the nucleus, and that centromeres are not a suitable label for direct validation. Adding absolute positional restraints from images is shown to be useful in better determining chromatin structure in synthetic tests. Finally, the presence of a FRET acceptor near a fluorescent protein is shown to improve its photophysical properties.

Spontaneous atomic emission is not a process of the isolated atom but rather a cooperative effect of the atom and the vacuum field. It is now well established that spontaneous radiative decay rates can be enhanced or suppressed through the effect of cavities comprising various types of discrete, reflective-mirror type, boundaries. The cavity effect is generally understood in terms of a cavity-induced modification of the vacuum spectral energy density. Recently, interest has grown in the possibility that systems characterized by distributed periodic boundary conditions, such as a spatially varying index of refraction, might be effective in controlling radiative atomic processes. A semi-classical theory is given that enables an estimate of the size of the lifetime modification of a two-level radiator contained within a three-dimensionally incomplete photonic bandgap structure called a fiber-Bragg grating. Following this is an exploration of a specific system and its effect upon radiative spontaneous emission. It is found through fluorescence line narrowing and frequency hole burning measurements that the observation of lifetime modification of the specific system is complicated due to intra and inter Stark energy migration. A lifetime modification measurement then shows that no change in lifetime is observed beyond the error bars on the measurement results. The tuning and coherence properties of a short-external-cavity diode laser that may be useful for future time-dependent spectroscopic measurements are examined using a fiber-based, self-heterodyne technique. Coherence properties during active frequency scans are characterized through analysis of time-dependent heterodyne beat signals at the output of a fiber interferometer.

As a high-resolution wide-field near-surface microscopy, total internal reflection fluorescence microscopy (TIRFM) has been widely applied for the study of biomolecules. Unlike those costly, sample consuming and time consuming traditional detection assays, the application of TIRFM enable the direct quantification of biomolecules in a sample pretreatment and enrichment free fashion. Taking advantages of the TIRFM imaging system, in this thesis we have applied the TIRFM imaging system to directly quantify the content of different cancer associated biomarkers. Four different detection approaches for direct cancer biomarkers quantification with the aid of TIRFM were herein presented respectively. In Chapter 2, a direct quantification of nasopharyngeal carcinoma associated miRNAs was described. In the assay, five different miRNAs were chosen as the target analytes, which hybridized with the synthetic complementary LNA, probe in solution. The duplex was labeled with intercalating fluorescence dye YOYO-1 and the signal was then detected by the TIRFM-EMCCD imaging system. The LNA probe exhibited a high binding affinity towards the complementary target miRNAs and a limit of detection of 8 pM was achieved. Since the LOD is far below the reported concentration of miRNAs found in body fluids, this developed assay is of high potential to serve as a tool for non-invasive detection of miRNAs for early disease diagnosis. In Chapter 3, an advanced single-molecule based assay for direct circulating miRNAs detection was developed. The assay was demonstrated to be capable of differentiating the expression of a nasopharyngeal carcinoma (NPC) up-regulator hsa-mir-205 (mir-205) in serum collected from patients of different stages of NPC. To overcome the background matrix interference in serum, locked nucleic acid modified molecular beacon (LNA/MB) was applied as the detection probe to hybridize, capture and detect target mir-205 in serum matrix with enhanced sensitivity and specificity. A detection limit of 500 fM was achieved. The as-developed method was capable of differentiating NPC stages by the level of mir-205 quantified in serum with only 10 μL of serum and the whole assay can be completed in an hour. The experimental results agreed well with reported and while the quantity of mir-205 determined by our assay was found comparable to that of quantitative reverse transcription polymerase chain reaction (qRT-PCR), supporting that this assay can be served as a promising non-invasive detection tool for early NPC diagnosis, monitoring and staging. In chapter 4, a self-assembled protein nanofibril based online pre-concentrating sensor was developed. This solution-based hybridization assay was applied to quantified the amount of target miRNAs, mir-196a. Biotinylated locked nucleic acid (LNA) of complimentary sequence was served as the probe to capture the target miRNA analyte. The target hybridization duplex was immobilized on the backbone of the nanofibril through the biotin-streptavidin interaction. The quantification was achieved by the fluorescence intensity measured with total internal reflection fluorescence microscopy. A detection limit of 1 pM was achieved with trace amount of sample consumption. This assay showed efficient single-base mismatch discrimination. The applicability of quantifying circulating mir-196a in both normal and cancer patient’s serums was also demonstrated. In chapter 5, a magnetic nanoparticles based sandwich immunosensor with carbazole-based cyanine as the fluorescence labeling dye for the direct quantification of prostate cancer related antigen, PSA, was developed. Taking benefit of the magnetic property of the nanoparticles, the target sandwich immunocomposites can be easily online separated from the sample matrix. The as-developed assay can efficiently discriminate the target PSA from other disease related antigens and achieve a LOD of 400 fM (13 pg/mL) and a LOQ of 2 pM (0.66 ng/mL). As the whole detection assay can be completed in 1 h with only 10 μL of sample, this assay is fast and cost effective and of high potential for early disease and cancer diagnosis, staging and monitoring