Every cell and tissue biology research program has needs for microscopic visualization of research specimens. While some of these needs are best addressed by traditional, routine light and confocal microscopy of thin, fixed samples, recent decades have seen an explosion of very sophisticated and demanding new microscopy procedures for in vivo imaging (e.g. live cell and deep tissue imaging, simultaneous expression of multiple fluorescent proteins, FRET and FRAP) that are now a required part of contemporary cell and tissue research. To be applied successfully, these methods require a versatile, advanced imaging system that incorporates 2-photon excitation, rapid imaging and environmental control. The demands of such state-of-the-art microscopy methods for both specialized expertise and specialized equipment often outstrip the resources available to individual research laboratories and as such are best met as a shared instrument resource housed in a shared imaging facility. This proposal requests funds to purchase a Prairie Technologies Ultima, 2-photon microscope equipped with the Coherent Chameleon Ultra II ultra-fast pulsed laser, for imaging, with high spatial and temporal resolution, live biological samples. This 2-photon live cell imaging system will be a shared resource, located in a well-established, multi-user, light and electron microscopy facility at Stanford University: the Cell Sciences Imaging Facility (http://taltos.stanford.edu). This facility is accessible to Stanford University's entire research community as well as surrounding biotech companies. The requested 2-photon live cell imaging system will support NIH funded projects from 8 major users. These projects investigate a wide range of NIH supported topics and model systems including, 1) analysis of cellular expression patterns during development (C. elegans, Kim);2) angiogenesis and anti-angiogenic therapy of cancer (mice, zebra fish, Kuo);3) pathogenesis of basal cell carcinoma of the skin (Drosophila, Oro);4) molecular mechanisms of viral replication, assembly and egress from infected cells (mice, human tissue, Arvin);5,6) characterization and modeling of the neuronal mechanisms underlying the development of epilepsy (rat, Huguenard, Prince);7) development and functional specificity of glial cells in the mammalian retina and brain (Barres) and 8) cellular dynamics regulating pancreatic development (mice, Seung Kim). These studies investigate critical functional and structural questions in a variety of model organisms and human tissues and cover areas of biomedical research with implications for diverse aspects of human health and disease, ranging from cancer and viral pathogenesis to understanding the cellular and molecular basis of epilepsy, sensory biology and organ development. All require 2-photon instrumentation for imaging live samples which is most effectively provided by the requested Prairie Technologies'Ultima 2- photon imaging system.
The studies currently being undertaken by the NIH user group researchers, investigate critical functional and structural questions in a variety of model organisms and human tissues and cover areas of biomedical research with implications for diverse aspects of human health and disease, ranging from cancer and viral pathogenesis to understanding the cellular and molecular basis of epilepsy, sensory biology and organ development. All require 2-photon instrumentation for imaging live samples over extended time periods which is most effectively provided by the requested Prairie Technologies Inc., Ultima 2-photon microscope equipped with the Coherent Inc. Chameleon Ultra II ultra-fast pulsed laser.
