The objective of the proposed research is to design an experimental setup to assess the ability of impedance measurements to characterize mouse embryonic stem cell (ESC) and embryoid body (EB) growth and differentiation. Existing quality assurance measurements used to stage the growth and differentiation of embryoid bodies are labor intensive and most often destructive to the cells, thus present methods are typically valid for a single time point. Bioimpedance measurements are non-invasive and non-destructive, presenting an alternative approach to this challenge. These measurements can be done continuously for real-time measurements on the changes in embryoid body growth and differentiation. A system capable of making bioimpedance measurements of ESC and EB suspensions was designed along with a biocompatible test device to hold the cells and Ag-AgCl electrodes. The system uses a lock-in amplifier to record the magnitude and phase changes of the ESC and EB suspensions when a 1 Vpp signal sweeping frequencies from 100 Hz to 100 kHz is applied. The system performance was validated with a test case of 1 mL of 0.1 M KCl. Then experiments with cell culture media, ESCs, and EBs were performed, with varying concentrations of cells and EBs. Experimental results for single ESC suspensions showed promise in detecting a difference in cell concentration between 2 million and 4 million cells in 0.5 mL of media. Results for four day old EBs were ambiguous, and we conclude that a different experimental set up is required due to EB settling during experimentation.

Epigenetic differences between maternally inherited and paternally inherited chromosomes, such as CpG methylation, render the maternal and paternal genome functionally inequivalent, a phenomenon called genomic imprinting. This functional inequivalence is exemplified with imprinted genes, whose expression is parent-of-origin specific. The dosage of imprinted gene expression is disrupted in cells with uniparental disomy (UPD), which is an unequal parental contribution to the genome. I have derived mouse embryonic stem (ES) cell sub-lines with maternal UPD (mUPD) for mouse chromosome 6 (MMU6) to characterize regulation and maintenance of imprinted gene expression. The main finding from this study is that maintenance of imprinting in mitotic UPD is extremely variable. Imprint maintenance was shown to vary from gene to gene, and to vary between ES cell lines depending on the mechanism of loss of heterozygosity (LOH) in that cell line. Certain genes analyzed, such as Peg10, Sgce, Peg1, and Mit1 showed abnormal expression in ES cell lines for which they were mUPD. These abnormal expression levels are similar to that observed in ES cells with meiotically-derived full genome mUPD (parthenogenetic ES cells). Imprinted CpG methylation at the Peg1 promoter was found to be abnormal in all sub-lines with mUPD for Peg1. Two cell sub-lines which incurred LOH through mitotic recombination showed hypermethylation of Peg1, consistent with the presence of two maternal alleles. Surprisingly, a cell sub-line which incurred LOH through full chromosome duplication/loss showed hypomethylation of Peg1. The levels of methylation observed in these sub-lines correlates with expression, as the first two sub-lines showed a near-consistent reduction of Peg1, while the latter showed Peg1 levels close to wild-type. Altogether these results suggest that certain imprinted genes, like Peg1 and Peg10, have stricter imprinting maintenance, and as a result show abnormal expression in UPD. This strict imprint maintenance is disrupted, however, in UPD incurred through full chromosome duplication/loss, possibly because of the trisomic intermediate stage which occurs in this mechanism.

Copies of author's previously published articles inserted. / Bibliography: leaves 117-130. / v, 130, [51] leaves, [33] leaves of plates : ill. (chiefly col.) ; 30 cm. / Title page, contents and abstract only. The complete thesis in print form is available from the University Library. / The general aim of this thesis is to develop methods for the identification of markers for pluripotent cell subpopulations in the developing mouse embryo. A screen for mouse embryonic stem (ES) cell markers is carried out, to identify transcripts that are differentially expressed between ES cells and X cells, and to investigate pluripotent cell heterogeneity during early development. The study demonstrates the potential to identify and characterise molecular heterogeneity within the developing pluripotent cell pool in vivo, via the controlled progression and analysis of pluripotent cells in vitro. / Thesis (Ph.D.)--University of Adelaide, Dept. of Biochemistry, 1997

Diss. (sammanfattning) Stockholm : Karolinska institutet, 2007. / Härtill 4 uppsatser.

Thesis (Ph.D.) -- University of Adelaide, Dept. of Molecular Biosciences (Biochemistry), 2001. / Includes bibliographical references (leaves 142-168).

Diss. (sammanfattning) Stockholm : Karolinska institutet, 2007. / Härtill 4 uppsatser.

Dissertation (Ph.D.)--University of South Florida, 2006. / Includes vita. Includes bibliographical references (leaves 154-187).. Also available online.

Thesis (Ph. D.)--Rutgers University, 2007. / "Graduate Program in Biomedical Engineering." Includes bibliographical references (p. 45-48).

Thesis (M. Phil.)--University of Hong Kong, 2010. / Includes bibliographical references (leaves 95-117). Also available in print.

Thesis (M.S.)--University of Florida, 2005. / Typescript. Title from title page of source document. Document formatted into pages; contains 50 pages. Includes Vita. Includes bibliographical references.