This proposal seeks to acquire a two-photon excitation imaging system for the Center for Cell Imaging at Yale University School of Medicine. Specifically, we would upgrade one of the Center's confocal microscopes to have two-photon excitation capabilities. Two photon laser excitation imaging has several properties that make it unique, even relative to point scanning confocal imaging. Two photon imaging permits more highly localized excitation of fluorophores, the use of dyes that excite at lower wavelengths, much deeper sample penetration, less photobleaching, and less cytotoxicity than conventional confocal microscopy. These features improve spatial resolution, especially in thicker tissue preparations such as intact organs, and make it possible for the first time to photolyse caged compounds or photobleach dyes in highly localized, submicron-sized intracellular regions. No two-photon imaging system currently is available at the School of Medicine although a number of investigators here require this technology to advance their research efforts. The range of projects that would make immediate use of this technology include flash photolysis of caged second messengers to examine regulation of Cai2+ signals at the subcellular level, photolysis of caged oligonucleotides to perform in situ PCR at the single cell level, immunochemistry of intact embryos and larvae, examination of acid production in intact gastric glands, examination of intercellular Cai2+ waves in intact, perfused livers, and distribution and trafficking of GFP-labeled proteins in tissues (including brain slices) and in whole brains of transgenic animals. In addition, the Center for Cell Imaging is the only such imaging resource available to all investigators at the School of Medicine. Therefore, acquisition of a two-photon imaging system by the Center would insure that any investigator at our school with a current or future need for this imaging modality would have access to it.
| Rogart, Jason N; Nagata, Jun; Loeser, Caroline S et al. (2008) Multiphoton imaging can be used for microscopic examination of intact human gastrointestinal mucosa ex vivo. Clin Gastroenterol Hepatol 6:95-101 |
