Core A (Imaging) is vital for the MRI imaging of fetuses in utero and teratomas, and ultimately for tracking transplanted stem cells. In vitro real-time and laser-scanning confocal, quantitative FRAP, wide-field and electron microscopy are also provided to all three subprojects to evaluate the dynamics of live cell indicators of pluripotency in hESC, nhpESC and PG cells in order to better define characteristics of the pluripotent state. The imaging core provides exceptional high-sensitivity detection of stem cell dynamics in vitro, in utero and in vivo, essential for each of the research projects. To ensure maximum utility of the Imaging Core, we will combine the strengths of in vivo noninvasive MRI with in vitro imaging by conventional, confocal, and electron microscopy. At the macroscopic whole animal level, we will utilize new small animal imaging technologies, such as MRI. The primary roles of the Imaging Core will be to: perform longitudinal MRI studies of pregnant monkeys and monitor viability and phenotypic differences of the chimeric fetus and placenta throughout pregnancy;employ high-resolution 3D MRI of fixed and live ES-derived NHP embryos to examine stem cell contributions;and monitor teratoma formation in vivo.
Aim 1. The imaging core will provide total body non-invasive functional imaging to all three projects. The core is responsible for tracking cell lineage contributions in ES and EG chimeric embryos and the fates of transplanted hES cells. The core will also use high resolution MRI to map fetal development and teratoma formation, and perform magnetic resonance microscopy on excised teratomas prior to sectioning. This core will also work towards developing and optimizing techniques for the tracking of stem cells in vivo including SPIO labeling of ES and EG cells for MRI.
Aim 2. The imaging core will provide instrumentation, oversight and guidance for fixed, immunocytochemistry, live cell confocal, real-time dynamic and electron microscopy for assaying cell structure and dynamics in pluripotent and differentiated human embryonic stem cells, especially live and fixed pluripotent marker assays to better characterize undifferentiated human and nonhuman primate ES and EG cells. This core will also perform fundamental studies of ES and EG cell contributions to chimeric embryos and imprinting status of ES and EG cells, as well as immunocytochemistry and immunohistochemistry of teratomas and embryoid bodies.
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